PATH 1 ALL Lectures Final1

Cell injury- Part 1
Cellular adaptations
Objectives
• Cellular adaptation- reversible phenomenon
• Hypertrophy- physiological and pathological examples, describe the
morphology & mechanism
• Hyperplasia- physiological and pathological examples, describe the
morphology &mechanism
• Atrophy - physiological and pathological examples, describe the
morphology, mechanism
• Metaplasia - physiological and pathological examples, describe the
morphology, mechanism
• Autophagy
Overview
Normal Reversible
Cell Adaptation cells Injury
Further
Injury Injury
More
Injury
Irreversible
Cell Death Cell Injury
Cellular adaptations
• When cell encounters physiological stress or pathological stimuli, they
undergo adaptations achieving a new steady state and preserve
viability and function.
• The principal adaptive response are-
1. Hypertrophy
2. Hyperplasia
3. Atrophy
4. Metaplasia
Hypertrophy
Definition: increase in size of
cells, resulting in an increase
in size of the organ
- no new cells, but larger cells
- increased production of
cellular proteins
Types:
Physiologic
Pathologic
Hypertrophy
Physiologic hypertrophy:
- Hypertrophy of muscle cells
(body builders)
- Physiologic growth of
uterus in pregnancy
(Remember: both hypertrophy and
hyperplasia can occur together)
Physiologic hypertrophy of the uterus during pregnancy. A,

Gross appearance of a normal uterus (right) and a gravid uterus
(left). B, Small spindle-shaped uterine smooth muscle cells
from a normal uterus (left) compared with large plump cells in
gravid uterus (right).
Hypertrophy
• Pathologic
hypertrophy:
Smooth muscle
hypertrophy
associated with
Prostatic hyperplasia
Urinary bladder: note, thickening of the bladder wall

(hypertrophy), enlarged Prostate (hyperplasia)
Hypertrophy
Pathologic hypertrophy:
- left ventricular hypertrophy
associated with
hypertension (increase in
afterload)
Remember: in myocardial growth

caused by exercise or pregnancy,
cardiac structure and function are
normal and there is no association
with heart failure
9
Hypertrophy
Mechanisms of hypertrophy:
(reference to Heart)
Three steps
1) Actions of mechanical sensors
- these come into play following increased
workload
- Growth Factors: TGF-B, insulin like growth
factor 1 (IGF1), fibroblast growth factor
- vasoactive agents: a-adrenergic agents,
endothelin-1, angiotensin II
2) Signal transduction pathways
- PI3K/AKT pathway: involved with physiologic
type
- G-protein-coupled receptor pathway: important
for cardiac hypertrophy
Hypertrophy
Mechanisms of hypertrophy (cont’d):

Three steps (continued)
3) Transcription pathways
- GATA4, nuclear factor of activated T-cells
(NFAT), myocyte enhancer factor 2 (MEF2)
- increase the synthesis of contractile protein
Hypertrophy
Associated mechanisms
1) switch of contractile proteins from adult to fetal or neonatal forms
- α isoform of myosin heavy chain is replaced by β isoform
β isoform:
- slower, more energetically economical contraction
2) atrial natriuretic factor (ANF)
- gene expression for ANF in both atrium and ventricles in embryonic heart
- soon after birth, these are down regulated
- in hypertension, the gene for ANF is re-expressed
- this helps in salt secretion by kidney, therefore the blood volume decreases
Normal Histology
Increase in cell size, and nuclear size
Hypertrophy 13
Concentric & eccentric cardiac hypertrophy
Concentric
hypertrophy Eccentric hypertrophy
Pressure over load Volume over load.
Caused By: Caused by valve
hypertension regurgitations
Narrow chamber and Dilated chamber/thick
thick wall wall
Hypertrophy (mechanism cont’d)
Regressive changes
- beyond a point these changes become maladaptive
- regressive changes: lysis and loss of myofibrillar contractile elements, myocyte
death may occur
- therefore, remodeling may lead to increased apoptosis, fibrosis, and dilatation of
the failing heart
- cardiac fibrosis contributes to both systolic and diastolic dysfunction, becomes a
setting for electrical disturbances (arrhythmia)
15
Hypertrophic cardiomyopathy (HCM):
In green
vEtiology: 50% of cases

autosomal dominant.
vGenetic: autosomal dominant β-
myosin heavy chain mutation.
Hypertrophic cardiomyopathy (HCM):
- autosomal dominant disease of the heart muscle
- characterized by a small left ventricular cavity and marked hypertrophy of the
myocardium with myofibril disarray
- mutation affecting the encoding of β-myosin heavy chain protein, which is a major
component of the cardiac sarcomere (disease of the sarcomere)
- affected genes include β-myosin heavy chain, myosin-binding protein C, cardiac troponin
T and I, α-tropomyosin, actin, titin, and myosin light chains
Gross:
- demonstrates asymmetric septal hypertrophy with a small left ventricular cavity
- mural endocardium may be thickened by fibrous tissue, and if left ventricular outflow
tract obstruction is present
- epicardial coronary arteries are normal
Micro:
- myocardial disarray consists of short runs of severely hypertrophied fibers
- interrupted by connective tissue
- there are large and bizarre nuclei and fibrosis present
- this disorganization results in a "whorling" of muscle fibers that is characteristic of HCM
18
Hyperplasia
Definition:
- Increase in number of cells in an organ or tissue
- Though, this is morphological remember that increase in size of the organ is
accompanied by increased function (example: endocrine organs)
Types:
Physiologic Hyperplasia
- divided into :
i) Hormonal hyperplasia- proliferation of glandular epithelium of female breast
at puberty, pregnancy
ii) Compensatory hyperplasia- regeneration of liver following partial

hepatectomy
19
Hyperplasia
Pathologic Hyperplasia
- Excessive hormonal stimulation or
growth factors acting on target cells
Examples: Endometrial Hyperplasia

- Due to the action of unopposed estrogen
on endometrium
- Post-menopausal women treated with Endometrium: proliferation of glands,
hormone replacement therapy (HRT) reduced stroma
- This is associated with anovulatory cycles
20
Pathophysiology of PCOD
Genetic, obesity, sedentary life style Dylipidemia, Type 2 DM, CAD
Increase in GnRH pulse Insulin resistance
↑LH: FSH
Anovulation
Theca cells- androstenediones & Testosterone Arrest in antral follicle development

Polycystic ovaries
Hirsutism, acne (string of pearls)
Adipose tissue (aromatase) Estrones & estradiols Endometrial hyperplasia

Hyperplasia
Endometrial Hyperplasia:
- defined as an increased proliferation of the
endometrial glands relative to the stroma, resulting
in an increased gland-to-stroma ratio when
compared with normal proliferative endometrium
- inactivation of the PTEN tumor suppressor gene is a
common genetic alteration in both endometrial
hyperplasias and endometrial carcinomas
- PTEN encodes a lipid phosphatase that is an
important negative regulator of phosphatidylinositol
3-kinase: PI3K/AKT growth-regulatory pathway
- when PTEN function is lost, the PI3K/AKT pathway
becomes overactive
22
Endometrium: normal
(proliferative phase)
Endometrium: Hyperplasia 23
Hyperplasia (examples cont’d)
Examples:
ii) prostatic hyperplasia
- hyperplasia of glandular
and stromal elements of
prostate
- associated with aging
- prostatic levels of
dihydrotestosterone
(DHT) remain high with Prostate: note glandular crowding
aging, even though
peripheral levels of
testosterone decrease
24
Hyperplasia (examples cont’d)
ii) prostatic hyperplasia
- most commonly affects the inner peri-urethral zone of the prostate, producing
nodules that compress the prostatic urethra
- on microscopic examination, the nodules exhibit variable proportions of stroma
and glands
- hyperplastic glands are lined by two cell layers, an inner columnar layer and an
outer layer composed of flattened basal cells
25
Prostate: Normal Histology
Prostate: Benign Prostatic

Hyperplasia (BPH)
26
Prostate: normal histology
27
Hyperplasia
iii) Endocrine glands:

- may be either diffuse or focal
Examples:
- parathyroid hyperplasia (in
ch. renal failure)
- thyroid hyperplasia in
Grave’s disease
- islets of Langerhans seen in
child of Diabetic mother
Grave’s Disease: hyperplasia
28
Thyroid: normal histology
Thyroid Hyperplasia: Grave’s disease 29

Hyperplasia
Mechanism of hyperplasia:
- increased local production of growth factors
- these engage cell receptors
- activation of particular intracellular signaling pathways
- that stimulate cell proliferation of mature cells
- sometimes, increase in new cells from tissue stem cells (example: partial
hepatectomy)
Hyperplasia is different from neoplasia (Cancer)
30
Atrophy
Definition:
- results from decrease in cell size and
number
- adaptive response
31
Atrophy
Mechanisms of Atrophy:
- imbalance between protein synthesis and degradation
- increased catabolism of cell organelles and cytoskeletal
proteins
- achieved through stimulation of ‘ubiquitin-proteasome’
pathway (target specific proteins)
- another mechanism is also known to play a role: Autophagy
Autophagy:
- appearance of autophagic vacuoles in target cells
- considered to be “self eating”
- thereby reducing nutritional requirements
- sometimes cell debris remain within the vacuoles resist
digestion
- these residual bodies are important: Lipofuscin granules
Atrophy
Types:
Physiologic: during
embryogenesis, notochord
and thyroglossal duct
Aging- gonads, vaginal
atrophy, heart-brown
atrophy, thymus, brain,
Brown atrophy
Atrophy
Causes of atrophy (pathologic):
i. Decreased workload: prolonged
immobilization of limb
ii. Loss of innervation: skeletal muscle atrophy
iii. Diminished blood supply (insufficient to kill
cells)
iv. Inadequate nutrition: marasmus ATROPHY
v. Loss of endocrine stimulation:

Hypopituitarism ( Sheehan’s adrenal
cortex-ZF), menopause
vi. Pressure: thick secretions in cystic fibrosis
cause atrophy of pancreas
34
Japanese cell biologist Yoshinori
Autophagy Ohsumi won the Nobel Prize in Medicine
in 2016 for his research on how cells
recycle and renew their content, a
process called autophagy
Definition: a process by which a cell eats
up its own contents
Types
1)chaperone-mediated
2)microautophagy: inward invagination of
lysosomal membrane for delivery)
3)macroautophagy: referred as autophagy,
as it is the major form of autophagy
Mechanisms:
Physiologic
- as a survival mechanism under various
stress condition
3
5
Autophagy
Mechanisms:
Pathologic
-cancer
-neurodegenerative disorders: Alzheimer, Huntington
-infectious diseases: Mycobacteria
-Inflammatory Bowel Disease
3
6
Metaplasia
Definition:
- reversible change in which one
adult cell type is replaced by
another adult cell type
Intestinal metaplasia
37
Metaplasia (types)
i) columnar to squamous
example: squamous metaplasia of bronchial mucosal lining
in chronic smokers
Tracheo-bronchial: Normal histo Metaplasia: squamous 38

Metaplasia (types)
i) columnar to squamous (cont’d)

Squamous cell carcinoma of lung
- commonly found in men and is strongly associated with smoking
- precursors lesions that give rise to invasive squamous cell carcinoma
are well characterized
- squamous cell carcinomas are often antedated by squamous
metaplasia in the bronchial epithelium,
39
Metaplasia (types cont’d)
ii) squamous to columnar:
- in patients with chronic esophageal injury from GERD, Barrett's metaplasia
develops when mucus-secreting columnar cells replace reflux-damaged
esophageal squamous cells
Diagnosis of Barrett’s esophagus:
- endoscopically, the gastroesophageal junction is identified as the most proximal
extent of gastric folds, and the columnar mucosa is salmon-colored and coarse,
in contrast to the pale, glossy esophageal squamous mucosa
- extent of esophageal columnar metaplasia determines whether long-segment
or short-segment Barrett's esophagus (≥3 cm or <3 cm of columnar metaplasia,
respectively) is diagnosed
40
Metaplasia
Barrett’s esophagus:
- both GERD, and Barrett’s esophagus has an
increased risk for adenocarcinoma of distal
portion of esophagus
- central obesity, which might contribute to
Barrett's carcinogenesis by promoting GERD
Endoscopy: shows tongues of reddened mucosa, indicative of Barrett’s

Histology: remember, its intestinal type of epithelium (goblet cells)
- eosinophils are seen
- if neutrophils are seen, it suggests a severe disease
41
Barrett’s esophagus
42
Metaplasia (types cont’d)
ii) squamous to columnar :
GERD
- current definition of the disorder is “a condition which
develops when the reflux of stomach contents causes
troublesome symptoms (i.e., at least two heartburn episodes
per week) and/or complications”
- with respect to the esophagus, the spectrum of injury includes
esophagitis, stricture, and the development of columnar
metaplasia in place of the normal squamous epithelium
(Barrett's esophagus)
43
Esophagus: normal histology
Mucus glands
Inflammation
Barrett’s esophagus 44
Metaplasia
iii) intestinal metaplasia

- seen in pylorus and antrum
- in the presence of Helicobacter
pylori
- characterized histologically by the
presence of goblet cells
Organism is conc
superficial mucous
Warthin Stany silver stain
45
Metaplasia
iii) intestinal metaplasia (cont’d):
- H. pylori infection most often presents as a
predominantly antral gastritis with normal or
increased acid production
- when inflammation remains limited to the antrum,
increased acid production results in greater risk of
duodenal peptic ulcer
- in other patients gastritis may progress to involve the
gastric body and fundus
- this multifocal atrophic gastritis is associated with
patchy mucosal atrophy, reduced parietal cell mass
and acid secretion, intestinal metaplasia, and
increased risk of gastric adenocarcinoma
46
Gastric mucosa: Normal
Gastric mucosa: Intestinal metaplasia 47

Metaplasia
iv) Transitional epithelium to
squamous
- chronic inflammation of
urinary bladder mucosa
due to Schistosoma
hematobium
Urinary bladder: note, squamous metaplasia of the mucosa. Note, presence of

numerous Schistosoma hematobiium in the lamina propria
48
Metaplasia
Schistosoma haematobium
infections in endemic areas
(Egypt, Sudan)
- the ova are deposited in
the bladder wall and incite a
chronic inflammatory
response that induces
progressive mucosal
squamous metaplasia and
dysplasia and, in some
instances, neoplasia
49
Urinary bladder: Normal histology
Urinary bladder: squamous metaplasia 50

Metaplasia
v) Connective tissue metaplasia
- formation of cartilage, bone, or
adipose tissue
- in tissues which normally does
not possess them
Myositis ossificans:
- seen following intramuscular
hemorrhage (hematoma)
- 4 to 7 weeks following injury
- common sites: pectoralis major,
biceps, thigh muscles
Myositis ossificans: note – extra skeletal calcification 51

52
Metaplasia
VI) Vitamin A deficiency:
Keratomalacia:
- vitamin A and retinoids play an
important role in the orderly
differentiation of epithelium
- deficiency of vitamin A leads to
metaplastic change leading to
keratomalacia Keratomalacia: squamous metaplasia
- also, squamous metaplasia of
upper respiratory and Please read foot notes
sometimes even that of
urinary tract are noted
53
Metaplasia
Vitamin A:
- activation of retinoic acid receptors (RARs) by their ligands causes
the release of corepressors and the obligatory formation of
heterodimers with another retinoid receptor, known as the
retinoic X receptor (RXR)
- both RAR and RXR have three isoforms, α, β, and γ
- the RAR/RXR heterodimers bind to retinoic acid response
elements located in the regulatory regions of genes that encode
receptors for growth factors, tumor suppressor genes, and
secreted proteins
- through these effects, retinoids regulate cell growth and
differentiation, cell cycle control, and other biologic responses
- All-trans-retinoic acid, a potent acid derivative of vitamin A, has
the highest affinity for RARs compared with other retinoids
54
Metaplasia
Mechanisms of metaplasia:
- reprogramming of stem
cells
- differentiation of stem cells
along a particular lineage
- brought about by various
cytokines, growth factors
55
Cellular adaptation summary
• Hyperplasia- increase in number of cells-
• Hypertrophy- Increase Size- increased mechanism- growth factors and hormones
protein synthesis or mRNA.
1. Physiological – Breast (puberty & pregnancy)
1. Physiogical eg- Sk muscle (body builders),
2. Pathological-
uterus (Hypertrophy and hyperplasia)
• Endometrial hyperplasia- PCOD- glandular
2. Pathological- eg Urinary bladder (BPH), proliferation (risk carcinoma)
LVH (HTN, myocardial infarction, valvular • BPH- normal aging process- DHT- both
abnormalities) glandular and stomal hyperplasia
• Parathyroid hyperplasia- ch renal failure
• Switch of alpha myosin to beta myosin.
• Graves disease
• Re-expression of ANP receptors. • Islet of Langerhans hyperplasia – gestational
DM
• Metaplasia- one adult cell replaced by other

• Atrophy – decrease in cell size- Ubiquitin adult cell.- reprogramming of stem cells
proteosomal degradation and autophagy. 1. Columnar to squamous- respiratory tract (vit A
1. Physiological- embryonal (thyroglossal duct and deficiency, smoking, pollution)
notochord. Aging- brown atrophy- lipofuchin) 2. Squamous to columnar- Barret's esophagus
2. Pathological- loss of innervation, decreased blood
3. Intestinal metaplasia- stomach- H pylori
supply, decreased nutrition, disuse atrophy,
deceased endocrinal stimulation- sheehans infection
syndrome, cystic fibrosis- pancreatic acinar cells 4. Transitional to squamous- schistostoma
hematobium – urinary bladder
5. Connective tissue metaplasia- myositis
ossificans
• Hypoxic Cell injury: Important causes
• Hypoxic Cell injury: reversible injury- Morphology - all tissue vs. Liver/Heart
• Reversible cell injury (hypoxic): Clinical causes, morphology of the cell (all except liver and Heart), and biochemical alteration of the cells.
• Irreversible cell injury: Morphology of the cytoplasm and nucleus, biochemical alteration.
• explain the Mechanisms of cell injury; mitochondrial damage, influx of calcium ions, accumulation of free radicals, defects in membrane permeability, and damage to DNA and proteins
• Free radical injury: generation of oxygen derived free radical and how they are removed. Example of free radical injury
• Deactivation of free radical after radiation injury
• Cytoskeleton damage: Learn some examples
• mechanism of Cyanide poisoning, CO poisoning. Acetaminophen overdose, Carbon tetrachloride exposure. Role of free radicals in Iron overload states, Ischemia-Reperfusion injury, Retinopathy of
prematurity.
• understand Hypoxia, and Ischemia. They should be able to give clinical examples for Hypoxia, and Ischemia and should know the difference between these terms.
• - Radiology, gross, and microscopic findings for the clinical examples must be explained. Myocardial infarction, Ischemic bowel disease, Iron deficiency anemia, Methemoglobinemia, CO poisoning,
TAO, etc. Students should be able to understand ‘watershed’ areas and their vulnerability to ischemia/hypoxia.
Cell Injury- Reversible and
Irreversible Injury
Overall view
Normal Reversible
Cell Adaptation cells Injury
Further
Injury Injury
More
Injury
Irreversible
Cell Death
Cell Injury
Causes of cell injury
Anemia, CO, cyanide,
methamoglobinemia,
cardiorespiratory failure Genetic
derangements
Hypoxia Poisons-
Acetaminophen(
Chemical tylenol), CCl4
Thrombosis, embolism, agents
venous occlusion
Ischemia Immunological
reactions –
autoimmune
conditions
Physical
agents
Nutritional Infectious
Mechanical trauma, agents
imbalance
extremes of temperature
, change in atmospheric PEM, anorexia
pressure, radiations, nervosa, type 2 DM,
electrical shock obesity,
atherosclerosis
Ischemia Vs Hypoxia
Ischemic and Hypoxic cell injury
What is ischemia?
- ischemia results from hypoxia induced by reduced blood flow:
most commonly by mechanical obstruction (example:
thrombotic occlusion)
- ischemia can also be due to reduced venous drainage
How is ischemia different from hypoxia?
- in hypoxia energy production continues in the form of
anaerobic glycolysis
- in ischemia, there is no aerobic or anaerobic glycolysis.
Obstruction leads to complete lack of oxygen, and substrates
5
Examples of Hypoxia
Ischemic and Hypoxic cell injury
Examples of Hypoxia
1) Anemia: low Hb leads to less oxygen
delivery
2) CO poisoning: CO avidly combines
with Hb to form carboxyhemoglobin.
3) Methemoglobinemia: iron is in
oxidized state (Fe3). Normally
methemogloblin is less than 1%.
Causes: congenital(NADH reductase
deficiency) , drugs (sulfa containing,
Nitrates). Treatment: Methylene blue
Methemoglobinemia:
(intravenous) chocolate color
6
Examples of Ischemic cell injury
Examples of Ischemic cell injury
1) Thrombotic, and embolic occlusion
2) Venous (testicular torsion, Budd-Chiari
syndrome)
3) Peripheral artery disease (atherosclerosis
associated with diabetes, hypertension,
cigarette smoking-)
Thrombosis/Embolism
- thrombi can obstruct arteries or veins
- atherosclerosis is the major cause of arterial
thromboses
- atherosclerosis is related to endothelial injury
- eventual obstruction is from thrombus
formation
- thrombosis can injure tissue by local vascular
occlusion or distant embolization
7
examples of ischemic diseases:
Buerger disease (Thromboangiitis Obliterans

TAO)
- is a non-atherosclerotic segmental
inflammatory disease that most commonly
affects the small and medium-sized arteries,
and veins of the arms and legs
- use of or exposure to tobacco is central to the
initiation and progression of the disease
- extremely high prevalence of thromboangiitis
obliterans in India among people of low
socioeconomic class who smoke bidis
(homemade cigarettes with raw tobacco)
- inflammatory thrombi affect both the arteries
and the veins
- occlusive, highly cellular, inflammatory
thrombus, with less inflammation in the walls of
the blood vessels
Note: ischemic ulcers on toes. Arrows
show thrombophlebitis
8
Tissues susceptible to hypoxia:
Tissues susceptible to hypoxia:

i) Watershed areas shared between two
blood vessels
- example: area shared between anterior
cerebral and middle cerebral arteries
(global hypoxia)
- area shared between superior and
inferior mesenteric arteries
ii) Subendocardial tissue (left ventricle in

MI)
iii) Renal cortex and medulla

- straight portion of proximal tubule in
cortex
iv) Purkinje cells in cerebellum 9

Early stage- reversible cell injury
• In early stages or mild forms of injury the
functional and morphological changes are
reversible if the damaging stimulus is
removed.
.
Mechanism of reversible cell
injury
-(hypoxia, metabolic injury best

seen in liver)
Cell swelling following loss of volume regulation (↓ ATP)
-when many cells are affected, it produces pallor, increased
turgor, and increased weight of the organ
-on microscopy: small clear vacuoles in cytoplasm
-fatty change (hypoxia, metabolic injury best seen in liver)
Microscopic feature of reversible cell injury
Ultrastructural changes
1. Plasma membrane
alterations: blebbing, blunting,
swelling of RER, and loss of
microvilli
2. Mitochondrial changes:
swelling
3. Dilation of ER:
4. Nuclear alterations:
Renal tubules: normal
Interstitial odema
Sloughing of tubular cells
Cellular swelling-
increase in nuclear
size
Renal tubules: cell swelling 14

Irreversible cell injury
Irreversible cell injury:

- with continuing injury , injury becomes irreversible: cell death
- critical biochemical event is not clear
- there are two types of cell death: necrosis, and apoptosis
Necrosis: results from damage to cell membranes, and loss of
homeostasis. Leakage of lysosomal enzymes, and leakage of
cellular contents
Apoptosis: cell suicide. There is nuclear dissolution, without
complete loss of cell membrane integrity. Cellular content do not
leak out. Apoptotic cells are cleared by macrophages.
15
2 important phenomenon of
irreversibility
1. Inability to reverse the
mitochondrial
dysfunction
2. Profound disturbance in
membrane function
amorphous densities in mitochondria –

calcium and proteins
Best seen in ischemic heart
Mitochondria with amorphous densities
(electron microscope) 16
Mechanism of irreversible cell
injury
1. Influx of Calcium
2. Mitochondrial damage
3. Accumulation of oxygen derived free radicals
(oxidative stress)
4. Defect in membrane damage
5. Damage to DNA and Proteins
1) Influx of calcium
- cytosolic calcium is kept

very low, compared to
extracellular
- (~0.1 µmol) compared
with extracellular levels of
1.3 mmol),
- ischemia and certain
toxins, causes increase in
cytosolic calcium
Influx of intracellular calcium and loss of
calcium homeostasis
• Release of calcium from mitochondria and ER
• Increases in calcium activates a number of
enzymes:
1. ATPases: depletes ATP
2. Phospholipases: damages cell membrane
3. Proteases: cell membrane damage/ cytoskeletal
proteins damage
4. Endonucleases: fragmentation of DNA
5. Caspases: leading to apoptosis
19
Endonuclease activation
Normal Pyknosis Karyorrhexis Karyolysis
20
Mitochondrial damage
2) Mitochondrial damage
Ø Mitochondria can be damaged by: increases in

cytosolic calcium, oxidative stress, activation of
phosholipase A
1. Formation of high conductance (mitochondrial
permeability transition) channel . One of the structural
component: cyclophilin D is targeted by
immunosuppressive drug (cyclosporine)
2. If stimuli persists, these pores become permanent
3. leakage of cytochrome c- Promotes apoptosis thru
capsases.
4. Formation of reactive oxygen species
22
3) Accumulation of oxygen free radicals
Examples of free radical injury

- chemical and radiation injury
- liver necrosis by drugs-
acetaminophen , CCl4
- ischemia-reperfusion injury
- retinopathy of prematurity
- iron overload
- cellular aging
- microbial killing by phagocytes-
inflammation
Properties of principle free
radicals
How are free radicals removed?
“Radical scavenging system”
- Antioxidants: vitamins A, C, E
- Trace elements binding proteins: Iron and
Copper binding proteins (ferritin and
ceruloplasmin)
- Enzymes: catalase, superoxide
dismutases, glutathione peroxidase
25
26
Ischemia-Reperfusion
injury:
- Exacerbation of cell injury resulting from
reperfusion into ischemic tissue( Heart:
myocardial infarction) is called reperfusion
injury. Thrombolytic
therapy
27
Ischemia-Reperfusion injury
- Restoring blood flow to ischemic tissue can help recovery but,
at the same time may prove injurious if blood flow is not
restored within a ‘window period’
- for example: restoration of blood flow to ischemic
myocardium is optimum within the first 3 hours
(approximation)
- following this ‘window period’ cells may sustain injury,
thereby extending the area of injury
- causes for this problem: 1) Oxidative stress, and release of
ROS (re-oxygenation leads to formation of ROS, due to
incomplete reduction of oxygen by injured mitochondria- by
parenchymal and infiltrative leukocytes ). 2) intracellular
calcium overload. 3) Inflammatory cytokines
28
Chemical Injury
Chemical Injury
Conversion to toxic metabolites:

- formation of reactive toxic metabolites by P-450 system
- thereby generating free radicals (ROS)
- examples: Acetaminophen (Tylenol) poisoning, and carbon
tetra chloride poisoning.
- both of these produce: Massive hepatic necrosis
29
Massive Hepatic Necrosis: Acetaminophen or CCl4 poisoning
Hemorrhage
Necrosis
Architecture is
preserved,
Inflammation
hepatocytes shows
absence of nuclei
30
4) Membrane damage
-Ischemia: ATP depletion, calcium mediated activation of
phospholipases
-Bacterial toxins, viral toxins, lytic components: direct damage of
membrane
-Physical, and chemical agents: mostly ROS
Clinical correlates
• The loss of membrane integrity allows
intracellular enzymes to leak out , which
can be measured in blood.
• Detection of these proteins in the
circulation serves as a clinical marker of
cell death and organ injury.
Clinically important examples
• Myocardial injury
1. Creatine phosphokinase-MB isoenzyme
2. Lactate dehydrogenase (LDH)
3. Troponin
• Hepatitis- transaminase (AST ad ALT)
• Pancreatitis- amylase and lipase
• Biliary tract obstruction- alkaline
phosphatase
Morphological evidence of
cytoskeletal damage
Mallory bodies in alcoholic
hepatitis-Ultrastucturally Mallory
bodies are composed of
aggregates of intermediate
(cytokeratin) filaments
Neurofibrillary tangles in
Alzheimer disease-
Microscopically there are
extracellular neuritic plaques
containing Aβ amyloid, silver
staining neurofibrillary
tangles in neuronal cytoplasm
35
Morphological evidence of cytoskeletal damage
Clinical correlation
3) damage to microtubules
- Kartagener’s syndrome (primary
ciliary dyskinesia)
- disorganization of microtubules
- leading to immobilization of cilia
in respiratory tract and, inhibition
of sperm motility
36
Chest X-ray:Situs inversus: is a term
used to describe the inverted position of
chest and abdominal organs.
It is called situs inversus totalis when

there is a total transposition of abdominal
and thoracic viscera (mirror image of
internal organs normal positioning).
Situs inversus is usually associated with
dextrocardia
3-5% incidence of congenital heart dis,
most commonly transposition of great
vessels. Of these patients, 80% have a
(R) sided aortic arch.
Up to 20% of patients with situs inversus

can have Kartagener’s Synd.
37
Damage to DNA and Proteins
5) Damage to DNA and Proteins
- cells have mechanisms that repair
damage to DNA, but if DNA
damage is too severe to be
corrected
- then the cell initiates a suicide
program that results in death by
apoptosis
- causes: exposure to drugs,
radiation Liver: apoptosis (arrow)
38
- Myelin figures- Whorl like structure –
originating from damaged cell membrane
Biochemical ↑Ca in the cell, ↑free radical, lipid
peroxidation – Membrane damage
Light Cytoplam: eosinophilic

microscopy Nucleus: pyknosis, Karyolysis, Karyorrhesis
Electron Myelin figures, amorphous densities in

microscopy mitochondria –calcium and proteins (Electron
microscopic finding)
Markers Hepatitis- ↑AST, ↑ALT
MI- ↑CK-MB, LDH, Troponin
Acute pancreatitis- ↑lipase &amylase
41
Hypoxic injury becomes
irreversible
• The vulnerability of cell to hypoxia varies
with the tissue or cell type.
1. 3-5 min for neurons
2. 20-40 min for myocardial cells and 1-
2hrs hepatocytes
3. Many hrs for skeletal muscle.
CELL DEATH: NECROSIS OR
APOPTOSIS
• Persistant or excessive injury causes the
cell to pass the “ point of no return” into
irreversible injury and cell death.
• 2 types of cell death
1. Necrosis
2. Apoptosis
CELL DEATH NECROSIS
• Necrosis is the major pathway of cell
death in commonly encountered injuries.
• Membrane damage è lysosomes leaks
out èenter cytoplasmè digest the cell.
• èCellular content leaks out sideè elicits
inflammation.
• Death of a tissue (many cells)

NECROSIS
Morphology:
• Cytoplasm: more eosinophilic (loss of
cytoplasmic mRNA and protein denaturation),
appear glassy, homogenous appearance.
• Nuclear changes; karyolysis ( loss of
basophilia, pyknosis, karyorrhexis).
• On electron microscopy, amorphous densities in
mitochondria (only seen in ischemic cardiac
myocytes
• Always pathologic, associated with
inflammation 5
Types of necrosis
1. Coagulative necrosis
2. Liquefactive necrosis
3. Caseous necrosis
4. Gangrenous necrosis
5. Fibrinoid necrosis
6. Fat necrosis
Coagulative necrosis
• Cause: sudden loss of blood supply to an
organ( Ischemia).
– Preservation of tissue/cell architecture
– Protein denaturation and loss of cytoplasmic RNA.
• Nuclear changes: pyknosis, karyorrhesis,
karyolysis
– Karyolysis: loss of nuclear basophilia.
Coagulative Necrosis- Micro
After coagulative
NORMAL RISSUE / Necrosis
CELLS
(Cell outlines are
visible, tissue
structure maintained
)
Typical Coagulative Necrosis : Kidney
1. wedge shaped, 2. well demarcated, 3. pale
Case: elderly, history of MI

or atherosclerosis, with
sudden flank pain,
Hematuria!
Angiogram- thrombus in
renal artery
localized area of ischemia: infarct

Coagulative necrosis: Kidney (Wedge
shaped)
Cell outline is preserved

(Tombstone)
1
0
Pale infarct
Occlusive thrombi
Atherosclerosis (Atheroma)
coronary occlusion by thrombi - pale area of infarct

- coagulative necrosis
Tombstone (Ghost cells)
1
1
Fate of coagulative necrosis : fibrosis/scar of the area
after 7 weeks.
Locate the scar!

Liquefactive Necrosis
Caused by
-focal bacterial, or fungal (abscess)
-also seen in CNS ischemia
Mechanism
-release of proteases by activated leukocytes directly damages cells.
The enzymatic action leads to digestion of dead cells (pus) - Heterolysis
- in CNS ischemia: not known. release of proteolytic enzymes-lysozyme from
brain cells (brain infarct)-Autolysis
Morphology
-localized area: abscess
-in brain ischemia: dissolution of neural tissue without inflammation
Clinical
-lung abscess
-in CNS ischemia: infarct area show dissolution, and later cystic space
1
3
Brain infarct- wedge shape Fate: cavity
pale, soft, and swollen gelatinous formation
Cavity formation by
macrophage after 1-3 months
Liquefactive Necrosis : Lung abscess - GROSS
• Aspiration of infective material (the most
.
frequent cause): acute alcoholism, coma,
anesthetized.
• Antecedent primary lung infection: Post-
pneumonic abscess formations are usually
associated with S. aureus, K. pneumoniae.
• Septic embolism: vegetations of infective
bacterial endocarditis on the right
• side of the heart
PUS = Necrotic
parenchymal cells and
Neutrophils.
Walling- off the abscess

X- Ray of lung Abscess- air fluid level
Caseous Necrosis
Caused by
-Tuberculosis
Mechanism
-it is an unique form of coagulative
necrosis. Activated T-lymphocytes
destroy macrophages containing
bacilli. The bacilli is within the caseous
necrotic material, however, cannot
divide as it is anoxic.
Morphology
-caseous necrosis: “cheese-like”
-on microscopy: caseating granuloma
(discussed in Ch. inflammation)
Clinical
-caseous material will calcify (calcified
hilar LN in x-ray)
-Identification of caseous necrosis is
diagnostic for Tuberculosis 1
7
Cottage cheese like
Caseous necrosis: TB
Calcified hilar LN
1
8
Granulomas with caseation
Amorphous
eosinophilic
appearance
Fat Necrosis
Caused by
-most common cause is 'binging’ alcohol leads to acute
pancreatitis
-Acute Pancreatitis (Enzymatic fat necrosis)

Mechanism
-alcohol injures pancreatic acinar cells. Injured acinar cells
release activated enzymes (amylase, lipase)
-fat destruction, following release of activated pancreatic lipases
into substance of pancreas and peritoneal cavity
-activated lipases split triglyceride esters. Released fatty acids
combine with calcium to form chalky white areas (fat
saponification)
2
0
Fat Necrosis
Morphology
-on histology, fat necrosis is identified as ‘shadowy’ cells, and
surrounding inflammation. Calcification may be present.
Shadowy necrotic fat cells

Fat necrosis (by Lipase) in omentum
Dystrophic
2
Calcification 1
Acute Pancreatitis: note calcification along the pancreas.
2
2
Clinical- acute pancreatitis
-Presents as acute abdomen (medical

emergency). Cullen, and Grey Turner
signs (see image)
-may be fatal due to shock (acute
hemorrhagic pancreatitis)
-calcification in plain x-ray of Cullen’s sign
abdomen (not seen in all cases)
-labs: elevated serum amylase, and
lipase
Traumatic fat necrosis: trauma to
breast fat (female). Seat-belt
restrainers in automobile accident.
On mammogram, mimics breast
cancer
Grey Turner’s sign

Fibrinoid Necrosis
Caused by
1) Immune vasculitis- Poly arthritis nodosa (vasculitis)/SLE
2) Malignant hypertension- BP= 200/120 mm of Hg
Mechanism
Immune vasculitis: Ag-Ab complex deposits on blood vessel
wall- leads to endothelial injury, inflammation, and formation
of fibrin- damage to endothelium: vascular leakage permits
cells and fibrin to leak out- imparts a bright pink color to the
involved areas.
2
4
Mechanism of Immune vasculitis
2
5
Malignant hypertensionBP=
200/120 mm of Hg
Kidney change = Gross of kidney: pin point
cortical hemorrhage on kidney-
aka- Malignant flea bitten appearance.
Nephrosclerosis Micro:
Hyperplastic arteriolosclerosis
(onion-skin):
Fibrinoid necrosis ( necrotizing
arteriolitis)
Indications of malignant transformation are:
Renal failure with hematuria
Retinal hemorrhages and exudates, with or
without papilledema.
Palpitation, blurring of vision, headache. 27
Two features of malignant hypertension
Hyper plastic arteriolosclerosis Fibrinoid necrosis :

(onion-skin or deposits of eosinophilic
concentric laminated fibrilary material in the
thickening of the walls) Intima
referred to as
necrotizing arteriolitis
Fibro muscular
Hyperplasia, duplication
of elastic tissue and
smooth muscles. 28
Gangrenous Necrosis
Caused by
-slowly developing occlusion of blood vessel
-typically seen with diabetic foot
Mechanism
-form of coagulative necrosis developing over a long period of time
(atherosclerotic narrowing)
Morphology- 2 forms:
1. Dry gangrene
2. Wet gangrene
2
9
Dry gangrene
Dry gangrene: peripheral arterial
disease in diabetic patient. Slowly
developing, leads to sluggish venous
drainage.
-ultimately produces: line of
demarcation, and blackening of tissue
(accumulation of iron sulphide)
-red cells lyse to release Hb which is
converted by bacteria to iron sulphide
-remember, bacteria does not survive
and therefore bacteria is not a major
problem Dry Gangrene: note “line of
demarcation”
Wet Gangrene
Morphology
Wet gangrene: is a combination of
coagulative necrosis and liquefactive
necrosis
Examples of wet gangrene:
1)Diabetic foot: small abrasions not felt by
the patient allows access for
microorganisms
2)Inguinal hernia: incarceration of small Wet Gangrene: note creamy yellow pus
bowel loop within inguinal canal
3)volvulus: twisting of a segment of
intestine
4)mesenteric ischemia: diabetic patients,
bowel ischemia may arise with
atherosclerotic narrowing of mesenteric
vessels
5)intussusception: most seen in pediatric
age group. Telescoping of a segment of
bowel due to obstruction.
3
1
• A 40-year-old woman has the sudden onset of severe abdominal
pain. On physical examination she has diffuse tenderness in all
abdominal quadrants, with marked guarding and muscular rigidity.
She has laboratory findings that include serum AST of 43 U/L, ALT of
30 U/L, LDH 630 U/L, and lipase 415 U/L (elevated). An abdominal
CT scan reveals peritoneal fluid collections and along with
enlargement of the pancreas. Which of the following cellular
changes is most likely to accompany these findings?
• A Coagulative necrosis
• B Dry gangrene
• C Fat necrosis
• D Apoptosis
• E Liquefactive necrosis C
CELL DEATH: APOPTOSIS
• Apoptosis: an active, energy dependant,
programmed cell death.
• Single cells death.

What is apoptosis ?
• Definition : Individual programmed Cell
death.
Cells in a DEATH OF ONE

tissue. CELL
Morphological appearance
• Cells shrinks in size
and dense eosinophilic
cytoplasm-acidophilic
body.
• Nuclear chromatic
condensation –
fragmentation of
nucleus- karyorrhexis.
• Cytoplasmic membrane
blebs – breakdown into
fragments(apoptotic
bodies).
Causes of apoptosis
Physiologic situations:
Syndactyly
- programmed cell destruction during
embryogenesis (loss of müllerian
structures in male fetus)
- hormone-dependent involution in adults:
endometrial cell breakdown in menstrual
cycle
- cell deletion in proliferating cell population
(intestinal crypt epithelia)
37
Apoptosis in pathologic situations
- Cell death following injury: radiation,
cytotoxic anticancer drugs
- Viral induced cell injury: viral B
hepatitis(councilman body)
38
Apoptosis in pathologic situations
- Accumulation of misfolded proteins:
degenerative diseases of brain
- Pathologic atrophy in parenchymal organs

following duct obstruction: pancreas, parotid
gland, kidney
- Cell death in tumors (cytotoxic CD 8 T
lymphocytes)
Apoptosis
Mechanisms of Apoptosis
- two phases: i) initiation phase
ii) execution phase
- Induced by distinct stimuli – converge to
activate caspases
Initiation phase:
- two pathways are involved
40
Mechanism of apoptosis
Mitochondrial pathway
Mechanism of Apoptosis
The intrinsic (mitochondrial) pathway:

- results from increased mitochondrial
permeability and release of pro-apoptotic
molecules
- growth factors stimulate production of anti-
apoptotic members of Bcl-2 family
43
• when cells lose stimuli or subjected to stress,
Bcl-2 is lost from mitochondrial membrane
• now, replaced by pro-apoptotic factors: Bak,
Bax and Bim
• leads to increased mitochondrial permeability.
• Cytochrome c leaks out into the cytoplasm of
mitochondria.
- cytochrome c binds to Apaf-1(apoptotic protease
activating factor-1)
- activation of caspase 9, leading to apoptosis
•
44
- Cytochrome c leaks out into the cytoplasm
of mitochondria.
- cytochrome c binds to Apaf-1(apoptotic
protease activating factor-1)
- activation of caspase 9, leading to apoptosis
45
Role of TP53(p53) gene
Role of p53: activation of p53 by DNA

damage initiates apoptotic signaling thru’ Apoptosis occur if the
mitochondria
- p53 increases production of pro apoptotic
DNA damage is
mitochondrial proteins (Bax) beyond Repair.
Extrinsic pathway
Extrinsic (death-receptor) pathway
- ‘death receptors’ are present on surface of
many cells
- ‘death receptors’ are members of Tumor
Necrosis Factor (TNF)
- ‘death receptors’: TNFR1 and Fas
49
- Fas and TNFR1 receptors are integral
membrane proteins
- When these death receptors are bound,
there is activation of caspase 8, leading to
apoptosis
- This pathway can be blocked by FLIP
(some viruses produce FLIP)
Cytomegalovirus, Herpes virus- kaposi
Sarcoma
50
The execution phase:
- Final phase of apoptosis is mediated by a
proteolytic cascade
- Caspases is divided into 2 groups:
- Initiator caspases (caspase-8, caspase-9)
- Executioners (caspase-3, caspase-6)
51
Removal of dead cells
Mechanism of removal of dead
cells
1. In healthy cells, phosphatidylserine is
present on the inner leaflet of the plasma
membrane, but in apoptotic cells this
phospholipid “flips” out and is expressed
on the outer layer of the membrane,
where it is recognized by several
macrophage receptors.
Mechanism of removal of dead
cells
2. Apoptotic bodies are coated by
thrombospondin, an adhesive glycoprotein
that is recognized by phagocytes.
Electrophoresis
5
7
Necroptosis
• Necroptosis:
• This form of cell death is a hybrid that shares aspects
of both necrosis and apoptosis. The following
features characterize necroptosis:
• 1) Morphologically, and to some extent biochemically,
it resembles necrosis, both characterized by loss of
ATP, swelling of the cell and organelles, generation of
ROS, release of lysosomal enzymes and ultimately
rupture of the plasma membrane
• 2) It is triggered by genetically programmed signal
transduction events that culminate in cell death. In
this respect it resembles programmed cell death,
which is considered the hallmark of apoptosis. In
sharp contrast to apoptosis, the genetic program that
drives necroptosis does not result in caspase
activation and hence it is also sometimes referred to
as “caspase-independent” programmed cell death.
Intracellular Accumulations
1
Dr. T. Krishna MD, www.mletips.com
Feature of alteration in cellular metabolism
Main categories
1. Normal substances but accumulated in
excess –Lipids, Carbohydrates, Proteins, and Water
2. Abnormal substances even in small
quantities
ØExogenous- Mineral – carbon, Infectious
agents
ØEndogenous - a product of abnormal
synthesis, abnormal metabolism
2
Feature of alteration in cellular metabolism
Main categories
1. Normal substances but accumulated in
excess
• Lipids - Triglycerides, Cholesterol
• Carbohydrates –Glucose/Glycogen, Glycoproteins,
Glycolipids
• Proteins
• Water
3
Main categories
2. Abnormal substances even in small quantities
– Exogenous
• Mineral – carbon
• Infectious agents
– Endogenous - a product of
• Abnormal synthesis
• Abnormal metabolism
4
Accumulations can be
• Transient/ Permanent
• Harmless/ Toxic
• Intracytoplasmic (Phoagolysosomes) / Intranuclear
• Intrinsic (produced by cell) / Extrinsic (produced
elsewhere)
5
Dr.T.Krishna MD, www.mletips.com
Hyaline change
• Can be intra or extracellular
• Homogenous, pink, glassy, acelluar material in H&E
stained sections
• Non-specific marker of disease/injury
• Examples
– Intracellular – Mallory bodies, Russell bodies
– Extracellular – Hyaline arteriolosclerosis of benign
HTN, DM, elderly people

Hyaline change
Mallory bodies
Hyaline arteriolosclerosis

Major Types (based
on mechanisms)
1. The normal endogenous
substance in excess
2. Abnormal Endogenous
substance
3. Normal Endogenous
substance due to defects
in enzyme metabolism
4. Abnormal Exogenous
substances
8
1. The normal
endogenous
substance in excess
• Inadequate rate of
metabolism
(↑production or ↓rate of
removal)
• Examples
– Liver - Fatty change
(in alcoholics)
– Kidney tubules –
protein droplets (in
proteinuria) 9
Normal Liver
Dr.T.Krishna MD, www.mletips.com webpath

Fatty Liver

Kidney tubules – protein droplets (in proteinuria)
Normal Tubule Tubule with Protein droplets

2. Abnormal Endogenous
substance
• Genetic mutations lead to
à defects in folding,
transport, or degradation of
abnormal protein
– Examples
• Liver – α 1AT defects in protein folding,
deficiency transport
• CNS – storage
disorders
13
α1-antitrypsin (A1A) deficiency
Normal Histology
Dr. T. Krishna MD, www.mletips.com PAS +ve & Diastase Resistant

Fatty Liver

3. Normal Endogenous
substance due to defects
in enzyme metabolism
• Accumulate in Lysosomes
• Examples –
Glycogen/Lipid storage
disorders
Lack of Enzyme
16
Tay-Sachs Disease (GM2 Gangliosidosis)
H& E (ballooning of Neuron)
Dr. T. Krishna MD, www.mletips.com Electron Microscopy

4. Abnormal Exogenous
substances
• Substances can't be
degraded or metabolized
• Examples
– Carbon – Coal worker’s
Pneumoconiosis
– Silica – Silicosis
Ingestion of Indigestible
materials
18
Normal Histology of lung
Dr. T. Krishna MD, www.mletips.com Coal worker’s Pneumoconiosis

Clinical Course
• Reversible
– if an overload is stopped/controlled
• Irreversible
– if it is due to an inherited disorder (lead to
tissue damage and death)
20
Types
– Triglycerides (TGLs)
– Cholesterol & its Esters
– Phospholipids – Myeline figures (of
necrotic cells)
Disorders
– Lysosomal storage disorders – deposits
of complex lipids & carbohydrates
21
Steatosis ( AKA Fatty change)
– Liver – (MC organ involved) Why?***
– Heart
– Muscle
– Kidney
***Major organ involved in lipid

metabolism -Liver
22
Mechanism of Hepatic Lipid metabolism
Adipose tissue /Dietary fats
1
2
1
2
Mechanism of Hepatic Lipid metabolism
• Sources of Hepatic Free Fatty Acids(FFA)
1. Adipose tissue /Dietary fats
2. Acetates of Liver
• Hepatic FFA metabolized into
1. Triglycerides (TGL) by esterification
• TGL bind with Apoprotein àLipoproteins
(LP)
• LP- export to blood
2. Covert to Cholesterol & Phospholipids (PL)
3. Ketone bodies by oxidation
(Ketoacidosis in Type1 DM – life-threatening)
24
Causes of Hepatic Steatosis
Starvation,
DM, Obesity Anoxia/Hypoxia
CCl4 , PM
DM,
Obesity

Causes
• Toxins-
1. Alcohol (MCC of in developed
countries)
1. Hepatotoxin
2. acts at mitochondrial and microsomal level
3. leads to ↑synthesis or ↓breakdown of lipids
2. Carbon Tetrachloride (CCl4)à
Ø ↓Apoprotein synthesis
• Protein Malnutrition (PM) à
Ø ↓Apoprotein synthesis
26
Causes
• DM (MCC in developed countries) à
– ? Mechanism
• Obesity (MCC in developed countries)-
– ? Mechanism
• Anoxia/Hypoxia (due to anemia) à
– ↓FFA oxidation
• Starvation à
– ↑mobilization from peripheral adipose tissue
27
Morphology -Normal Liver
Reddish-brown, rubbery
1. Lobular
Architecture,
2. Trabecular
pattern
3. Zonation (1,2,3)

Morphology - Fatty Liver
1. uniformly
enlarged (2 to 4
times of normal),
2. pale yellow
3. Greasy
perinuclear cytoplasmic
vacuoles (Liver looks
like adipose tissue) –
can be
1. Microvesicular
2. Macrovesicular

Fatty change

Morphology of Hepatic Steatosis
1. Gross – (Macroscopic)
2. Microscopic - (HPE- Histo Pathologic
Examination) with routine H & E stain
and others
3. Ultra – structural - (EM- Electron
Microscopy)
31
1. Gross – (Macroscopic)
• Organ damage depends on the severity of accumulation
1. Mild – no significant change
2. Moderate - ↑the size of the liver; yellowish
discoloration
• what is the normal color of the Liver?
• Reddish - brown
3. Severe / Extreme - ↑↑↑ size (2 to 4 times of normal;
• what is the normal weight of liver?); soft; greasy
• 1500gms
32
Gross – (Macroscopy) of Fatty Liver
Normal
Fatty change

2. Microscopic - (HPE- Histo Pathologic
Examination)
• How fatty change takes place?
1. Starts with the formation of small, membrane-
bound structures (LIPOSOSOMES) close to
ER – Endoplasmic Reticulum); appear as
perinuclear cytoplasmic vacuoles -
Microvesicular
2. Macrovesicular àPush nucleus to the
periphery (eccentric nuclei)
3. Adjacent cells rupture and produce Fatty
cysts
34
Microscopy of Fatty Liver
Normal
Fatty change Stain for Fat

3. Morphology{ Ultra – structural - (EM)} of Fatty Liver
Normal CCl4 Poisoning
36
• TIGERED effect à moderate, prolonged Hypoxia (from
Anemia) à Accumulation of Cholesterol
Alternating bands of
1. Light (yellow– affected myocardium)
2. Dark (normal myocardium)
37
Cholesterol & its Esters
• Cholesterol is mainly used for cell membrane
synthesis, but not produced in excess
• Under normal circumstances, cells don’t accumulate
cholesterol
• Accumulation of Cholesterol and its Esters – always
pathologic
ØDisorders
1. Atherosclerosis (AS)
2. Xanthomas
3. Cholesterolosis
4. Niemann –Pick Disease Type “C”
38
1. Coronary Artery – Atherosclerosis (Gross)
39
Coronary Artery Normal
lumen
40
1. Coronary Artery Atherosclerosis
lumen
41
Atherosclerosis Coronary Artery with complication
Cholesterol clefts
Plaque
Thrombus
42
Dr. T. Krishna MD, www.mletips.com webpath

1. Atherosclerosis (AS)
• Plaque
• The most characteristic lesion of Atherosclerosis
• Plaque is an intimal extension narrows lumen
• Plaque contains Foam cells that engulf cholesterol &
its esters (LDL, VLDL)
• Foam cells –
– 1. Macrophages of Blood Monocytes enter into intimal
layers;
– 2. Smooth muscle cells – SMC, from media
• Extracellular cholesterol looks like needle-shaped crystals
43
Atherosclerosis (AS)
44
2. Xanthomas
• Hereditary or Acquired Hyperlipidemias
• Intracellular cholesterol accumulation leads to à sub-
epithelial clusters of foam cells forming tumors or Xanthomas
• Common sites – Skin, Tendons
• Xanthelesma – at inner sides of eyes
Xanthomas- Skin
45
3. Cholesterolosis
• Foam cell collections in Lamina propria of
Gall bladder
• Mechanism – Unknown
Gall bladder Cholesterolosis
46
Cholesterol & its Esters)
4. Niemann –Pick Disease Type “C”
• Lysosomal storage disorder
• Mutations in enzyme lead to à impaired
Cholesterol trafficking
• Involves multiple organs
47
4. Niemann –Pick Disease Type “C”)
48
4. Niemann –Pick Disease Type “C”)
49
2. Proteins
• Light Microscopy (LM ) – Cytoplasmic rounded
eosinophilic droplets or vacuoles
• Electron Microscopy (EM) – appear as
amorphous/Fibrillar/crystalline structures
• Physiologic – Russell bodies in Plasma cells (next slide)
• Pathologic
1. Renal - Nephrotic syndrome
2. Defective intracellular transport / Secretion - α1-AT
deficiency
3. Accumulation of cytoskeletal proteins
4. Accumulation of Abnormal proteins-AMYLOIDOSIS
50
B. Proteins
Physiological
• Plasma cells (RUSSEL bodies- cytoplasmic
immunoglobulin secretions); enlarged ER with large
homogenous inclusions -
51
RUSSEL bodies Dr.T.Krishna MD, www.mletips.com
B. Proteins
• Pathologic
1. Renal - Nephrotic syndrome
Ø disorders causing massive proteinuria (Nephrotic
syndrome)
Ø Protein accumulate as pink, hyaline, Intracytoplasmic
droplets in PCT epithelium
Ø Reversible
52
Kidney tubules – protein droplets (in proteinuria)
Normal Tubule Tubule with Protein droplets
53
B. Proteins
• Pathologic
2. Defective intracellular transport / Secretion
α1Anti-Trypsin (α1-AT) deficiency
• Proteolytic enzyme synthesized and secreted by the liver
• Mutations of the protein lead to à slow folding &
accumulation of partially folded α1-AT in ER of Hepatocytes
• Organs involved
– Lung – an imbalance of Protease (Neutrophilic elastase)
and Anti protease lead to Emphysema (Panacinar?)
– Liver- ER stress àHepatitis, Cirrhosis
54
α1-antitrypsin (A1A) deficiency
Normal Histology
Diastase Sensitive = Glucose or Glycogen (simple

sugars/carbohydrates)
Diastase Resistant = Glycoprotein , Glycolipid 55
(Complex carbohydrates)
PAS +ve & Diastase Resistant
B. Proteins
• Pathologic
3. Accumulation of cytoskeletal proteins
• Alcoholic liver disease(ALD) - Mallory
bodies(Cytokeratin in hepatocytes)
• Alzheimer’s disease – with Neurofibrillary
tangles(neurofilaments)
56
B. Proteins
• Alcoholic liver disease(ALD) - Mallory
bodies(Cytokeratin in hepatocytes)
Fatty change
Mallory bodies
57
B. Proteins
• Alzheimer’s disease – with Neurofibrillary
tangles(neurofilaments)
58
2. Proteins
• Pathologic
4. Accumulation of Abnormal proteins-
AMYLOIDOSIS
• AMYLOIDOSIS – extracellular*** accumulation
of abnormal protein;
• Due to mis-folded or abnormal proteins
• Included in Proteinopathies/ Protein Aggregation
diseases
59
Amyloid protein under normal light
Congo Red stain
60
(Salmon Pink in light microscopy)
Amyloid protein under normal light
Congo Red stain
(Salmon Pink in light microscopy) 61

Amyloid protein – under polarized light
Congo Red stain
62
Apple Green Birefringence under Polarized Light
3. Glycogen
• Intracytoplasmic readily available energy source
• Excessive intracellular deposits of glycogen à Glycogen
Storage Disorders
• How glycogen appears in cells ?
– clear vacuoles within the cytoplasm.
• Why we need to take special precaution in tissue processing to
demonstrate glycogen/glucose ?
– glycogen/glucose are water soluble {(most commonly used
fixative – formalin (10% aqueous solution of
formaldehyde)} can’t used
– absolute alcohol is best fixative for glycogen/glucose in
tissues***
63
3. Glycogen
• What are best stains to demonstrate
glycogen/glucose in tissues ?
• glycogen/glucose in Haematoxylin and Eosin (H
&E) appears as nonspecific, hyaline material
• Best stains for glycogen/glucose in tissues are
1. Best carmine*** or
2. PAS ( another tissue section before staining treated
with diastase should be used as control)
64
PAS+ Diastase Negative
65
Self study

Cytoskeletal proteins
Types of cytoskeletal proteins
1. Thin /Actin filaments ( 6-8nm)
2. Intermediate filaments (10nm)
3. Thick filaments(15nm)
4. Microtubules (20-25nm)

Intermediate filaments (10nm)
•Intermediate filaments –flexible fibers protect

cells from shearing forces
•Types
1. Keratin – of the epithelium
2. Neurofilaments (neurons)
3. Desmin (muscle)
4. Vimentin (connective tissue)
5. Glial filaments (Glial cells)

Intracellular Accumulations
1
Pigments
Ø Pigments
ØColored substances
ØMC is carbon à causes coal worker’s
pneumoconiosis (CWP)
2
Pigments
Ø Exogenous
1. Carbon
2. Tattoo
Ø Endogenous
1. Lipofuscin
2. Melanin
3. Bilirubin
4. Hemosiderin
Ø MC is carbon à causes coal worker’s
pneumoconiosis (CWP)
3
Pigments
1. Carbon
ØMC exogenous pigment
Øair pollutant
ØAnthracosisà blackening of lungs, Lymph nodes
ØCoal worker’s pneumoconiosis (CWP) à in coal
miners, serious lung disease
2. Tattoo
Ø Pigmentation of the skin (pigment laden
macrophages in dermis without inflammatory
response)
4
Pigments
ØEndogenous
Ø1. Lipofuscin - yellow-brown, insoluble
Ølipochrome, wear & tear pigment
Øcomposed of polymers of lipids and phospholipids
Øproducts of lipid peroxidation of polyunsaturated lipids
ØHarmless
Ø seen in liver and heart of aging patients, also insevere
malnutrition and cancer cachexia
ØPresence indicate evidence of free radical injury & lipid
peroxidation
ØLocation -Perinuclear, Lysosomal
5
Pigments - Lipofuscin
Location -Perinuclear, Lysosomal
6
Pigments - Lipofuscin
Location -Perinuclear, Lysosomal

7
Pigments
ØEndogenous cont’d….
Ø 2. Melanin (normal black pigment)
Ø homogentisic acid
Øblack pigment seen in Alkaptonuria
ØAlkaptonuria is a rare metabolic disease
Ø Ochronosis – pigmentation of skin, connective
tissue, and cartilage
Ø 3. Bilirubin ; Major pigment of bile; Non iron
hemoglobin derived pigment in patients with
Jaundice
8
homogentisic acid
9
Bilirubin
Pigments
Ø 4. Hemosiderin
ØHemoglobin - derived, golden yellow-to-brown
pigment
Ørepresents aggregates of ferritin micelles
ØHemochromatosis - Persistent overload of all
organs but mainly liver, pancreas, heart, and
endocrine organs
ØComplications
ØLiverà Cirrhosis (Fibrosis)
ØHeartà Failure (Cardiomyopathy)
Ø Pancreas- Diabetes mellitus
11
Hemosiderin
H&E stain
Prussian Blue stain
12
Pathologic Calcification
• Abnormal tissue deposition of calcium salts,
along with smaller amounts of iron,
magnesium, and other mineral salts.
• Two forms of pathologic calcification.
1. Dystrophic calcification deposition
2. Metastatic calcification
13
Feature Dystrophic Metastatic Calcification
Calcification
Status of tissue dying tissues normal (viable) tissues
Serum calcium Normal Hypercalcemia
levels
Calcification
Derangements in Absent 1.Present (like Hyperparathyroidism from
parathyroid tumors, Paraneoplastic syndromes)
calcium
2. Destruction of bone by tumors of bone
metabolism marrow (e.g., multiple myeloma, leukemia) or
diffuse skeletal metastasis (e.g., breast
cancer), accelerated bone turnover (e.g., Paget
disease), or immobilization
3. vitamin D–related disorders- a) vitamin D
intoxication,
b) sarcoidosis (macrophages activate a
vitamin D precursor)
c) Idiopathic hypercalcemia of infancy
(Williams syndrome- abnormal sensitivity to
vitamin D)
4. renal failure (retention of PO4 à, secondary
hyperparathyroidism)
Calcification
Location intracellular, Widespread & in multiple
extracellular, or in organs (like gastric mucosa,
both kidneys, lungs, systemic
arteries et.,)
Pathogenesis Formation of • all of these tissues excrete
hydroxylapatite acid
just like that of • therefore have an internal
bone. alkaline compartment
• predisposes them to
metastatic calcification
Precipitating factors Hypercalcemia Hypercalcemia (both for initiation and
progression)
Feature Dystrophic Metastatic
Calcification Calcification
Examples/Causes Areas of necrosis Less common are
Atherosclerosis 1. aluminum intoxication (in chronic
psammoma bodies (lamellated renal dialysis pts.)
calcification of single cells in 2. milk-alkali syndrome (milk or
Papillary carcinomas) antacid like calcium carbonate)
Asbestos bodies ( calcium and
iron salts form exotic, beaded,
dumbbell-shaped )
Feature Dystrophic Metastatic
Calcification Calcification
Appearance
Gross fine, white granules or resemble those of
clumps as gritty dystrophic calcification.
deposits
Microscopy basophilic, amorphous resemble those of
(H&E stain) granular or like dystrophic calcification.
clumped appearance
Significance sign of previous cell
injury
Dystrophic

Dystrophic
psammoma bodies

Metastatic

Self study

Cytoskeletal proteins
Types of cytoskeletal proteins
1. Thin /Actin filaments ( 6-8nm)
2. Intermediate filaments (10nm)
3. Thick filaments(15nm)
4. Microtubules (20-25nm)

Intermediate filaments (10nm)
•Intermediate filaments –flexible fibers protect

cells from shearing forces
•Types
1. Keratin – of the epithelium
2. Neurofilaments (neurons)
3. Desmin (muscle)
4. Vimentin (connective tissue)
5. Glial filaments (Glial cells)

1
} General
◦ Definition
◦ Purpose
◦ Components
◦ Regulating factors
} Historical points
} Acute Inflammation
◦ Major components
◦ General
◦ Stimuli
◦ Events
Vascular
Cellular
◦ Termination
◦ Chemical mediators
◦ Morphologic patterns
◦ Summary
2
} Cell injury à Inflammation à Repair
} Most important feature*** - vascular response à extracellular
accumulation of fluid & blood leukocytes
} Purpose
} Mainly protective but
} Can be harmful (like chronic Infl. Diseases - Rheumatoid arthritis
(RA), Atherosclerosis (AS), type 2 DM, Alzheimer’s and cancer)
} Main components
} Vascular events
} Cellular events
} Divided into
} Acute inflammations
} Chronic inflammations
} Mediators of both acute & Chronic inflammation
} Mainly chemical factors
3
} Definition= “Complex host response to injurious
agents such as microbes and damaged, necrotic,
cells that consists of vascular responses, migration
and activation of leukocytes, and systemic
reactions”
} Why inflammation?
◦ mainly protective
} What happens if there is no inflammation?
◦ Infections never stop
◦ Wounds never heal
◦ Tissue damage à permanent
} Is inflammation always protective?
◦ No (lead to Acute and Chronic Infl. Disorders)
4
} Cardinal signs of inflammation (Celsus , Roman,
first century AD)
1. Rubor- redness
2. Tumor- swelling
3. Calor- heat
4. Dolor - pain
5. Loss of function (functio laesa by Virchow in 19th
century)
} Metchnikoff - discovered phagocytosis (cellular
event)
} Sir Thomas Lewis
◦ Triple response – 1. red line, à2. flare around
that line, then à3. wheal (Histamine mediate the
vascular changes of inflammation – Vascular
event)
5
Components:
1. Alterations in vascular caliber results in increase in blood flow
2. Structural changes in the microvasculature
3. Emigration of the leukocytes
6
} Pus
ØPurulent inflammatory exudate
ØRich in leukocytes (Neutrophils – pus cells), cellular
debris, microbes
} Edema
ØExcess of fluid in the interstitial or serous cavities
ØExudate or Transudate
} Features
ØMinutes to hours
ØExtravasation of Cells (Neutrophils)
} Termination of inflammation
ØRemoval of offending agent
ØExhaustion of secreted mediators
ØAnti-inflammatory mechanisms
7
} Hallmark of acute inflammation - Edema***
8
} Exudate } Transudate
} inflammatory extra } Fluid with low protein
vascular fluid & high content (Mainly
protein concentration albumin) & Ultrafiltrate
of plasma due to osmotic
or hydrostatic imbalance
} specific gravity > 1.020 } specific gravity < 1.012
} cellular debris present } Very few or no cells
} Indicates altered } No increase in vascular
permeability of small permeability
blood vessels in the area
of injury

Components
10
Stimuli produce Acute inflammatory
reaction
} Infections and microbial toxins
} Trauma (blunt and penetrating)
} Physical and chemical agents(thermal injury,
irradiation)
} Tissue necrosis (any cause)
} Foreign bodies (dirt, sutures)
} Immune reactions (hypersensitivity reactions)
11
} Vascular Reactions
} Changes in Vascular Flow and Caliber
} Vasoconstriction of arterioles followed by
vasodilatation à opening of new capillary beds result
in increased blood flow (heat, redness)
} Mediated by histamine, nitric oxide - on vascular SMC
} Increased permeability of microvasculature lead
to
} Exudation of protein-rich fluid into the extra vascular
tissues (edema)àred cell stasis & Neutrophil
collection along the vascular endothelium
12
13
14
1. EC gaps in Venules
} MCmechanism of vascular
leakage
} Immediate, transient response
} Affecting venules 20 to 60 μ.m.
2. Direct EC injury (cell

necrosis and detachment)
***
} Severe burns or lytic bacterial
infections ***
} Immediate, sustained response
} Affects all levels of the
microcirculation
15
3. Leukocyte-mediated EC injury
} Leukocytes damage
endothelium by free radicals &
Proteolytic enzymes
} Cause endothelial injury or
detachment
} venules and capillaries of lung
& kidney
4. Increased transcytosis
} via Vesiculo - vacuolar
organelle
} located close to intercellular
junctions
} VEGF -↑number and the
size of organelle
16
5. Leakage from new blood
vessels (angiogenesis)
} New vessel are leaky
} VEGF àangiogenesis
17
Leukocyte migration & Phagocytosis
18
Steps in Extravasation
} In the lumen of the vessel
◦ Margination - Accumulation of WBC along endothelium
◦ Rolling-slow movement on endothelium and temporary
adhesion
◦ Adhesion to endothelium
◦ Pavmenting - formation of layer of WBC along
endothelium and firm adhesion
} Transmigration across the endothelium or diapedesis
} Migration into extra cellular space
} Migration in interstitial tissues toward a chemotactic
stimulus
19
Adhesion and transmigration are controlled by
} Adhesion molecules –
} Selectins
} Ig super family
} Integrins
} Mucin-like Glycoproteins
} Chemical mediators
◦ cytokines
◦ chemo attractants
20
Leukocyte migration & Phagocytosis
21
Thank You
22
1
Adhesion molecules
1. Selectins
2. Integrins
3. Ig Super family
4. Mucin like Glycoproteins (gp)
Type of cell response
Clinical Significance
Chemotaxis
Leukocyte Activation
Phagocytosis
Clinical Examples
Defects in Leukocyte Functions
2
Specific adhesion between cell-cell & cell-ECM is essential
for
} Embryogenesis
} Tissue repair
} Immune responses
} Inflammatory responses
Different genes encode proteins with specific adhesive
functions
Adhesion molecules are
1. Selectins
2. Immunoglobulin (Ig) super family
3. Integrins
4. Mucin -like gps’
3
} Adhesion of leukocytes to Endothelial Cells (EC)
} C- Lectin (calcium-dependent lectin domain)bind to sialylated
oligosaccharides
} single-chain transmembrane glycoproteins
} Three types (E,P,L)
} Important in rolling*** (of cellular events)
L- selectin (L-leukocyte)à is for
◦ homing of Lymphocytes in lymph nodes(LN)
◦ Adhesion of Neutrophils to EC
E- Selectins (E –Endothelium)à
} Expressed only on cytokine-activated EC
} For homing of T cells to peripheral sites,
} ***Hallmark of acute cytokine-mediated inflammation
P-Selectins (Platelets & EC :W-P bodies),
} Following stimulation, this selectin translocated within
minutes to the endothelial cell surface (next slide)
} Sialylated – oligosaccharides react with sialic acid or its derivatives
4
EC EC
Cell? within minutes

Ligand for E
selectin
E selectin
1-2 hours
EC EC

Clinical significance
} Leukocyte Adhesion Deficiency-2 (LAD-2) à
defect in adhesion due to mutations of ligand to E -
Selectins
} knockout mice =
} L –selectin deficiency à poorly formed LN
} E or P selectin deficiency à mild impairment of
leukocyte recruitment
} Deficiency Both E&P Selectins à severe
impairment of leukocyte recruitment & risk of
infection
7
“Integrate” signals from extracellular ligands with
cytoskeleton
} 30 structurally homologous proteins
} Mediate firm adhesion between leukocytes & EC
} Heterodimeric {(dimer = Two) cell surface proteins (α and β)}
} Sub - families are based on β subunits
} Integrins on the leukocytes
1. VLA-4 (β1- containing) and
2. LFA-1, Mac-1 (β2- containing)
} Ligands on the EC (Ig super family)
1. Vascular cell adhesion molecule -1 (VCAM-1à for β1 containing
Integrins)
2. Intercellular adhesion molecule -1 (ICAM-1à for β2 containing
Integrins)
8
?
Enter from site injury into

blood vessel
Chemokines act on what cells?
9
LFA-1
Enter from site injury into

blood vessel
Chemokines act on what cells?
Both Leukocytes and EC

10
Immunoglobulin family
} Endothelial adhesion molecules
◦ ICAM-1 (intercellular adhesion molecule 1)

◦ VCAM-1 (vascular cell adhesion molecule 1)
} Ligands for Integrins on leukocytes
} ***PECAM-1 (Platelet Endothelial Cell Adhesion Molecule) or

CD31 for diapedesis (transmigration of Leukocytes across EC) ,
expressed on both EC & leukocytes
Mucin-like glycoproteins
} Heparan sulfate
} ligands for CD44 (leukocyte adhesion molecule )
} leukocytes are retained at the site where they are needed by
◦ Integrins
◦ CD44 (binding with Hyaluronic acid)
11
Type of leukocyte emigration depends on age of the
inflammatory response
} Neutrophils à during the first 6 to 24 hours
} Mononuclear cells (Lympho, Mono, Plasma cells) àin 24 to

48 hours
} Exceptions
} Eosinophils in hypersensitivity reactions
} Pseudomonas infection = Neutrophils last over 2 to 4 days

} Viral & Tuberculosis infections =lymphocytes
Clinical significance***
} leukocyte adhesion deficiency type 1 (LAD1)
◦ defect in synthesis of β2 chain of Integrins

} Leukocyte adhesion deficiency type 2 (LAD2)
◦ absence of sialyl -Lewis X (ligand for E-selectin)

12
13
14
15
} Chemotaxis - Emigration extravasated leukocytes
(N, M) in ECM toward site of injury
Chemo attractants
} Exogenous and Endogenous substances
} MC exogenous ones =bacterial products
} Endogenous can be = C5a, Leukotrienes B4 (LTB4) &
cytokines (chemokine –IL-8) bind to seven-
transmembrane G-protein-coupled receptors
(recognize microbes)
Process
} phospholipase C {(PLCγ) and phosphoinositol-3
kinase (PI3K) - second messengers} à cause
} increase cytosolic calcium à lead to
} polymerized actin at filopodia 16
L
E
U
K
O
C
Y
T
E
17
Steps in Phagocytosis
} Recognition and attachment of the particle to
leukocyte
◦ By Mannose & scavenger receptors expressed on
the leukocyte surface
◦ Opsonization (on microbes) increases the affinity of
the process
} Engulfment
◦ Extensions pseudopods around the particle à
phagosome à Phoagolysosome
(Pagosome+lysosome)
19
Steps in Phagocytosis
} Killing or degradation by
Ø 1.Oxygen-dependent mechanisms (mainly – next slide)
Ø 2. Oxygen-independent mechanisms***
◦ Bactericidal permeability increasing protein (BPI)- against
gram-negative bacteria
◦ Lysozyme- hydrolyzes the muramic acid–N-
acetylglucosamine bond of bacteria cell wall
◦ Lactoferrin - iron-binding protein
◦ Major basic protein (MBP) of Eosinophils - Cytotoxic to
many parasites
◦ Defensins - cationic arginine -rich granule peptides that are
toxic to microbes
◦ Cathelicidins - antimicrobial proteins found in neutrophils
and other cells
◦ Other enzymes- elastase
20
hypochlorite
21
Acute Chronic
Acute respiratory distress syndrome Arthritis
Acute transplant rejection Asthma
Asthma Atherosclerosis
Glomerulonephritis Chronic lung disease
Reperfusion injury Chronic rejection
Septic shock
Vasculitis
Disease Defect
Genetic
Leukocyte adhesion deficiency 1 β chain of CD11/CD18 integrins
Leukocyte adhesion deficiency 2 Fucosyl transferase required for synthesis of
sialylated oligosaccharide (receptor for selectin)
Chronic granulomatous disease Decreased oxidative burst
X-linked NADPH oxidase (membrane component)
Autosomal recessive NADPH oxidase (cytoplasmic components)
Myeloperoxidase deficiency Absent MPO-H2O2 system
Chédiak-Higashi syndrome Protein involved in organelle membrane docking and
fusion
Acquired
Thermal injury, diabetes, malignancy, Chemotaxis
sepsis, immunodeficiencies
Hemodialysis, diabetes mellitus Adhesion

Leukemia, anemia, sepsis, diabetes, Phagocytosis and microbicidal activity
neonates, malnutrition
Very Important Table
2 4
3
Phagosome
1
respiratory burst
24
Chédiak - Higashi
Peripheral blood =Giant granules in neutrophils

25
Bone Marrow Suppression
} MCC of Defect in Leukocyte Functions
Chédiak - Higashi syndrome =AR inheritance

} Pts present with neutropenia, albinism, nerve defects, bleeding disorders,
recurrent infections
} Defect in degranulation, delayed microbial killing
} Peripheral blood =Giant granules in neutrophils
} Mechanism = Reduced transfer of lysosomal enzymes to phagocytic
vacuoles in phagocytes
Chronic granulomatous disease

} Congenital defects in bacterial killing (Oxygen-dependent ) defects in the
genes encoding NADPH oxidase
◦ 1. X-linked ( defect in plasma membrane-bound components)- MC
◦ 2. Autosomal recessive (defect in cytoplasmic components)
26
Acute
Inflammation
Vascular Events Cellular Events
Caliber & Adhesion &

Permeability Chemotaxis
Flow Migration
Activation
Phagocytosis
Recognition &
Attachment
Engulfment
Killing &
Degradation
Thank You
28
1
} General
} Cell Derived Mediators
◦ Vasoactive amines
Histamine
Serotonin
◦ Arachidonic acid (AA) metabolites
Prostaglandins (also Thromboxanes)
Leukotrienes
Lipoxins
} Plasma Derived Mediators
◦ Complements
◦ Kinins
◦ Coagulation/ fibrinolysis components
} Others
2
3
4
Sources
} Plasma-derived ( complement proteins,
Kinins, clotting factors)
◦ Present in the form of precursor forms
} Cell-derived (Platelets, Neutrophils,
Monocytes, mesenchymal , epithelial, mast
cells)
◦ 1. Preformed (sequestered in intracellular
granules)à need to be secreted (example -
histamine in mast cell granules)
◦ 2. Synthesized de novo (examples-
prostaglandins, cytokines)
5
Sources
} Triggering factors = microbial products or host
proteins
} Act by = binding to specific receptors on target cells
◦ Direct action (example - lysosomal proteases) or
mediate oxidative damage
◦ act on = one or few target cell types
} Effects =different effects on different target cell types
} Cascade = One mediator can stimulate the release of
others
} Life span= Activated and released mediators are
short-lived
} Effects = Most mediators àcause harmful effects
6
Histamine and Serotonin
A } Plasma-derived Present in preformed forms
B
C in cells
} First mediators to be released during
inflammation
Histamine
} Richest in mast cells (also in Basophils and
PLTS)
} Mast cells à located in ECM close to
blood vessels
} Stimuli for histamine release ànumerous
(histamine-releasing proteins (HRP) of
WBC, Cytokines, Trauma, Anaphylotoxins,
substance P etc.,)
} Effects
◦ Dilation of arterioles, constriction of
large arteries
◦ Increase vascular (venular) permeability
A. Venule by inducing EC gaps
( Immediate & Transient response)
B. Mast cell
Serotonin
C. Adipose tissue } Source – PLTS, Enterchromaphin cells
} Release – during IgE mediated reactions
àPAF àPLTS aggregation à release 7
HPETE – Hydroperoxyeicosatetraenoic acid
HETE – Hydroxyeicosatetraenoic acid

} AA metabolites
} Also called autacoids (or short range
hormones)
} Important for inflammation & hemostasis
} AA present as esterified form in membrane
phospholipids
} Release of AA à by action of phospholipase
A2 (PLA2)
} PLA2 is activated by à several stimuli
9
} AA metabolites cont’d….
◦ 1. prostaglandins {PGs which include
Thromboxanes (TXs)} by action of
cyclooxygenases (COX-1 & COX-2) on AA
◦ 2. Leukotrienes (LTs ) & Lipoxins (LPs) by action
of Lipoxygenases on AA
} AA metabolites bind to G-protein coupled receptors
(GPCRs) à mediate almost every step in
inflammation
} ***AA metabolites (PGE2) Present in
inflammatory exudate
} Anti – inflammatory medications act on the
above enzymes
10
AA
COX pathway
} Initiated by COX-1 and COX-2
} Produce PGS ( includes TXs)
} Important PGs in inflammation à
PGE2, D2, I2, TXA2
} TXA2 is mainly in – PLTS (PLTS
have TX synthase)
} Effects; ***PGE2 à Hyperalgesic
(↑↑pain)
◦ PGE2, D2 à Vasodilation &
edema, Fever
◦ COX-1 important in
inflammation & hemostasis
(PGs, TXs)
◦ ***COX-2 important only
inflammation (by PGs)
} ***Clinical significance =
Imbalance of PGI2, TXA2
àthrombi formation in CVS
& CNS (MI & Stroke)
11
AA
LPX ( Lipoxygenase ) pathway
} 5-LPX (PMNs)àproduce 5-
HPETE (from AA) à LTs
(A4,B4, C4, D4, E4)
} 12-LPX (PLTS) àproduce
Lipoxins LXA4, LXB4
} LTs = play role in pathogenesis
of bronchial asthma
} ***LXs= endogenous negative
regulators of Leukotriene action
} Fish Fats = poor substrates for
cyclooxygenases &
Lipoxygenases
} LT Inhibitors – Zileuton
PETE – Hydroperoxyeicosatetraenoic acid } LTR (LT receptor) inhibitors -

HETE – Hydroxyeicosatetraenoic acid Montelukast
12
13
Action Metabolite
Vasoconstriction Thromboxane A2, leukotrienes

C4, D4, E4
Vasodilation PGI2, PGE1, PGE2, PGD2
Increased vascular Leukotrienes C4, D4, E4

permeability
Chemotaxis, leukocyte Leukotriene B4, HETE, lipoxins

adhesion
} Complements
} Kinins
} Clotting systems
15
} Inactive plasma proteins labeled as numeric (C1 – C9)
} Critical part of Early steps in complement pathway activation is activation of
C3 by C3 convertase (C3 convertase by 3 pathways)
◦ 1. classic ( triggered by fixation of C1 to Ag-Ab complex)
◦ 2. alternative {Triggered by microbial surface molecules (endotoxin,
or LPS), complex polysaccharides, cobra venom, and others (absence
of antibody)}
◦ 3. Lectin (triggered by lectin binds to carbohydrates on microbes which
activates C1)
} Critical part of Late steps is formation of C5 convertase ( which finally forms
C5-C9 or MAC complex)
} Autocatalytic
} Participate in both Cell Mediated Immunity (CMI) & Antibody Mediated
Immunity (AMI)
} ***Effects
1. Vascular effects
} Anaphylotoxins = C3a, C5a(also C4a) stimulate histamine release from
mast cells
} Leukocyte adhesion, chemotaxis, and activation = C5a ( Neutrophils,
Chemotaxis, other WBC àM, E, & B)
} Phagocytosis =C3b and iC3b (inactive C3b)
17
***Regulation of complement pathway
} DAF (Decay-accelerating factor ) = decays C3 & C5
convertases
} Factor-l = cleaves C3b
} CINH = inhibits C1 binding with immune complex
} CD59 (membrane inhibitor of reactive lysis) = inhibits MAC

complex
} Disorders = result in increased susceptibility to infections
} Defective C3 à susceptibility to infections & FATAL
} Defective alternative pathway à ↓resistance to infections
} Defective C2 & C4 à autoimmune diseases
} ***Defective late components à defective formation of the

MAC (Neisseria organisms)
18
***Paroxysmal nocturnal hemoglobinuria
} Mutations in the genes lead to lack of phosphatidylinositol
linkages of Membrane proteins (DAF and CD59) à Un-
controlled complement activation
***Hereditary Angioneurotic Edema

} Due to C1 inhibitor (C1INH) Deficiency
} Episodic edema in Skin, Extremities, Larynx, Intestinal

mucosa
} C1INH =inhibitor of C1, Kallikrein, and Coagulation factor
XII
19
2
C5
1
C5a
- 3
Kininase
& XII
ACE
XIIa
HMWK – High Molecular Weight Kininogen

} Ultimate step in the pathway = Bradykinin (vasoactive
nonapeptide) production
} ***Effects of Bradykinin
1. Increases vascular permeability
2. Contraction of smooth muscle
3. Dilation of blood vessels
4. Pain when injected into skin
21
Thrombin= link between the coagulation system and inflammation
} 1.Converts fibrinogen to generate an insoluble fibrin clot
} 2.Binds to PAR (protease-activated receptors) à
} PAR = G protein-coupled receptors (on PLTS, EC, SMC& other cells)
} Thrombi + PAR interaction lead to
1. Mobilization of P-selectin
2. Production of chemokines
3. Expression of endothelial adhesion molecules for leukocyte integrins
4. COX-2
5. Production of PGs
6. Production of PAF and NO
7. EC shape change
Factor XIIa (Hageman factor) initiates four systems
1. Complement
2. Kinin
3. Clotting
4. Fibrinolysis
22
PAF
} Derived from & activates PLTS,
} Sources = blood cells (PLTS, Mono, PMN, B), EC
} Effects = VC (VD in very low conc.), Broncho-constriction, ↑vascular
permeability (edema),
} Inhibited by = acetyl hydrolases
} *** Important role in Necrotising Enterocolitis
Cytokines
} Sources = activated macrophages, EC, Epithelium etc.,
} Types = macrophage derived ( TNF-α, IL-1), T cell derived
(Lymphotoxins)
} Effects = EC activation & ↑ thrombogenicity of EC, Acute phase response
( Fever, loss of appetite, slow wave sleep, steroid secretion),
} TNF-α = PMN priming, cachexia, shock etc.,
Neuropeptides
} Substance P, Neurokinin A
} Initiate & propagate inflammatory response
23
Chemokines
} Chemo attractants of WBC
} 40 types & 20 receptors
} 4 major groups with two forms (cell surface bound or soluble forms)
} Act by binding with 7-GPCR
} Most important ones= CXCR-4, CCR-5 (also act as co-receptors for HIV-1
virus)
NO
} Act via cGMP (what are other examples act via cGMP ?)
} Has very short life (seconds)
} Synthesis = from L- arginine & the enzyme is nos
} Types of nos (i- inducible, E- endothelial, n-neuronal)
} Effects
} 1. VD, ↓PLTS adhesion & aggression, ↓mast cell degranulation
} 2. Inflammation = ↑NO production à ↓replication of pathogens
( including tumor cells)
} 3. Microbicidal = by producing free radicals
} Clinical
} ↓ NO production à ↑ microbial replication,
} Defective EC synthesis of NO = seen in AS, DM, HTN
24
25
26
27
1. Classical activation
} Stimuli - microbial products and T-cell cytokines
(IFN-γ)
} Result - strong microbicidal activity,
2. Alternative activation
Stimuli –cytokines from TH2 cells (IL-4 and IL-13)
Result –Tissue repair and fibrosis
Significance of Macrophage activation
} Different stimuli activate them to secrete mediators
of inflammation as well as inhibitors of the
inflammatory response
} Helps in both amplify and control inflammatory
reaction.
28
1. Serous inflammation
2. Fibrinous inflammation
3. Purulent inflammation
4. Ulcer
29
1.Serous Inflammation
◦ With exudate of cell poor fluid (no
organisms, no leukocytes)
◦ Can be due to
inflammatory conditions -burns /viral
infections (blisters) or
Non –inflammatory conditions – Heart
failure, Kidney or liver disorders (
◦ AKA known as effusions in Pleura
(pleural effusion), Pericardium
(Pericardial effusion , Peritoneum (?)
30
31
– due to
1. large vascular leaks
2. Local Procoagulant stimulus
} Affect body cavities (Pleura, pericardium) and Meninges
32
} Characterised by - pus, exudate of neutrophils, liquefied
necrotic material, edema fluid
} Abscess – localised collection of purulent inflammatory
tissue (PUS)
33
} 4. Ulcer
◦ Local defect or excavation of surface of organ or tissue
◦ Caused by shedding or sloughing of inflamed necrotic tissue
◦ Best examples – oral, skin, peptic ulcers
What is the difference between ulcer and erosion?
Erosion – superficial (not beyond muscularis mucosae)
Ulcer – Deep beyond muscularis mucosae
34
Oxygen Leukocytes Endothelial damage,
metabolites tissue damage
PAF Leukocytes, mast cells Bronchoconstriction,

leukocyte priming
IL-1 and TNF Macrophages, other Acute-phase reactions,

endothelial activation
Chemokines Leukocytes, others Leukocyte activation
Nitric oxide Macrophages, Vasodilation, cytotoxicity

endothelium

Vasodilation Prostaglandins
Nitric oxide
Histamine
Increased vascular permeability Vasoactive amines
C3a and C5a (through liberating

amines)
Bradykinin
Leukotrienes C4, D4, E4
PAF
Substance P

Chemotaxis, leukocyte recruitment C5a
and activation
Leukotriene B4
Chemokines
IL-1, TNF
Bacterial products
Fever IL-1, TNF
Prostaglandins
Pain Prostaglandins
Bradykinin
Tissue damage Neutrophil and macrophage
lysosomal enzymes
Oxygen metabolites
Nitric oxide

} Complements
} Kinins
} Clotting systems
38
Thank You
39
Chronic inflammation (CI)
• Inflammation of prolonged duration (weeks to

months to years) in which :
– inflammation, tissue destruction and attempts
at repair proceed simultaneously.
• Most often results from persistence of an injury
causing agent.
• Predominant cell type is mononuclear, which
includes
– macrophages (key cells),
– lymphocytes and plasma cells.
Win 21 A Jalan 1
Chronic inflammation (CI)
• Chronic inflammation: the characteristics:
1. Tissue destruction.
• by offending agent/inflammatory cells
2.Infiltration with mononuclear cells
• Macrophages, lymphocytes and plasma cells
3.Replacement of damaged tissue by CT (repair)
• due to proliferation of fibroblasts and new vessels
(fibrosis & angiogenesis).
• Chr. inflammation Unlike acute inflammation has:
– Mononuclear cell infiltrate
– Induration (hardening) of affected tissue (due to
fibrosis)
– Less swelling (less exudate)
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– Less hyperemia (redness)
Three characteristic histologic features of CI
Fibrosis
Tissue destruction
Chronic inflammatory cells
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Mononuclear
cells
Chronic
Win 21 endometritis = Chronic
A Jalaninflammation of endometrium
4
Acute Inflammation
Acute Vs Chronic inflammation
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Chronic Inflammation
Causes of chronic inflammation
1. Following acute 3. Prolonged exposure to toxic
inflammation. agents.
– Silica (silicosis)
2. Infections with certain – Lipids (atherosclerosis)
organisms – Silicone in breast implants
a. Viral infections 4. Autoimmune diseases
b. M. tuberculosis – Rheumatoid arthritis
(tuberculosis) – Systemic lupus
c. Treponema pallidum erythematosus (SLE)
(syphilis)
d. Fungal infections
(histoplasma)
e. Parasitic infections
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Cells of chronic inflammation
• Different from acute inflammation !!
• Key cells:
– Monocytes and macrophages**
• Other cells include
– Lymphocytes
– Plasma cells and
– Eosinophils
• Dominant cellular player in chronic inflammation is the
macrophage
Belongs to the:
– MONONUCLEAR PHAGOCYTE SYSTEM
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Mononuclear Phagocyte System
Consists of :
• Circulating blood monocytes and Tissue macrophages (2
types):
1. Fixed macrophages
Kupffer cells (liver)
Sinus Histiocytes (spleen)
Bone (osteoclast)
2. Wandering macrophages
Microglia (CNS)
Alveolar Macrophages (lung)
• Remember that tissue macrophages are derived from
blood monocytes.
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• During inflammation macrophages are recruited from the
blood (circulating monocytes).
• Chemotactic factors: MCP-1.
• At the site of injury monocyte transforms in to a larger cell
the macrophage : capable of phagocytosis.
• These can become activated macrophages
– Due to action of cytokines etc.
• Activated macrophages
– Greater ability to kill ingested organisms
• Increased lysosomal enzymes, NO and ROS
– Help in tissue repair.
• Secrete growth factors
– Activation signal :
• IFN gamma by CD4 T cells
• Bacterial endotoxins, AIL-4,
Win 21 Jalan IL-13 9
sel
es
v Tissue Macrophage
ood
Bl
Endotoxin
IFN γ
Activated
Activated macrophage CD4 T cell
Inflammation & Fibrosis

Tissue injury
•IL-1, TNF •Growth factors:

•Toxic oxygen metabolites PDGF, FGF, TGF beta
•Acid and neutral proteases: •Angiogenesis factor etc
Win 21 •degrade elastic and A Jalan 10
•collagen fibers.
Functions of macrophages
1. Phagocytosis
– Receptor for IgG and C3b
2. Microbicidal activity
– No O2 dependent - MPO system
– Have Oxygen derived free radicals
3. Process antigens
– Act as Ag presenting cells (APC).
4. Scavenger cells
5. Secrete Cytokines :
– IL-1 and TNF
6. Secrete growth factors (e.g. PDGF)
7. Have proteolytic enzymes
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EM Macrophage
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Lymphocyets : in chronic inflammation
• Lymphocytes: Primary inflammatory cells in

– Viral infections
– Type IV hypersensitivity reaction.
– Bordetella Pertussis infection
• Whooping cough
• Two types of lymphocytes:
1. T cells
• CD 4 (Helper T cells)
• CD 8 (Cytotoxic T cells)
2. 21B Cells
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T cells
• Interact with macrophages in chronic
inflammation
• Activated by ag presented by macrophages
• Activated lymphocytes produce IFN gamma
• IFN gamma activates macrophages
• Activated macrophages produce IL-1 and TNF
that activates lymphocytes and other cells.
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Lymphocyte
IL-12, TNF,IL-1
Activated
Activated macrophage
Lymphocyte
Ag presentation
IFN γ
Win 21
Macrophage
A Jalan 16
• Plasma cells :
– Derive from activated B cells
– Blue cytoplasm and eccentrically situated
nucleus
• Functions:
– Antibody production
– IgM produced on first exposure in acute
inflammation
– IgG synthesized after 10-14 days
– IgG is the main immunoglobulin of chronic
inflammation
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Plasma cells
Win 21 Ink blue Acytoplasm
Jalan 18
Eccentrically situated nucleus
Plasma cell: EM
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Other Cells of chronic inflammation
• Eosinophils: found in
– parasitic infections
– immune reactions mediated by IgE.
– Chemo-attractant:
• eotaxin
– Eosinophil granules contain:
• major basic protein-toxic to parasites
(also causes epithelial cell necrosis).
Win 21 A Jalan 20
Patterns of chronic inflammation
1. Chronic nonspecific inflammation

2. Granulomatous inflammation
• Chronic nonspecific inflammation: Characterized by

– Infiltration by mononuclear cells e.g,
macrophages, lymphocytes and plasma cells.
– Inflammatory response not specific for any
particular etiological agent.
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Chronic nonspecific inflammation
(endometritis)
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Granulomatous inflammation
• Definition: specialized form of chronic

inflammation characterized by formation of
granulomas.
– Granulomas: are localized collections of
modified/activated macrophages
(epithelioid cells).
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Types of granulomas
• Two:
1. Caseating granulomas and
2. Non-caseating granulomas
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Caseating granulomas
• Exhibit central caseous necrosis

• Characteristic of tuberculosis and fungal
infections (e.g.histoplasmosis)
• Caseous (“cheese-like”) material:
– is due to lipid released from the cell walls
of dead pathogens.
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Caseating granulomas: tuberculosis
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Caseous necrosis
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Tubercular
A Jalan
Granuloma 28
Noncaseating granulomas
• Lack central necrosis.

• Common etiologies include:
– Reaction to foreign material (e.g. suture)
• Foreign body granuloma
– Sarcoidosis
– Beryllium exposure
– Crohn disease, and
– Cat scratch disease
Win 21 A Jalan 29
Granuloma without necrosis
= non-caseating granuloma
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Composition of granulomas
• Epithelioid cells
– IFN gamma transforms macrophages à epithelioid cells (
hallmark cell of granuloma*)
• Multinucleated giant cells
– Formed by fusion of epithelioid cells
– Langhans type giant cells (TB)
– Foreign body type of giant cells
– May be present ; but not necessary*
• Lymphocytes, Plasma cells
• Central caseous necrosis
– Seen in granulomas due to tuberculosis, fungal
infections*.
– Rare in other granulomatous diseases.
• Outermost rim of fibrous tissue
– If healing has begun
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Caseating granuloma: tuberculosis
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Langhans
Granuloma Giant Cell
Lymphocytic
Rim
Caseous
Necrosis
Epithelioid
Macrophage
Win 21 A Jalan 34
Tuberculosis: Granuloma
Rim of
lymphocytes
Epithelioid cells
Langhan
Giant cell
Win 21 A Jalan 35
Non-caseating granuloma:Sarcoidosis
Granuloma
Giant cell
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Non-caseating granuloma: Foreign body
Suture Foreign body

material type giant cell
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Non-caseating granuloma: Foreign body
vegetable material
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Examples of granulomatous diseases
Disease Cause Reaction
Tuberculosis M.tuberculosis Caseating*
Fungal infections Histoplasma Caseating*
Sarcoidosis Unknown Non-caseating

Silicosis,Berylliosis Silica,beryllium Non-caseating
Foreign body Suture,Breast Non-caseating

prosthesis
Leprosy M.Leprae Non-caseating
Syphilis T.Pallidum Non-caseating (Gumma)
Cat scratch disease Gram negative bacillus Non-caseating (Stellate

granuloma)
Schistosomiasis
Win 21 S.hematobium
A Jalan Non-caseating 39
Steps involved in granuloma formation
1. Macrophages process and present antigen via

MHC class II to CD4 helper T cells.
2. Interaction leads macrophages to secrete
IL-12, inducing CD4 helper T cells to
differentiate into TH1 subtype.
3. TH1 cells secrete IFN-y, which converts
macrophages to epithelioid cells (=activated
macrophages).
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Blood
vessel
INF γ Epithelioid cell

= activated
macrophage
Monocyte
Win 21 Granuloma
A Jalan 42
Outcome of chronic inflammation
1. Development of secondary amyloidosis.

– due to production of serum associated
amyloid (SAA) protein by the liver
2. Chronic inflammation à repair by fibrosis:
• Fibrosis can result in:
1. Intestinal obstruction by fibrous bands
(adhesions)
2. Induction of cancer (scar cancer of
lung)
3. Stricture (narrowing) formation.
Win 21 A Jalan 43
Question
• A 40-year-old woman had bilateral silicone breast
implants placed two years ago. During that time,
there is increased firmness with slight deformity on
the left. The implants are removed, and there is
evidence for leakage of the implant contents on the
left. Which of the following cell types would be most
characteristic of the inflammatory response in this
situation:
• A Mast cell
• B Eosinophil
• C Giant cell
• D Neutrophil
•Win E
21
Plasma cell A Jalan 44
Question
• A 40-year-old woman has a chronic cough with fever
and weight loss. A chest radiograph reveals multiple
nodules from 1 to 4 cm in size, some of which
demonstrate cavitation in the upper lobes. A sputum
sample reveals the presence of acid fast bacilli.
Which of the following is the most important cell
that participates in the development her lung lesions:
• A Macrophage
• B Fibroblast
• C Neutrophil
• D Mast cell
• E Platelet
Win 21 A Jalan 45
Comparison of Acute and Chronic Inflammation
FEATURE ACUTE INFLAMMATION CHRONIC INFLAMMATION
Pathogenesis Microbial pathogen, burn, From Acute inflammation,
trauma foreign body, autoimmune
disease, tuberculosis, leprosy
Primary cell involved Neutrophil Monocytes.Macrophages (key
cells), B and T lymphocytes,
plasma cells, fibroblasts
Scar tissue Absent Present
Onset Immediate Delayed
Duration Few days Weeks, months, years
Outcome Resolution, abscess Scar tissue formation,
formation, chronic disability, amyloidosis
inflammation, healing by
fibrosis
Main immunoglobulin IgM IgG
Peripheral blood leukocyte Neutrophilic leukocytosis Monocytosis
response
Win 21 A Jalan 46
Self Study
Win 21 A Jalan 47
Win 21 Acute Inflammation
A Jalan 48
Multiple tuberculous granulomas
Win 21 A Jalan 49
Granuloma
Langhans giant cell

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Two spherules of Coccidioides immitis
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Cat scratch disease
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Lymphatics and lymphnodes in inflammation
• Filter the Extravascular fluid

• Represent a secondary line of defense.
• Lymphatics:
– Help to drain edema fluid during inflammation
– Can get inflamed : lymphangitis
• Lymphnodes:
– May get inflamed = lymphadenitis.
– May get enlarged = lymphadenopathy
– If fail to filter the microbes à circulation à
bacteremia.
Win 21 A Jalan 53
• Critical to the survival of an organism is the ability to repair the damage
caused by toxic insults and inflammation.
• The inflammatory response to microbes and injured tissues not only serves to
eliminate these dangers but also sets into motion the process of repair.
• Repair, sometimes called healing, refers to the restoration of tissue
architecture and function after an injury.
• It occurs by two types of reactions:
• regeneration of the injured tissue and
• scar formation by the deposition of connective tissue
Inflammation also sets into motion the process of repair.
Remember if it were not for inflammation- our wounds would not heal!
Mechanisms of tissue repair: regeneration and scar formation.
• After mild injury, which damages the epithelium but not the underlying tissue,
resolution occurs by regeneration, but
• after more severe injury with damage to the connective tissue, repair is by
scar formation.
Regeneration.
• Some tissues are able to replace the damaged cells and essentially
return to a normal state; this process is called regeneration.
• Regeneration occurs by proliferation of residual (uninjured) cells that
retain the capacity to divide, and by replacement from tissue stem cells.
• It is the typical response to injury in the rapidly dividing epithelia of the
skin and intestines, and some parenchymal organs, notably the liver.
Labile cells:
divide actively during life to replace lost cells
are capable of regeneration after injury
include cells of the epidermis and gastrointestinal mucosa, cells
lining the surface of the genitourinary tract and hematopoietic cells of bone
marrow.
Stable cells
characteristically undergo few divisions but are capable of division
when activated
are capable of regeneration following injury
include hepatocytes, renal tubular cells, parenchymal cells of many
organs and numerous mesenchymal cells (e.g. smooth muscle, cartilage,
connective tissue, endothelium, osteoblasts)
Permanent cells
are incapable of division and regeneration
includes neurons and myocardial cells
are replaced by scar tissue (typically fibrosis ; gliosis in CNS) after
irreversible cell injury and cell loss.
In 3rd degree burn: there is loss of skin, basement membrane and connective
tissue infrastructure and hence it can not be restored to normal. Healing then
occurs by replacement of damaged tissue by fibrous tissue or scar tissue.
Scar formation.
• If the injured tissues are incapable of regeneration, or if the supporting
structures of the tissue are severely damaged, repair occurs by the laying
down of connective (fibrous) tissue, a process that results in scar formation.
• Although the fibrous scar cannot perform the function of lost parenchymal
cells, it provides enough structural stability that the injured tissue is usually
able to function.
• The term fibrosis is most often used to describe the extensive deposition of
collagen that occurs in the lungs, liver, kidney, and other organs as a
consequence of chronic inflammation, or in the myocardium after extensive
ischemic necrosis (infarction).
• If fibrosis develops in a tissue space occupied by an inflammatory exudate, it
is called organization (as in organizing pneumonia affecting the lung).
• Note:
• After many common types of injury, both regeneration and scar formation
contribute in varying degrees to the ultimate repair.
• Both processes involve the proliferation of various cells and close interactions
between cells and the ECM.
Repair by Scar Formation
•Tissues can be repaired by regeneration with complete restoration of form and
function, or by replacement with connective tissue and scar formation.
•Repair by connective tissue deposition involves angiogenesis, migration and
proliferation of fibroblasts, collagen synthesis, and connective tissue remodeling.
•Repair by connective tissue starts with the formation of granulation tissue and
culminates in the laying down of fibrous tissue.
•Multiple growth factors stimulate the proliferation of the cell types involved in
repair.
•TGF-β is a potent fibrogenic agent; ECM deposition depends on the balance
among fibrogenic agents, the metalloproteinases (MMPs) that digest ECM, and
the TIMPs.
This cross section through the heart demonstrates the left ventricle on the left.
Extending from the anterior portion and into the septum is a large recent
myocardial infarction. The center is tan with surrounding hyperemia. The
infarction is "transmural" in that it extends through the full thickness of the wall.
Microscopic features of MI
Dense polymorphonuclear leukocytic infiltrate in area of 2- to 3-day-old MI.
Granulation tissue gross appearance: Reddish and granular
Do not confuse with Granuloma or granulomatous disease
Hand, healing by secondary intention - Clinical presentation Wounds with
a large tissue defect heal by secondary intention. Note the red granular
appearance of the granulation tissue. A whitish-greenish-yellow neutrophilic
exudate represents an inflammatory response to bacterial invasion of the
wound.
Granulation tissue : microscopy
Circle: Fibroblasts
Arrows: new blood vessels
Definition: Granulation tissue is a type of highly vascular tissue that is composed of
blood vessels and activated fibroblasts.
Blood vessels derive from preexisting blood vessels and de novo from EC precursors
recruited from the bone marrow (i.e., angiogenesis).
Important growth factors in angiogenesis include vascular endothelial cell growth factor
(VEGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), platelet-
derived growth factor (PDGF), and TGF-β.
Vascularization is essential for normal wound healing.
Granulation tissue is the precursor of scar tissue.
Granulation tissue is essential for repair by connective tissue.

Angiogenesis;
Critical for WOUND healing, tumor growth and development of collateral circulation.
Two processes
Vasculogenesis
primitive blood vessel formation during embryonic
development from angioblasts
Angiogenesis or neovascularization
Formation of new blood vessels from preexisting vessels.
Enhanced by
fibronectin and
basic fibroblast growth factor. (BFGF)
Vascular endothelial growth factor (VEGF)
Fibroblast migration and proliferation

Most important GROWTH FACTOR is TGF-β
Enzyme: Lysyl oxidase necessary for cross linking of collagen fibrils.
Copper is a cofactor on lysyl oxidase. deficiency of this metal results in
poor wound healing.
Hydroxylation of proline and lysine (necessary for collagen synthesis)
requires presence of vitamin C.
Vitamin C deficiency is associated with poor wound healing.
Remember: type IV collagen is found in basement membrane.

Metalloproteinases (MMPs); Zn necessary for their activity
Different types: Interstitial collagenases, Gelatinases, Stromelysins etc.
Degrade collagen, fibronectin etc.
Produced by: fibroblasts, macrophages, neutrophils, synovial cells etc.
Inhibition of secretion: TGF-β and steroids.
Inactivated by TIMPS (tissue inhibitors of metalloproteases) that prevent
uncontrolled tissue degradation
Healing of Skin Wounds
Cutaneous wound healing is a process that involves both epithelial regeneration
and the formation of connective tissue scar.
Depending on the nature and size of the wound, the healing of skin wounds is
said to occur by first or second intention.
Major differences between healing by primary intention and secondary intention:
Primary intention: wound edges are apposed

Secondary intention:
wound is left open
Myofibroblasts are important in closing wound
takes much longer to heal
Healing by First Intention
•One of the simplest examples of wound repair is the healing of a clean,
uninfected surgical incision approximated by surgical sutures.
•This is referred to as primary union, or healing by first intention.
•The incision causes only focal disruption of epithelial basement membrane
continuity and death of relatively few epithelial and connective tissue cells.
•As a result, epithelial regeneration is the principal mechanism of repair.
•A small scar is formed, but there is minimal wound contraction.
•The narrow incisional space first fills with fibrin-clotted blood, which then is
rapidly invaded by granulation tissue and covered by new epithelium.
•The steps in the process are well defined:
•Within 24 hours, neutrophils are seen at the incision margin, migrating toward
the fibrin clot. Basal cells at the cut edge of the epidermis begin to show
increased mitotic activity. Within 24 to 48 hours, epithelial cells from both edges
have begun to migrate and proliferate along the dermis, depositing basement
membrane components as they progress. The cells meet in the midline beneath
the surface scab, yielding a thin but continuous epithelial layer.
•By day 3, neutrophils have been largely replaced by macrophages, and
granulation tissue progressively invades the incision space. Collagen fibers are
now evident at the incision margins, but these are vertically oriented and do not
bridge the incision. Epithelial cell proliferation continues, yielding a thickened
epidermal covering layer.
•By day 5, neovascularization reaches its peak as granulation tissue fills the
incisional space. Collagen fibrils become more abundant and begin to bridge the
incision. The epidermis recovers its normal thickness as differentiation of surface
cells yields a mature epidermal architecture with surface keratinization.
•During the second week, there is continued collagen accumulation and
fibroblast proliferation. The leukocyte infiltrate, edema, and increased vascularity
are substantially diminished. The long process of “blanching” begins,
accomplished by increasing collagen deposition within the incisional scar and the
regression of vascular channels.
•By the end of the first month, the scar consists of a cellular connective tissue,
largely devoid of inflammatory cells, covered by an essentially normal epidermis.
However, the dermal appendages destroyed in the line of the incision are
permanently lost. The tensile strength of the wound increases with time.
Steps in wound healing by first intention.
Remember fibronectin also acts as an Opsonizing agent.
What are the other opsonising agents??
Answer : IgG and C3b
Fibronectin derived from: macrophages/fibroblasts/endothelial
cells.
Collagenase important in remodeling (zinc is a cofactor). Zinc deficiency
causes poor wound healing.
Metalloproteinases (collagenases containing zinc) replace type III collagen

with type I collagen (strong collagen), which increases the tensile strength
of the wound to ≈70% to 80% of the original after ≈3 months. Scar tissue
after 3 months is primarily composed of acellular connective tissue that is
devoid of inflammatory cells and adnexal structures and is surfaced by an
intact epidermis.
Important collagen types:

Type I : high tensile strength
bone, skin, tendons
Type II: cartilage
Type III: initial type in wound repair
Type IV: Basement membrane
Type X: epiphyseal plate
Healing by Second Intention
•When cell or tissue loss is more extensive, such as in large wounds, at sites of
abscess formation, ulceration, and ischemic necrosis (infarction) in parenchymal
organs, the repair process is more complex and involves a combination of
regeneration and scarring.
•In second intention healing of skin wounds, also known as healing by
secondary union, the inflammatory reaction is more intense, and there is
development of abundant granulation tissue, with accumulation of ECM and
formation of a large scar, followed by wound contraction mediated by the action
of myofibroblasts.
Steps in wound healing by second intention .
Note the large amount of granulation tissue and wound contraction.
Wound contraction is a feature of healing by second intention. The cells
involved are the myofibroblasts.
Secondary healing differs from primary healing in several respects:

• A larger clot or scab rich in fibrin and fibronectin forms at the surface of the
wound.
• Inflammation is more intense because large tissue defects have a greater
volume of necrotic debris, exudate, and fibrin that must be removed.
• Larger defects require a greater volume of granulation tissue to fill in the gaps
and provide the underlying framework for the regrowth of tissue epithelium.
• A greater volume of granulation tissue generally results in a greater mass of
scar tissue.
• Secondary healing involves wound contraction.
• Within 6 weeks, for example, large skin defects may be reduced to 5% to 10%
of their original size, largely by contraction.
• This process has been ascribed to the presence of myofibroblasts, which are
modified fibroblasts exhibiting many of the ultrastructural and functional
features of contractile smooth muscle cells.
wound.
Hand, wound healed by secondary intention, same case as previous
image, eight months later - Clinical presentation As the granulation tissue
matures, the myofibroblasts contract to close the wound. They then become
mature fibroblasts, which produce collagen, forming a fibrous scar. The
contraction that takes place can lead to decreased mobility.
Wound Strength
• Carefully sutured wounds have approximately 70% of the strength of normal
skin, largely because of the placement of sutures.
• When sutures are removed, usually at 1 week, wound strength is
approximately 10% of that of unwounded skin, but this increases rapidly over
the next 4 weeks.
• The recovery of tensile strength results from collagen synthesis exceeding
degradation during the first 2 months, and from structural modifications of
collagen (e.g., cross-linking, increased fiber size) when synthesis declines at
later times.
• Wound strength reaches approximately 70% to 80% of normal by 3 months
and usually does not improve substantially beyond that point.
FACTORS THAT INFLUENCE TISSUE REPAIR
•Tissue repair may be altered by a variety of influences.
•Variables that modify healing may be extrinsic (e.g., infection) or intrinsic to the
injured tissue. Particularly important are infections and diabetes.
•Infection is clinically the most important cause of delay in healing; it prolongs
inflammation and potentially increases the local tissue injury.
•Nutrition has profound effects on repair; protein deficiency, for example, and
especially vitamin C deficiency inhibit collagen synthesis and retard healing.
•Glucocorticoids (steroids) have well-documented anti-inflammatory effects, and
their administration may result in weakness of the scar because of inhibition of
TGF-β production and diminished fibrosis. In some instances, however, the anti-
inflammatory effects of glucocorticoids are desirable. For example, in corneal
infections, glucocorticoids are sometimes prescribed (along with antibiotics) to
reduce the likelihood of opacity that may result from collagen deposition.
•Mechanical variables such as increased local pressure or torsion may cause
wounds to pull apart, or dehisce.
•Poor perfusion, due either to arteriosclerosis and diabetes or to obstructed
venous drainage (e.g., in varicose veins), also impairs healing.
•Foreign bodies such as fragments of steel, glass, or even bone impede healing.
•The type and extent of tissue injury affects the subsequent repair. Complete
restoration can occur only in tissues composed of stable and labile cells; injury to
tissues composed of permanent cells must inevitably result in scarring, as in
healing of a myocardial infarct.
•The location of the injury and the character of the tissue in which the injury
occurs are also important. For example, inflammation arising in tissue spaces
(e.g., pleural, peritoneal, or synovial cavities) develops extensive exudates.
Subsequent repair may occur by digestion of the exudate, initiated by the
proteolytic enzymes of leukocytes and resorption of the liquefied exudate. This is
called resolution, and generally, in the absence of cellular necrosis, normal tissue
architecture is restored. In the setting of larger accumulations, however, the
exudate undergoes organization: Granulation tissue grows into the exudate, and
a fibrous scar ultimately forms.
Most common infection that delays wound healing is by Staphylococcus
aureus.
Vitamin C:
responsible for hydroxylation of proline and lysine.
Deficiency of vitamin C (scurvy) produces structurally weak collagen à
poor wound healing.
Copper:
cofator on lysyl oxidase which is responsible for cross
linking of collagen fibrils. Deficiency of this element responsible for poor
wound healing and predisposes to dissecting aortic aneurysm (since
elastic tissue is also weak)
Glucocorticoids :
-occasionally used with antibiotics to prevent scar tissue
formation (e.g. bacterial meningitis).
-inhibit wound healing by impairing collagen synthesis.
Glucocorticoids occasionaly used with antibiotics to prevent scar formation ( e.g.
bacterial meningitis).
Glucocorticoids
Glucocorticoids: prevent scar formation
a.Interfere with collagen formation and decrease tensile strength
b.Useful clinically in preventing excessive scar formation
(1)Dexamethasone is used along with antibiotics to prevent scar formation in
bacterial meningitis.
(2)Plastic surgeons inject high potency steroids into wounds to prevent
excessive scar tissue formation.
Aberrations of cell growth and ECM production may occur even in what begins
as normal wound healing. For example, the accumulation of exuberant amounts
of collagen can give rise to prominent, raised scars known as keloids. There
appears to be a heritable predisposition to keloid formation, and the condition is
more common in African-Americans. Healing wounds may also generate
exuberant granulation tissue that protrudes above the level of the surrounding
skin and hinders re-epithelialization. Such tissue is called “proud flesh” in old
medical parlance, and restoration of epithelial continuity requires cautery or
surgical resection of the granulation tissue.
Abdomen, poor wound healing, dehisced surgical wound - Clinical
presentation Wound dehiscence is the disruption of apposed surfaces of a
wound (most often abdominal), due to inadequate healing. Many factors, such
as infection, inadequate blood supply, diabetes, and mechanical stress can
impair healing.
Definition: Keloids are raised scars that grow beyond the borders of the original
wound.
Develop in 15% to 20% of African Americans, Asians, and Hispanics, suggesting
a genetic predisposition.
Often refractory to medical and surgical intervention.
Definition: Hypertrophic scars are raised scars that remain within the confines of
the original wound. Frequently regress spontaneously.
Normal scars have collagen bundles that are randomly arrayed (not all in the
same direction), whereas keloids and hypertrophic scars have stretched
collagen bundles arranged in the same plane as the epidermis.
Head and neck, keloids - Clinical presentation Keloids, nodular masses of
hyperplastic scar tissue, occur when the wound healing process runs
unchecked. They are more common in people of African descent. Surgical
excision typically leads to repeated keloid formation.
Keloid microscopy
Note hyperextensible skin and extremely hypermobile joint
Cell regeneration depends on factors that stimulate parenchymal cell division
and migration.
Stimulatory factors include loss of tissue and production of growth factors
C
C
A
C
Repair in other tissues
1.Liver
a.Mild injury (e.g., hepatitis A)
•Regeneration of hepatocytes with restoration to normal is possible if the
cytoarchitecture is intact.
b.Severe or persistent injury (e.g., hepatitis C)
(1)Regenerative nodules develop that show twinning of liver cell plates (two cells
thick); a double line of hepatocytes is present, and nuclei seem to run in parallel.
(2)Portal triads are not present in regenerative nodules.
(3)Increased fibrosis occurs around the regenerative nodules, which leads to
cirrhosis of the liver if the injurious agent is not removed.
2.Lung
a.Type II pneumocytes are the key repair cells of the lung.
b.Replace damaged type I and type II pneumocytes and synthesize surfactant.
3.Brain
a.Astrocytes proliferate in response to an injury (e.g., brain infarction).
•Proliferation of astrocytes is called gliosis.
b.Microglial cells (macrophages) are scavenger cells that remove debris (e.g.,
myelin).
4.Peripheral nerve transection

a.Without innervation, muscle atrophies in ~15 days.
b.After nerve transection, there is distal degeneration of the axon and myelin
sheath and proximal axonal degeneration up to the next node of Ranvier.
Wallerian degeneration: distal degeneration of the axon
(1)Macrophages and Schwann cells phagocytose axonal/myelin debris.
(2)Nerve cell body undergoes central chromatolysis.
(a)Nerve cell body swells.
(b)Nissl bodies (composed of rough endoplasmic reticulum and free ribosomes)
disappear centrally, and the nucleus moves to the periphery.
(3)Schwann cells proliferate in the distal stump.
Peripheral nerve transection: Schwann cell is key in reinnervation
(4)Axonal sprouts develop in the proximal stump and extend distally using the
Schwann cells for guidance.
(5)Regenerated axon grows 2 to 3 mm/day.
(6)Axon becomes remyelinated.
(7)Muscle is eventually reinnervated.
5.Heart
a.Cardiac muscle is permanent tissue.
b.Damaged muscle is replaced by noncontractile scar tissue.
6.Skeletal muscle postexercise

a.After exercise, there is damage to the sarcomeres in skeletal muscle.
•A sarcomere is the basic unit of a muscle and gives skeletal muscle its striated
appearance.
b.Satellite cells are stem cells that repair and form new myofibers in sarcomeres
that have been damaged by mechanical strain.
Skeletal muscle postexercise: myofiber repair by satellite cells (muscle stem
cells)
Amyloid: fibrillar protein; deposited in interstitial tissue; pressure atrophy
Matthias Schleiden a German Botanist was the first to use the term “amyloid” in
botany, for starch, referring to“starch-like”.
German pathologist Rudolf Virchow (1804 – 1881) applied it in the medical
literature in 1854.
He used the word “Amyloid” to describe some deposits in nervous system which
showed the same color reaction with iodine and sulfuric acid, i.e. a change from
brown to blue, typical to starch
In the later years it was found that the chemical nature was unrelated to starch
but proteins!
Yet.. The name “amyloid” still remains Probably because of Virchow was
considered as one of the most valued scientists of his time In the later years it
was found that the chemical nature was unrelated to starch but proteins!
Cross beta pleated sheet pattern is responsible for Congo red staining
Electron microscopy:
Linear, non-branching fiber with hollow cores.
Amyloid: linear filament; β-pleated sheet
Amyloid: Congo red +; apple green birefringence when polarized

Amyloidosis of the kidney.
The glomerular architecture is almost totally obliterated by the massive
accumulation of amyloid.
Cardiac amyloidosis.
The atrophic myocardial fibers are separated by structureless, pink-
staining amyloid (arrows).
Amyloidosis:
A:A section of the liver stained with Congo red reveals pink-red
deposits of amyloid in the walls of blood vessels and along sinusoids.
B:Note the yellow-green birefringence of the deposits when observed by
polarizing microscope.
Derived from three major precursor proteins:
a.Immunoglobulin light chains, with λ light chains more frequently involved than κ
light chains
•Light chains in urine are called Bence Jones proteins.
b.Serum amyloid A (SAA) protein, which is an acute phase reactant synthesized
and released by the liver in inflammation
c.Amyloid precursor protein (APP); gene located on chromosome 21
Major precursor proteins: λ light chains, SAA, APP (beta amyloid precursor
protein)
Other important precursor proteins include:

a.Transthyretin, which is a normal carrier protein for thyroxine and retinoic acid
(vitamin A)
b.β2-Microglobulin, which is the light chain component of the MHC
c.Prion proteins, which normally maintain neuronal membranes
The basic mechanism in the pathogenesis of amyloidosis is abnormal folding
of proteins or misfolded proteins.
Under normal circumanstances, these abnormal or misfolded proteins are
degraded by proteasome pathway intracellularly and by the macrophages
extracellularly.
In Amyloidosis, these control mechanisms fail or there may be mutations which
favor misfolding which further leads to accumulation and aggregation to form
fibrils.
So, the pathogenesis can be broadly categorized into two mechanisms.
Normal proteins, when produced in abnormal numbers.
Production of normal amounts of mutant proteins
This is explained in the illustration above.
Immunocyte dyscrasias (primary amyloidosis)
Plasma cell disorders (e.g., multiple myeloma, other monoclonal plasma cell
dyscrasias [10% of all monoclonal gammopathies]
AL (designation for amyloid derived from immunoglobulin light chains, particularly
λ light chains)
Reactive systemic amyloidosis (secondary amyloidosis)

Chronic inflammation: rheumatoid arthritis (MC), ankylosing spondylitis,
inflammatory bowel disease (Crohn disease, ulcerative colitis), tuberculosis,
leprosy, osteomyelitis, renal cell carcinoma, Hodgkin lymphoma, heroin abusers
(“skin popping”)
AA (designation for amyloid derived from serum amyloid A protein)
Aβ2m (designation for amyloid derived from β2-microglobulin). Occurs in
patients with chronic renal failure who are on dialysis.
Hemodialysis- associated amyloidosis:

Amyloid derived from β2-microglobulin
Type of amyloid: Aβ2M
May cause carpal tunnel syndrome and joint disease.
Aβ (designation for amyloid derived from amyloid precursor protein, which is
coded for by chromosome 21)
CNS:
Alzheimer’s disease
Β amyloid protein coded by chromosome 21.
Amyloid toxic to neurons
MC COD in Down’s syndrome patients >40 years
Familial Mediterranean fever
Autosomal recessive
Increased production of IL-1
Fever, inflammation of serosal membranes (pleura, peritoneum, synovium)
AA (designation for amyloid derived from serum amyloid protein)
Type of amyloid: AA (= amyloid derived from serum amyloid

associated protein- SAA)
High SAA levels à deposits as AA amyloid
Wide spread amyloidosis.
Kidney involvement
a.Most common overall organ involved
b.Glomeruli, interstitial tissue, arteries, and arterioles are all involved.
c.Proteinuria in the nephrotic range leads to generalized pitting edema and cavity
effusions.
• Kidney: proteinuria with nephrotic syndrome
•Cardiac involvement
•A.Restrictive cardiopathy is present because of infiltration of amyloid between
myocardial fibers.
•(1)Ejection fraction is frequently preserved.
•(2)Produces a diastolic dysfunction type of left-sided heart failure.
•B.Conduction defects are very common.
• Heart: restrictive cardiomyopathy, diastolic dysfunction LHF, conduction
defects
•Gastrointestinal involvement
fat.
•B.Macroglossia (enlarged tongue) leads to problems with speech and
swallowing.
• Gastrointestinal: malabsorption, macroglossia
•Liver involvement
•A.Hepatomegaly is a common finding in systemic amyloidosis.
•B.Pressure atrophy of hepatocytes; however, functional impairment is
uncommon
• Liver: hepatomegaly, functional impairment uncommon
•Spleen involvement
•A.Common in the systemic type of amyloidosis
•B.If white pulp (splenic lymphoid follicles) is involved, the splenic surface looks
like it is impregnated with grains of sand (called a sago spleen).
•C.If red pulp is involved, the splenic surface has a waxy appearance (called a
lardaceous spleen).
• Spleen: splenomegaly; white pulp sago spleen, red pulp lardaceous
spleen
•In hemodialysis-associated amyloidosis, musculoskeletal involvement is

common.
••Clinical findings include carpal tunnel syndrome, destructive arthropathy, bone
cysts, and fractures.
• Hemodialysis-associated amyloidosis: carpal tunnel syndrome
•Nervous system involvement

•Dementia (Alzheimer disease), peripheral neuropathies (paresthesias, muscle
weakness), and disabling autonomic neuropathies may occur.
Carpal tunnel syndrome:

formed by the carpal bones and flexor retinaculum
Immunoelectrophoresis
to detect light chains in primary amyloidosis.
Kidney involvement
a.Most common overall organ involved
b.Glomeruli, interstitial tissue, arteries, and arterioles are all involved.
c.Proteinuria in the nephrotic range leads to generalized pitting edema and cavity
effusions.
•Kidney: proteinuria with nephrotic syndrome
•Cardiac involvement
•A.Restrictive cardiopathy is present because of infiltration of amyloid between
myocardial fibers.
•(1)Ejection fraction is frequently preserved.
•(2)Produces a diastolic dysfunction type of left-sided heart failure.
•B.Conduction defects are very common.
•Gastrointestinal involvement
•A.Diarrhea is of the malabsorptive type with loss of carbohydrates, proteins, and
fat.
•B.Macroglossia (enlarged tongue) leads to problems with speech and
swallowing.
•Gastrointestinal: malabsorption, macroglossia
•Liver involvement
•A.Hepatomegaly is a common finding in systemic amyloidosis.
•B.Pressure atrophy of hepatocytes; however, functional impairment is
uncommon
•Liver: hepatomegaly, functional impairment uncommon
•Spleen involvement
•A.Common in the systemic type of amyloidosis
•B.If white pulp (splenic lymphoid follicles) is involved, the splenic surface looks
like it is impregnated with grains of sand (called a sago spleen).
•C.If red pulp is involved, the splenic surface has a waxy appearance (called a
lardaceous spleen).
•Spleen: splenomegaly; white pulp sago spleen, red pulp lardaceous
spleen
•In hemodialysis-associated amyloidosis, musculoskeletal involvement is

common.
••Clinical findings include carpal tunnel syndrome, destructive arthropathy, bone
cysts, and fractures.
•Hemodialysis-associated amyloidosis: carpal tunnel syndrome
•Nervous system involvement

•Dementia (Alzheimer disease), peripheral neuropathies (paresthesias, muscle
weakness), and disabling autonomic neuropathies may occur.
Hyperemia and congestion both stem from increased blood volumes within tissues, but
have different underlying mechanisms and consequences.
• Hyperemia is an active process in which arteriolar dilation (e.g., at sites of
inflammation or in skeletal muscle during exercise) leads to increased blood flow.
• Affected tissues turn red (erythema) because of increased delivery of
oxygenated blood.
• Congestion is a passive process resulting from reduced outflow of blood from a
tissue.
• It can be systemic, as in cardiac failure, or localized, as in isolated venous
obstruction.
• As a result of increased hydrostatic pressures, congestion commonly leads to
edema.
• In long-standing chronic passive congestion, the associated chronic hypoxia
may result in ischemic tissue injury and scarring.
• In chronically congested tissues, capillary rupture can also produce small
hemorrhagic foci; subsequent catabolism of extravasated red cells can leave
residual telltale clusters of hemosiderin-laden macrophages.
Hyperemia versus congestion.
In both cases there is an increased volume and pressure of blood in a given tissue with
associated capillary dilation and a potential for fluid extravasation. In hyperemia,
increased inflow leads to engorgement with oxygenated blood, resulting in erythema.
In congestion, diminished outflow leads to a capillary bed swollen with deoxygenated
venous blood and resulting in cyanosis.
Morphology.
• The cut surfaces of congested tissues are often discolored due to the presence of
high levels of poorly oxygenated blood.
• Microscopically, acute pulmonary congestion exhibits engorged alveolar capillaries
often with alveolar septal edema and focal intra-alveolar hemorrhage.
• In chronic pulmonary congestion the septa are thickened and fibrotic, and the alveoli
often contain numerous hemosiderin-laden macrophages called heart failure cells.
• In acute hepatic congestion, the central vein and sinusoids are distended;
centrilobular hepatocytes can be frankly ischemic while the periportal hepatocytes—
better oxygenated because of proximity to hepatic arterioles—may only develop
fatty change.
• In chronic passive hepatic congestion the centrilobular regions are grossly red-brown
and slightly depressed (because of cell death) and are accentuated against the
surrounding zones of uncongested tan liver (nutmeg
• Microscopically, there is centrilobular hemorrhage, hemosiderin-laden macrophages,
and degeneration of hepatocytes.
• Because the centrilobular area is at the distal end of the blood supply to the liver, it
is prone to undergo necrosis whenever the blood supply is compromised.
RHF= right heart failure
LHF= left heart failure
What is the brown pigment that is derived from hemoglobin?
Hemosiderin
The alveolar septa appear thickened in some areas by fibrous connective tissue (red
arrows), and the alveolar spaces contain numerous hemosiderin-laden macrophages
(black arrows) . A few alveolar spaces contain pink edema fluid.
Here is an example of a "nutmeg" liver seen with chronic passive congestion of the
liver. Note the dark red congested regions that represent accumulation of RBC's in
centrilobular regions.
Microscopically, the nutmeg pattern results from congestion around the central veins.
There is also atrophy & disorganization of the hepatic cords surrounding the central
vein (V).
This is usually due to a "right sided" heart failure.
Centrilobular necrosis with degenerating hepatocytes and hemorrhage.
Causes of Hemorrhage:
trauma, Erosive cancer, atherosclerosis,
hypertension à retinal and Cerebral hemorrhage à blindness and ~
death respectively
bleeding diathesis = increased bleeding tendency
seen in scurvy, thrombocytopenia and deficiency of
clotting factor
A: Petechial hemorrhages – Brain
Petechiae (pinpoint hemorrhages) represent bleeding from small
vessels and are classically found when a coagulopathy is due to a low platelet count.
They can also appear following sudden hypoxia.
B: The skin has numerous purpural hemorrhages. In this patient the lesions are
caused by Neisseria meningitidis.
What is purpura fulminans??
C: The blotchy areas of hemorrhage in the skin are called ecchymoses (singular
ecchymosis), or also as areas of purpura.
Ecchymoses are larger than petechiae. They can appear with coagulation disorders.
D:A localized collection of blood outside the vascular system within tissues is known
as a hematoma.
Here is a small hematoma under the toenail following trauma, which has a bluish
appearance from the deoxygenated blood within it.
A:Punctate petechial hemorrhages of the colonic mucosa, a consequence of
thrombocytopenia.
B, Fatal intracerebral hemorrhage. Even relatively inconsequential volumes of
hemorrhage in a critical location, or into a closed space (such as the cranium), can have
fatal outcomes.
• Critical to the survival of an organism is the ability to repair the damage
caused by toxic insults and inflammation.
• The inflammatory response to microbes and injured tissues not only serves to
eliminate these dangers but also sets into motion the process of repair.
• Repair, sometimes called healing, refers to the restoration of tissue
architecture and function after an injury.
• It occurs by two types of reactions:
• regeneration of the injured tissue and
• scar formation by the deposition of connective tissue
Inflammation also sets into motion the process of repair.
Remember if it were not for inflammation- our wounds would not heal!
Mechanisms of tissue repair: regeneration and scar formation.
• After mild injury, which damages the epithelium but not the underlying tissue,
resolution occurs by regeneration, but
• after more severe injury with damage to the connective tissue, repair is by
scar formation.
Regeneration.
• Some tissues are able to replace the damaged cells and essentially
return to a normal state; this process is called regeneration.
• Regeneration occurs by proliferation of residual (uninjured) cells that
retain the capacity to divide, and by replacement from tissue stem cells.
• It is the typical response to injury in the rapidly dividing epithelia of the
skin and intestines, and some parenchymal organs, notably the liver.
Labile cells:
divide actively during life to replace lost cells
are capable of regeneration after injury
include cells of the epidermis and gastrointestinal mucosa, cells
lining the surface of the genitourinary tract and hematopoietic cells of bone
marrow.
Stable cells
characteristically undergo few divisions but are capable of division
when activated
include hepatocytes, renal tubular cells, parenchymal cells of many
organs and numerous mesenchymal cells (e.g. smooth muscle, cartilage,
connective tissue, endothelium, osteoblasts)
Permanent cells
are incapable of division and regeneration
includes neurons and myocardial cells
are replaced by scar tissue (typically fibrosis ; gliosis in CNS) after
irreversible cell injury and cell loss.
In 3rd degree burn: there is loss of skin, basement membrane and connective
tissue infrastructure and hence it can not be restored to normal. Healing then
occurs by replacement of damaged tissue by fibrous tissue or scar tissue.
Scar formation.
• If the injured tissues are incapable of regeneration, or if the supporting
structures of the tissue are severely damaged, repair occurs by the laying
down of connective (fibrous) tissue, a process that results in scar formation.
• Although the fibrous scar cannot perform the function of lost parenchymal
cells, it provides enough structural stability that the injured tissue is usually
able to function.
• The term fibrosis is most often used to describe the extensive deposition of
collagen that occurs in the lungs, liver, kidney, and other organs as a
consequence of chronic inflammation, or in the myocardium after extensive
ischemic necrosis (infarction).
• If fibrosis develops in a tissue space occupied by an inflammatory exudate, it
is called organization (as in organizing pneumonia affecting the lung).
• After many common types of injury, both regeneration and scar formation
contribute in varying degrees to the ultimate repair.
• Both processes involve the proliferation of various cells and close interactions
between cells and the ECM.
Repair by Scar Formation
•Tissues can be repaired by regeneration with complete restoration of form and
function, or by replacement with connective tissue and scar formation.
•Repair by connective tissue deposition involves angiogenesis, migration and
proliferation of fibroblasts, collagen synthesis, and connective tissue remodeling.
•Repair by connective tissue starts with the formation of granulation tissue and
culminates in the laying down of fibrous tissue.
•Multiple growth factors stimulate the proliferation of the cell types involved in
repair.
•TGF-β is a potent fibrogenic agent; ECM deposition depends on the balance
among fibrogenic agents, the metalloproteinases (MMPs) that digest ECM, and
the TIMPs.
This cross section through the heart demonstrates the left ventricle on the left.
Extending from the anterior portion and into the septum is a large recent
myocardial infarction. The center is tan with surrounding hyperemia. The
infarction is "transmural" in that it extends through the full thickness of the wall.
Microscopic features of MI
Dense polymorphonuclear leukocytic infiltrate in area of 2- to 3-day-old MI.
Granulation tissue gross appearance: Reddish and granular
Do not confuse with Granuloma or granulomatous disease
wound.
Granulation tissue : microscopy
Circle: Fibroblasts
Arrows: new blood vessels
Definition: Granulation tissue is a type of highly vascular tissue that is composed of
blood vessels and activated fibroblasts.
Blood vessels derive from preexisting blood vessels and de novo from EC precursors
recruited from the bone marrow (i.e., angiogenesis).
Important growth factors in angiogenesis include vascular endothelial cell growth factor
(VEGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), platelet-
derived growth factor (PDGF), and TGF-β.
Vascularization is essential for normal wound healing.
Granulation tissue is the precursor of scar tissue.
Granulation tissue is essential for repair by connective tissue.

Angiogenesis;
Critical for WOUND healing, tumor growth and development of collateral circulation.
Two processes
Vasculogenesis
primitive blood vessel formation during embryonic
development from angioblasts
Angiogenesis or neovascularization
Formation of new blood vessels from preexisting vessels.
basic fibroblast growth factor. (BFGF)
Vascular endothelial growth factor (VEGF)
Fibroblast migration and proliferation

Most important GROWTH FACTOR is TGF-β
Enzyme: Lysyl oxidase necessary for cross linking of collagen fibrils.
Copper is a cofactor on lysyl oxidase. deficiency of this metal results in
poor wound healing.
Hydroxylation of proline and lysine (necessary for collagen synthesis)
requires presence of vitamin C.
Vitamin C deficiency is associated with poor wound healing.
Remember: type IV collagen is found in basement membrane.

Metalloproteinases (MMPs); Zn necessary for their activity
Different types: Interstitial collagenases, Gelatinases, Stromelysins etc.
Degrade collagen, fibronectin etc.
Produced by: fibroblasts, macrophages, neutrophils, synovial cells etc.
Inhibition of secretion: TGF-β and steroids.
Inactivated by TIMPS (tissue inhibitors of metalloproteases) that prevent
uncontrolled tissue degradation
Healing of Skin Wounds
Cutaneous wound healing is a process that involves both epithelial regeneration
and the formation of connective tissue scar.
Depending on the nature and size of the wound, the healing of skin wounds is
said to occur by first or second intention.
Major differences between healing by primary intention and secondary intention:
Primary intention: wound edges are apposed

Secondary intention:
wound is left open
Myofibroblasts are important in closing wound
takes much longer to heal
Steps in wound healing by first intention.
Remember fibronectin also acts as an Opsonizing agent.
What are the other opsonising agents??
Answer : IgG and C3b
Fibronectin derived from: macrophages/fibroblasts/endothelial
cells.
Collagenase important in remodeling (zinc is a cofactor). Zinc deficiency
causes poor wound healing.
Metalloproteinases (collagenases containing zinc) replace type III collagen

with type I collagen (strong collagen), which increases the tensile strength
of the wound to ≈70% to 80% of the original after ≈3 months. Scar tissue
after 3 months is primarily composed of acellular connective tissue that is
devoid of inflammatory cells and adnexal structures and is surfaced by an
intact epidermis.
Important collagen types:

Type I : high tensile strength
bone, skin, tendons
Type II: cartilage
Type III: initial type in wound repair
Type IV: Basement membrane
Type X: epiphyseal plate
Healing by Second Intention
•When cell or tissue loss is more extensive, such as in large wounds, at sites of
abscess formation, ulceration, and ischemic necrosis (infarction) in parenchymal
organs, the repair process is more complex and involves a combination of
regeneration and scarring.
•In second intention healing of skin wounds, also known as healing by
secondary union, the inflammatory reaction is more intense, and there is
development of abundant granulation tissue, with accumulation of ECM and
formation of a large scar, followed by wound contraction mediated by the action
of myofibroblasts.
Steps in wound healing by second intention .
Note the large amount of granulation tissue and wound contraction.
Wound contraction is a feature of healing by second intention. The cells
involved are the myofibroblasts.
Secondary healing differs from primary healing in several respects:

• A larger clot or scab rich in fibrin and fibronectin forms at the surface of the
wound.
• Inflammation is more intense because large tissue defects have a greater
volume of necrotic debris, exudate, and fibrin that must be removed.
• Larger defects require a greater volume of granulation tissue to fill in the gaps
and provide the underlying framework for the regrowth of tissue epithelium.
• A greater volume of granulation tissue generally results in a greater mass of
scar tissue.
• Secondary healing involves wound contraction.
• Within 6 weeks, for example, large skin defects may be reduced to 5% to 10%
of their original size, largely by contraction.
features of contractile smooth muscle cells.
wound.
Hand, wound healed by secondary intention, same case as previous
image, eight months later - Clinical presentation As the granulation tissue
matures, the myofibroblasts contract to close the wound. They then become
mature fibroblasts, which produce collagen, forming a fibrous scar. The
contraction that takes place can lead to decreased mobility.
Wound Strength
• Carefully sutured wounds have approximately 70% of the strength of normal
skin, largely because of the placement of sutures.
• When sutures are removed, usually at 1 week, wound strength is
approximately 10% of that of unwounded skin, but this increases rapidly over
the next 4 weeks.
• The recovery of tensile strength results from collagen synthesis exceeding
degradation during the first 2 months, and from structural modifications of
collagen (e.g., cross-linking, increased fiber size) when synthesis declines at
later times.
• Wound strength reaches approximately 70% to 80% of normal by 3 months
and usually does not improve substantially beyond that point.
FACTORS THAT INFLUENCE TISSUE REPAIR
•Tissue repair may be altered by a variety of influences.
•Variables that modify healing may be extrinsic (e.g., infection) or intrinsic to the
injured tissue. Particularly important are infections and diabetes.
•Infection is clinically the most important cause of delay in healing; it prolongs
inflammation and potentially increases the local tissue injury.
•Nutrition has profound effects on repair; protein deficiency, for example, and
especially vitamin C deficiency inhibit collagen synthesis and retard healing.
•Glucocorticoids (steroids) have well-documented anti-inflammatory effects, and
their administration may result in weakness of the scar because of inhibition of
TGF-β production and diminished fibrosis. In some instances, however, the anti-
inflammatory effects of glucocorticoids are desirable. For example, in corneal
infections, glucocorticoids are sometimes prescribed (along with antibiotics) to
reduce the likelihood of opacity that may result from collagen deposition.
•Mechanical variables such as increased local pressure or torsion may cause
wounds to pull apart, or dehisce.
•Poor perfusion, due either to arteriosclerosis and diabetes or to obstructed
venous drainage (e.g., in varicose veins), also impairs healing.
•The type and extent of tissue injury affects the subsequent repair. Complete
restoration can occur only in tissues composed of stable and labile cells; injury to
tissues composed of permanent cells must inevitably result in scarring, as in
healing of a myocardial infarct.
•The location of the injury and the character of the tissue in which the injury
occurs are also important. For example, inflammation arising in tissue spaces
(e.g., pleural, peritoneal, or synovial cavities) develops extensive exudates.
Subsequent repair may occur by digestion of the exudate, initiated by the
proteolytic enzymes of leukocytes and resorption of the liquefied exudate. This is
called resolution, and generally, in the absence of cellular necrosis, normal tissue
architecture is restored. In the setting of larger accumulations, however, the
exudate undergoes organization: Granulation tissue grows into the exudate, and
a fibrous scar ultimately forms.
Most common infection that delays wound healing is by Staphylococcus
aureus.
Vitamin C:
responsible for hydroxylation of proline and lysine.
Deficiency of vitamin C (scurvy) produces structurally weak collagen à
poor wound healing.
Copper:
cofator on lysyl oxidase which is responsible for cross
linking of collagen fibrils. Deficiency of this element responsible for poor
wound healing and predisposes to dissecting aortic aneurysm (since
elastic tissue is also weak)
Glucocorticoids :
-occasionally used with antibiotics to prevent scar tissue
formation (e.g. bacterial meningitis).
-inhibit wound healing by impairing collagen synthesis.
Glucocorticoids occasionaly used with antibiotics to prevent scar formation ( e.g.
bacterial meningitis).
Glucocorticoids
Glucocorticoids: prevent scar formation
a.Interfere with collagen formation and decrease tensile strength
b.Useful clinically in preventing excessive scar formation
(1)Dexamethasone is used along with antibiotics to prevent scar formation in
bacterial meningitis.
(2)Plastic surgeons inject high potency steroids into wounds to prevent
excessive scar tissue formation.
Aberrations of cell growth and ECM production may occur even in what begins
as normal wound healing. For example, the accumulation of exuberant amounts
of collagen can give rise to prominent, raised scars known as keloids. There
appears to be a heritable predisposition to keloid formation, and the condition is
more common in African-Americans. Healing wounds may also generate
exuberant granulation tissue that protrudes above the level of the surrounding
skin and hinders re-epithelialization. Such tissue is called “proud flesh” in old
medical parlance, and restoration of epithelial continuity requires cautery or
surgical resection of the granulation tissue.
Abdomen, poor wound healing, dehisced surgical wound - Clinical
presentation Wound dehiscence is the disruption of apposed surfaces of a
wound (most often abdominal), due to inadequate healing. Many factors, such
as infection, inadequate blood supply, diabetes, and mechanical stress can
impair healing.
Definition: Keloids are raised scars that grow beyond the borders of the original
wound.
Develop in 15% to 20% of African Americans, Asians, and Hispanics, suggesting
a genetic predisposition.
Often refractory to medical and surgical intervention.
Definition: Hypertrophic scars are raised scars that remain within the confines of
the original wound. Frequently regress spontaneously.
Normal scars have collagen bundles that are randomly arrayed (not all in the
same direction), whereas keloids and hypertrophic scars have stretched
collagen bundles arranged in the same plane as the epidermis.
Head and neck, keloids - Clinical presentation Keloids, nodular masses of
hyperplastic scar tissue, occur when the wound healing process runs
unchecked. They are more common in people of African descent. Surgical
excision typically leads to repeated keloid formation.
Keloid microscopy
Note hyperextensible skin and extremely hypermobile joint
Cell regeneration depends on factors that stimulate parenchymal cell division
and migration.
Stimulatory factors include loss of tissue and production of growth factors
C
C
A
C
Repair in other tissues
1.Liver
a.Mild injury (e.g., hepatitis A)
•Regeneration of hepatocytes with restoration to normal is possible if the
cytoarchitecture is intact.
b.Severe or persistent injury (e.g., hepatitis C)
(1)Regenerative nodules develop that show twinning of liver cell plates (two cells
thick); a double line of hepatocytes is present, and nuclei seem to run in parallel.
(2)Portal triads are not present in regenerative nodules.
(3)Increased fibrosis occurs around the regenerative nodules, which leads to
cirrhosis of the liver if the injurious agent is not removed.
2.Lung
a.Type II pneumocytes are the key repair cells of the lung.
b.Replace damaged type I and type II pneumocytes and synthesize surfactant.
3.Brain
•Proliferation of astrocytes is called gliosis.
b.Microglial cells (macrophages) are scavenger cells that remove debris (e.g.,
myelin).
4.Peripheral nerve transection

a.Without innervation, muscle atrophies in ~15 days.
b.After nerve transection, there is distal degeneration of the axon and myelin
sheath and proximal axonal degeneration up to the next node of Ranvier.
Wallerian degeneration: distal degeneration of the axon
(1)Macrophages and Schwann cells phagocytose axonal/myelin debris.
(2)Nerve cell body undergoes central chromatolysis.
(a)Nerve cell body swells.
(b)Nissl bodies (composed of rough endoplasmic reticulum and free ribosomes)
disappear centrally, and the nucleus moves to the periphery.
(3)Schwann cells proliferate in the distal stump.
Peripheral nerve transection: Schwann cell is key in reinnervation
(4)Axonal sprouts develop in the proximal stump and extend distally using the
Schwann cells for guidance.
(5)Regenerated axon grows 2 to 3 mm/day.
(6)Axon becomes remyelinated.
(7)Muscle is eventually reinnervated.
5.Heart
a.Cardiac muscle is permanent tissue.
b.Damaged muscle is replaced by noncontractile scar tissue.
6.Skeletal muscle postexercise

a.After exercise, there is damage to the sarcomeres in skeletal muscle.
•A sarcomere is the basic unit of a muscle and gives skeletal muscle its striated
appearance.
b.Satellite cells are stem cells that repair and form new myofibers in sarcomeres
that have been damaged by mechanical strain.
Skeletal muscle postexercise: myofiber repair by satellite cells (muscle stem
Normal fluid homeostasis: involves
maintaining vessel wall integrity and intravascular pressure à
failure à edema (interstitial accumulation of fluid)
maintaining blood as liquid in a clot free state in uninjured vesselà
failure à thrombosis à embolism à can obstruct blood flow à infarction.
failure to clot à hemorrhage
extensive hemorrhage à hypotension à shock and death
Edema, congestion, hemorrhage, thrombosis, embolism, infarction and

shock are the hemodynamic disorders
Normal fluid homeostasis: involves
maintaining vessel wall integrity and intravascular pressure à
failure à edema (interstitial accumulation of fluid)
maintaining blood as liquid in a clot free state in uninjured vesselà
failure à thrombosis à embolism à can obstruct blood flow à infarction.
failure to clot à hemorrhage
extensive hemorrhage à hypotension à shock and death
Edema, congestion, hemorrhage, thrombosis, embolism, infarction and

shock are the hemodynamic disorders
According to Starling’s law the normal fluid exchange between the blood
vessels and the interstitial tissue is dependent upon two opposing forces
the hydrostatic pressure (which drives fluid out)
the oncotic pressure (which keeps fluid in) , oncotic pressure is the
function of the albumin concentration in plasma.
Also remember !!
Total body water = 60% of body weight.
Distributed in two compartments:
Intracellular fluid
Extracellular fluid.
Extracellular fluid compartment divided into two minor compartments:
Interstitial fluid
Blood plasma
Generalised edema
is mainly due to hypoproteinemia (renal disease).
Look for edema around the eyes = Periorbital edema.
Pitting edema: digital pressure produces temporary indentation of the skin
. Most commonly seen in ankles of patients with Congestive heart failure.
An abscess is an enclosed collection of liquefied tissue, known as pus,
somewhere in the body. It is the result of the body’s defensive reaction to
foreign material.
Tissue swelling here is due to accumulation of exudate (pus).
It it non-pitting.
Pitting edema due to transudate.
Non pitting edema due to exudate, lymphatic fluid and glycosaaminoglycans.
Pitting edema: digital pressure produces temporary indentation of the skin

. Most commonly seen in ankles of patients with Congestive heart failure.
Lymphedema: nonpitting edema due to lymphatic obstruction.
The capillary system and mechanisms of edema formation.
A.Normal.The differential between the hydrostatic and oncotic pressures at the
arterial end of the capillary system is responsible for the filtration into the
interstitial space of approximately 14 mL of fluid per minute.
This fluid is reabsorbed at the venous end at the rate of 12 mL/min. It is also
drained through the lymphatic capillaries at a rate of 2 mL/min.
B.Hydrostatic edema.If the hydrostatic pressure at the venous end of the
capillary sys-tem is elevated, reabsorption decreases. As long as the lymphatics
can drain the surplus fluid, no edema results. If their capacity is exceeded, how-
ever, edema fluid accumulates.
Left ventricular failure: hydrostatic pressure in pulmonary vein overrides
pulmonary capillary oncotic pressure à transudate
Volume overload due to Excess infusion of IV fluids can also
raise HP and cause edema.
C.Oncotic edema.Edema fluid also accumulates if reabsorption is diminished by
decreased oncotic pressure ofthe vascular bed, owing to a loss of albumin.
Malabsorption syndromes like Celiac disease also associated with
hypoproteinnemia
E.Lymphedema.Lymphatic obstruction causes the accumulation of interstitial
fluid because ofinsufficient reabsorption and deficient removal of proteins, the
latter increasing the oncotic pressure of the fluid in the interstitial space.
Elephantiasis : a term used to denote massive leg edema caused by obstruction
of inguinal lymphatics and lymphnodes by filarial worms in filariasis
Cause : Filarial worm infection ( Wuchereria bancrofti)
Lymphogranuloma venereum: C. trachomatis
Peau d’ orange = “orange peel appearance”: due to blockage of lymphatics by

cancer cells. Seen in Breast Cancer
Renin-angiotensin-aldosterone system:
Renal artery occlusion or dietary salt restriction leads to increased renal

secretion of renin.
Renin converts angiotensinogen to angiotensin I
Angiotensin I is converted to Angiotensin II by Angiotensin Converting Enzyme
(ACE).
Angiotensin II acts in several ways to increase the blood pressure
causes vasoconstriction à increasing TPR à increase BP
(BP=COXTPR)
Stimulates aldosterone secretion from adrenal à acts on renal
tubules to increase Na absorption à increase in total body fluid volume à
increase in BP
Increases sympathetic output à increases HR and SV à increase
in BP
D.Inflammatory and traumatic edema.Edema, either local or systemic, results if
the vascular bedbecomes leaky following injury to the endothelium.
GAG= glycosaaminoglycan
A: Here is an example of fluid collection into a body cavity, or an effusion.
This is a right pleural effusion (in a baby). Note the clear, pale yellow
appearance of the fluid. This is a serous effusion. Extravascular fluid
collections can be classified as follows:
Exudate: extravascular fluid collection that is rich in protein and/or cells.
Fluid appears grossly cloudy.
Transudate: extravascular fluid collection that is basically an ultrafiltrate
of plasma with little protein and few or no cells. Fluid appears grossly
clear.
B: The milky white fluid shown here in the peritoneal cavity represents a
chylous ascites. This is an uncommon fluid accumulation that can be due
to blockage of lymphatic drainage, in this case by a malignant lymphoma
involving the mesentery and retroperitoneum.
Here is simple edema, or fluid collection within tissues. This is "pitting" edema
because, on physical examination, you can press your finger into the skin and
soft tissue and leave a depression.
Note the finger indentations within the edematous tissue (arrows).

What is the pathogenesis of pulmonary edema?
Left ventricular failure (eg, caused by a myocardial infarct) causes pump failure,
and secondarily there is impaired flow of blood from the lung to the left atrium.
This causes increased hydrostatic pressure in pulmonary alveolar capillaries and
subsequent transudation of fluid into alveoli.
Pulmonary edema in other cases may also result from damage to alveolar
capillaries (eg, in adult respiratory distress syndrome).
How does this type of edema differ from that seen in acute inflammation?
The fluid in pulmonary edema is a transudate (ie, it is protein poor, has low
specific gravity, and does not contain inflammatory cells). Edema in inflammation
is an exudate.
A) lungs is congested and (red) and shiny (edema). B) Note frothy fluid
indicative of edema
Lung, pulmonary congestion and edema - Medium power
The alveolar septa are prominent, due to marked congestion of the capillaries.
The alveolar lumens contain pale-staining edema fluid.
3. Impaired lymph drainage:
Lymph can not flow through fibrotic liver – diverted to peritoneal
cavity
4. Increased retention of sodium and water
1. Increased fluid in peritoneal cavity à decreased intravascular
volume à activation of renin angiotensin aldosterone system
à retention of sodium and water
2. Decreased metabolism of aldosterone
Note the flattened gyri and narrow sulci in the edematous brain as compared to
the normal brain.
MI = myocardial infarction = heart attack
Gross or naked eye examination of heart of the patient reveals:
a thrombus (blood clot) in the coronary artery and
an area of dead myocardial tissue (necrosis)
Death of tissue due to blockage in blood supply is known as INFARCTION
A) lungs is congested and (red) and shiny (edema). B) Note frothy fluid
indicative of edema
• Hemostasis can be defined simply as the process by which blood clots form at sites
of vascular injury.
• Hemostasis is essential for life and is deranged to varying degrees in a broad range
of disorders, which can be divided into two groups.
• In hemorrhagic disorders, characterized by excessive bleeding, hemostatic
mechanisms are either blunted or insufficient to prevent abnormal blood loss.
• By contrast, in thrombotic disorders blood clots (often referred to as thrombi) form
within intact blood vessels or within the chambers of the heart.
• Thrombosis has a central role in the most common and clinically important forms of
cardiovascular disease (e.g. myocardial infarction)
Thrombosis: is pathologically predisposed by many conditions, including venous stasis usually
from immobilization; CHF; polycythemia; sickle cell disease; visceral malignancies and the use
of oral contraceptives, especially in association with cigarette smoking.
Thrombosis
▪Thrombus development usually is related to one or more components of the Virchow triad:
▪Endothelial injury (e.g., by toxins, hypertension, inflammation, or metabolic products)
associated with endothelial activation and changes in endothelial gene expression that favor
coagulation
▪Abnormal blood flow—stasis or turbulence (e.g., due to aneurysms, atherosclerotic plaque)
▪Hypercoagulability, either primary (e.g., factor V Leiden, increased prothrombin synthesis,
antithrombin III deficiency) or secondary (e.g., bed rest, tissue damage, malignancy, or
development of antiphospholipid antibodies [antiphospholipid antibody syndrome]) or
antibodies against platelet factor IV/heparin complexes [heparin-induced thrombocytopenia])
▪Thrombi may propagate, resolve, become organized, or embolize.
▪Thrombosis causes tissue injury by local vascular occlusion or by distal embolization.
Normal Hemostasis vs. Thrombosis
Normal hemostasis
Maintains blood flow in a fluid, clot free state in normal vessels.
Induces localized hemostatic plugs at sites of vascular injury.
Thrombosis
Pathologic process
Formation of blood clots (thrombus) in uninjured vessels or vessels with
relatively minor injuries
Thrombus may form in the arterial system including heart chambers and in
venous system.
Question: What is the thrombus made of?

Fibrin, platelets, and entraped blood cells.
Question: What causes arterial thrombosis? ...venous thrombosis?
Arterial thrombosis is caused by endothelial damage (eg, atherosclerosis
or vasculitis);
venous thrombosis is caused by stasis (sluggishness) of blood flow.
Both types of vessels are affected in hypercoagulable states such as
antithrombin or protein C deficiency.
Thrombosis results from the interaction of platelets, endothelial cells
and the coagulation cascade.
Virchow triad in thrombosis.
Endothelial integrity is the single most important factor.
Note that injury to endothelial cells can affect local blood flow and/or
coagulability; abnormal blood flow (stasis or turbulence) can, in turn, cause
endothelial injury.
The elements of the triad may act independently or may combine to cause
thrombus formation.
The primary abnormalities that lead to thrombosis are

(1) endothelial injury,
(2) stasis or turbulent blood flow, and
(3) hypercoagulability of the blood (the so-called Virchow triad)
Thrombosis is one of the scourges of modern man, because it underlies the

most serious and common forms of cardiovascular disease.
Endothelial injury usually results in arterial thrombosis.
Increased homocysteine levels: seen in vitamin B12 and folic acid deficiency.
Myocardial infarction (endocardial damage)
Intact endothelial cells resist formation of thrombus
Endothelial Injury
• Endothelial injury leading to platelet activation almost inevitably underlies thrombus formation in the
heart and the arterial circulation, where the high rates of blood flow impede clot formation.
• Notably, cardiac and arterial clots are typically rich in platelets, and it is believed that platelet adherence
and activation is a necessary prerequisite for thrombus formation under high shear stress, such as
exists in arteries.
• This insight provides part of the reasoning behind the use of aspirin and other platelet inhibitors in
coronary artery disease and acute myocardial infarction.
• Obviously, severe endothelial injury may trigger thrombosis by exposing vWF and tissue factor.
• However, inflammation and other noxious stimuli also promote thrombosis by shifting the pattern of
gene expression in endothelium to one that is “prothrombotic.”
• This change is sometimes referred to as endothelial activation or dysfunction and can be produced by
diverse exposures, including physical injury, infectious agents, abnormal blood flow, inflammatory
mediators, metabolic abnormalities, such as hypercholesterolemia or homocystinemia, and toxins
absorbed from cigarette smoke.
• Endothelial activation is believed to have an important role in triggering arterial thrombotic events.
• The role of endothelial cell activation and dysfunction in arterial thrombosis:
• Procoagulant changes.
• Endothelial cells activated by cytokines downregulate the expression of thrombomodulin, already
described as a key modulator of thrombin activity. This may result in sustained activation of thrombin,
which can in turn stimulate platelets and augment inflammation through PARs expressed on platelets
and inflammatory cells. In addition, inflamed endothelium also downregulates the expression of other
anticoagulants, such as protein C and tissue factor protein inhibitor, changes that further promote a
procoagulant state.
• Antifibrinolytic effects.
Alternations in Normal Blood Flow
• Turbulence contributes to arterial and cardiac thrombosis by causing endothelial injury or
dysfunction, as well as by forming countercurrents that contribute to local pockets of stasis.
• Stasis is a major contributor in the development of venous thrombi.
• Normal blood flow is laminar such that the platelets (and other blood cellular elements) flow
centrally in the vessel lumen, separated from endothelium by a slower moving layer of
plasma.
• Stasis and turbulence therefore:
• Promote endothelial activation, enhancing procoagulant activity and leukocyte adhesion, in
part through flow-induced changes in the expression of adhesion molecules and pro-
inflammatory factors
• Disrupt laminar flow and bring platelets into contact with the endothelium
• Prevent washout and dilution of activated clotting factors by fresh flowing blood and the inflow
of clotting factor inhibitors
• Altered blood flow contributes to thrombosis in several clinical settings.
• Ulcerated atherosclerotic plaques not only expose subendothelial vWF and tissue factor but
also cause turbulence.
• Aortic and arterial dilations called aneurysms result in local stasis and are therefore fertile
sites for thrombosis
• Acute myocardial infarctions result in areas of noncontractile myocardium and sometimes in
cardiac aneurysms; both are associated with stasis and flow abnormalities that promote the
formation of cardiac mural thrombi.
• Rheumatic mitral valve stenosis results in left atrial dilation; in conjunction with atrial
fibrillation, a dilated atrium is a site of profound stasis and a prime location for thrombosis.
• Hyperviscosity (such as is seen with polycythemia vera; increases resistance to flow and
causes small vessel stasis, and the deformed red cells in sickle cell anemia impede blood
flow through small vessels, with the resulting stasis also predisposing to thrombosis.
Normal blood flow is laminar (i.e. cellular elements flow centrally in the vessel lumen,
separated from endothelium by a plasma clear zone).
Hyperviscosity syndrome (polycythemia ) and deformed RBC (sickle cell anemia) result
in small vessel stasis and also predispose to thrombosis.
Hypercoagulability
• Hypercoagulability (also called thrombophilia) can be loosely defined as any disorder of the
blood that predisposes to thrombosis.
• Hypercoagulability has a particularly important role in venous thrombosis and can be divided
into primary (genetic) and secondary (acquired) disorders.
• Of the inherited causes of hypercoagulability, point mutations in the factor V gene and
prothrombin gene are the most common.
• Approximately 2% to 15% of Caucasians carry a single-nucleotide mutation in factor V that is
called the factor V Leiden, after the city in The Netherlands where it was discovered.
• Among individuals with recurrent DVT, the frequency of this mutation is considerably higher,
approaching 60%.
• The mutation results in a glutamine for arginine substitution at amino acid residue 506 that
renders factor V resistant to cleavage and inactivation by protein C.
• As a result, an important antithrombotic counterregulatory pathway is lost.
• Indeed, heterozygotes have a five-fold increased relative risk of venous thrombosis, and
homozygotes have a 50-fold increase.
• A single nucleotide change (G20210A) in the 3′-untranslated region of the prothrombin gene
is another common mutation (1% to 2% of the population) associated with hypercoagulability.
It leads to elevated prothrombin levels and an almost three-fold increased risk of venous
thrombosis.
• Elevated levels of homocysteine contribute to arterial and venous thrombosis, as well as the
development of atherosclerosis.
• The prothrombotic effects of homocysteine may be due to thioester linkages formed between
homocysteine metabolites and a variety of proteins, including fibrinogen.
• Marked elevations of homocysteine may be caused by an inherited deficiency of cystathione
β-synthetase.
• Rare inherited causes of primary hypercoagulability include deficiencies of anticoagulants
such as antithrombin III, protein C, or protein S; affected individuals typically present with
venous thrombosis and recurrent thromboembolism beginning in adolescence or early
Q:How do the various natural anticoagulants act?
A:There are three natural anticoagulants:
(1) The protein C system generates active protein C that inactivates cofactors V and VIII. Protein
C itself is activated by thrombin after the latter binds to thrombomodulin on the endothelium.
(2) Antithrombin III is activated by binding to heparin-like molecules on the endothelium;
activated antithrombin causes proteolysis of active factors IX, X, and XI, and thrombin.
(3) Plasmin cleaves fibrin. It is derived from its circulating precursor, plasminogen, by the action
of tissue plasminogen activator, which is synthesized by endothelial cells.
OC pills:hypercoagulable blood
decreased antithrombin III
increased fibrinogen, V, VIII
Hypercoagulable States
Primary (Genetic)
Common
• Factor V mutation (Arg to Gln substitution in amino acid residue 506 leading to resistance to
activated protein C; factor V Leiden)
• Prothrombin mutation (G20210A noncoding sequence variant leading to increased prothrombin
levels)
• Increased levels of factors VIII, IX, XI, or fibrinogen (genetics unknown)
Rare
• Antithrombin III deficiency
Very Rare
• Homozygous homocystinuria (deficiency of cystathione β-synthetase)
• Secondary (Acquired)
High Risk for Thrombosis
• Prolonged bed rest or immobilization
• Myocardial infarction
• Atrial fibrillation
• Tissue injury (surgery, fracture, burn)
• Cancer
• Prosthetic cardiac valves
• Disseminated intravascular coagulation
• Heparin-induced thrombocytopenia
• Antiphospholipid antibody syndrome
Lower Risk for Thrombosis
• Cardiomyopathy
• Nephrotic syndrome
• Hyperestrogenic states (pregnancy and postpartum)
• Oral contraceptive use
• Sickle cell anemia
• Smoking
• Thrombi can develop anywhere in the cardiovascular system and vary in size and shape depending on the involved site and the
underlying cause.
• Arterial or cardiac thrombi usually begin at sites of turbulence or endothelial injury, whereas venous thrombi characteristically occur at
sites of stasis.
• Thrombi are focally attached to the underlying vascular surface, particularly at the point of initiation.
• From here, arterial thrombi tend to grow retrograde, while venous thrombi extend in the direction of blood flow; thus both propagate
toward the heart.
• The propagating portion of a thrombus is often poorly attached and therefore prone to fragmentation and embolization.
• Thrombi often have grossly and microscopically apparent laminations called lines of Zahn, which are pale platelet and fibrin deposits
alternating with darker red cell–rich layers.
• Such laminations signify that a thrombus has formed in flowing blood; their presence can therefore distinguish antemortem clots from
the bland nonlaminated clots that occur postmortem.
• Thrombi occurring in heart chambers or in the aortic lumen are designated mural thrombi.
• Abnormal myocardial contraction (arrhythmias, dilated cardiomyopathy, or myocardial infarction) or endomyocardial injury
(myocarditis or catheter trauma) promotes cardiac mural thrombi , while ulcerated atherosclerotic plaque and aneurysmal dilation are
the precursors of aortic thrombi.
• Arterial thrombi are frequently occlusive; the most common sites in decreasing order of frequency are the coronary, cerebral, and
femoral arteries.
• They typically consist of a friable meshwork of platelets, fibrin, red cells, and degenerating leukocytes. Although these are usually
superimposed on a ruptured atherosclerotic plaque, other vascular injuries (vasculitis, trauma) may be the underlying cause.
• Venous thrombosis (phlebothrombosis) is almost invariably occlusive, with the thrombus forming a long luminal cast.
• Because these thrombi form in the sluggish venous circulation, they tend to contain more enmeshed red cells (and relatively few
platelets) and are therefore known as red, or stasis, thrombi.
• Venous thrombi are firm, are focally attached to the vessel wall, and contain lines of Zahn, features that help distinguish them from
postmortem clots (see later).
• The veins of the lower extremities are most commonly involved (90% of cases); however, upper extremities, periprostatic plexus, or
the ovarian and periuterine veins can also develop venous thrombi. Under special circumstances, they can also occur in the dural
sinuses, portal vein, or hepatic vein.
• Postmortem clots can sometimes be mistaken for antemortem venous thrombi.
• However, clots that form after death are gelatinous and have a dark red dependent portion where red cells have settled by gravity
and a yellow “chicken fat” upper portion, and are usually not attached to the underlying vessel wall.
• Thrombi on heart valves are called vegetations.
• Bloodborne bacteria or fungi can adhere to previously damaged valves (e.g., due to rheumatic heart disease) or can directly cause
valve damage; in either case, endothelial injury and disturbed blood flow can induce the formation of large thrombotic masses
(infective endocarditis)
• Sterile vegetations can also develop on noninfected valves in persons with hypercoagulable states, so-called nonbacterial thrombotic
endocarditis.
• Less commonly, sterile verrucous endocarditis (Libman-Sacks endocarditis) can occur in the setting of systemic lupus
erythematosus.
Stasis of blood:
procoagulants released (tissue thromboplastin or tissue factor) from
damaged endothelium cause localized activation of the coagulation system
fibrin is produced, which forms a mesh around RBCs, platelets and white
blood cells in the stagnant blood within the vessel to produce a venous thormbus
Coagulation factors are used up in formation of venous thrombi

fibrinogen (I), prothrombin (II), V, VIII are consumed
Heparin and warfarin prevent formation of venous thrombi since these drugs are
anticoagulants.
Not prevented by aspirin.
Venous thrombi in Superficial veins (ex Saphenous vein) à congestion, edema and
pain à varicose ulcer
DVT = deep vein thrombosis

Usually asymptomatic unlike superficial vein thrombosis
Anticoagulalants like Heparin and warfarin inhibit the formation of venous thrombi. They
do not dissolve dissolve the thrombus, they only prevent further formation of a
thrombus.
The fibrinolytic system (plasmin) is responsible for dissolution of the thrombus.
Dr Fogel’s Slide
Venous thrombus. The femoral vein has been opened to reveal a large thrombus within
the lumen.
Factors causing endothelial injury:
atherosclerosis (most important)
smoking : chemical damage endothelial cells
hypertension
vasculitis
Increased plasma homocysteine : folate deficiency most common cause
Aspirin prevents formation of arterial thrombi by inhibiting platelet cyclooxegenaseà
decreased production of thromboxane A2 (platelet aggregator)
Ticlopidine is used if patient is allergic to aspirin (side effect à neutropenia)
Heparin and Warfarin do not prevent arterial thrombi composed of platelets.
Q: What are other causes of arterial thrombosis?
Arterial thrombosis is caused by injury to the endothelium.
In addition to atherosclerosis, other causes are vasculitis , Hypertension, trauma,
increased plasma homocysteine.
Most common site for arterial thrombi: coronary arteries

Most common site for venous thrombi” deep veins of the calf.
• Thrombi often have grossly and microscopically apparent laminations called lines of
Zahn, which are pale platelet and fibrin deposits alternating with darker red cell–rich
layers.
• Such laminations signify that a thrombus has formed in flowing blood; their presence
can therefore distinguish antemortem clots from the bland nonlaminated clots that
occur postmortem.
A: This is coronary thrombosis, one of the complications of atherosclerosis. The dark
red thrombus is seen in the anterior descending coronary artery.
The thrombus occludes the lumen and produces ischemia and/or infarction of the
myocardium.
B: A coronary thrombosis is seen microscopically occluding the remaining small lumen

of this coronary artery
Q: What therapeutic agent can be used to lyse the clots in coronary vessels?
A:Thrombolysis can be accomplished by tissue plasminogen activator (t-PA) or
streptokinase; both cause fibrinolysis by generating plasmin.
Coronary artery, atherosclerosis - Low power
This epicardial coronary artery is almost completely occluded by atherosclerotic plaque.
The plaque is heavily lipid laden and has ruptured, releasing thrombogenic substances
into the narrow lumen.
A thrombus has occluded the tiny lumen that remains.
Questions:
What are some of the clinical manifestations associated with the presence of an
unstable atheromatous plaque?
Answer:
Unstable angina, acute myocardial infarction, sudden cardiac death.
Note the thrombus (T) is attached to the vessel (V) wall (arrow head)
Lines of Zahn alternating areas layer of RBCs and fibrin/platelets are present
These are "lines of Zahn" which are the alternating pale pink bands of platelets with
fibrin and red bands of RBC's forming a true thrombus.
• Thrombi often have grossly and microscopically apparent laminations called lines of
Zahn, which are pale platelet and fibrin deposits alternating with darker red cell–rich
layers.
• Such laminations signify that a thrombus has formed in flowing blood; their presence
can therefore distinguish antemortem clots from the bland nonlaminated clots that
occur postmortem.
Clinical note:
Aspirin prevents formation of arterial thrombi by inhibiting platelet
cyclooxegenaseà decreased production of thromboxane A2 (platelet
aggregator)
Ticlopidine is used if patient is allergic to aspirin (side effect à neutropenia)
Heparin and Warfarin do not prevent arterial thrombi composed of platelets.
What is the role played by aspirin in preventing thrombus formation? What stage
of hemostasis is affected by aspirin?
A: Aspirin prevents thrombogenesis by inhibiting platelet aggregation.
This is achieved by inhibition of cyclooxygenase, thereby preventing the generation of
thromboxane A2.

(1) The protein C system generates active protein C that inactivates cofactors V and
VIII. Protein C itself is activated by thrombin after the latter binds to thrombomodulin
on the endothelium.
(2) Antithrombin is activated by binding to heparin-like molecules on the endothelium;
activated antithrombin causes proteolysis of active factors IX, X, and XI, and
thrombin.
(3) Plasmin cleaves fibrin. It is derived from its circulating precursor, plasminogen, by
the action of tissue plasminogen activator, which is synthesized by endothelial cells.
• Thrombi occurring in heart chambers or in the aortic lumen are designated
mural thrombi.
• Abnormal myocardial contraction (arrhythmias, dilated cardiomyopathy, or
myocardial infarction) or endomyocardial injury (myocarditis or catheter trauma)
promotes cardiac mural thrombi , while
• ulcerated atherosclerotic plaque and aneurysmal dilation are the precursors of aortic
thrombi.
Mural thrombi: causes include

MI, dilated cardiomyopathy, myocarditis, mitral stenosis
Mural thrombi.Thrombus in the left and right ventricular apices
Heart, rheumatic mitral valve, atrial changes – Gross
The mitral valve has been reduced to a narrow orifice.
The left atrium is markedly dilated.
Thrombi have formed on the atrial wall.
Question: What is the basis of thrombosis in this setting?
Answer: In the dilated atrium with a stenotic mitral valve, the blood stagnates. Hence,
stasis is a factor in thrombogenesis. In addition, there may be endocardial damage
resulting from regurgitant jets; this, in turn, can also predispose to thrombosis.
Question: What EKG change might you find in this patient?
Answer: Atrial fibrillation.
Question: Why is patient’s with mitral stenosis feel short of breath? Why is left atrium
enlarged? Why are lungs abnormal? Why do they get short of breath at night? Do they
develop heart failure? What kind?
Answer:
The patients with MS
Feel short of breath because they have mitral stenosis.
There is fusion of the commissures and fibrotic closure of the mitral valve.
As the cross-sectional area of the orifice of the affected valve grows smaller, there is
progressive obstruction to blood flow from the left atrium to the left ventricle.
Valve orifices smaller than 2 cm2 are usually symptomatic.
Blood begins to back up first into the left atrium, which becomes enlarged.
Pressure rises in the atrium and pulmonary vascular tree.
Elevated pulmonary vascular pressure leads to pulmonary edema and explains
dyspneic symptoms.
Eventually, the pressure backs up into the right heart. With longstanding severe mitral
stenosis, permanent structural changes in the pulmonary vasculature may limit flow
even after the valve is fixed.
They develop left heart failure
Aneurysm of abdominal aorta.
. A, External view, gross photograph of a large aortic aneurysm that ruptured; the
rupture site is indicated by the arrow.
B, Opened view, with the location of the rupture tract indicated by a probe. The wall of
the aneurysm is exceedingly thin, and the lumen is filled by a large quantity of layered
but largely unorganized thrombus.
• Thrombi on heart valves are called vegetations.
• Bloodborne bacteria or fungi can adhere to previously damaged valves (e.g., due to
rheumatic heart disease) or can directly cause valve damage; in either case,
endothelial injury and disturbed blood flow can induce the formation of large
thrombotic masses (infective endocarditis)
• Sterile vegetations can also develop on noninfected valves in persons with
hypercoagulable states, so-called nonbacterial thrombotic endocarditis.
• Less commonly, sterile verrucous endocarditis (Libman-Sacks endocarditis) can
occur in the setting of systemic lupus erythematosus.
ARF= Acute Rheumatic fever

SLE= Systemic lupus erythematosus
Acute Bacterial Endocarditis: large friable and irregular vegetations (arrow) are present
on the margin of the mitral valve.
The small pink vegetation on the rightmost aortic cusp margin represents the typical
finding with non-bacterial thrombotic endocarditis (NBTE, or so-called "marantic
endocarditis"). This is a non-infective form of endocarditis. It tends to occur in persons
with a hypercoagulable state (such as Trousseau syndrome, a paraneoplastic
syndrome associated with malignancies) and in very ill persons.
Bottom right:
Marantic endocarditis. Sterile platelet–fibrin vegetations are seen on the leaflets of a
structurally normal mitral valve.
These vegetations are rarely over 0.5 cm in size. However, they are friable and very
prone to embolize.
The rheumatic fever phase of rheumatic heart disease (RHD) is marked by small, warty
vegetations along the lines of closure of the valve leaflets. Infective endocarditis (IE) is
characterized by large, irregular masses on the valve cusps that can extend onto the
chordae
Nonbacterial thrombotic endocarditis (NBTE) typically exhibits small, bland vegetations,
usually attached at the line of closure. One or many may be present.
Libman-Sacks endocarditis (LSE) has small or medium-sized vegetations on either or
both sides of the valve leaflets.
• Postmortem clots can sometimes be mistaken for antemortem venous thrombi.
• However, clots that form after death are gelatinous and have a dark red dependent
portion where red cells have settled by gravity and a yellow “chicken fat” upper
portion, and are usually not attached to the underlying vessel wall.
Q:What is the difference between a postmortem clot and a thrombus?

A:Postmortem clots
are not attached to endothelium;
they are gelatinous, rubbery, dark red at the bottom and yellowish on
top.
Thrombi
are attached to endothelium and are
traversed by pale grey fibrin strands that can be seen on cut section;
they are more firm but fragile.
Fate of the Thrombus
• If a patient survives the initial thrombosis, in the ensuing days to weeks thrombi undergo some combination of the following four events:
1. Propagation. Thrombi accumulate additional platelets and fibrin.
2. Embolization. Thrombi dislodge and travel to other sites in the vasculature.
3. Dissolution. Dissolution is the result of fibrinolysis, which can lead to the rapid shrinkage and total disappearance of recent thrombi. In contrast, the
extensive fibrin deposition and cross-linking in older thrombi renders them more resistant to lysis. This distinction explains why therapeutic administration
of fibrinolytic agents such as t-PA (e.g., in the setting of acute coronary thrombosis) is generally effective only when given during the first few hours of a
thrombotic event.
4. Organization and recanalization. Older thrombi become organized by the ingrowth of endothelial cells, smooth muscle cells, and fibroblasts. Capillary
channels eventually form that reestablish the continuity of the original lumen, albeit to a variable degree. Continued recanalization may convert a
thrombus into a smaller mass of connective tissue that becomes incorporated into the vessel wall. Eventually, with remodeling and contraction of the
mesenchymal elements, only a fibrous lump may remain to mark the original thrombus.
• Occasionally the centers of thrombi undergo enzymatic digestion, presumably as a result of the release of lysosomal enzymes from trapped leukocytes and
platelets. In the setting of bacteremia, such thrombi may become infected, producing an inflammatory mass that erodes and weakens the vessel wall. If
unchecked, this may result in a mycotic aneurysm.
• Clinical Features
• Thrombi come to clinical attention when they obstruct arteries or veins, or give rise to emboli.
• The clinical presentation depends on the involved site.
• Venous thrombi can cause painful congestion and edema distal to an obstruction, but are mainly of concern due to their tendency to embolize to the lungs.
• Conversely, although arterial thrombi can also embolize and cause downstream infarctions, the chief clinical problem is more often related to occlusion of a
critical vessel (e.g., a coronary or cerebral artery), which can have serious or fatal consequences.
• Venous Thrombosis (Phlebothrombosis).
• Most venous thrombi occur in the superficial or deep veins of the leg.
• Superficial venous thrombi typically occur in the saphenous veins in the setting of varicosities.
• Such thrombi can cause local congestion, swelling, pain, and tenderness, but rarely embolize.
• Nevertheless, the associated edema and impaired venous drainage predispose the overlying skin to the development of infections and ulcers (varicose
ulcers).
• Deep venous thrombosis (DVT) involving one of the large leg veins—at or above the knee (e.g., the popliteal, femoral, and iliac veins)—is more serious
because such thrombi more often embolize to the lungs and give rise to pulmonary infarction.
• Although DVTs may cause local pain and edema due to venous obstruction, these symptoms are often absent due the opening of venous collateral
channels. Consequently, DVTs are asymptomatic in approximately 50% of affected individuals and are recognized only in retrospect after embolization.
• Lower extremity DVTs are often associated with hypercoagulable states.
• Common predisposing factors include bed rest and immobilization (because they reduce the milking action of the leg muscles, resulting in stasis), and
congestive heart failure (also a cause of impaired venous return).
• Trauma, surgery, and burns not only immobilize a person but are also associated with vascular insults, procoagulant release from injured tissues,
increased hepatic synthesis of coagulation factors, and decreased t-PA production.
• Many elements contribute to the thrombotic diathesis of pregnancy, including decreased venous return from leg veins and systemic hypercoagulability
associated with the hormonal changes of late pregnancy and the postpartum period.
• Tumor-associated inflammation and coagulation factors (tissue factor, factor VIII), as well as procoagulants (e.g., mucin) released from tumor cells, all
contribute to the increased risk of thromboembolism in disseminated cancers, so-called migratory thrombophlebitis or Trousseau syndrome.
• Regardless of the specific clinical setting, advanced age also increases the risk of DVT.
• Arterial and Cardiac Thrombosis.
• Atherosclerosis is a major cause of arterial thromboses because it is associated with loss of endothelial integrity and with abnormal blood flow
• Myocardial infarction can predispose to cardiac mural thrombi by causing dyskinetic myocardial contraction and endocardial injury and rheumatic heart
disease may engender atrial mural thrombi by causing atrial dilation and fibrillation.
• Both cardiac and aortic mural thrombi are prone to embolization.
Fate of a thrombus
Q: What are the various fates of thrombi?
Propagation, embolism, dissolution, and organization with recanalization.
Q: Which of these fates is clinically most significant in the arterial circulation vs.
the venous circulation?
The most significant problem with arterial thrombi is propagation leading

to luminal obstruction, resulting in infarction of the tissue supplied.
Important examples include myocardial and cerebral infarction.
In contrast, the most significant problem with venous thrombi is the
possibility of potentially fatal embolization into the pulmonary circulation.
Dr Fogels Slide
Dr Fogels Slide
Low-power view of an artery with an old thrombus. A, H&E-stained section. B, Stain for
elastic tissue. The original lumen is delineated by the internal elastic lamina (arrows)
and is totally filled with organized thrombus, now punctuated by a number of
recanalized channels (white spaces).
Thrombosis
▪Thrombus development usually is related to one or more components of the Virchow
triad:
▪Endothelial injury (e.g., by toxins, hypertension, inflammation, or metabolic products)
associated with endothelial activation and changes in endothelial gene expression that
favor coagulation
▪Abnormal blood flow—stasis or turbulence (e.g., due to aneurysms, atherosclerotic
plaque)
▪Hypercoagulability, either primary (e.g., factor V Leiden, increased prothrombin
synthesis, antithrombin III deficiency) or secondary (e.g., bed rest, tissue damage,
malignancy, or development of antiphospholipid antibodies [antiphospholipid antibody
syndrome]) or antibodies against platelet factor IV/heparin complexes [heparin-induced
thrombocytopenia])
▪Thrombi may propagate, resolve, become organized, or embolize.
▪Thrombosis causes tissue injury by local vascular occlusion or by distal embolization.
Hypercoagulability
• Hypercoagulability (also called thrombophilia) can be loosely defined as any disorder
of the blood that predisposes to thrombosis.
• Hypercoagulability has a particularly important role in venous thrombosis and can be
divided into primary (genetic) and secondary (acquired) disorders.
• Of the inherited causes of hypercoagulability, point mutations in the factor V gene
and prothrombin gene are the most common.
• Approximately 2% to 15% of Caucasians carry a single-nucleotide mutation in factor
V that is called the factor V Leiden, after the city in The Netherlands where it was
discovered.
• Among individuals with recurrent DVT, the frequency of this mutation is considerably
higher, approaching 60%.
• The mutation results in a glutamine for arginine substitution at amino acid residue
506 that renders factor V resistant to cleavage and inactivation by protein C.
• As a result, an important antithrombotic counterregulatory pathway is lost.
• Indeed, heterozygotes have a five-fold increased relative risk of venous thrombosis,
and homozygotes have a 50-fold increase.
• A single nucleotide change (G20210A) in the 3ʹ-untranslated region of the
prothrombin gene is another common mutation (1% to 2% of the population)
associated with hypercoagulability. It leads to elevated prothrombin levels and an
almost three-fold increased risk of venous thrombosis.
1
• Elevated levels of homocysteine contribute to arterial and venous thrombosis, as well
as the development of atherosclerosis.
• The prothrombotic effects of homocysteine may be due to thioester linkages formed
between homocysteine metabolites and a variety of proteins, including fibrinogen.
• Marked elevations of homocysteine may be caused by an inherited deficiency of
cystathione β-synthetase.
• Rare inherited causes of primary hypercoagulability include deficiencies of

anticoagulants such as antithrombin III, protein C, or protein S; affected individuals
typically present with venous thrombosis and recurrent thromboembolism beginning
in adolescence or early adulthood.
1
(1) The protein C system generates active protein C that inactivates cofactors V and VIII.
Protein C itself is activated by thrombin after the latter binds to thrombomodulin on
the endothelium.
(2) Antithrombin III is activated by binding to heparin-like molecules on the
endothelium; activated antithrombin causes proteolysis of active factors IX, X, and
XI, and thrombin.
(3) Plasmin cleaves fibrin. It is derived from its circulating precursor, plasminogen, by
the action of tissue plasminogen activator, which is synthesized by endothelial cells.
Hypercoagulable States
Primary (Genetic)
Common
• Factor V mutation (Arg to Gln substitution in amino acid residue 506 leading to
resistance to activated protein C; factor V Leiden)
• Prothrombin mutation (G20210A noncoding sequence variant leading to increased
prothrombin levels)
2
• Increased levels of factors VIII, IX, XI, or fibrinogen (genetics unknown)
Rare
• Antithrombin III deficiency
• Protein C deficiency
• Protein S deficiency
Very Rare
• Homozygous homocystinuria (deficiency of cystathione β-synthetase)
• Secondary (Acquired)
High Risk for Thrombosis
• Prolonged bed rest or immobilization
• Myocardial infarction
• Atrial fibrillation
• Tissue injury (surgery, fracture, burn)
• Cancer
• Prosthetic cardiac valves
• Disseminated intravascular coagulation
• Heparin-induced thrombocytopenia
• Antiphospholipid antibody syndrome
Lower Risk for Thrombosis
• Cardiomyopathy
• Nephrotic syndrome
• Hyperestrogenic states (pregnancy and postpartum)
• Oral contraceptive use
• Sickle cell anemia
• Smoking
2
Hepatic vein thrombosis is also k/a Budd Chiari syndrome.
Clues for hereditary thrombosis syndromes:

Inherited causes of hypercoagulability should be considered in patients younger than
50 years of age who present with thrombosis and no other obvious explanation for an
acquired prothrombotic state.
Recurrent DVTs and Recurrent PE occurring at early age.
3
Mutation in factor V gene, which renders factor V resistant to inactivation by protein C.
This mutation is known as Leiden mutation.
The factor V Leiden mutation is the most common inherited cause for
hypercoagulability.
4
5
Antiphospholipid antibodies include:
Anticardiolipin antibody and lupus anticoagulant
Antiphospholipid syndrome (APS or APLS) or antiphospholipid antibody syndrome is a

disorder of coagulation that causes blood clots (thrombosis) in both arteries and veins
as well as pregnancy-related complications such as miscarriage, stillbirth, preterm
delivery, or severe preeclampsia. The syndrome occurs due to the autoimmune
production of antibodies against phospholipid (aPL), a cell membrane substance. In
particular, the disease is characterised by antibodies against cardiolipin (anti-cardiolipin
antibodies) and β2 glycoprotein I.
The term "primary antiphospholipid syndrome" is used when APS occurs in the absence
of any other related disease. APS is commonly seen in conjunction with other
autoimmune diseases; the term "secondary antiphospholipid syndrome" is used when
APS coexists with other diseases such as systemic lupus erythematosus (SLE). In rare
cases, APS leads to rapid organ failure due to generalised thrombosis and a high risk of
death; this is termed "catastrophic antiphospholipid syndrome" (CAPS).
6
Lupus anticoagulants are the most common cause of prolonged APTT
7
Cardiolipin from a cow heart is used as an antigen in the VDRL test for syphilis.
Cardiolipin is an important component of the inner mitochondrial membrane, where it

constitutes about 20% of the total lipid composition.
The name ‘cardiolipin’ is derived from the fact that it was first found in animal hearts.
It was first isolated from beef heart in the early 1940s.
8
Lupus anticoagulant is present in 10-30% of patients with SLE, though not all of these
patients exhibit the clinical syndrome.
This anticoagulant causes an acquired hypercoaguable state in vivo via uncertain
mechanism ( platelet activation and endothelial cell inhibition)repeated second or third
trimester miscarriages are characteristic of patients with the APLA syndrome.
This may in part be due to APLA mediated inhibition of t-PA activity necessary for
trophoblastic invasion of the uterus
In vitro these antibodies cause prolongation of APTT (In fact APLA syndrome is the MCC
of prolonged APTT)
9
10
11
12
13
Q:What are the major similarities between a myocardial and a cerebral infarct?
A:The major similarity is in the etiology. Both types of infarcts are commonly caused by
thrombotic occlusion of the arteries supplying them. Thrombi usually form on the same
underlying disease process (ie, atherosclerotic arterial disease). Also, the early
histologic reactions, such as neutrophilic infiltration and granulation tissue formation,
are common to both.
Q:What are the major differences between a myocardial and a cerebral infarct?
A myocardial infarct typically features coagulative necrosis, which heals by fibrosis and
leaves behind a fibrous scar. In contrast, a cerebral infarct is typically liquefactive
necrosis, in which dead tissue is digested without being replaced by fibrosis, leaving
behind a cystic, cavitary lesion.
Question: What is the mechanism of formation of hemorrhagic infarcts in brain?

Brain infarcts can be pale or hemorrhagic. Hemorrhagic infarcts are due to arterial
occlusion followed by reperfusion. Examples are embolic occlusion followed by
fragmentation of emboli or occlusive vasospasm that later is relieved.
Acute myocardial infarct. A cross-section of the left ventricle reveals a sharply
circumscribed, soft, yellow area of necrosis in the posterior free wall (arrows).
Myocardial infarct. Transverse sections of ventricular myocardium show
(A) Reperfused infarct. Reperfusion is typically associated with hemorrhage
Healed infarct.
In C, a white scar (arrowhead) is evident in the anterior ventricular septum.
Kidney: pale infarct
Here is hemorrhagic pumonary infarction in a patient with a pulmonary
thromboembolus to a medium sized pulmonary artery.
Such infarctions tend to be based on the pleural surface and be roughly wedge-
shaped in cross section.
The small intestine is infarcted.
The dark red to grey infarcted bowel contrasts with the pale pink normal bowel at
the bottom.
Some organs such as bowel with anastomosing blood supplies, or liver with a dual
blood suppy, are hard to infarct.
This bowel was caught in a hernia and the mesenteric blood supply was constricted
by the small opening to the hernia sac.
Hemorrhagic infarction intestine
On the left is the bluish external surface of an ovary.
On the right is the cut surface which is extremely hemorrhagic.
Arrows point to fallopian tubes.
Torsion Etiology
• Frequently associated with benign or occasionally malignant ovarian enlargement
• Unusually mobile adnexa
• Increasing incidence in patients undergoing ovulation induction
Pathogenesis
• Ovary twists obstructing first venous return resulting in intense congestion
• Eventually obstructs arterial inflow
Examples of infarcts. A, Hemorrhagic, roughly wedge-shaped pulmonary infarct. B,
Sharply demarcated white infarct in the spleen.
Septic infarct. A myocardial abscess (arrow) within the left ventricular free wall was due
to infection with Staphylococcus aureus.
DIC results in decreased circulating fibrinogen and factor V and VIII levels.
Platelet count is decreased
Prothrombin time and aPTT are prolongrd due to activation of both extrinsic and
intrinsic coagulation pathways.
Damage to RBCs as they pass through microthrombi causes microangiopathic
hemolytic anemia, seen on peripheral blood smear as schistocytes.
Subsequent production of plasmin from plasminogen activates the secondary

fibrinolysis (thrombolysis) . Opens up the occluded blood vessels and releases
fibrin degradation products (FDPS) and D-Dimers.
Gram negative sepsis: release of TNF alpha à increased tissue factor release
from endothelium
AML M3 : cytoplasmic granules in neoplastic promyelocytes contain
thromboplastin à activates clotting cascade
Pancreatic adenocarcinoma : secretes mucous à activates coagulation cascade
Obstetric complications:
Placenta previa : placental tissue contains tissue factor à
activates coagulation cascade
amniotic fluid embolism : contains tissue factor.
retained dead fetus à RELEASE OF THROMBOPLASTIN FROM
PLACENTA.
Microorganisms: damage endothelium à release of tissue factor
Note:
A close monitoring of platelet count and fibrinogen level is helpful in early
identification of the onset of DIC in high risk patietns.
You should suspect DIC in any “sick patient” who has both an elevated PT and PTT.
Besides a rise in both PT and APTT, platlelts and fibrinogen levels decrease, while fibrin
degradation products, particularly D-dimer, are elevated (indicates lyses of cross linked
fibrin) in DIC
DIC:
Pateints bleed
Coagulation cascade is activated
PT and APTT are prolonged
Low fibrinogen and increased FDP
TTP/HUS
Usually do not bleed
Only platelets are activated
Normal PT and APTT
Normal fibrinogen
(B) acute (arrow) and healed (arrowhead) infarcts.
Reperfusion is typically associated with hemorrhage as in A (arrow)and B (arrow).
Neoplasia
Dr.T.Krishna MD, www.mletips.com 1

Lecture Plan
Ø General
Ø Nomenclature
Ø Characteristics of Benign and the maligant tumors
• Differentiation & Anaplasia
• Local Invasion
• Metastasis
Ø Epidemiology of Cancer
• Impact of Cancer
• Environmental factors
• Age
• Acquired Predisposing factors
• Genetic Predisposition, Interactions between Env. & Inherited factors
Ø Molecular basis of Cancer – Role of Genetic & Epigenetic Alterations
• ESSENTIAL ALTERATIONS FOR MALIGNANT TRANSFORMATION
• SELF-SUFFICIENCY IN GROWTH SIGNALS: ONCOGENES
• INSENSITIVITY TO GROWTH INHIBITION AND ESCAPE FROM SENESCENCE: TUMOR
SUPPRESSOR GENES
• EVASION OF APOPTOSIS
• LIMITLESS REPLICATIVE POTENTIAL: TELOMERASE
• ANGIOGENESIS
• INVASION AND METASTASIS
• GENOMIC INSTABILITY—ENABLER OF MALIGNANCY
• STROMAL MICROENVIRONMENT AND CARCINOGENESIS
• METABOLIC ALTERATIONS: THE WARBURG EFFECT
2
• DYSREGULATION OF CANCER-ASSOCIATED GENES
Lecture Plan
Ø Molecular Basis of Multistep Carcinogenesis
Ø Carcinogenic Agents and Their Cellular Interactions
Ø RADIATION CARCINOGENESIS
Ø MICROBIAL CARCINOGENESIS
Ø Host Defense against Tumors—Tumor Immunity
Ø TUMOR ANTIGENS
Ø ANTITUMOR EFFECTOR MECHANISMS
Ø IMMUNE SURVEILLANCE AND ESCAPE
Ø Clinical Aspects of Neoplasia
Ø GRADING AND STAGING OF TUMORS
Ø LABORATORY DIAGNOSIS OF CANCER

General

ØSecond leading cause of death in USA
Ø“When will there be a cure for cancer?”
ØGood news à cancer mortality (men and

women)↓ in last decade

Nomenclature

ØNeoplasiaà “New growth “
ØTumor àswelling caused by inflammation
ØOncologyà the study of tumors or Neoplasia
ØCancerà all malignant tumors
ØCancer (Latin) = crab
ØClonal àsingle population of cells
ØTreatment response
– Usually expressed asà 5 year survival rate

ØComponents of tumors (both Benign & Malignant)
ØParenchyma –
Øclonal neoplastic cells
ØClassification and behavior based on Parenchyma
ØStroma -
Øreactive & made of connective tissue & Vessels
ØDetermines Growth & spread of tumors
ØScant stroma & Abundant Parenchyma
à neoplasm is Soft & fleshy
ØAbundant stroma & Scant Parenchyma
àDesmoplasiaà Abundant collagen
ØExample for Desmoplastic tumor - Ca. Breast
(Scirrhous= stony hard )
Components of tumors
Parenchyma
Stroma

Desmoplastic tumor - Ca. Breast
(Scirrhous)
Parenchyma?
Stroma?
Which is abundant?
2

Neoplasia- Nomenclature
Benign
Mesenchymal Epithelial
Easy =Cell origin + oma Not easy
• Chondro + Oma = Chondroma • Adenoma à benign epithelial tumor
derived from or present with
• {Cartilage}
glandular/acinar pattern
• Fibro + Oma = Fibroma
• Papilloma à finger-like or warty
• {Fibroblast} projections from the epithelial
• Lipo + Oma = Lipoma surfaces (MC - skin)
• {Fat/ lipocyte / adipocyte} • Polyp à fist-like projection from
mucosal epithelial surfaces( MC -
colon)
• Cystadenoma: forms large cystic
mass (ovaries)
• Papillary cystadenoma: papillary
patterns that protrude into the cystic
spaces (ovaries)
Leiomyo = Smooth muscle cell
Oma= Benign Tumor of Mesenchymal cells
Skeletal is called? Dr.T.Krishna MD, www.mletips.com 12

Adenoma= benign epithelial tumor
Benign
Chondroma Adenoma - Thyroid

Malignant
Mesenchymal Epithelial
Not easy =Carcinoma (any germ layer)
Easy =Sarcoma
• Adenocarcinomaà glandular growth
Greek = “fleshy” ( little stroma) pattern
Ø Chondro + sarcoma = • Squamous cell carcinomaà squamous
Chondrosarcoma {Cartilage} cell differentiation
Ø Fibro + sarcoma = Fibrosarcoma • Specify organ of origin àRenal cell
{Fibroblast} adenocarcinoma, Bronchogenic
Ø Lipo + sarcoma = Liposarcoma • Undifferentiated/ poorly differentiated
{Fat/ lipocyte / adipocyte} àCan’t determine tissue of origin
Ø Rhabdomyo +sarcoma = • Mixed tumors àPleomorphic adenoma
Rhabdomyosarcoma
– Divergent differentiation of a single
{striated muscle} germ line of parenchymal cells
(salivary gland)
• Teratoma à From Totipotential cells
(gonads), > one germ layer { Dermoid
cyst/Ovarian Cystic Teratoma)
Malignant
Rhabdomyosarcoma Adenocarcinoma - Colon

Squamous cell = Keratin
Carcinoma= Malignancy of Epithelial cells
Anaplastic= Undifferentiated
Cancer= Malignancy
3 1
Mixed tumors àPleomorphic adenoma

1. Myxoid= resembles mucous (appear like cartilage)
2. Epithelial
3. Vessel
a
1
2 b
3
c
Cystic = resemble space with lining (Example-urinary bladder)

Teratoma= Tissues from >one germ layer
Dermoid cyst (Ovary)
a. Skin, b. Glands, C. Nerve tissue
Cyst
1
1. Wall
2. Lining
3. Contents

Malignant tumors
Feature Sarcoma Carcinoma

(Mesenchymal) (Epithelial)
Arise from mesenchymal Arise from epithelial
Origin tissue tissue
Usually larger since harder Usually smaller since
Mass size to detect them in deep easier to detect on
tissue!!! surface
Takes longer to diagnose Takes shorter period of
Time of time to diagnose from
diagnosis onset
Goes directly to vascular Goes through lymphatics
Route of system/ Blood vessels (i.e. first – longer route to
Metastasis blood) vascular system
Worse prognosis Good/worse
Prognosis
Hamartoma
disorganized but benign-appearing masses composed of cells indigenous to the
particular site.

Choristoma
congenital anomaly
Heterotopic rest of cells. (pancreatic tissue in the submucosa of the GIT).

The education what we need is not
amount of information put into our brain,
undigested all our lives.
The education what we need is

life- building, Man – making & Character-
making assimilation of Ideas.
Swami Vivekananda
Neoplasia - 2

Characteristics of Benign and the
Malignant tumors
Differentiation & Anaplasia

Local Invasion
Metastasis

ØMajority of cases
ØBenign or Malignant tumors are differentiated based on
morphology (Cellular & Nuclear) – this is part
differentiation
ØDifferentiation and Anaplasia
Ø Rate of growth
Ø Local invasion
ØMetastasis

Ø Differentiation and Anaplasia
ØDifferentiation
Øextent to which tumor cells resemble normal
parenchymal cells (morphologically and functionally)
ØBenign tumors à well-differentiated cells
ØMalignant tumors à range from
ØWell-differentiated (Grade-1)
ØModerately differentiated (Grade-2)
Ø Poorly differentiated (Grade-3)
ØUndifferentiated or Anaplastic (Grade-4)
ØHallmark of malignancy = Anaplasia
ØWhat are the features seen in Anaplasia ? (next slide)
Ø Differentiation and Anaplasia
ØHallmark of malignancy
Ø ***“ Anaplasia”
Adeno/Gland No Adeno/Gland
formations formations

Well differentiated adenocarcinoma:
shows glandular pattern, however cells
exhibit features of malignancy
Anaplastic tumors are difficult to categorize as

they can be epithelial derived or mesenchymal
derived. To distinguish, Pathologist used stains,
and for more definite diagnosis, use immuno-
histo- chemical (IHC) stains.
Poorly differentiated adenocarcinoma

(anaplastic): shows poor differentiation
and bears no semblance to parent tissue
- requires immunohistochemistry to
confirm epithelial origin
- cytokeratin stain positive: epithelial
origin (Foot note) 6
Well differentiated - Benign
Lipoma - Gross
Lipoma -
Microscopy

Adipose tissue Lipoma

Liposarcoma - Gross
Liposarcoma -
Microscopy

Well-differentiated - Benign
Well Differentiated - Malignant

Ø What is Anaplasia ?
Ø 1. Pleomorphism (pleo= poly, morphism = variance)
Ø a. Cellular pleomorphism = size and shape
Ø b. Nuclear pleomorphism = ↑DNA content,
Hyperchromatic, irregular chromatin distribution
Ø 2. Nuclear: Cytoplasmic ratio (N:C)
ØA. Prominent nucleoli
Ø 3. Mitoses
ØAtypical or abnormal (tripolar, quadripolar, ring)
Ø 4. Loss of Polarity
ØLoss of orientation (as compared to other cells)
Ø 5. Tumor Giant cell
Øvery large or cells with multiple nuclei
Ø 6. Hemorrhages
Ø 7. Central Necrosis
Anaplastic
2A
1a
2
1b

What is Anaplasia ? Cont’d…
Ø 8. Dysplasia à
Ø Disordered growth of Pleomorphic epithelium
Ø Loss of uniformity of cells & architecture
Ø Loss of Polarity/Orientation
Ø If severe = carcinoma in situ (premalignant)
Ø Uterine Cervix
Ø Barrett's Esophagus
Ø lung in smokers
Ø 9. Secretions
Ø Both benign & malignant Tumors can have secretions
Ø Secrete normally (like normal function of the cells)
Ø Squamous cell Ca.à keratin
Ø Hepato - cellular carcinoma à Bile
Ø Secrete hormones / like substances
Ø↑ blood levels of secretions à help in Diagnosis
ØNew or unexpected production (paraneoplastic)
ØSmall (Oat) cell ca. of Lungà ACTH like peptide (Cushing's
syndrome),ADH Dr.T.Krishna MD, www.mletips.com 13
Carcinoma in situ
Ectocervix
Normal Carcinoma In Situ
Basement membrane

Carcinoma
Carcinoma in
in situ
situ
Ectocervix
Lower Magnification Higher Magnification
Perinuclear halo
Loss of polarity

Carcinoma in situ Vs. Invasive
Carcinoma in situ Carcinoma
Ectocervix
CIS Squamous cell carcinoma
Basement membrane (BM)

Keratin pearl
Where is BM?

Metaplasia
Ø Replacement of one type of cell with another type?
Ø (death of native tissue and the stems programmed to
produce different cell type)
Ø Barrett's Esophagus
Ø Squamous Metaplasia of lung in smokers
Ø Associated with tissue damage, repair, and
regeneration
Ø Why metaplasia is needed?
Ø Metaplastic cells withstand the adverse environment
Ø Acts as fertile soil for development of Neoplasia
ØBarrett's Esophagus à Adenocarcinoma of Esophagus
ØSquamous Cell Carcinoma ofwww.mletips.com
Dr.T.Krishna MD, Lungs in Smokers 17
Metaplasia
Reprogramming
of Stem
cell

Rates of growth
ØSignificance
ØBy the time of diagnosis, neoplasm completes major
part of life cycle
ØHelps only in treatment aspects
ØFactors determine growth
ØDoubling time of tumor cells
ØCells lost in process (apoptosis)
ØGrowth fraction - GF
ØProportion of proliferating cells
Ø1. Rapidly growing à 20% GF (Leukemia, Lymphoma,
Lung Ca. - Small cell type)
Ø2. Slow growingà 5% GF (Cancers of Colon, Breast)
Growth Fraction - determinants
Clonal
1. Tumor Doubling time

2. Tumor cell death (apoptosis)
Rate of growth
Cell division (doubling

time)= 3days
One cancer 1. Karyokinesis -1 day
cell 2. Cytokinesis-2days
30 divisions ? days
1gram
Just difficult to Tumor
diagnose Heterogeneity
10 divisions
? days
1kg
Metastasize
and die

Rates of growth –Tumors cont’d…
ØGeneral Rules
ØBenign & well-differentiated malignant tumors àslow
growing
ØModerately, poor, and un-differentiated malignant
tumorsà have faster or erratic growth
ØExceptions
ØLeiomyoma à growth spurt during pregnancy
ØSpontaneous remission or benign transformation of
malignancies àNeuroblastomas, Malignant melanoma

ØLocal invasion
ØNext to metastasis, Local invasion is the most reliable feature of
malignant tumors
Feature Benign Malignant

invasion no yes
course slow growing, cohesive Infiltrate & invade
expansible masses &, àdestroy surrounding
do not infiltrate, tissue
invade, or metastasize
to distant sites
Capsule Present (except in Absent

Hemangioma)
Significance easily palpable & difficult to remove

easily removable Need wide excision

Local Invasion
capsule
Fibroadenoma -Breast Carcinoma -Breast

Metastasis
Ødiscontinuous spread of tumor
ØThe most reliable feature to differentiate malignant
from benign tumors
ØAll cancers can metastasize (few exceptions)
Ø Exceptions
ØGliomas à tumors of glial cells in CNS
ØBasal cell carcinoma( of skin) à rarely metastasize
ØMalignancies with ↑ likelihood of metastasis
ØMore aggressive
ØMore rapidly growing
ØLarger neoplasms
Ø Significance of Metastasis
Ø↓cure rate
Ø30% of malignancies metastasize at the time of diagnosis

Metastasis
1. Multiple
2. Superficial
3. Central Umbilicated (like umbilicus)

Metastasis
Ø Pathways of spread
Ø1. Seeding of body cavitiesà peritoneal (ovarian Ca.)
Ø2. Lymphatic spread = MC in carcinomas
Øfollows the natural routes of drainage
ØIf bypassed à“skipped metastasis” (Malignant melanoma)
Ø3. Hematogenous spread = MC in early stages of
sarcomas
ØVeins > arteries (arteries- thicker walls)
ØLiver and lungs are most frequently involved
Ø Liver àAll portal area drainage flows to the liver
Ø Lungs àAll caval blood flows to the lungs
Ø4. Malignancies with high tendency for invasion of veins
ØRenal cell carcinoma àrenal vein
ØHepatocellular carcinomaà hepatic vein
Ø Best strategy is of Cancer Rx.
ØPrevention of metastasis (most beneficial)
Benign Vs. Malignant

Differentiation &Anaplasia
Stem cell
Proliferate & Differentiated Proliferate & undifferentiated
Benign Malignant
For Practice

Tumor from Adiopcytes
Degree of Differentiation ?

Finger or Fist?
Papilloma or Polyp?

Finger or Fist?
Papilloma or Polyp?

Benign tumor of Cartilage?
Chondroma

Benign tumor of fibroblasts in Ovary ?
Follicular cyst
Fibroma

Malignant tumor of stomach
Adenocarcinoma

Derived from single germ layer
Glandular
Cartilage
Myxoid
areas
Pleomorphic adenoma 37
Derived from > one germ layer
Teratoma
Demoid cyst in ovary

Different sizes and shapes of Nuclei and cells
Pleomorphism

Nuclear : Cytoplasmic ratio
High N:C ratio

Dark Nuclei
Hyperchromasia

Mitosis
Tripolar Tetra polar

Mitosis
Bipolar – normal or abnormal
Stellate mitosis

Polarity/Orientation

Size of tumor cells?
Tumor Giant cells

Strength is Life.
Fear is Death.
Swami Vivekananda
Neoplasia – Part 3

Epidemiology of Cancer
1. Impact of Cancer
2. Geographic & Environmental
factors
3. Age
4. Genetic Predisposition,
Interactions between Env. &
Inherited factors
5. Acquired(NONHEREDITARY)Pre-
disposing factors
Epidemiology of Cancer
• Study of cancer patterns in populations
• Benefits of knowledge about Epidemiology of
cancers
– smoking and lung cancer
– high dietary fat and low fiber in the development of
colon cancer in Western world and Africa
– Relation between causes of cancer & environmental,
racial (possibly hereditary), and cultural influences
– Knowledge about pre-neoplastic disorders (last slide)
à gives clues to the pathogenesis of cancer

1. Impact of Cancer

1. Impact of Cancer
Cancer incidence
Ø ***2nd MCC of all the deaths in USA (BEQ)
Ø MC cancers in men
Ø Prostate – 25%
Ø Lung - 15%
Ø Colon and rectum – 10%
Ø MC cancers in women
Ø Breast – 26%
Ø Lung -14%
Ø Colon and rectum – 10%
Ø ***MC cancer deaths (in both sexes)
Ø LUNG CANCER (M Vs. F = 31% Vs. 26%) (BEQ)

Cancer Incidence
(BEQ)
(BEQ)

Cancer Deaths
(BEQ) (BEQ)

1. Impact of Cancer
ØRisk in dying with cancer in USA
Øone in five chance
ØCancer deaths (in 2008)
Ø 23% of all mortality
Ø> 50% of cancer diagnoses and cancer
deaths in USA
Ølung, female breast, prostate, colon/rectum
ØAge-adjusted cancer death rate among
sexes (in last 50 years)
Øsignificantly ↑ but
1. Impact of Cancer
Øsex-specific in cancer death rates
Ø↓by 40% due to
ØDecreased use of tobacco – Lung cancer
Øimproved detection and treatment - in
colorectal, female breast, and prostate ca.
Ø***↓ incidence of cancer –
Øcervical cancer, stomach cancer (BEQ)
Ø↑ cancer death rates in
ØLung Ca. in women
Ø liver and intrahepatic bile duct in men
{(hepatitis C virus (HCV)}
Racial Difference
ØCancer mortality rates between white
and black Americans
Ødifferent
ØAfrican Americans
Ø*** largest decline in cancer mortality
ØHispanics
Ølower frequency of the MC tumors in USA
Øhigh frequency of stomach, liver, uterine cervix,
and gallbladder Ca. , & childhood Leukemias
Racial Difference bothers

2. Geographic & Environmental factors
The only certain way of avoiding cancer is ?
not to be born

2. GEOGRAPHIC AND ENVIRONMENTAL FACTORS
ØEnvironmental factors
ØSignificant role in MC sporadic Ca.
ØRisk from env. causes – 65%
ØHeritable factors
ØRisk is on average – 32%
ØStomach carcinoma
Ø 7-8X higher in Japan (in Japan Vs. in USA)
Øcomparing mortality rates for
Japanese immigrants (next slide)
2. GEOGRAPHIC AND ENVIRONMENTAL FACTORS
Japanese in Japan
2nd Generation
Japanese in USA

2. ENVIRONMENTAL FACTORS
ØImmigration
ØCa. Stomach in Japanese's immigrants to USA
ØSun exposure Skin cancers & Ultraviolet rays
ØNew Zealand & Australia
ØOccupational
ØAsbestos, Vinyl chloride, 2-naphthylamine
ØObesity
Ø52% & 62% ↑ risk of cancers in Males &
Females respectively
ØAlcohol
ØGIT cancers
ØCigarette smoking
ØLung cancers
ØSexual Practices
Ø Cervical Cancers
2. . ENVIRONMENTAL FACTORS
BEQ***

2. OCCUPATIONAL FACTORS BEQ***

3. Age

Age
ØIncidence of cancers with age

Ø↑↑
ØMC cancer age group
Ø≥ 50 yrs.
ØMC cancers in Infants & Children –
ØBlastomas
ØMC cancers in Older Children (± 15
years)
ØAcute leukemia & CNS neoplasms (2nd
MCC of deaths)
4. Genetic Predisposition

4. GENETIC PREDISPOSITION TO CANCER
Ø“My mother and father both died of cancer. Does that
mean I am doomed to get it?”
ØHow to answer this question ?
ØBest Example - lung cancer
Ørelated to cigarette smoking (Environmental)
Ølung cancer deaths – 4X common in non - smoking
relatives (parents and siblings) of lung cancer patients
as compared to nonsmoking relatives of controls
(Genetic)
Ø***Risk of inherited mutations predispose to cancer
Ø<10%

4. GENETIC PREDISPOSITION TO CANCER
BEQ***

Feature Inherited cancer syndromes Familial cancers (AD but AR syndromes and
(AD) multifactorial) defective DNA repair
Specific phenotype Yes No -

specific sites and tissues
Li-Fraumeni - exception
Unique features Inheritance of a single 1. Early age chromosomal or DNA
mutant gene à greatly ↑ 2. affected close instability (Xeroderma
risk of cancer relatives (risk – 2- pigmentosum, Ataxia
3X) telangiectasia, Bloom
3. multiple or bilateral syndrome, Fanconi
cancers Anemia)
4. transmission pattern
– not clear
Examples 1. Retinoblastoma with 1. BRCA1 & BRCA2 - HNPCC –
Osteosarcoma*** BEQ breast or ovarian
1. inactivation of a
2. Familial adenomatous cancers
DNA mismatch repair
polyposis 2. p16 tumor
gene
3. Li-Fraumeni syndrome suppressor gene
(TSG) - familial 2. ***MC cancer pre-
4. Multiple Endocrine
Neoplasia types 1 and 2 melanomas disposition syndrome
(MEN-1 and MEN-2) (Familial association is 3. cancer of the colon,
5. hereditary nonpolyposis 20% in both) the small intestine,
colon cancer (HNPCC) endometrium, and
ovary
5. NONHEREDITARY PREDISPOSING
CONDITIONS

5. NONHEREDITARY PREDISPOSING CONDITIONS
Proliferations
ØFertile soil for the cancer development
ØProliferations can be –
ØRegenerative – in HBV infection of liver
ØMetaplastic – Barrett’s Esophagus
ØHyperplastic – Endometrial (in Tamexifen Rx.
For Breast Ca.)
ØDysplastic – Ectocervix,
ØWhy proliferations make fertile soil for Cancers?
ØProliferating cells that accumulate the
genetic lesions required for carcinogenesis
Chronic Inflammation and Cancer
ØVirchow in 1863
ØCancer develops at sites of chronic inflammation by
ØExamples
Øulcerative colitis,
Ø Helicobacter pylori gastritis,
Ø viral hepatitis, and
Øchronic pancreatitis
ØPrecise mechanisms –
Ø? (unknown)
ØPossibilities –
ØImmune response may become maladaptive,
promoting tumorigenesis
ØExample - cyclooxygenase-2 (COX-2)à induced by
inflammatory stimuli, and ↑in colon and other cancers
ØCOX-2 inhibitors for cancer treatment is an active area of
research
Chronic Inflammation and Cancer
↑ Growth factors, cytokines, chemokines
cause genomic stress ↑ stem cells pool

and mutations

Precancerous Conditions
1.Chronic atrophic gastritis of pernicious anemia
2.Solar keratosis of the skin
3.Chronic ulcerative colitis (inflammatory bowel disease)
4.Leukoplakia of
A. Oral cavity
B. Vulva
C. penis
ØMajority of these lesions no malignant neoplasm emerges
ØBut there is increased risk of cancers
Ø***Biopsy is a must
ØVillous adenoma of the colonà risk of malignancy 50%
ØMost benign neoplasms do not become cancerous
“Everything is easy when you are busy”
“Nothing is easy when you are lazy”
Swami Vivekananda
Neoplasia

Molecular basis of cancer
Lecture Plan
1. FUNDAMENTAL PRINCIPLES
2. ESSENTIAL ALTERATIONS FOR MALIGNANT TRANSFORMATION
a. SELF-SUFFICIENCY IN GROWTH SIGNALS: ONCOGENES
b. INSENSITIVITY TO GROWTH INHIBITION AND ESCAPE
FROM SENESCENCE: TUMOR SUPPRESSOR GENES
c. EVASION OF APOPTOSIS (APOPTOTIC GENES)
d. GENOMIC INSTABILITY— (DNA REPAIR GENES)
e. LIMITLESS REPLICATIVE POTENTIAL: TELOMERASE
f. ANGIOGENESIS
g. INVASION AND METASTASIS
3. STROMAL MICROENVIRONMENT AND CARCINOGENESIS
4. METABOLIC ALTERATIONS: THE WARBURG EFFECT
5. DYSREGULATION OF CANCER-ASSOCIATED GENES

Ø Primary target of cellular damage
Ø Certain genes
Ø Most important step in carcinogenesis
Ø Non - lethal genetic damage
Ø Neoplasia is a
Ø multi step Process
Ø result of Non-lethal cellular
damage (multiple mutations)
Ø Tumors are
Ø Monoclonal – How to know? (next slide)
Ø To determine tumor clonality
Ø MC method used - analysis of methylation patterns
adjacent to polymorphic locus of ***AR gene
(Androgen Receptor –AR)
Ø why?
Ø Frequency of such polymorphisms of AR gene > 90%
Ø If all the tumor cells show acquired cytogenetic
aberrations (for e.g., a translocation)à monoclonal
Ø Markers of clonality in B- and T-cell lymphomas
Ø Study of Ig receptor (in B cell) and T-cell receptor
gene (T-cell) rearrangements

Principal regulatory genes are main targets
Ø 1.Protooncogenes
Ø Proto-oncogene are growth promoting à normal
Ø Oncogenes are due to mutations in Proto-oncogene à
abnormal
Ø Mutation of one allele of proto-oncogenes behave as
Autosomal Dominant à transform cells to neoplastic (unlike
TSG)
Ø 2.Anti-oncogenes / Tumor-suppressor genes (TSG)
Ø Mutation of both normal alleles of TSG is a must for
transformation
Ø Exception à mutation of one TSG allele and LOF lead haplo-
insufficiency
Ø Growth inhibiting genes - p53 & Rb
Ø Suppress cell proliferation.
Ø Mutations of TSG à release cells from growth suppression &
lead to hyper-proliferation
Ø 3.Apoptotic genes
Ø Regulate programmed cell death
Ø Act as either dominant or TS genes
Ø 4. Genes regulating damaged DNA
Ø Behave in unique fashion (unlike the other three types of
genes)
Ø Mutations of DNA repair genes à
Ø do not directly transform cells
Ø Cells with DNA-repair genes (called ***mutator
phenotype)à predispose cells to mutations in the
genome (Proto-oncogenes, TSG, Apoptotic genes)
Ø Others - microRNAs (miRNAs)
Ø Do not encode proteins but behave like Proto-oncogenes,
TSG by affecting translation of other genes
Role of Genetic & Epigenetic Alterations
Ø How we got his information ?
Ø DNA sequencing and others
Ø ***Non-lethal Genetic damage
Ø Initial damage and a must to develop cancer
Ø Caused by Environmental, inherited, spontaneous and random
(bad luck)
Ø Clonal expansion of cell with non-lethal genetic damage
Ø Tumors are monoclonal/Clonal
Ø All cells in the tumor share same set of mutations

Ø Four classes of Genes (mentioned in previous slides)
Ø The principle targets of cancer – causing mutations
1. Proto- Oncogenes à Oncogenes (GOF mutations, like AD type)
2. Tumor suppressor Gene mutations à LOF , AR type, some times –
haplo-insufficiency)
3. Apoptosis –regulating gene mutations - imbalance
1. less death /increased survival (GOF àof Anti – Apoptotic genes
or
2. More death/ decreased survival(LOF à of Pro-apoptotic genes)
4. DNA repair gene mutations à LOF,
1. Makes the cells unable detect and fix the non-lethal genetic
damage
2. Cause s acceleration of additional mutations,
3. Called Mutator phenotype or Genomic instability
Ø Epigenetic aberrations (important in cancer treatment)
Ø Changes are reversible (genetic ones are irreversible)
1. DNA methylations à silence gene expression (of TSG)
2. Histone modifications à effects of gene expression of cancer cells
Ø Hallmarks of Cancer à Essential alterations
2. Essential Alterations
1. Self sufficiency in growth signals
Ø No need of external stimuli
2. Insensitivity of growth inhibitory signals
Ø TGFβ inhibits proliferation of normal cells
Ø Cyclin - Dependent Kinase Inhibitors (CDKI)
3. Evasion of Apoptosis
Ø Carcinoma cells resistant apoptosis by
Ø inactivation of p53)
Ø activation of anti-apoptotic genes
4. Defects in DNA repair
Ø NER (Nucleotide Excision Repair)
5. Limitless replication potential
Ø telomerase reactivation
6. Sustained angiogenesis
Ø VEGF
7. Ability to invade & metastasis
Ø ↑cancer deaths
2(a).Self sufficiency in growth signals
Oncogenes

Proto-oncogenes & Oncogenes
Protooncogenes Oncogenes
Proteins oncoprotein
Growth Factors No need of Growth Factors

(Hormones etc.)
Normal Growth Malignant Growth
Oncoprotein àsimilar functions as their normal counterparts

but make cells self-sufficient to grow
D
Figure 7.25

2(a). Oncogenes
Normal Cellular counterpart of Genes Homologous
to Oncogenes
1. Growth Factors
2. Growth Factor Receptors
3. G Proteins
4. Kinases – Tyrosine & Cyclin dependent (major
ones)
5. Gene Regulatory Phosphoproteins
6. Transcription

1. Oncogenes :Growth Factors
Proto-oncogene Mechanism Associated tumors

1. PDGF-β CHAIN over expression astrocytoma and
Osteosarcoma
2. TGF-α with EGF cause over expression of Astrocytomas (TGF-α)
growth factor genes
3. HGF ( its receptor- over expression Carcinomas of the
c-MET) thyroid
4. FGF
Proto – Oncogenes
HST 1 over expression STOMACH CANCER
INT 2 (FGF3) Amplification bladder, breast, and

melanoma
Growth Factors
Ø In normal and cancer cells
Ø GF’s stimulate proliferation of cells
Ø In normal cells
Ø GFs’ have Paracrine action
Ø In cancer cells
Ø GFs’ are autocrine
Ø Glioblastomas express both PDGF & PDGFR TK
Ø Sarcomas over express TGFα and its receptor – EGFR
Ø Commonly GF genes are not altered or mutated
Ø Other Oncoproteins cause over expression or ↑secretion of GFs’

TK Signal Pathway
Growth Factor
RTK (Receptor Tyrosine Kinase)
RAS
MAPK PI3/AKT
If RAS mutations are present in a tumor, RTK mutations will be absent

These are mutually exclusive
Very important in Lung cancer treatment strategy.
Receptor Tyrosine Kinase (RTK)
Normal
1. Most important
2. Transmembrane protein à Following stimulation by
GFs’, RTK phosphorylates (from GDP to GTP state)
and dimerise
3. Recruit signalling pathway à RAS
Mutated RTK
1. TK activity is GF – independent
2. TK continues to send signals down stream even in the
absence of GFs’

3. How RTK mutations takes place?
A. Point mutations:
• ERBB! à encodes EGFR ,
• mutations of ERBB1 à activation of EGFR (RTK)
• Lead to Lung Adenocarcinoma
B. Amplifications lead to over expression:
• ERBB2 encode HER2 (RTK);
• mutations of ERBB2à activation of HER2 (RTK)
• Lead yo Breast Ca.
• Anti – bodies to HER2 used to Rx. Breast Ca.
3. Gene rearrangements:
• deletion on Ch.5: lead to fusion of part of ALK gene with that of EML4
• Lead to Fusion/ Chimeric gene and it has RTK activity
Ø In early Lung cancer àRx with inhibitors of EGFR/ALK is
effective
Ø In Advanced Cancers à Rx. resistance to inhibitors of
EGFR/ALK is due to mutations
Dr.T.Krishna of other RTK(MET)
MD, www.mletips.com 19
Ø In

Ø In

Growth Factor Receptors

2.
1. S

2.
1. S

D

2.
1. S

D

2(b). Insensitivity of growth
inhibitory signals
TGFβ inhibits proliferation of
normal cells
Cyclin - Dependent Kinase
Inhibitors (CDKI)

Molecular basis of cancer - Summary
29
De
God’s love for is Unconditional

Retinoblastoma
Feature Inherited/ Sporadic

Familial
Age Childhood Adult
Frequency 40% 60%
Bilateralism Yes No
Second Yes No
malignancy
Time of 1st hit Germ cells Somatic cells
P53 mechanism
DNA damage
( direct or indirect)
↑p53
G1 arrest
degrade
Successful Repair No repair
MDM2 ↑ BAX & ↓Bcl-2
apoptosis
Evasion of Apoptosis
↑proapoptotic ↓antiapoptotic
genes cell survival genes
BCL-2 – anti - apoptotic
q85% of B-cell Follicular lymphomas (NHL)
qt(14;18)(q32;q21) translocation
qBCL-2 gene from 18q21 translocated to 14q32
qover expression of the BCL-2
Clinical Significance
MC Non Hodgkin’s Lymphoma(NHL) in adult population of USA
= Follicular lymphomas
indolent (slow growing), low grade
Invasion and Metastasis
Phase 1
Invasion of the ECM
phase 2
Vascular
dissemination
&
Homing of tumor cells
Neoplasia

Lecture Plan
f. ANGIOGENESIS

Ø Primary target of cellular damage
Ø Certain genes
Ø Most important step in carcinogenesis
Ø Non - lethal genetic damage
Ø Neoplasia is a
Ø multi step Process
Ø result of Non-lethal cellular
damage (multiple mutations)
Ø Tumors are
Ø Monoclonal – How to know? (next slide)
Ø To determine tumor clonality
Ø MC method used - analysis of methylation patterns
adjacent to polymorphic locus of ***AR gene
(Androgen Receptor –AR)
Ø why?
Ø Frequency of such polymorphisms of AR gene > 90%
Ø If all the tumor cells show acquired cytogenetic
aberrations (for e.g., a translocation)à monoclonal
Ø Markers of clonality in B- and T-cell lymphomas
Ø Study of Ig receptor (in B cell) and T-cell receptor
gene (T-cell) rearrangements

Principal regulatory genes are main targets
Ø 1.Protooncogenes
Ø Proto-oncogene are growth promoting à normal
Ø Oncogenes are due to mutations in Proto-oncogene à
abnormal
Ø Mutation of one allele of proto-Oncogenes behave as
Autosomal Dominant à transform cells to neoplastic (unlike
TSG)
Ø 2.Anti-oncogenes / Tumor-suppressor genes (TSG)
Ø Mutation of both normal alleles of TSG is a must for
transformation
Ø Exception à mutation of one TSG allele and LOF lead haplo-
insufficiency
Ø Growth inhibiting genes - p53 & Rb
Ø Suppress cell proliferation.
Ø Mutations of TSG à release cells from growth suppression &
lead to hyper-proliferation
Ø 3.Apoptotic genes
Ø Regulate programmed cell death
Ø Act as either dominant or TS genes
Ø 4. Genes regulating damaged DNA
Ø Behave in unique fashion (unlike the other three types of
genes)
Ø Mutations of DNA repair genes à
Ø do not directly transform cells
Ø Cells with DNA-repair genes (called ***mutator
phenotype)à predispose cells to mutations in the
genome (Proto-oncogenes, TSG, Apoptotic genes)
Ø Others - microRNAs (miRNAs)
Ø Do not encode proteins but behave like Proto-Oncogenes,
TSG by affecting Dr.T.Krishna
translation of other genes
MD, www.mletips.com 7
Ø How we got his information ?
Ø DNA sequencing and others
Ø ***Non-lethal Genetic damage
Ø Initial damage and a must to develop cancer
Ø Caused by Environmental, inherited,
spontaneous and random (bad luck)
Ø Clonal expansion of cell with non-lethal genetic
damage
Ø Tumors are monoclonal/Clonal
Ø All cells in the tumor share same set of
mutations
Ø Four classes of Genes (mentioned in previous slides)
Ø The principle targets of cancer – causing mutations
1. Proto- Oncogenes à Oncogenes (GOF mutations, like AD type)
2. Tumor suppressor Gene mutations à LOF , AR type, some times –
haplo-insufficiency)
3. Apoptosis –regulating gene mutations - imbalance
1. less death /increased survival (GOF àof Anti – Apoptotic genes
or
2. More death/ decreased survival(LOF à of Pro-apoptotic genes)
4. DNA repair gene mutations à LOF,
1. Makes the cells unable detect and fix the non-lethal genetic
damage
2. Cause s acceleration of additional mutations,
3. Called Mutator phenotype or Genomic instability
Ø Epigenetic aberrations (important in cancer treatment)
Ø Changes are reversible (genetic ones are irreversible)
1. DNA methylations à silence gene expression (of TSG)
2. Histone modifications à effects of gene expression of cancer cells
Ø Hallmarks of Cancer à Essential alterations
2. Essential Alterations
1. Self sufficiency in growth signals
Ø No need of external stimuli
2. Insensitivity of growth inhibitory signals
Ø TGFβ inhibits proliferation of normal cells
Ø Cyclin - Dependent Kinase Inhibitors (CDKI)
3. Evasion of Apoptosis
Ø Carcinoma cells resistant apoptosis by
Ø inactivation of p53)
Ø activation of anti-apoptotic genes
Ø NER (Nucleotide Excision Repair)
5. Limitless replication potential
Ø telomerase reactivation
6. Sustained angiogenesis
Ø VEGF
7. Ability to invade & metastasis
Ø ↑cancer deaths
2(a).Self sufficiency in growth signals
Oncogenes

Proto-Oncogenes & Oncogenes
Protooncogenes Oncogenes
Proteins oncoprotein
Growth Factors No need of Growth Factors

(Hormones etc.)
Normal Growth Malignant Growth
Oncoprotein àsimilar functions as their normal counterparts

but make cells self-sufficient to grow
2(a). Oncogenes
Normal Cellular counterpart of Genes Homologous
to Oncogenes
1. Growth Factors
3. G Proteins
4. Kinases – Tyrosine & Cyclin dependent (major
ones)
5. Gene Regulatory Phosphoproteins
6. Transcription

1. Oncogenes :Growth Factors
Proto-oncogene Mechanism Associated tumors

1. PDGF-β CHAIN over expression astrocytoma and
Osteosarcoma
2. TGF-α with EGF cause over expression of Astrocytomas (TGF-α)
growth factor genes
3. HGF ( its receptor- over expression Carcinomas of the
c-MET) thyroid
4. FGF
Proto – Oncogenes
HST 1 over expression STOMACH CANCER
INT 2 (FGF3) Amplification bladder, breast, and

melanoma
1. Growth Factors
Ø In normal and cancer cells
Ø GF’s stimulate proliferation of cells
Ø In normal cells
Ø GFs’ have Paracrine action
Ø In cancer cells
Ø GFs’ are autocrine
Ø Glioblastomas express both PDGF & PDGFR TK
Ø Sarcomas over express TGFα and its receptor – EGFR
Ø Commonly GF genes are not altered or mutated
Ø Other Oncoproteins cause over expression or ↑secretion of GFs’

TK Signal Pathway
Growth Factor
RTK (Receptor Tyrosine Kinase)
RAS
MAPK PI3/AKT
If RAS mutations are present in a tumor, RTK mutations will be absent

These are mutually exclusive
Very important in Lung cancer treatment strategy.
Normal
1. Most important
2. Transmembrane protein à Following stimulation by
GFs’, RTK phosphorylates (from GDP to GTP state)
and dimerise
3. Recruit signalling pathway à RAS
Mutated RTK
1. TK activity is GF – independent
2. TK continues to send signals down stream even in the
absence of GFs’

3. How RTK mutations takes place?
A. Point mutations:
• ERBB1 à encodes EGFR ,
• mutations of ERBB1 à activation of EGFR (RTK)
B. Amplifications lead to over expression:
• ERBB2 encode HER2 (RTK);
• mutations of ERBB2à activation of HER2 (RTK)
• Lead yo Breast Ca.
• Anti – bodies to HER2 used to Rx. Breast Ca.
3. Gene rearrangements:
• deletion on Ch.5: lead to fusion of part of ALK gene with that of EML4
• Lead to Fusion/ Chimeric gene and it has RTK activity
Ø In early Lung cancer à
Ø Rx with inhibitors of EGFR/ALK is effective
Ø In Advanced Cancers à
Ø Rx. resistance to inhibitors of EGFR/ALK is due to mutations of other
RTK(MET)

3. Signal Transduction Pathways
RAS
• Point mutations – MC mutations involving proto-oncogenes in
humans
• Types (studied in Retroviruses) - à H, K , N
• Members of family of membrane associated small G proteins
• Bind to GTP in active and GDP in inactive forms(GDP à GTP is
Phosphorylation)
• Up to 20% of human tumors have RAS mutations
– Pancreatic, Cholangiocarcinoma – 90%
– Colon, Endometrial Ca.--> 50%
– Lung (Adeno), Myeloid LEukemias -à 30%
• RTKàGDPRAS to GTPRASà MAKP/PI3/AKT signal pathwaysà
activate Cytoplasmic & TFs’

RAS cont’d…
GAPs’ à GTPase activating proteins)
• Convert GTP to GDP RAS (inactivate) – protect from
uncontrolled RAS activity
• Point mutations of RAS lead to
– Low GTPase activity à Uncontrolled RAS activityà proliferative
activity
– GOF mutations of RAS or LOF of GAP has same effect
• NF-1 (Neurofibromatosis -1 à LOF of GAP

• MAPK –BRAF (GOF) mutations
BRAF
• Serine/Threonine Kinase
• Acts at top of MAPK pathway
• Mutations of BRAFà activate TFs’
– 100% cases of Hairy cell Leukemia (B cell neoplasm)
– 80% of Benign Nevi(Mole)
– 60% Melanoma (Specific Anti -BRAF antibodiesà Very
successful,
– Oncogene addiction – only melanomas with BRAF
mutations show best treatment response

PI3K –
• like BRAF, activates Serine/Threonine Kinase
• has two sub-units (regulatory, catalytic)
• Mechanism: RTKà RASà PI3Kà AKT
• AKT acts on
1. mTOR – sensor of cellular nutirion status; if activatedà
protein and lipid synthesis
2. BAD à TF; Pro-apoptotic (positively regulated by AKT)
3. FOXO à TF; Pro-apoptotic (negatively regulated by AKT)
• PI3K is negatively regulated by PTEN (TSG)
• PI3K and PTEN mutations –
– Most common PI3K mutations (catalytic subunit)à GOF
mutations in Breast Ca. à 30% cases
– PTEN mutationsà LOF Endometrial Ca.
• Rx. Strategies against RAS àunsuccessful
– 1.Complex RAS structure
– 2. Complex functionality
• Rx. Strategies against BRAF (MAPK)àmost
successful
– Oncogene addiction à Rx. Is best in BRAF
mutation associated melanomas only

NonRTK (NRTK)
1. BCR:ABL reciprocal translocations
• ABL +BCR onCh.22 forms fusion/chimeric gene (Ch.22 –
Philadelphia chromosome)
• Fusion gene à TK activity à CML, ALL (Leukemias)
• Oncogene addiction is also seen in ABL:BCR (Targeted
Rx)
• Treatment failure : Stem cells with ABL:BCR persist
even after Rx.
• 2. JAK/STAT pathway
• Activate TFs’
• JAK2 mutations à release Hemopoietic stem cells from
GF dependence
– Myelo Proliferative Disorders (MPD) – Polycythemia Vera,
Essential Thrombocythemia, Myelofibrosis etc.,

4. Transcription Factors (TFs’)
• Mutations lead to expression of pro-growth
genes, Cyclins
• Examples of TFs à MYC, MYB,JUN, FOS, RET
• MC and important à MYC
MYC
• Master transcription regulator of cell growth
• Expressed in all eukaryotic cells
• Immediate and early response gene
• RAS/MAPKà cause rapid transient induction
of MYC
• Myc concentrations- under strict control ( by
SNPs’) Dr.T.Krishna MD, www.mletips.com 28
• SNP
1.Increase Conc. Of MYC in Ca. Prostate, Ovary
2.Also activate expression of many genes ( up regulate –
Cylcin-D, rRNA gene expression and processing)
3.Upregulate metabolic reprogramming and Warburg effect
(on of the hallmarks of cancer)
4.Burkitt’s Lymphomaà Fastest growing human tumor
Ø MYC translocations - Always involved in Burkitt's
5. Also increase Telomerase expression à tumor cell
immortality
6. MYC + other TFs’ à reprogram somatic cells to Pleuripotent
stem cells in cancers
• MYC cell translocations àBurkitt's, Some T, B cell tumors
• MYC amplificationsà Ca. Breast, Colon, Lung, others
• N,L Myc amplifications àNeuroblastomas, Small cell Ca. of
Lung

5. Cyclins/CDK
Cyclins/CDK
• Cylcin + CDKà cell division
– (cyclic nature of production and degradation)
• CDKI à inhibit cell division
• Cell Cycle check points
– G1/S à GOF mutations in CyclinD and CDK4
oncogenes
• Cyclin mutations
– Translocationsà Lymphomas
– Amplificationsà Solid tumors
• CDK4 amplificationsà Melanomas, Sarcomas,
Glioblastomas
6. CDKI
• CDKI à inhibit cell division
• P16CDKN2A ( CDKIs’) germ line mutations
àinhibit CyclinD+CDK4 complex
– 75% - Ca. Pancreas
– 55% - Glioblastomas
– 50% -Ca. Esophagus
– 35% - ALL
– 20% -Non small cell Lung cancers, soft tissue
sarcomas, Bladder ca.

Cyclins/CDK

Neoplasia

Lecture Plan
f. ANGIOGENESIS

Malignant Transformation
Essential Alterations
1. Self sufficiency in growth signalsà no need of
external stimuli
2. Insensitivity of growth inhibitory signalsà
TGFß
3. Evasion of Apoptosisà Ca. cells resistant
apoptosis ( inactivation of p53)
5. Limitless replication potential à telomerase
reactivation
6. Sustained angiogenesisà VEGF
7. Ability to invade & metastasis à ↑cancer
deaths
EVASION OF APOPTOSIS
Ø Cells with genomic injury are directed to death
Øto avoid accumulation of mutations
Ø Triggers for apoptosis
ØMany (Loss of membrane integrity, DNA damage etc.)
Ø Pathways of Apoptosis
ØExtrinsic - CD95/Fas & FAS ligand
ØIntrinsic – starts with leak of cytochrome c from
mitochondria
Ø Apoptosis regulating Proteins
ØPro -apoptotic proteins BAX and BAK etc.
ØAnti-apoptotic proteins - BCL2 of BCL2 family and BCL-XL
ØBalancing Proteins - BAD, BID, and PUMA
Ø Caspases cause apoptosis (C- cytokine proteases,
aspase - cleave after aspartic acid)
Caspases
Small number (3) –
executioners
Big numbers (8,9) -
Imitators
6
How tumors evade Apoptosis?
ØFLIP (protein produced by tumors)
Øprevents activation of Caspase 8
ØOver expression of BCL2 of BCL2 family (anti-
apoptotic) due to
Ø***t(14;18)(q32;q21) translocation
Ø seen in 85% of follicular lymphomas (B-cell)
ØTumor grows by reduced cell death rather than explosive
cell proliferation à indolent or slow growing
Øp53 -pro-apoptotic gene
ØMediated through activation of BAX

How tumors evade Apoptosis?
ØFLIP (protein produced by tumors)
Øprevents activation of Caspase 8
ØOver expression of BCL2 of BCL2 family (anti-
apoptotic) due to
Ø***t(14;18)(q32;q21) translocation
Ø seen in 85% of follicular lymphomas (B-cell)
ØTumor grows by reduced cell death rather than explosive
cell proliferation à indolent or slow growing
Øp53 -pro-apoptotic gene
ØMediated through activation of BAX

external stimuli
TGFß
reactivation
deaths
Defects in DNA repair (AR)
Normal DNA repair
Ø Mismatch repair - HNPCC
Ø Nucleotide e excision repair (NER) - XP
Ø Recombination repair – Bloom’s
If DNA repair is defective (Mutator Phenotype)
Ø genomic instability syndromes
Ø increased risk of developing cancer
Ø But DNA repair genes themselves are are not Oncogenic
1. Hereditary Nonpolyposis Cancer Syndrome (HNPCC)
Ø Mechanism à Affect DNA mismatch repair genes -MSH2 (2p16)
and MLH1 (3p21) genes ("spell checkers.“)
Ø Mutations à replication error (microsatellite instability)
Ø Effects à familial carcinomas (cecum and proximal colon) & endometrial
and ovarian cancers
Ø HNPCC Differs from APC
Ø Cancers not arise in adenomatous polyps
Ø Only 2% to 4% of all colonic cancers
Ø Develop colon cancers at a much younger age
10
Defects in DNA repair
2. Xeroderma pigmentosum (XP)
Ø Normally àUV light (sun) à pyrimidine cross-links àaction of NER à normal
DNA replication
Ø In XP à Defective nucleotide excision repair (NER)à prevent normal DNA
replicationà↑ risk(2000 times) for skin cancers
3. BRCA Genes (1 &2)
Ø Both involved in transcription regulation
Ø Mutations lead to breast & other cancers
Ø BRCA-1 gene à breast cancer, ovarian, prostate and colon cancers
Ø BRCA-2 geneà male breast cancers
4. Ataxia Telangiectasia-
Ø Hypersensitivity to DNA-damaging agents (AT)
Ø ATM gene à senses DNA double-strand breaks & phosphorylates p53
Ø ATM gene is heterozygous 1% of the population
Ø Other disorders Defects in DNA Repair
Ø Bloom syndrome - helicase (fixes DNA repair by homologous
recombination) is defective
Ø Fanconi's anemia -13 genes that make up the Fanconi anemia
complex
11
LIMITLESS REPLICATIVE POTENTIAL:
TELOMERASE

TELOMERASE
Ø Senescence-Cells lose their ability to divide
Ø shortening of telomeres at the ends of chromosomes (double-
stranded DNA breaks) after 60 to 70 doublings
Ø Result in cell cycle arrest mediated by p53 and RB
Ø Mitotic catastrophe
Ø due to genomic instability from the repeated bridge-fusion-
breakage cycles at chromosomal ends
Ø Result in massive cell death
Ø Not happing in tumors – why?
Ø tumors to grow indefinitely due to reactivation of telomerase
Ø 85% to 95% of cancers show up-regulation of the enzyme telomerase
Ø Other mechanism - alternative lengthening of telomeres by DNA
recombination.
13
Telomere & Telomerase
ANGIOGENESIS

ANGIOGENESIS
Ø Angiogenesis =essential for supplying nutrients
Ø If tumors are not vascularizedà grow up to only 1 to 2 mm
in diameter
Ø Stimulus of Angiogenesis – Hypoxia
ØHypoxia stimulates HIF1α (Transcription factor) à
acitvates production of VEGF and bFGF
Ø Angiogenesis is requisite for growth & metastasis
Ø Tumor Vessels are
ØTortuous & irregular
ØLeaky (due to VEGF)
ØGrow continuously (unlike normal angiogenesis)
17
ANGIOGENESIS
Ø Angiogenic factors
Ø VEGF (important ) & bFGF
Ø Notch activation à branching and density of the new vessels
Ø Produced by Tumor cells & or inflammatory cells
Ø Angiogenic switch
Ø Normal cells à p53 à Thrombospondin -1 à↓VEGF & HIF-1
Ø Tumor cells à Angiogenic (inactivate p53)
Ø Tumor cells also produce anti- Angiogenic factors
Ø Clinical evidence
Ø ↑serum and urinary levels of VEGF & bFGF in cancer patients
Ø Angiogenic inhibitors (Endostatin, Tumstatin)
Ø anti-VEGF monoclonal antibody, bevacizumab à cancer Rx
Ø Antibodies to inhibit Notch activation
18
INVASION AND METASTASIS

external stimuli
TGFß
reactivation
deaths
Invasion and Metastasis
Phase 1
phase 2
1. Invasion of the ECM
Steps in Invasion of the ECM
1. Detachment ("loosening up") of the tumor cells from each other
à down-regulation of E-cadherin & or mutations in catenins
2. Attachment to matrix componentsà receptor-mediated
attachment of tumor cells to laminin and fibronectin
3. ***Degradation of ECMà Collagenases (MMP9
and MMP2) àcleave type IV collagen
4. Migration of tumor cells
1. invasive carcinomas, melanomas, and sarcomas à high levels

of these Collagenases
2. In situ lesions and adenomas of breast and colon àmuch less
Collagenases
3. Inhibition of collagenase activity à greatly reduces metastases
2. Vascular dissemination & homing of tumor cells
Tumor cells escape from immune defenses by àaggregate in
clumps
Endothelial adhesion à adhesion molecules (Integrins, laminin
receptors)
• Homing à***CD44 adhesion molecule (T lymphocytes)
used to migrate to selective sites in the lymphoid tissue
Ø Egress: Anatomy and Tropism
– Prostatic carcinoma à bones
– Bronchogenic carcinomas à adrenals and brain
– Neuroblastomas à liver and bones
Ø Organ tropism is due to:
– Tumor cells express adhesion molecules à ligands
expressed on the EC of the target organ
– target organs release chemo attractants à recruit
tumor cells to the site (insulin-like growth factor I& II )
– Target tissue unpermissive environment (unfavorable
soil) for the growth of a tumor (protease inhibitors) à
prevent establishment of a tumor colony)
Over expression of CD44 àfavor metastatic spread

E-cadherins are connected to β-catenin and
the actin cytoskeleton
down-regulation of E-cadherin expression in
colon and breast,
matrix metalloproteinases (MMPs), cathepsin

D, and urokinase plasminogen activator
MMP2 or
MMP9
cytokines, growth factors (e.g., IGFs I and II)

hepatocyte growth factor–scatter factor
24
TO BUILD KNOWLEDGE BASE

Neurofibromatosis
Café au lait spots
Lisch nodule 27
Neoplasia

Lecture Plan
f. ANGIOGENESIS

2(b). INSENSITIVITY TO GROWTH INHIBITION
AND
ESCAPE FROM SENESCENCE:
TUMOR SUPPRESSOR GENES

Tumor suppressor genes (TSG)
Ø TSGs’ are involved in
Ø1. Cell proliferation inhibition
Ø2. Cell differentiation
ØPrevent uncontrolled growth
ØLOF mutations lead to uncontrolled growth
ØRB and p53 are classic examples
ØOncogene expression in cell without mutations of
TSG leads to
ØQuiescence, or permanent cell cycle arrest, and then
ØGrowth -inhibitory pathways àcells into apoptosis
ØWhenever there is Genotoxic stress
ØTSGs’à arrest proliferation
1. RB gene (chromosome locus 13q14)
Ø First TSG discovered
Ø Mutations of RB gene causes Retinoblastoma
Ø Knudson “two-hit” hypothesis of oncogenesis
ØExplain inherited and sporadic Retinoblastoma (next
slide)
ØFamilial retinoblastoma –
Ø inherited as AD trait.
Ø Child is heterozygous at the RB locus and perfectly normal
Ø loss of heterozygosity leads to Retinoblastoma
Ø Study of Retinoblastoma helped to learn about several other
genes that act similarly

1. RB
ØUbiquitous nuclear phosphoprotein
ØRegulates the cell cycle
Øactive hypophosphorylated state in quiescent cells
Øinactive hyperphosphorylated state in G1/S cell cycle
transition
7
RB effects
Ø A. Blocks E2F-mediated transcription in two ways
Ø1. sequesters E2F
Ø2. recruits chromatin-remodeling proteins that bind to
Cyclin E
Ø1.histone deacetylases
Ø2. Histone methyltransferases
Ø B. Controls stability of p27 (cell cycle inhibitor)
Ø C. induce senescence
Ø ***Germline loss or mutations à retinoblastoma and
Osteosarcoma
Ø RB mutations are noticed in in Glioblastomas, small-cell
carcinomas of lung, breast cancers, and bladder carcinomas
Ø RB is a member of pocket proteins(also include p107 and
p130)
8
1. RB
Ø why the loss of RB is not more common in human tumors?
Ø Loss of normal cell cycle control is central to malignant
transformation
Ø One of key regulators of the cell cycle (p16/INK4a, cyclin D, CDK4)
is dysregulated in majority of cancers
Ø Function of RB is disrupted even if the RB gene itself is not
mutated
Ø Animal and human DNA viruses Bind and inhibit activities of RB
Ø Binding takes place at E2F site of RB pocket
ØSimian virus 40
ØPolyomavirus large T antigens
Øadenoviruses EIA protein
Ø***HPV E7 of HPV type 16 binds stronger with RB à Cervical
cancers
9
Knudson

Retinoblastoma
Feature Inherited/ Sporadic

Familial
Age Childhood Adult
Frequency 40% 60%
Bilateralism Yes No
Second Yes No
malignancy
Time of 1st hit Germ cells Somatic cells
2. p53 (chromosome 17p13.1)
Ø Guardian of the Genome, “molecular policeman”
Ø MC target for genetic alteration in human tumors (> 50% of human
tumors contain mutations of p53 )
Ø Homozygous loss of p53 occurs in every type of cancer
Ø *** Mutations of both p53 alleles are acquired in somatic cells (unlike
RB)
Ø ***Li-Fraumeni syndrome
Ø inheritance mutations of one p53 allele.
Ø 25-fold greater chance of malignant tumor by age 50
Ø MC tumors are sarcomas, breast cancer, leukemia, brain tumors,
and carcinomas of the adrenal cortex (what is the difference
between RB & p53 in this aspect?)
Ø 80% of p53 point mutations present in human cancers are located
Ø in the DNA-binding domain of the protein
12
Ø p53 functions can be inactivated by
Ø somatic
Ø inherited mutations
Ø E6 protein of HPV (degradation of p53)
Ø MDM2 and MDMX stimulate the degradation of p53
Øover expressed in malignancies
Ø p53 blocks neoplastic transformation by
Ø 1. Activation of temporary cell cycle arrest (quiescence)
Ø 2. Induction of permanent cell cycle arrest (senescence)
Ø 3. Triggering of programmed cell death (apoptosis)
Ø p53 triggered transcription of genes
Ø 1. genes cause cell cycle arrest
Ø 2. genes cause apoptosis
13
Ø p53 activates transcription of the mir34 family of miRNAs (mir34a–
mir34c)
Ø Blocking mir34 severely affect p53 response in cells
Ø Ectopic expression of mir34 (without p53 activation) can induce
growth arrest and apoptosis
Ø Targets of mir34s
Ø pro-proliferative -cyclins, and
Ø anti-apoptotic -BCL2
Ø Regulation of miRNA is crucial for p53 response
Ø How p53 senses DNA damage & determines the adequacy of DNA
repair ?
Ø Through ataxia-telangiectasia mutated (ATM) and ataxia-telangiectasia and
Rad3 related (ATR)
Ø ATM and ATR phosphorylatep53 and DNA-repair proteins àpause in the cell
cycle
14
Following DNA damage
ØPreventing entry of cells into G1 phase by
ØP21 which inhibits cyclin-CDK complexes and
phosphorylation of RB
Ø1. late in the G1 phase by p53-through
Ø transcription of the CDK inhibitor CDKN1 A (p21)
ØInducing GADD45 (growth arrest and DNA damage)
Ø2. If DNA damage is reparable
Øp53 ↑MDM2à lead to p53 destruction àreleasing cell
cycle block
Ø3. If DNA damage is irreparable
Øp53-induced senescence causes permanent cell cycle
arrest àapoptosis Dr.T.Krishna MD, www.mletips.com 15
P53 mechanism
DNA damage
( direct or indirect)
↑p53
G1 arrest 1
degrade
3
Successful Repair No repair
2
MDM2 ↑ BAX & ↓Bcl-2 senescence
apoptosis
+ PUMA

P53 & therapeutic implications.
1. Tumors that retain normal p53
Ø respond to Irradiation and chemotherapy (RT & CT) by
Ø Preventing entry of cells into G1 phase
Ø induce DNA damage & apoptosis
Ø ***testicular teratocarcinomas & childhood ALL
2. Tumors with mutant p53
Ø Resistant to RT & CT
Ø ***lung & colorectal cancers
Research is going on to increase normal p53 activity in
tumor cells
Ø P63, p73 belong to p53, but of more tissue specificity
Ø P63 – Stratified Squamous cell differentiation,
Ø p73 is -strong pro-apoptotic
Ø p53-p63-p73 network à chemoresistance & poor prognosis
3. APC/β-Catenin Pathway
Ø Main function àdown-regulate growth-promoting signals
Ø Germ-line mutations of APC (Ch.5q21) àFAP (familial adenomatous polyposis)
Ø 1000s’ of colonic adenomatous polyps by teens or 20s’
Ø polyps undergoes malignant transformation à Colonic Adenocarcinoma
Ø After several additional mutations
Ø Adenoma à Adenocarcinoma (***multistep process)
Ø strong hereditary predisposition,
Ø homozygous loss of the APC gene noted in
Ø 80% of non-familial colo-rectal-carcinomas
Ø sporadic adenomas

3. APC/β-Catenin Pathway àcomponent of the WNT signaling pathway
Ø Important function of APC is
Ø down-regulate β-catenin
Ø In absence of WNT signaling àAPC degrades of β-catenin
Ø by ubiquitin + Proteasome pathway
Ø In presence of WNT signalingà Blocks APC-AXIN-GSK3β destruction complex
Ø β-catenin to translocates into nucleus &cause cellular proliferation by c-MYC, cyclin D1
Ø loss of APC à Cell behaves like under continuous WNT signaling
Ø Mutations in the β-catenin & E-cadherin lead to loss of cell-cell contact between
and β-catenin
Ø 50% of Hepatoblastoma
Ø 20% of HCC
Ø Germline mutations of the E- cadherin
Ø Ca. stomach
Ø ↓E- cadherin by SNAIL
Ø epithelial-mesenchymal transition
Ø ***metastasis
4. INK4a/ARF
Ø Also called CDKN2A gene
Ø Encodes two protein products – function as TSG
Ø Mutation or silencing à impacts RB and p53 pathways
(how?)
Ø1. p16/INK4a CDKI, ( blocks cyclin D/CDK2-
phosphorylation of RB)
Ø2. p14/ARF (activates the p53 pathway by inhibiting
MDM2)
Ø p16 - most important for induction of senescence
ØMutations of p16 bladder, head and neck tumors, acute
lymphoblastic leukemias, and cholangiocarcinomas
ØCa. Cervix - p16/INK4a silenced by hypermethylation
without mutationDr.T.Krishna MD, www.mletips.com
5. The TGF-β Pathway
Ø Potent inhibitor of proliferation (normal epithelial, endothelial, and hematopoietic
cells)
Ø Repression of c-MYC, CDK2, CDK4, and Cyclins A and E
Ø TGF-β receptors I and II are part of serine-threonine kinase complex
Ø Activation leads tot phosphorylation of receptor SMADs (R-SMADs)
Ø R-SMADs in nucleus + SMAD-4 à transcription of CDKIs p21 and p15/INK4b.
Ø All the above changes à↓ phosphorylation of RB and ?
Mutations in the TGF-β signaling pathway
Ø Affect the type II TGF-β receptor or interfere SMAD molecules
Ø 1. type II TGF-β receptor mutationsà ca. colon, stomach, and endometrium
Ø 2. inactivation of SMAD4à pancreatic ca.
Ø ***100% of pancreatic cancers
Ø 83% of colon cancers mutation of at least one component of TGF-β
Ø Tumor cells use TGF-β–induced program for immune system suppression/evasion

or promotion of angiogenesis (through loss of p21 and/or persistent expression of
c-Myc)
6. PTEN (TSG)
Ø PTEN (Phosphatase and tensin homologue)
Ø Brake on pro-survival/pro-growth PI3K/AKT pathway
Ø chromosome 10q23
Ø Cowden syndrome,
ØAutosomal dominant disorder
ØMutations of PTEN
ØPresent with benign growths, (of the skin appendages),
and epithelial cancers (breast, endometrium, and
thyroid)
Ø Acquired loss of PTEN function
ØUp- regulate PI3K/AKT signaling àvarious cancers

7. NF1
Ø One mutant allele of the NF1 gene à Neurofibromatosis type 1
Ø Neurofibromin -protein product of NF1 gene
Ø GTPase-activating domainà regulates signal transduction through
RAS
8. NF2
Ø mutations in the NF2 gene à Neurofibromatosis type 2
Ø Neurofibromin 2 or merlin- protein product of NF 2 gene
1. Has structural similarity to red cell membrane cytoskeletal protein 4.1
2. Lack of protein – makes cells insensitive to normal growth arrest signals generated
by cell-to-cell contact
3. Salvador-Warts-Hippo (SWH) tumor suppressor pathway which controls organ size
Ø Mutations à***bilateral schwannomas of the acoustic nerve
Ø Somatic mutations à meningiomas and ependymomas

9. VHL von Hippel-Lindau (VHL) gene
Ø Germline mutations (ch. 3p) cause
Ø Hereditary RCC, pheochromocytomas, hemangioblastomas of the
central nervous system, retinal angiomas, and renal cysts
Ø Also sporadic RCC
Ø VHL protein
Ø part of a ubiquitin ligase complex
Ø In presence of oxygen,
Ø HIF1α (hypoxia-inducible transcription factor 1α) binds to VHL
protein
Ø leading to ubiquitination + proteasomal degradation
Ø In hypoxic states
Ø HIF1α escapes degradation à translocate to the nucleus and
growth/angiogenic factors, (VEGF) and PDGF

10. Patched (PTCH) – TSG
Ø PTCH1 and PTCH2
Øencode a cell membrane protein (PATCHED)
Ø Receptor for Hedgehog family
Ø Hedgehog/PATCHED pathway regulates
ØTGF-β and PDGFRA and PDGFRB
Ø Gorlin syndrome
ØMutations in PTCH
ØNevoid basal cell carcinoma syndrome
Ø Basal cell carcinoma -20% to 50% of sporadic cases are
ØDue to mutations in PTCH
Ø50% of mutations caused by UV exposure
?
Neoplasia

Lecture Plan
f. ANGIOGENESIS

STROMAL MICROENVIRONMENT AND
CARCINOGENESIS
Ø Neoplasms have
Ø parenchymal cells
Ø Stroma (ECM)
Ø ECM produces Growth factors
Ø PDGF, TGF-β, and bFGF
Ø Act on growth of tumor cells in a Paracrine way
• Do tumor cells depend on stroma for proliferation, survival, or
metastases?
– If yes, we can target the stroma to treat tumors
1. immune/inflammatory cells à cytokines
– Promote development of cancer
– Also promote survival and progression of cancer cells
2. Tumor macrophages to secrete metastasis promoting factors
STROMAL MICROENVIRONMENT AND
CARCINOGENESIS
3. Fibroblasts secrete matrix and cause Desmoplasia
4. Experiments on mice suggest that stroma can drive
genetic changes to promote carcinogenesis.
5. Genes expressed in stromal cells control tumor
behavior
6. Best studied tumor about role of stromal cells in
tumor growth is human breast cancer

METABOLIC ALTERATIONS: THE WARBURG
EFFECT
THE WARBURG EFFECT
Ø Even in the presence of ample oxygen, cancer cells shift their
glucose metabolism to glycolysis (most cancer cells instead
rely on aerobic glycolysis)
Ø Warburg effect used in
– PET scan (positron emission tomography)
– 18F-fluorodeoxyglucose(non-metabolizable derivative of
glucose) is injected
– preferentially taken up into tumor cells and actively
dividing normal (bone marrow) cells
– Most tumors are PET-positive
– Rapidly growing tumors are intensely positive
METABOLIC ALTERATIONS: THE WARBURG EFFECT
THE WARBURG EFFECT CONT’D…

• Why tumors prefer glycolysis generates just 2
ATP molecules per molecule of glucose?
– Not clear
– Attractive theory - altered metabolism gives a growth
advantage in the hypoxic tumor microenvironment
– hypoxic tumor microenvironment à stimulates
HIF1α which in turn stimulates angiogenesis

DYSREGULATION OF CANCER-ASSOCIATED GENES
Chromosomal Changes
Ø Specific chromosomal abnormalities are noted in
§ most leukemias and lymphomas
§ many sarcomas
§ increasing number of carcinomas
1. Aneuploidy and chromosomal instability are
§ initiating events in tumor growth
Ø Aneuploidy and structural abnormality of chromosomal are
§ late events in cancer progression
Ø Purpose of study of chromosomal changes
§ Identification of Oncogenes (e.g., BCL2, ABL) and TSG (e.g., APC, RB)
§ Diagnostic & predictive of clinical course (Examples - ABL oncogene
in CML and c-MYC in Burkitt lymphoma)
1. Translocations
Ø MC chromosomal rearrangements (oncogene)
§ A. Follicular Lymphoma
§ MC lymphoid (B cell) malignancy with t(14;18) and BCL2 gene
activation
§ B. Burkitt lymphoma
§ lymphoid (B cell) tumor with translocations t(8:14) and MYC gene over
expression
Ø Translocations allow formation of hybrid fusion genes that encode
chimeric proteins
§ in hematopoietic tumors, sarcomas, and carcinomas – (Example -
Philadelphia chromosome, of CML)
Ø Translocations well known in
§ leukemias/lymphomas and sarcomas
§ Not in carcinomas – why?
§ carcinomas have complex karyotypes
§ 50% of Prostatic carcinomas - translocation of androgen-regulated
gene, TMPRSS2 (21q22), and one of three ETS family of TFs’
9
Very
important
Dr.T.Krishna MD, 10
www.mletips.com
2. Deletions
Ø 2nd MC structural abnormality in tumors
§ More common in non- hematopoietic solid
tumors (translocations more common in
hematopoietic tumors)
§ Deletion lead to loss of particular TSG
(translocations – Oncogenes)
§ Deletions of RB gene (chromosome 13q14) à ?
tumor
§ Deletions of 17p, 5q, and 18q à colorectal cancers
§ Deletion of 3p- extremely common in small-cell
lung carcinomas
11
3. Gene Amplification
• Activation of proto-oncogenes lead to overexpression
• detected by molecular hybridization with appropriate DNA
probes
• Two mutually exclusive patterns are noted
– double minutes
– homogeneous staining region (HSR) – is due to insertion of
the amplified genes into new chromosomal locations
(appear homogeneous in a G-banded karyotype)
• N-MYC amplified in 25% to 30% of Neuroblastomas
– poor prognosis
– gene is present both in double minutes and homogeneous
staining regions
• ERBB2 (also called ?) amplification seen in breast cancers,
– antibody therapy directed against this receptor is effective12
4. Epigenetic Changes
• Reversible, heritable changes in gene expression
• Epigenetic Changes occur without mutation
• post-translational modifications of
– DNA methylation
– Histones
13
4. Epigenetic Changes - DNA methylation
• In normal, differentiated cells,
– Most of DNA is compact and in heterochromatin form
– due to DNA methylation and histone modifications (not
expressed)
• In cancer cells
– Almost complete DNA hypomethylation (expressed) and
– selective promoter-localized hypermethylation
• TSG is silenced by hypermethylation of promoter sequences not by
mutation in CDKN2A of colon and gastric cancers
• Silencing of p16/INK4a lead to inactivation of p53 and Rb pathways
(to important check points in cell cycle)
– TSG silencing by methylation also seen in
• BRCA1 in breast cancer,
• VHL in renal cell carcinomas,
• MLH1 mismatch-repair gene in colorectal cancer.
14
5. miRNAs and Cancer
§ Small (22 nucleotides size) noncoding, single-stranded RNAs
§ Introduced into the RNA-induced silencing complex
§ Mediate post-transcriptional gene silencing
§ In normal cells
§ miRNAs control cell growth, differentiation, and cell survival
§ in many cancers
§ Amplifications and deletions of miRNA identified
§ Cause ↑ expression of Oncogenes or ↓expression of TSG
§ upregulation of RAS and MYC Oncogenes has detected in lung tumors
& certain B-cell leukemias
§ down-regulation or deletion of certain miRNAs In some leukemias and
lymphomas results in increased expression of BCL2
§ miRNA do the above by negative feed back (read foot note)
15
Number of
Number of miRNA?
miRNA?
leukemias
and
lymphomas In Breast and
results in Brain tumors
increased
expression of
BCL2
Dr.T.Krishna MD, 16
www.mletips.com
Molecular Basis of Multistep Carcinogenesis
GOF of Oncogenes and LOF of TSGs’

Dr.T.Krishna MD, 17
www.mletips.com
Test your knowledge

Child with Ocular pathology
1. The gene affected ?

2. Which Hypothesis?
3. Second cancer ?

Teenager with intestinal lesion

2. % risk of cancers if untreated?
3. Cancers arise from where?

Young patient with GI Cancer
A
C

Invasion & Metastasis
1. Enzymes involved?

Teenager with intestinal lesion


Neoplasia

Carcinogenic Agents and Their Cellular
Interactions
Lecture Plan
1. Chemical Carcinogenesis
2. RADIATION CARCINOGENESIS
A. Ultraviolet Rays
B. Ionizing Radiation
3. MICROBIAL CARCINOGENESIS
A. Oncogenic RNA Viruses
i. Human T-Cell Leukemia Virus Type 1
B. Oncogenic DNA Viruses
i. Human Papillomavirus
ii. Epstein-Barr Virus
iii. Hepatitis B and C Viruses
C. Helicobacter pylori
1. Chemical Carcinogenesis
Ø Sir Pott (1750)
Ø Scrotal skin cancer in chimney sweeps àchronic
exposure to soot
Ø Steps Involved in Chemical Carcinogenesis
Ø Initiation - caused by sufficient dose of a carcinogenic agent (initiator)
Ø Progression - induce tumors only in initiated cells (Promoters)
memory
4
Initiators & Promoters
Initiator Promoters
àcarcinogenic agent à Changes are reversible
ànot sufficient for tumor ànon- tumorigenic by
formation by themselves themselves
à permanent DNA à not affect DNA directly
damage (Mutations) à induce tumors in
à rapid and irreversible initiated cells &
àhas “memory” reversible
Chemical Carcinogenesis
Initiators
Ø Natural and Synthetic
Ø Carcinogenic potency of a chemical is determined by
Ø inherent reactivity of its electrophilic nature
Ø Balance between metabolic activation and inactivation
reactions.
Ø Two Categories: Direct acting & Indirect acting
Ø Direct acting –not need metabolic conversion
§ highly reactive electrophiles (have electron-deficient
atoms)
§ react with nucleophilic (electron-rich) sites in the cell
(DNA, RNA, and proteins)
§ DNA = primary target
§ Reactions = non=enzymatic
§ interaction = nonlethal
Initiators cont’d…
Ø Indirect acting - requires metabolic conversion
1. Carcinogens use cytochrome P-450 gene
• CYP1A1àpolymorphic form of P-450 (10% of whites)
• Metabolizes BENZ(O)PYRENE (of tobacco smoke)à
carcinogen (lung cancer)
2. Glutathione –s-transferase
• Detoxification of polycyclic aromatic hydrocarbons
(PAH) - aromatic amines and azo dyes
• 50% of whites à enzyme polymorphism
• ↑risk of Bladder & Lung cancers in smokers

8
Molecular Targets
Ø DNA damage àprimary target for chemical carcinogens
Ø AMES TEST
– ability of chemical to induce mutations in the bacterium
S. typhimurium à if MUTATION is presentà positive score
– Environmentally induced insults to DNA + defect in DNA
repair (inherited) à↑risk of cancers
Molecular fingerprinting or signature mutation” of
tumors
Ø Example: Hepatocellular carcinoma (HCC) & p53 (at 249=
G:C::T:A) )mutations
– HCC Africa and China à linked to Aflatoxin B1 & p53
mutations are frequent
– HCC in other parts à follows chronic HBV infection &
p53 mutations are much less frequent

• Initiated Cell
– carcinogen-altered cell
– must undergo at least one cycle
– Then change in DNA becomes fixed or
permanent
• Promotion of Chemical Carcinogenesis
– promoters (such as phorbol esters, hormones,
phenols, and drugs)
– proliferation and Clonal expansion of initiated
(mutated) cells
• Promoters & Tumorogenesis
– Phenobarbital àblockage of the TGF-β pathway
– phorbol ester (activator of protein kinase C) à
signal transduction pathways
Chemical carcinogens= Initiators
• Alkylating Agents= Weak carcinogens, direct acting
– Anti – Cancer Drugs (cyclophosphamide, chlorambucil,
busulfan)
– damage DNA (anticancer effect) but also render it
carcinogenic
• Polycyclic Aromatic Hydrocarbons =
– ***Most potent carcinogens known, indirect acting
• Smoking – Tobacco
• animal fats à broiling meats
• smoked meats and fish.
• Aromatic Amines àβ-naphthylamine aniline (dye and rubber
industries) àCause Transitional Cell Cancer (TCC) of Urinary
bladder (glucoronidase)
• Azo Dyesà found in food coloring (butter yellow & scarlet
red)
Chemical carcinogens=Promoters
• Endogenous à hormones and Bile salts
• Exogenous
1. Diethylstilbestrol (used for Threatened Abortion)
I. Postmenopausal mothers à endometrial carcinoma
II. Offspring (daughters) exposed in- uteroà vaginal cancer
2. Fat = colon cancer
3. Estrogen = liver tumors
4. Tobacco and viral infection = tissue damage and reactive
hyperplasia
5. Tobacco – has both initiators and promoters
LOF= Loss of Function mutations
•haplo-insufficiency= reduces levels or activity of protein (protective )à lead to release of breaks à causes Tumor cell proliferation and survival,
• dosage of the gene or two copies (of TSG) are required for normal function

13
Radiation Carcinogenesis
Radiation Carcinogenesis= UVB, Ionizing Radiation
• Synergism with other carcinogens
• 1. UVB (280-320nm)
• Also causes mutation in Oncogenes and TSG (RAS & p53)
• Induction of cutaneous cancers
– Carcinogenicity:
• Pyrimidine dimers in DNA
• Higher risk in Defective NER repair genes– Xeroderma
pigmentosum
2. Ionizing Radiation
– Electromagnetic (x-rays & γ rays) and particulate (α particles, β particles,
protons, neutrons) à all CARCINOGENIC
• Order of Vulnerability:
– High risk = Hematological (Leukemia), Thyroid
– Intermediate risk= Breast, lungs, salivary gland
– Resistant =GIT, Skin, bone
Microbial carcinogenesis
DNA Oncogenic Viruses: HPV, EBV, HBV, Kaposi
Sarcoma, Herpes Virus
• Human Papilloma virus (HPV):
– Squamous Cell Carcinoma à Cervix and Anogenital, Oral
and Laryngeal regions
– HPV serotypes =16/18, 31/33/35/51 ( High risk group)
– Integration of viral DNA à malignant transformation
– Over expression of the E6 and E7 Viral proteins (mainly
16 , 18 )
– E6 inactivate p53 by enhancing its degradation through
ubiquitin-dependent proteolysis)
– E7 disable pRb (& p53) by disrupting the E2F/RB complex
and promoting the degradation of RB

• Epstein Barr Virus (EBV)
– Herpes family
– self-limited disorder àinfectious mononucleosis
– Pathogenesis of human tumors
– Burkett Lymphoma
– B-cell Lymphomas (NHL) in immunosuppressed
individuals (HIV)
– Hodgkin’s Disease
– Naso -Pharyngeal Carcinoma (100% EBV association)
– Viral genesà dysregulate normal proliferative and
survival signals in infected cells
• Examples:
• Latent membrane protein-1(Mimicking CD40)
• à activates the NFκB and JAK/STAT signaling
pathways
• à promotes B-cell survival and proliferation
16
• HBV
– Carcinogenic effect à indirect and multifactorial
– HBx Protein à binds to p53 and interferes with its growth suppressing
activities
• RNA Oncogenic Viruses
• Human T-Cell Leukemia Virus Type 1 (HTLV-1)
– Non – Neoplastic
• Tropical Spastic Paraparesis à Demyelinating neurological
disorder
• Uveitis & Arthritis
• Endemic in Japan & Caribbean
• Tropism for CD4+ T-CELLS
– Neoplastic = 1% ( in 1% of infected)
• TAX region à Activate the transcription of host cell genes
• HCV
– Pathogenesis of HCC - ?
– chronic Hepatitisà Regeneration à Neoplastic change 17
CD4+ tropism - ?another infection & ? drug
18
19
Burkitt’s Lymphoma
Endemic Sporadic
African elsewhere
Kids Adults
Mandible (Jaw) Abdomen
No visceral or Bone Yes
Marrow involvement
EBV infection + No

21
Helicobacter Pylori
Ø Non – Neoplastic = Chronic Gastritis (90% of
patients )
Ø Neoplastic
Ø1. Gastric Lymphomas (B-cell) or MALTomas
or Marginal Zone Lymphoma
ØTreatment with antibiotics à Regression of lymphoma
Ø2. Gastric Carcinomas
ØMechanism = H. pylorià Activate reactive T-
cellsà activate Polyclonal (infection) B-cells
→ Continue on with Monoclonal ( Malignant)
B-cells
22
Neoplasia

Host Defense against Tumors
Tumor Immunity
Lecture Plan
§ TUMOR ANTIGENS
§ Products of Mutated Genes.
§ Overexpressed or Aberrantly Expressed Cellular Proteins
§ Tumor Antigens Produced by Oncogenic Viruses
§ Oncofetal Antigens
§ Altered Cell Surface Glycolipids and Glycoproteins
§ Cell Type–Specific Differentiation Antigens
§ ANTITUMOR EFFECTOR MECHANISMS
§ IMMUNE SURVEILLANCE AND ESCAPE

TUMOR ANTIGENS
Ø Immune surveillance
Ø Normal function of the immune system
Ø Detect emerging malignant cells and destroy them
Ø Evidence
Ø Lymphocytic infiltrates around tumors and in lymph nodes draining
sites of cancer
Ø Tumor-specific T cells and antibodies in patients
Ø Increased incidence of some cancers in immunodeficient (risk is 200
times > in immunocompetent)
Ø Then, Why tumors still occur in in immunocompetent ?
Ø Immune surveillance is imperfect
Ø Cancer immunoediting
Ø Effects of the immune system in preventing tumor formation
Ø To shape immunogenic properties of tumors to select tumor cells that
escape immune elimination
TUMOR ANTIGENS
Classified into two categories (not perfect)
Ø 1. tumor-specific antigens
Ø Present only on tumor cells and not on any normal cells
Ø 2. tumor-associated antigens
Ø Present on tumor cells and also on some normal cells

TUMOR ANTIGENS
Products of Mutated Genes
Ø mutated RAS or p53 proteins - Potential tumor antigens

Ø In cancer patients - circulating CD4+ and CD8+ T cells against
the mutated proteins are present
Ø Oncoproteins are not major targets of tumor-specific CTLs in
most patients.

TUMOR ANTIGENS
Overexpressed or Aberrantly Expressed Cellular Proteins
Ø Normal cellular proteins that are abnormally expressed in tumor cells

Ø Produced at low levels in normal cells and Overexpressed in tumor cells
Ø Elicit immune response
Ø Best example – in human melanomas - tyrosinase
Ø T cells against peptides of tyrosinase are detected in melanoma patients
Ø Tyrosinase vaccine
Ø Why the Tyrosinase of normal cells can’t elicit immune response?
Ø Normal cells produce small amounts of tyrosinase
Ø Only few cells few cells produce tyrosinase

TUMOR ANTIGENS
Overexpressed or Aberrantly Expressed Cellular Proteins
MAGE Family of genes (cancer-testis antigen family)
MAGE-1 antigen (melanoma antigen-1)

ØMAGE is present but not expressed on the cell surfaces in an
antigenic form by normal testicular tissues (sperm do not
express MHC class I antigens)
ØMAGE is originally described in melanomas (37%), is also
expressed in lung, liver, stomach, and esophageal carcinomas
ØSimilar antigens are GAGE, BAGE, and RAGE
ØTargets for immunotherapyDr.T.Krishna MD, www.mletips.com 8
TUMOR ANTIGENS
Ø Mutated self proteins (due to radiation, chemicals) represent

antigens that never encountered by the immune system
Ø Can act as tumor antigens
Ø Recognized by CD8+ T cells

TUMOR ANTIGENS
ØMost potent antigens are proteins of DNA viruses - HPV & EBV
ØCTLs recognize these antigens and kill virus-infected cells
ØVaccines against HPV antigens - effective in preventing cervical
cancers in young females

Oncofetal Antigens
Ø Proteins that are expressed at high levels on
1. Cancer cells
2. In normal developing (fetal)
3. But not adult tissues.
Ø Why not expressed on adult tissues?
Ø silenced/repressed during development
Ø Why expressed at high levels on cancer cells?
Ø De-repressed upon malignant transformation
Ø Importance
Ø Markers in tumor diagnosis (also increased in tissues and in the circulation
in inflammatory conditions)
Ø Not proven targets of antitumor immunity
Ø The two most important Oncofetal antigens are
1. Carcinoembryonic antigen (CEA) – GI cancers (Stomach, Colon)
2. α-fetoprotein (AFP) –HCC, Yolk Sac tumors of Gonads (non tumor
condition with ↑↑ levels?)
Altered Cell Surface Glycolipids and Glycoproteins
1. Tumors express higher levels of normal levels and/or

abnormal surface glycoproteins and Glycolipids
Ø Antibodies are raised in animals against these
glycoproteins and Glycolipids
Ø Used as Diagnostic markers and targets for therapy
Ø Examples
1. Melanomas express high levels of GM2, GD2, and GD3
2. Clinical trials of anti-GM2 and anti-GD3 antibodies and
immunization with vaccines containing GM2 in melanoma
patients

Altered Cell Surface Glycolipids and Glycoproteins
2. Mucins are high-molecular-weight glycoproteins
Ø Mucins contain “O-linked carbohydrate side chains” – can be
tumor-specific epitopes
Ø Mucins that are used in diagnostic and therapeutic studies
Ø ***CA-125 – ovarian ca.
Ø ***CA-19-9 – Pancreatic Ca.
Ø ***MUC-1; breast ca.
Ø MUC-1 - integral membrane protein (unique)
Ø Normally expressed only on the apical surface of breast ductal
epithelium and sequestered from the immune system
Ø Expressed in an unpolarized fashion and contains new, tumor-
specific form which induce both antibody and T-cell responses
(AMI & CMI)
Ø Candidates for tumor vaccines
Cell Type–Specific Differentiation Antigens
Differentiation antigens = molecules normally present on the cells of

origin are expressed by tumors
– These antigens are specific for particular lineages or differentiation stages
cell types
– Since these are normal self-antigens, they do not induce immune response
in tumor-bearing hosts
Significance
– Potential targets for immunotherapy
– Helps in identifying the tissue of origin of tumors
Examples
1. B cell lymphomas with surface marker- CD20
Ø Antibodies against CD20 are also used for tumor immunotherapy
2. Surface immunoglobulin of a clonal B-cell specific for particular
idiotype
Ø Highly specific diagnosticDr.T.Krishna
tumorMD, antigen for B-cell lymphomas and leukemias
www.mletips.com 15
ANTITUMOR EFFECTOR
MECHANISMS

ANTITUMOR EFFECTOR MECHANISMS
Cytotoxic T lymphocytes:
Ø protective role of CD8+ CTLs in humans against virus-
associated neoplasms (e.g., EBV- and HPV-induced tumors)
Ø In these cases, CD8+ T cells do not develop spontaneously in
vivo but can be generated by immunization with tumor
antigen–pulsed dendritic cells.
Natural killer cells:
Ø Capable of destroying tumor cells without prior sensitization
Ø First line of defense against tumor cells
Ø After activation with IL-2 and IL-15, NK cells can lyse a wide range of
human tumors
Ø T cells and NK cells provide complementary antitumor mechanisms
Ø Tumors that fail to express MHC class I antigens cannot be recognized by
T cells, but trigger NK cells
Ø NKG2D proteins expressed on NK cells (and some T cells) are important
activating receptors
Ø NKG2D proteins recognize antigens expressed on tumor
ANTITUMOR EFFECTOR MECHANISMS
Macrophages:
Ø Activated macrophages exhibit cytotoxicity against tumor cells
in vitro.
Ø T cells, NK cells, and macrophages may collaborate in
antitumor reactivity
Ø interferon-γ, a cytokine secreted by T cells and NK cells, is a
potent activator of macrophages.
Ø Activated macrophages may kill tumors by mechanisms
similar to those used to kill microbes or by secretion of TNF
Antibodies:
• No evidence of protective effects of antitumor antibodies against
spontaneous tumors
• Administration of monoclonal antibodies against tumor cells is in
practice
• Best example - monoclonal antibody against CD20, a B-cell surface
antigen, used for treatment of lymphomas
IMMUNE SURVEILLANCE AND ESCAPE

Is immune surveillance operate in vivo to prevent emergence of
neoplasms?
Ø Yes
Ø increased frequency of cancers in immunodeficient hosts
Ø 5% of persons with congenital immunodeficiency develop cancers (200
times more common than in immunocompetent individuals)
Ø Immunosuppressed transplant recipients and AIDS patients have an
increased incidence of malignancies.
Ø Majority are of these neoplasms are lymphomas( diffuse large B-cell
lymphomas)
Ø X-linked recessive immunodeficiency disorder - XLP (X-linked
lymphoproliferative syndrome)
Ø caused by mutations in the gene important in lymphocyte signaling
pathways
Ø affected boys develop an EBV infection, and later develop a chronic
fatal form of infectious mononucleosis or B-cell lymphoma
Ø Why immunocompetent hosts develop cancers?
Ø Tumor cells must develop mechanisms to escape or evade the
immune system
Sub clone without
tumor Ag
Where is MHC 1?
CTLA4
(T-cell inhibitory receptor)
TGFβ by tumor cells

Lack of co-stimulation:
Ø T cells requires two signals for activation
Ø 1. Foreign peptide (Antigen) presented by MHC molecules
and
Ø 2. Co-stimulatory molecules
Ø Tumor cells often do not express co-stimulatory
molecules and escape from T cell recognition
Ø Treatment strategies
Ø 1. Immunize patients with autologous tumor cells transfected
with the gene for the co-stimulatory molecule B7-1 (CD 80)
Ø 2. Autologous dendritic cells (dendritic cells express high
levels of co-stimulatory molecules) expanded in vitro and
pulsed with tumor antigens (e.g., MAGE1) are infused into
cancer patients
Antigen masking:
• Cell surface antigens of tumors may be hidden, or masked,
• Glycocalyx molecules, (sialic acid–containing
mucopolysaccharides) form the mask.
Apoptosis of cytotoxic T cells:
• Melanomas and HCC express FasL & kill Fas-expressing T cells
Paradoxically, the immune system may promote the growth of
tumors
1. Activated lymphocytes and macrophages produce growth
factors for tumor cells,
2. Regulatory T-cells and certain macrophages suppress host
response to tumors.
3. Enzymes (MMPs) enhance tumor invasion
Neoplasia

Clinical Aspects of Neoplasia
Cancer Cachexia
Paraneoplastic syndrome
Grading and Staging
Lab diagnosis of cancer
Tumor Markers
Cancer Cachexia
Ø Anorexia and weight loss
Ø Equal loss of both fat and lean muscle (unlike starvation)
Ø Elevated BMR (reduced food intake)
Ø Cytokines (TNF, IL-1, IFNγ) produced by tumor and host cells
are responsible
- Paraneoplastic syndrome

Paraneoplastic syndrome
Cushing Syndrome Small CC of lung ACTH or ACTH-like substance
Syndrome of Inappropriate Small CC of lung ADH. (Please read footnotes)

ADH secretion (SIADH)
Hypercalcemia Sq. C.C – lung, Breast ca. PTHrP
Carcinoid Syndrome HCC, Bronchial adenoma Serotonin, Bradykinin
Polycythemia Renal Cell Carcinoma (RCC) Erythropoietin

Myasthenia Lung carcinoma Immunological
Eaton-Lambert syndrome Small CC of lung Immunological
Acanthosis nigricans Gastric ca. Immunological
Hypertrophic Lung carcinoma Immunological

osteoarthopathy and
clubbing of fingers
Venous Thrombosis Pancreatic adenocarcinoma Mucins that activate
Trousseau Syndrome coagulation
Dr.T.Krishna MD, www.mletips.com See foot notes 4
Cancer Cachexia
Ø Anorexia and weight loss
Ø Equal loss of both fat and lean muscle (unlike starvation)
Ø Elevated BMR (reduced food intake)
Ø Cytokines (TNF, IL-1, IFNγ) produced by tumor and host cells
are responsible
Grading and Staging (next slides)

• Staging & Grading of Tumors
• Basis of Grading
– Degree of tumor cell differentiation and number of mitoses
within the tumor
– to judge the extent to which the tumor cells resemble or fail to
resemble their normal counterparts
• GRADING: I-IV
– GRADE I: well-differentiated (looks ALMOST like parent tissue)
– GRADE II: moderately differentiated
– GRADE III: poorly differentiated
– GRADE IV: undifferentiated (almost impossible to tell where
primary neoplasm originated)
• Basis of STAGING
• 1. UICC (Union Internationale Contre Cancer - TNM)
– Tumor size
– Lymph Nodes involvement
– Metastases
• 2. AJC (American Joint Committee)
– Divides all cancers into stages 0-IV
Grading
undifferentiated

Staging

Staging

Which is most important for Clinical Prognosis of
Tumors?
Staging
Exception – Prostate Ca.

Gleason Grading is important
Lab diagnosis
1. Immuno Histo Chemistry (IHC)
Ø known specific monoclonal antibodies are used
Ø For categorization of undifferentiated malignant tumors:
1)Poorly-differentiated & anaplastic Cancers
2)lymphomas,
3)melanomas,
4)sarcomas
Ø Cytokeratins: for epithelium derived tumors
Ø CD markers: for Lymphomas (CD20 - ? B cell or T cell)
Ø S-100 and HMB-45: will stain melanomas
Ø Vimentin, Desmin: for tumors of mesenchymal origin
Ø Vimentin (general stain), Desmin (muscle derived)
Ø GFAP (Glial Fibrillary Acidic Protein) : for Glial tumors
Immuno Histo Chemistry (IHC)
CD30 “+”
Lymphoma
? Carcinoma
? Sarcoma
Lymphoma?

• Lab Diagnosis of Neoplasms
1. Fine Needle Aspiration Cytology (FNAC)
2. Biopsy
3. Immunochemistry
4. Cytogenetic Analysis (Karyotyping)
• FISH (fluorescent in situ hybridization)
• Chromosome Painting
• Spectral Karyotyping
5. Recombinant DNA Technology
• PCR Assays
• Southern, Northern and Western blots
6. Others
Lab diagnosis Bone Marrow biopsy
IHC with PSA
2. Diagnosis of Origin of
Metastasis
Prostatic specific antigen (PSA)
in bone metastases.
Ø Presents with Low back pain,
Ø No prostate related
symptoms
Thyroglobulin in carcinoma
thyroid
Ø Presents with upper back
pain
Ø No thyroid related symptoms

Cytology (Pap) Smears
Diagnosis of cervix carcinoma
Normal smear Abnormal smear
large, flattened squamous cells pleomorphic, Hyper-chromatic

nuclei
no malignant cells Malignant cells

Tumor Markers: Hormones
Hormones
beta Hcg Hydatidiform mole, choriocarcinoma, testicular

tumor (Seminoma)
Calcitonin Medullary carcinoma thyroid
Catecholamines Pheochromocytoma
Ectopic hormones Paraneoplastic syndrome
Oncofetal Antigens
alpha-fetoprotein (AFP) HCC, (Yolk sac tumors or Gonads)
Carcinoembryonic antigen (CEA) Colonic cancers
Isoenzymes
Neuron Specific Enolase (NSE) Small cell carcinoma lung, Neuroblastoma
Specific proteins
Immunoglobulins Multiple myeloma
PSA Prostate cancer

Mucins
CA-125 Ovarian cancer
RBC Disorders. Part 1
Dr. Roy
Hematocrit (Hct)
Basic tests
RBC count: part pf complete blood counts (CBC)
Hemoglobin (Hb): normal in adults- 14 gm% to 17 gm%
Hematocrit (Hct): normal- 42% to 52% in men slightly lower in females
Rule of 3:
- Hb will be 3 times the RBC count
- Hct will be 3 times Hb value
For example: if the total red cell count is 5 million cells/mm3
Hb will be 5 x 3 = 15 gm%
Hct will be 15 x 3 = 45%
Basic Tests
How do you investigate blood disorders (anemia)?
(cont’d)
Peripheral Blood Smear
-examines red cell, leukocytes, and platelets
Red Blood Cells:
Color: Hb content
Size/shape: example- microcytic, macrocytic
Inclusions: example- Heinz bodies, malarial
parasites
Peripheral blood smear:
Normal
Basic test: red cell distribution with (RDW)
- spectrum of red blood cell
size
- the width widens in cases of
1) iron deficiency anemia 2)
megaloblastic anemia
Basic Test: red cell indices
MCV:
- refers to the average size of the red blood cells
- reported as femtoliters (fL)
- normal range for adults is typically 80 - 100 fL
MCH:
- refers to the average weight of hemoglobin in red blood cells
- reported as picograms (pg)
- normal range for adults is typically 26 - 32 pg
MCHC:
- refers to the average concentration of hemoglobin in red blood cells
- normal range for adults is typically 32 - 36 g/dL
Basic Test: reticulocyte count
Reticulocytes:
- are young RBCs containing RNA filaments in the cytoplasm
- newly released RBCs from the bone marrow
- effective erythropoiesis refers to an appropriate bone marrow response to anemia
- this correlates with an increase in the synthesis or release of reticulocytes from
the bone marrow
- Supravital stains detect the threadlike RNA filaments in the cytoplasm of immature
red cells
- reticulocyte becomes a mature red cells in 24 hours
Basic test: bone marrow evaluation
Bone marrow aspiration:
- is performed to obtain specimens
used to assess cellular morphology
- and to conduct specialized tests on
the bone marrow, such as f low
cytometry for immunophenotypic
analysis, cytogenetic studies, or
molecular studies
Basic Test: bone marrow evaluation
(cont’d)
Bone marrow aspiration:
-aspiration of particles of bone marrow thru’ suction
-via wide bore needle
Advantages:
1.rapid method of study
2.morphological details superior to that of biopsy
Procedure: sedation, local anesthetic infiltration to periosteum, bone penetrated in a
rotatory movement, remove stilette, attach syringe, aspirate, films are prepared on slides.
Stain with Romanovsky stains (Giemsa)
Basic test: bone marrow evaluation
Bone marrow biopsy:
- biopsy is often performed as part of
the aspiration procedure and can
provide more specific information
about the cellularity of the marrow
and the extent of disease
Basic Test: bone marrow evaluation
(cont’d)
Bone marrow trephine biopsy:
-used for detection an study of neoplastic diseases
-performed by using a specialized biopsy needle (Jameshidi & Swain)
-surgical procedure (requires operation theater)
-the core obtained is prepared for histological sectioning
Bone Marrow Aspiration
Bone Marrow Biopsy

Anemia
Complete Blood Counts
Total Red Cell Count
Total White Cell Count
Total Platelet Count
Hemoglobin (Hb)
Hematocrit
Red Cell Indices
-MCV
-MCH
-MCHC
Reticulocyte count
Anemia
(cont’d)
Leukocytes:
Differential Leukocyte count (%)
Neutrophils: 40 to 70%
Lymphocytes: 20 to 40%
Monocytes: 1 to 8%
Eosinophils: 1 to 6%
Basophils: less than 1%
Anemia
What is anemia?
-anemia is present when the Hb level in blood is below the lower extreme of
normal range for the sex of the individual
-normal range must include age, and sex
What is the normal range?
adult male: 14 to 17 gm%
adult female: 13 to 15 gm%
How is Hb measured?
- most common method is ‘Hemiglobincyanide method’
Anemia
What are the physiological adaptions?
-increased production of red cells occur when oxygen transport to the tissue is
impaired
What is “physiological anemia”?
-all infants experience a decrease in hemoglobin concentration after birth
-transition from a relatively hypoxic state in utero to a relatively hyperoxic state
-after birth leads to a decline in erythropoietin (EPO)
-in term infant, physiologic anemia is asymptomatic and seen is between 6 to 8
weeks after birth
Anemia
What is the physiological jaundice?
-newborns produce bilirubin at a rate of approximately 6 to 8 mg/kg/day
-this is more than twice the production rate in adults, primarily because of relative
polycythemia (in utero)
-in most infants, unconjugated hyperbilirubinemia reflects a normal transitional
phenomenon
-physiological jaundice normally clears by two weeks
What is breast milk jaundice?
-normally, breastfed newborn develops jaundice after the first 4-7 days of life
-but may persists longer than physiologic jaundice, and has no other identifiable cause
Anemia
Why do pregnant women have anemia?
-with normal pregnancy, blood volume increases, which results in a concomitant
hemodilution
-although red blood cell mass increases during pregnancy, plasma volume increases more,
resulting in a relative anemia
-iron requirements are greater in pregnancy than in the non-pregnant state
-each pregnancy requires 500 to 600 mg of iron for the fetus, and to cover blood loss
during parturition
-lactation causes further demands
Why do women have higher incidence of anemia?
- average monthly loss of iron from menstruation is 15-28 mg
Classification of Anemias
Based on MCV
Microcytic anemias: MCV less than 80
Macrocytic anemias: MCV more than 100
Normocytic anemias: MCV between 80 to 100

Microcytic Anemias
MCV: < 80
Microcytic Anemias: Causes
Iron deficiency anemia
Anemia of Chronic Disease
Sideroblastic Anemias
Thalassemias (β and α)
Iron Deficiency Anemia (IDA)
Incidence:
- most common disease globally
- 30% of world population. Approximately 2 billion (WHO stats)
- young children, and women in reproductive age group, pregnancy
- “iron deficiency is a late manifestation of prolonged negative iron balance”
Iron metabolism (General):
Iron stores in body
- total body iron content: 3 gm in females, and 6 gm in males
- distribution: Functional (80%), and Storage (20%)
Functional Iron
- Hemoglobin
- Myoglobin
Storage Iron
- Ferritin
- Hemosiderin
Recommended Dietary Allowance:
- 10 mg per day. Only 10% is absorbed from the gut.
Dietary Iron
- heme (meat products)
- non-heme (plant source)
Iron Absorption
- occurs in duodenum and jejunum
- heme iron absorption is best (20% of heme iron is absorbed)
- non-heme iron (only 1% to 2%)
Iron Balance
- iron balance is maintained by regulating the absorption of dietary iron in the
proximal duodenum
- “iron is never lost from body”. As there is no regulated pathway for iron excretion
- iron loss is limited to the 1 to 2 mg lost each day through the shedding of mucosal
and skin epithelial cells
Iron Absorption:
Physiology/Biochemistry
Regulation of iron absorption
- duodenal epithelial cell uptake of
heme and nonheme iron
- when the storage sites of the body
are replete with iron and
erythropoietic activity is normal,
plasma hepcidin balances iron uptake
and loss to and maintains iron
homeostasis by downregulating
ferroportin and limiting iron uptake
(middle panel). Hepcidin rises in the
setting of systemic inflammation or
when liver iron levels are high,
decreasing iron uptake and
increasing iron loss during the
shedding of duodenocytes (right
panel). Conversely, hepcidin levels
fall in the setting of low plasma iron,
primary hemochromatosis, or
ineffective hematopoiesis (left
panel), leading to a rise in iron
absorption. DMT1, Divalent metal
transporter 1.
Etiology:
Infants
- ingestion of non-fortified cow’s milk
- continuation of breast feeding beyond 6 months
Children
- malnutrition (poor countries)
- Meckel’s diverticulum
Adolescence
- females: menstruation
- Hookworm infestation
Adults
- in females- menorrhagia, pregnancy, lactation
- iron loss thru’ peptic ulcer, intestinal polyps, colonic cancer (chronic iron loss)
Pathogenesis:
- when supply fails to meet the demand
- this would shift the iron balance to negative
- deficit initially is corrected by mobilizing the iron stores
- its only when the tissue stores are exhausted
- that “iron deficiency anemia” manifests
Lab diagnosis:
CBC: mostly normal, though total RBC
count may be lowered
Hb: reduced (normal is 14 gm% to 17
gm%)*
Hematocrit: reduced (normal is 42% to
54% in adult males and 38% to 48% in
adult females)
Red cell indices: MCV, MCH, and MCHC
are reduced RDW: A) Normal B) Iron Deficiency Anemia
RDW (red cell distribution width):
increased
Peripheral Blood Smear:
- very important test
Interpretation of peripheral smear
RBC: anisocytosis (variation in shape), poikilocytosis
(variation in shape)
- microcytic and hypochromic
- target cells may be seen when severe
WBC: usually normal. Eosinophilia may be noted in
presence of Helminthic/parasitic diseases
Platelets: normal. Count may increase in infective states Peripheral Blood Smear
(reactive)
Note: microcytic, & hypochromic red cells.
Compare normal blood smear with
iron deficiency
Bone Marrow study:
- ideal but always necessary
- hypercellular marrow (increased
erythropoiesis)
- characteristic feature: “microcytic maturation”
- there is extra division seen in maturation
sequences
- loss of stainable iron, this can be quantified
using Perls Prussin Blue stain Bone marrow Prussian Blue: normal (grades 2, and 3)
Bone marrow film stained with Prussian blue: a) normal, b) iron deficiency
Lab diagnosis: Biochemical

1)Serum iron: low
2) TIBC (total iron binding capacity): increased in iron deficiency
4) Serum ferritin: decreased
Remember: serum ferritin may not be sensitive (infections/inflammation is associated with higher
ferritin levels)
3) Percentage saturation: percentage of transferrin molecules that are bound by iron (normal: 33%).
This is decreased in iron deficiency.
5) Free Erythrocyte Protoporphyrin (FEP): increased
Stages of Iron Deficiency Anemia
- absent iron stores decreased serum ferritin decreased serum
iron increased TIBC decreased iron saturation normocytic
normochromic anemia microcytic hypochromic anemia
Clinical features
- fatigue, light headedness, palpitation, pounding in the ears, decreased work
performance (school children)
- menstrual abnormalities
- sometimes angina pectoris (severe anemia), high-output heart failure
- pica (craving for unusual things. example: clay)
- pallor, glossitis (smooth red tongue)
- angular cheilitis, ‘burning sensation’ of tongue
- koilonychia (spoon shaped concavity of nails)
- growth impaired (children)
- hair loss (especially in females)
Angular cheilitis
koilonychia
Glossitis
Clinical features (cont’d)
Plummer-Vinson syndrome:
- Triad of findings
i) microcytic hypochromic anemia
ii) atrophic glossitis
iii) esophageal webs
- in the junction between hypopharynx, and esophagus: web of mucosa is seen,
leading to dysphagia
- risk of carcinoma (premalignant lesion)
Plummer-Vinson syndrome
Barium x-ray showing cervical web (arrow) and large

diaphragmatic hernia. The lower esophagus is kinked secondary
to the hernia.
Anemia of chronic disease (ACD)
Incidence
-very common
Etiology
Associated with
i)Chronic microbial infections: TB, osteomyelitis, pelvic inflammatory diseases,
bacterial endocarditis, lung abscess
ii)Chronic immune disorders: Rheumatoid arthritis, SLE, vasculitic syndromes
iii)Neoplasms: for example- Hodgkin lymphoma
Pathogenesis
- caused by IL-1, IL-6, and TNF, IFN gamma
- brought on by chronic infection/neoplasia
- these cytokines stimulate
Hepcidin: synthesized in liver. Hepcidin inhibits release of iron from the storage
pool (inhibits ferroportin function)
Erythropoietin (EPO): reduction in erythropoietin, leads to diminished bone
marrow production of red blood cells.
Hepcidin, and cytokines depress erythropoietin synthesis
Lab Diagnosis
Anemia: microcytic, hypochromic (when fully evolved)
MCV: is reduced
TIBC: is decreased or normal
serum ferritin: is increased or normal
serum iron: is reduced
Bone marrow
Cellularity: normal usually
macrophage iron: increased
*remember: serum ferritin is decreased in Iron deficiency anemia
- bone marrow hemosiderin is low or even absent in iron deficiency anemia
Sideroblastic anemia
Incidence:
- group of disorders of varying etiology
- characterized by pathologic iron deposits
in erythroblast mitochondria
- iron glutted mitochondria is seen as ring
sideroblasts
ringed sideroblasts (arrows)

Classification of Sideroblastic Anemia:
Hereditary: rare cause, X linked (ALA synthetase-2)
Acquired:
Primary
- Myelodysplastic Syndrome (MDS)
Secondary
- Chronic Alcoholism
- Drugs (Isoniazid, Chloramphenicol)
- Pyridoxine deficiency (Vit B6)
- Lead poisoning
Pathogenesis
- defect in heme synthesis within mitochondria
- iron is brought into mitochondria to form heme
- if protoporphyrin is absent, iron would remain within mitochondria
- these iron laden mitochondria appear as a ring around the nucleus of
developing red cells (precursors) in bone marrow
- “ringed sideroblasts”
- this leads to increased iron stores (bone marrow and macrophages)
Lab Findings
Anemia: microcytic hypochromic
MCV: reduced
Peripheral blood smear: dimorphic red cells may be present (normocytic +
microcytic)
- Pappenheimer bodies
Bone Marrow
Cellularity: erythroid hyperplasia, abnormal forms- “ring sideroblasts” (15 to
100%), increased hemosiderin
Special test: expression of mitochondrial ferritin
Sideroblastic Anemia: Pappenheimer bodies
Lab Findings (cont’d):
Biochemical
TIBC: reduced
Serum Ferritin: increased
Serum iron: increase
Urine Porphyrin: erythropoietic porphyria
Remember: order for serum lead, alcohol (when appropriate)
Clinical Features
- fatigue, pallor, palpitation. Slow, progressive onset.
- iron overload
Cause looked for: alcohol, INH, MDS
Remember: patients may be Vit B6 responsive, or Vit B6 resistant (on treatment
with Pyridoxine)
Lead poisoning (self-directed learning)
Introduction
- lead exposure may occur through contaminated air, food
and water
- flaking lead paint in older houses is an important source
of lead poisoning
- blood levels of lead in children living in older homes
containing lead-based paint or lead-contaminated dust
often exceed 5 µg/dL, the level at which the Centers for
Disease Control and Prevention (CDC) recommends that
measures be taken to limit further exposure
- treatment is needed only when blood lead levels are
≥45 µg/dL, however, subclinical lead poisoning may occur
in children with blood lead levels considerably below this
mark
Lead Poisoning
- lead is a readily absorbed metal
- that binds to sulfhydryl groups in
proteins and
- interferes with calcium metabolism,
- effects that lead to hematologic,
skeletal, neurologic, gastrointestinal, and
renal toxicities
Sources of lead:
- from mining, foundries, batteries, and
spray painting, which constitute
occupational hazards
- flaking lead paint in older houses and
soil contamination pose major hazards
to youngsters
Lead Poisoning
Coarse stippling
Lead line (Plumbism)
Increased density in epiphysis

Introduction to Hemolytic Anemias
(General)
Hemolytic Anemia: Site
site of red cell destruction
Intravascular hemolysis
-acute process
-destruction of red cells within circulation
-with release of free Hb
Examples: Incompatible blood transfusion, Malaria, Sepsis, PNH
Extravascular hemolysis
- exaggeration of the normal mechanism of removal of aging red cells. Spleen being the most
common site.
- red cells are recognized as abnormal by reticulo endothelial system, and therefore destroyed
(spleen)
- Example: Hereditary spherocytosis
Evidence of increased Hb breakdown
1) Hyperbilirubinemia & jaundice

- not a reliable guide to the rate of hemolysis
- bilirubin level depends upon the amount of
Hb broken down, but also..
- the ability of liver to excrete the increased
amount of bilirubin presented to it
- remember: absence of jaundice does not
exclude the diagnosis of hemolytic anemia
2) Plasma haptoglobin
- haptoglobin is synthesized in the
liver
- haptoglobin binds to free Hb in
plasma
- Hb is small enough to pass thru’ the
normal glomerulus
- Hb combined with haptoglobin is a
large molecule (150 kDa)
- thus cannot pass through the
glomerulus
2) Plasma haptoglobin (cont’d)

- following transient release of Hb into circulation,
- the plasma haptoglobin level falls & returns to normal after 3 to 6 days
- if there is persistent hemolysis, then plasma haptoglobin level remains low
- haptoglobin is reduced in both types of hemolysis- intravascular &
extravascular
- plasma haptoglobin is normally expressed in terms of Hb binding. Normal: 0.3
to 2.0 gm/L
Intravascular and Extravascular hemolysis: plasma haptoglobin is low
3) Plasma hemopexin
- binds to free heme
- does not bind to Hb
Mechanism:
- if large amounts of Hb is released, and plasma haptoglobin level is exceeded
- some of the unbound Hb is converted to methemoglobin
- methemoglobin disassociates into ferriheme and globin
- ferriheme binds to hemopexin
- ferriheme is not lost through glomerular filtration
Intravascular hemolysis: plasma hemopexin level is low
Evidence of intravascular hemolysis
Hemoglobinemia and hemoglobinuria
- normal level of free Hb in plasma is low: 0.6mg/dl.
- in intravascular hemolysis, when haptoglobin binding capacity is exceeded plasma Hb
increases to 100 to 200 mg/dl
- when plasma Hb is increased, it imparts a pink/red color to plasma (hemoglobinemia)
- when renal threshold for Hb reabsorption is exceeded, hemoglobinuria is noted
- imparts pink color to urine on examination
Remember: hemoglobinuria must be differentiated from hematuria
through microscopy of urine
Remember: urine may be red due to drugs: pyridium, rifampicin. Others: porphyrinuria,
myoglobinuria
Features of accelerated hematopoiesis
Reticulocyte count: increased (normal: 0.2 to 2.0%)

Peripheral blood smear: presence of nucleated red cells
Bone marrow: erythroid hyperplasia
Lactate Dehydrogenase (LDH): increased, non-specific. Indicates high cell turn-over
Extra medullary hematopoiesis: sites- liver, spleen
Gall bladder: cholelithiasis (pigment gall stones)
Skeletal system: poor growth. Enlarged skull (chipmunk facies)
Pigmented gallstones
Reticulocytes in peripheral blood
smear: polychromatophils
Reticulocytes: supravital stain

Thalassemia syndromes
Normal Hemoglobin molecule:
Structure of Heme:
-derived from porphyrin
-porphyrins are cyclic compounds
formed by 4 pyrrole rings
-an atom of iron is present, therefore
called as ferroprotoporphyrin
-the 4 rings will be called as: alpha,
beta, gamma, and delta
-heme consists of ’ferrous iron’
Adult Hb: most HbA- 2 alpha, 2 beta,
and HbA2- 2 alpha, 2 delta
Fetal Hb: HbF- 2 alpha, 2 gamma
Thalassemia syndromes
Alpha Thalassemia: deficient / or lack of synthesis of alpha (α) chain
Beta Thalassemia: deficient synthesis (b+ thalassemia), or
total absence (b0 thalassemia)
- diminished synthesis of structurally normal β globin chains, along with unimpaired
synthesis of α chains
- genetic mutation (splicing mutation involving introns, promoter region mutations, and
chain terminator mutations)
- in beta Thalassemia, the deficit in HbA (2 alpha, 2 beta) produces “under
hemoglobinized” hypochromic, microcytic red cells which has less oxygen bound to it.
There is also, diminished survival resulting from an imbalance between alpha, and beta
chains
Beta thalassemia
Incidence
-mostly seen in Mediterranean, African, and Southeast Asian population
-manifestations of the disease may not be apparent until a complete switch from
fetal to adult Hb synthesis occurs (by six-months)
-Beta thalassemia affects 1 or both of the beta-globin genes
-inherited as an autosomal recessive disorder
Beta thalassemia
Clinical Syndromes
Homozygous: in general patients having beta thalassemia alleles B+/B+, B+/B0, B0/B0
have severe, transfusion-dependent anemia: Beta thalassemia Major
Heterozygous: patients have one normal beta chain, and one beta thalassemia
gene (B/B+, B/B0). These patients have mild asymptomatic anemia: Beta
thalassemia Minor/Trait
Beta thalassemia
Pathogenesis
-lack of adequate HbA (a2 b2) production leads to imbalance between a & b chains
synthesis
-excess free a chains in comparison to reduced b chains:
-insoluble precipitates aggregates in rbc
-inclusions of a chains damages cell membrane , vulnerable to phagocytosis (extra-
vascular hemolysis)
-HbA synthesis is low, resulting in poorly hemoglobinized ‘microcytic hypochromic’
red cells
Pathogenesis:
- beta Thalassemia
Beta thalassemia major: Labs
Hb: low, severe anemia

RBC Indices: MCV decreased
Peripheral blood smear: microcytic, hypochromic, target cells, fragmented red cells, nucleated red cells
Reticulocyte count: increased
Biochemical: serum bilirubin: increased
Iron studies:
TIBC: normal/decreased
serum ferritin: increased
serum iron: increased
Hb Electrophoresis: HbA2, and HbF are increased
- low or absent HbA
β-thalassemia: Note: nucleated red cell,
hypochromia, target cells
Beta thalassemia major
Clinical features
- marked pallor, jaundice
- requires blood transfusion
- secondary hemochromatosis
- heart failure from iron overload
- growth retardation
- splenomegaly by 3 years
- skeletal system abnormalities
- gall stones
Treatment
- blood transfusion
- splenectomy may help
β-thalassemia: “crew cut” or “hair on end” appearance on skull x-ray.
Beta Thalassemia Minor
- more common than thalassemia major
- moderate reduction in the synthesis of β chain
- also referred to as thalassemia trait, normal life expectancy
- diagnosis made by chance, anemia is mild
- but becomes evident during pregnancy
- mild microcytic anemia, decreased Hb, decreased MCV, and normal to increased serum
ferritin levels, decreased TIBC
- Hb electrophoresis: mild decrease in HbA, and increase in HBF, and HBA2
Importance: differentiation from iron deficiency anemia
- iron deficiency anemia improves with iron therapy, whereas
- β Thalassemia trait or minor worsened by iron therapy
- estimation of serum iron, ferritin and transferrin are important
Alpha thalassemias
Incidence
-South East Asia, and China
-reduced or absent synthesis of α globin chains
-affects: Hb-A, Hb-A2 & Hb-F
-severity depends upon the number of α chains missing
-studies show reduced red cell invasion by Malarial parasites in thalassemia (beta
as well as alpha)
-this may explain the high prevalence of these disorders in Malaria prone areas
-Malaria may be responsible for maintaining the prevalence of
‘hemoglobinopathies’ in the world
Alpha thalassemias
Genetics and Clinical Types

- total number: 4 α globin chains
- caused by deletion of α globin chains
Clinical types
Silent carrier
- deletion of single α chain
α thalassemia trait
- deletion of two α chains
HbH disease
- deletion of three α chains
- severe hemolytic disease
Hydrops Fetalis:
- deletion of all four α chains
- incompatible with life
Alpha thalassemias
Hydrops fetalis
- deletion of all four α globin chains
- most severe form, South East Asia
- in fetus, excess gamma chains form tetramers (Hb Barts)
- very high affinity for oxygen, but no oxygen delivered to tissue
- decreased MCV, increased rbc count
- survival of fetus to term is mainly due to other embryonic hemogloblins (Hb Portland 1,
& Hb Portland 2)
- many pregnancies terminate unnoticed or early in gestation
- clinical picture similar to severe Rh-incompatibility (erythroblastosis fetalis)
Alpha thalassemias
Hydrops fetalis (cont’d)

- fetal distress occur in 3rd trimester
- intra uterine death
Findings in fetus
- severe anemia (Hb 3 to 8 gm%), pallor, generalized edema, massive
hepatosplenomegaly
- mother frequently has toxemia of pregnancy (hypertension, fluid retention with
or without proteinuria)
α-thalassemia: Note edema of face and abdominal distension (A). The thoracic
cavity is filled by the pericardial sac distended by effusion and cardiomegaly; there is
massive hepatomegaly (B).
“The weak can never forgive. Forgiveness is the attribute of the strong”
Mahatma Gandhi
RBC Disorder. Part 2
Dr. Roy
Macrocytic anemia
MCV: > 100
Megaloblastic anemia
What is megaloblastic anemia?
- it is a group of disorders that share common morphologic characteristics: large
cells with an arrest in nuclear maturation
- nuclear maturation is immature relative to cytoplasmic maturity
- these abnormalities are mostly due to impaired DNA synthesis
Is there a difference between megaloblastosis, and macrocytosis?

- yes, there is: increase in MCV indicates ‘macrocytosis’. Macrocytic anemia is
classified as
1) Megaloblastic (bone marrow changes present)
2) Non-megaloblastic (not seen with Vit B12/Folic acid, therefore bone marrow
changes are very different)
Types
Megaloblastic anemias:
- vitamin B12 deficiency
- Folic deficiency
Macrocytic anemias:
- Alcoholism
- Anemia of Renal failure
Megaloblastic anemia
Introduction:
- group of disorders characterized by the presence of distinctive morphologic
appearances of the developing red cells in the bone marrow
- the marrow is usually cellular and the anemia is based on ineffective
erythropoiesis
- the cause is usually a deficiency of either cobalamin (vitamin B12) or folate
- interaction between folate and B12 is responsible for the megaloblastic
anemia seen in both vitamin deficiencies
Vitamin B12 Deficiency
Decreased Intake
1) Inadequate diet, Vegan
Impaired Absorption
1) Intrinsic Factor deficiency: Pernicious Anemia
2) Gastrectomy
3) Malabsorption
4) Ileal resection
5) Parasite: tapeworm
6) Bacterial overgrowth: Diverticulosis
Increased Requirements
1) Pregnancy/Lactation
Vitamin B12
Sources:
- animal protein origin
- lesser amounts seen in eggs, cheese, & milk
- vegetables contain no vit B12
- vegans are at risk for vit B12 deficiency
- it will take few years before vit B12 stores are completely depleted
Absorption:
- active mechanism is mediated though gastric intrinsic factor, highly efficient mechanism
8
Vitamin B12
Transport:
- Vit B12 binds to transcobalamin II and transcobalamin I
- transcobalamin I is essentially a storage form
Tissue stores:
- liver is the main storage site
- smaller amounts stored in kidney, heart, and brain
9
Vitamin B12 Biochemistry
Odd chain fatty acid oxidation

Absorption of Vitamin B12 & Folic
Acid
13
Vitamin B12: Pernicious Anemia
Incidence:
- older age group (5th to 8th) decade of life
- familial pattern
- older than 60, more in women, Scandinavia
- may be associated with other autoimmune diseases: Hypothyroidism, Vitiligo
Pathogenesis:
- malabsorption of vitamin B12 in patients with pernicious anemia is due to intrinsic-factor
deficiency. Two mechanisms
1) progressive destruction and eventual loss of parietal cells from the gastric mucosa lead
to failure of intrinsic-factor production
2) blocking autoantibodies present in the gastric juice can bind to the vitamin B12 - binding
site of intrinsic factor, thereby preventing the formation of the vitamin B12 - intrinsic
factor complex
Gastric mucosa: normal
Gastric mucosa: chronic atrophic gastritis

Lab Diagnosis:
Complete Blood Counts: leukopenia, and thrombocytopenia
Hb: reduced
Red cells: macrocytic change (typically oval; ovalocytes), and Howell-Jolly bodies
White blood cells:
- hypersegmented neutrophils are seen
Reticulocyte Count: reduced
Red cell indices:

- increased MCV (› 100) in most cases MCV exceeds 120
Lab Diagnosis:
Bone Marrow:
- increased cellularity (erythroid)
- megaloblastic change seen in all erythroid precursors
- nucleus retains finely distributed chromatin, no chromatin clumping
- cytoplasm shows regular hemoglobinization (N/C asynchrony)
- granulocytic series: giant metamyelocytes
- platelets: abnormally large, multilobation of nuclei
Bone Marrow study

19
Lab Diagnosis
Biochemical Tests:
Serum bilirubin: mild increase
Serum cobalamin: decreased levels
Serum Homocysteine: increased (not specific)
Methylmalonic acid: increased in serum and urine.
*Seen only in cobalamin deficiency and not in Folate defn
Antibodies to IF: present in most cases
Schilling test: assays vit B12 absorption by measuring urinary radioactivity, after an oral
dose of radioactive Vit B12
Schilling Test
Description Radiolabeled Vit B12 measure in urine
Stage 1 Oral radiolabeled Vit B12 plus Normal excretion: Malnutrition, or increased demand
Intramuscular unlabeled Vit B12 injection * If decreased excretion, go to stage 2
Stage 2 Oral radiolabeled Vit B12 plus Intrinsic Normal excretion: Malabsorption due to Intrinsic
Factor Factor deficiency
*If decreased excretion, go to stage 3
Stage 3 Oral radiolabeled Vit B12 plus Antibiotics Normal excretion: Bacterial overgrowth
*If decreased excretion, go to stage 4
Stage 4 Oral radiolabeled Vit B12 plus Pancreatic Normal excretion: Lack of pancreatic enzymes (Ch.
enzymes Pancreatitis)
Gastric Biopsy
- chronic atrophic gastritis can be confirmed
by gastric biopsy
- Total (pentagastrin-resistant) achlorhydria,

the direct result of the loss of gastric
parietal cells, is diagnostic of pernicious
anemia because it is the only gastric lesion
that results in total achlorhydria
Chronic Atrophic Gastritis: please note intestinal

metaplasia (red arrow), inflammation (green arrow)
Clinical features:
- anemia (lemon tint), mild icterus, atrophic glossitis (smooth, sore (beefy)
Neurological:
- peripheral neuropathy: paresthesias and numbness
- lesions in the posterior and lateral columns of the spinal cord: subacute
combined degeneration
- cerebral manifestations: range from mild personality defects and memory loss to
frank psychosis (“megaloblastic madness”)
Neurological:
Lesion in the spinal cord:
- are a mixture of signs of a posterior column lesion (loss of
vibration and position sense, and sensory ataxia with positive
Romberg's sign)
- and those of a lateral column lesion (limb weakness, spasticity,
and extensor plantar responses)
Treatment
- the standard treatment is regular monthly intramuscular
injections of at least 100 μg of vitamin B12 to correct the
vitamin deficiency
- this treatment corrects the anemia and may correct the
neurologic complications if given soon after their onset
Vitamin B12 deficiency: Clinical Features
Folic Acid: Megaloblastic anemia
Incidence:
- more in economically poor nations
- other considerations: history of excessive alcohol intake with concurrent poor
diet intake (‘tea and toast’)
Pathogenesis:
- folates are present in natural foods and tissues as polyglutamates because these
forms serve to keep the folates within cells
- in plasma and urine, they are found as monoglutamates because this is the only
form that can be transported across membranes
- enzymes in the lumen of the small intestine convert the polyglutamate form to
the monoglutamate form of the folate, which is absorbed in the proximal jejunum
Pathogenesis:
- within the plasma, folate is present, mostly in the 5-methyltetrahydrofolate (5-
methyl THFA) form, and is loosely associated with plasma albumin in circulation
- the 5-methyl THFA enters the cell via a diverse range of folate transporters
- once inside, 5-methyl THFA may be demethylated to THFA, the active form
participating in folate-dependent enzymatic reactions
- cobalamin (B-12) is required in this conversion, and in its absence, folate is
"trapped" as 5-methyl THFA
Decreased Intake
1) Malnutrition
2) Infants/Elderly
3) Alcoholism
Malabsorption
1) Celiac Disease
Drug Inhibition
1) Methotrexate
2) 5-Fluorouracil
Increased utilization
1) Pregnancy, lactation
2) Disseminated malignancies
Lab diagnosis: (blood findings similar to Vit B12 deficiency)
Complete Blood Counts: leukopenia, and thrombocytopenia
Hb: reduced
Red cells: macrocytic change (typically oval; ovalocytes), and Howell-Jolly bodies
White blood cells:
- hypersegmented neutrophils are seen
Red cell indices:
- increased MCV (› 100) in most cases MCV exceeds 120
Bone Marrow: similar to Vit B12 deficiency
Lab diagnosis:
Serum Folate: reduced
Red blood cell folate level: reduced
Serum homocysteine: increased (similar to Vit B12)
Methylmalonic acid: normal (raised in Vit B12)
Folic acid: neural tube defect (NTD)
Incidence:
- spina bifida and anencephaly, the two most common forms of neural-tube
defects, occur in 1 in 1000 pregnancies in the United States
Embryology:
- development and closure of the neural tube are normally completed within 28
days (3rd and 4th week) after conception
- before many women are aware that they are pregnant
- closure of the neural tube occurs at several sites and that the clinical types of
neural-tube defects differ depending on the site
Embryology (cont’d):
Cranial defects
- incomplete closure of anterior neuropores
- cranial cleft formation (open defects) with involvement of skull and brain tissue
Spinal defects
- incomplete closure of posterior neuropores
- bone defects of vertebral arches (lumbar or sacral)
- possible herniation of spinal neural tissue and meninges
NTD: are classified as Open or Closed

Spina bifida occulta: occurs at S1, S2 or both. Usually
covered by normal skin
Meningocele: saccular herniation of meninges, and
CSF. Meningoceles are covered by normal skin.
Myelomeningocele: most common. These are usually
open defects, in which either meninges, or neural
tissue is exposed
Pathogenesis:
- women who gave birth to babies with neural-tube defects had low serum levels
of micronutrients, including some vitamins
- folate is closely linked to neural tube defects
Lab:
- monitoring pregnant women using alpha fetoprotein
- alpha fetoprotein in maternal serum is raised in neural tube defect
- detected best during 14th to 20th week of gestation
Non-megaloblastic anemia
Know the differences between ‘megaloblastic’ and ’non-megaloblastic’ anemias
In non-megaloblastic anemia:
- macrocytes are round rather than the typical ovalocytes
- hypersegmented neutrophils are absent
- pancytopenia not seen
- neurological features, and glossitis are not seen
- anemia may not manifest
- alcohol is the most common cause
Anemia of Renal failure
Pathogenesis:
Three factors: 1) reduced erythropoietin, 2) suppression of erythropoeisis, and 3)
shortened red cell survival
- of these, reduction in erythropoietin (EPO) is a major cause
Labs:
Peripheral blood smear: macrocytic red cells, ‘burr cells’
Platelets: qualitative defects due to uremia. Prolonged bleeding time
Treatment:
- Darbepoetin alfa (Aranesp)
- Epoetin (Epogen)
Anemia of chronic renal failure: Burr cells on peripheral blood smear
End-Stage Renal Disease (ESRD): CVS issues
Alcoholism
Introduction:
- as an indirect effect, alcohol can produce ‘megaloblastic’ anemia due to nutritional causes
(Folic acid, and Vit B12 deficiencies)
- as a direct effect, alcohol can act on bone marrow, and precursor cells to produce ‘non-
megaloblastic’ anemia
Pathogenesis:
- uncertain. Possible mechanisms include antibodies against acetaldehyde modified red cell
protein.
- remember, folate and Vit B12 levels remain normal in ‘non-megaloblastic’ anemia
Blood picture:
- macrocytes noted, tend to be thin, and rounded
Remember: life span of the red cells are not reduced
Normocytic Anemia
MCV: 80 to 100
Reticulocyte count
- reticulocyte count is used to estimate the degree of effective erythropoiesis

- reported as absolute reticulocyte count or as a reticulocyte percentage
- the normal reticulocyte count in adults is 0.5% to 2%
The problem
- when Hb drops in a healthy individual, reticulocyte count increases
- to factor in this problem, you need to correct the reticulocyte count
- you now use: corrected reticulocyte count, whenever Hb is low
Reticulocyte count
Corrected Reticulocyte count

Formula: reticulocyte% X (Hb/15)
- this formula helps to correct the Hb, and let you know if the patient is making
enough reticulocyte
- thereby, letting you know if the bone marrow is healthy
- less than 3% (corrected reticulocyte count) tells you that the bone marrow is not
responding
- more than 3% (corrected reticulocyte count) tells you that the bone marrow is
responding, thus healthy
Normocytic Anemia: corrected reticulocyte count < 3%
Aplastic Anemia
Intro
- there is reduction in number of hematopoietic stem/progenitor cells
- leads to an inability to produce normal numbers of mature cells
Etiology:
- please see following slide
Acquired Aplastic Anemia: Etiology
Idiopathic
Acquired stem cell defects
Immune mediated
Chemical Agents
Alkylating agents
Antimetabolites
Benzene
Chloramphenicol
Inorganic arsenicals
Idiosyncratic
Chloramphenicol
Phenylbutazone
Organic arsenicals
Carbamazepine
Penicillamine
Gold salts
Physical Agents
Whole-body irradiation
Viral Infections
Hepatitis (unknown virus)
Cytomegalovirus infections
Epstein-Barr virus infections
Herpes zoster (varicella zoster)
Inherited
Fanconi anemia
Telomerase defects
Aplastic anemia
Pathogenesis:
- there are two possible mechanisms
1)Extrinsic: immune mediated suppression of marrow progenitors
Mechanism
- stem cells may be antigenically altered by drug exposure/infection
- this provokes cellular immune response
- activated TH1 T cells produces: IL-1, TNF, IFN gamma
- suppress and kill hematopoietic progenitors
47
Aplastic anemia
Pathogenesis:
- 2) Intrinsic: due to stem cell abnormality
- characterized by karyotype aberration
- which transforms to myeloid malignancy
- example- Myelodysplastic syndrome (MDS) and Acute Myelogenous Leukemia
(AML)
48
Pathogenesis: Aplastic Anemia
Aplastic anemia
Lab diagnosis:
- pancytopenia
- absolute neutrophil count is reduced
Bone marrow
- marked reduction in cellularity (less
than 25%)
- increase fatty marrow
- “empty bone marrow”
50
Aplastic anemia
Clinical Importance:
- need to exclude other causes
- Myelodysplastic syndrome, leukemias, hairy cell leukemia
Clinical features:
- pallor, weakness, easy bruising, infections
51
“the best portion of a good man's life is his little, nameless, unremembered acts of kindness
and of love”
William Wordsworth, poet (1770-1850)
RBC Disorders.
Part 3
Dr. Roy
Normocytic Anemia
MCV: 80 to 100
Corrected Reticulocyte count > 3%
Causes:
Membrane defects:
- Hereditary Spherocytosis
- Paroxysmal Nocturnal hemogloblinuria (PNH)
Hemogloblinopathies:
- Sickle cell disease
Enzyme defects:
- G6PD Deficiency
- Pyruvate kinase deficiency
Membrane defects: Hereditary Spherocytosis
Introduction:
- inherited disorder due to intrinsic defects in the red cell membrane, mostly
Autosomal Dominant
- resulting in red cells which are spherical and less deformable (red cells become rigid)
- vulnerable to splenic sequestration and distortion
- in North European ancestry
4
Hereditary spherocytosis (HS)
Pathogenesis:
- mutation in spectrin, or
- mutation in ankyrin
Others:
- Band 3
- Protein 4.2
- loss of red cell membrane
- increased permeability to sodium
5
Hereditary Spherocytosis:
Pathogenesis
6
Labs:
Hb: reduced
MCHC: increased (this due to reduced volume of red
cells, and Hb remains constant)
RDW: increased
Hemolytic Anemia features: increased bilirubin, and
decreased Haptoglobin
Peripheral Blood smear: normocytic normochromic,
loss of central pallor
Spherocytes
Labs (cont’d):
Eosin 5-malemide (EMA) binding test:
- rapid test (2 hours)
- shows decreased binding between the dye
(EMA) and the red cell membrane
- this can been seen on flow cytometry
Labs (cont’d)
Osmotic fragility test:
- increased osmotic fragility
- spherocytes rupture easily
Other tests:
- increased serum bilirubin
- LDH: increased
- negative Coomb test
Osmotic fragility test
10
Clinical features/complications
- anemia, jaundice, splenomegaly, gall stones
Aplastic crises:
- associated with acute parvovirus infection
- virus kills red cell progenitors
- worsening anemia, reduced reticulocytes
Hemolytic crises:
- increased splenic destruction of spherocytes
- enlarged tender spleen
- worsening anemia, increasing jaundice and darkening of urine
Cholelithiasis
- pigment gall stones (hyperbilirubinemia)
11
Treatment:
- splenectomy (done after 7 years of age)
- prior immunization with polyclonal
pneumococcal vaccine
- antibiotics post splenectomy
*spherocytes can be still seen following

splenectomy
Also, Howell-Jolly bodies may be present
Pigment gallstones
12
Parvovirus infection (SDL)
- Parvovirus B19 (B19V) is a single-stranded DNA virus
- Parvovirus B19 has a unique tropism for human erythroid progenitor cells
- virus requires the P blood antigen receptor (also known as globoside) to enter
the cell
- virus is cytotoxic to host cells, tropism for rapidly dividing erythrocyte
precursors (particularly pronormoblasts and normoblasts),
- wherein they replicate to high titers), leads to the suppression of
erythrogenesis seen during infection
- no reticulocytes (immature erythrocytes) are available to replace aging or
damaged erythrocytes as they are cleared by the reticuloendothelial system
Please see footnotes
Paroxysmal nocturnal hemoglobinuria (PNH)
Introduction:
- rare, acquired disorder, chronic
- intravascular hemolysis, intermittent hemoglobinuria (occurs during the night)
Pathogenesis:
- hematopoietic stem cell acquire somatic mutation of PIGA (phosphatidylinositol glycan class A1)
- PIGA is located on X chromosome which encodes for an enzyme
- that is essential component of a biosynthetic pathway that generates GPI (glycosyl
phosphatidylinositol)
- GPI serves as a membrane anchor for more than 20 proteins
- among these proteins, includes DAF 14
Pathogenesis (cont’d)
1) Decay accelerating factor (DAF) or CD55
- normally DAF neutralizes complements that fixes onto red cells, neutrophils, and
platelets
- loss of DAF leads to greater binding of complement to red cells)
2) Membrane inhibitor of reactive lysis (MIRL) or CD59
3) C8 binding protein
- results in intravascular hemolysis due to activation of the membrane attack complex:
C5b-C9
15
Lab diagnosis:
Hb: low at the time of diagnosis
Serum haptoglobin: decreased
Serum hemopexin: decreased
Hemoglobinemia: present (pink to red plasma)
Coomb Test: negative
Urine: hemoglobinuria, hemosiderinuria
Bone Marrow: hyperplastic, reduced iron stores
16
Other tests:
Flow cytometry: detects granulocytes with missing anchors for
inhibitors of complement
Screening test: sucrose hemolysis test
Confirmatory test: HAM test
17
Clinical features:
i) Nocturnal hemoglobinuria
- passage of red/brown urine in the morning
- (respiratory acidosis during night time augments complement attachment to
cell)
- cyclic pattern
ii) Chronic hemolysis
- most common feature (hemosiderinuria)
- intravascular hemolysis
18
iii) Iron deficiency

- iron loss in urine
iv) Bleeding
- due to thrombocytopenia
v) *Risk of thrombosis (very important)

- due to release of aggregating factors from damaged platelets
- (hepatic vein/portal/cerebral)
Hepatic vein thrombosis: Budd-Chiari syndrome
vi) Development of acute myeloid leukemia (AML) is rare
19
Sickle cell anemia (HbS)
Introduction:
- point mutation at the 6th position of
the β globin chain leads to
- replacement of a glutamate residue
with a valine residue
20
Sickle cell anemia: Types
Heterozygous
- 8% of African Americans are heterozygous for HbS (sickle cell trait)
- 40 to 45% of the Hb is HbS
Homozygous
- in homozygous state, almost all Hb in red cells are HbS (90 to 95%).
Therefore the Hb in these cases would be:
HbS: 90 to 95%
HbA: absent
HbA2: 1 to 3%
HbF : more than 5%
21
Hemoglobin: Sickle Cell Disease
Hb Normal Sickle Cell Trait Sickle Cell Disease

HbA 95% to 98% 60% 0%
HbS 0% 40% to 45% up to 95%
HbF Less than 2% Less than 2% 5% to 25%

Pathogenesis:
- when deoxygenated, HbS undergoes polymerization
- with continued deoxygenation, aggregated HbS molecules form needle like fibers
within red cells
- producing distorted sickle cell or holly-leaf shaped structures
When sickle cells are present:
1) Chronic hemolysis (macrophage destruction)
2) Microvascular obstruction (rigid red cells)
3) Tissue damage (ischemic infarcts)
23
Sickle Cell Disease: Pathogenesis

Lab Diagnosis:
Hb: decreased

- normocytic normochromic
- sickled cells (10 to 15%)
25
Sickling test
- observation of sickling of red cells containing HbS
under cover slip
- when treated with 2% sodium metabisulfite
Hb electrophoresis: confirmatory
Prenatal diagnosis
- only in select cases
- recombinant DNA technique
- prepared from amniotic fluid cells
- obtained at 15th to 20th week
Neonatal screening:
- mandatory in USA 26
Clinical Features
1) Vaso-occlusive crises
- sickle red cells express higher than normal levels of adhesion molecules, and are
therefore sticky
- mediators release from granulocytes during inflammation up regulates
expression of adhesion molecules on endothelial cells
- this leads to stagnation of red cells in micro-circulation
‘Hand-foot’ syndrome (dactylitis)
- dorsa of hand and/or feet appear swollen, and painful
- micro-infarction of carpal and tarsal bones
Differential diagnosis: osteomyelitis 27
Clinical Features (cont’d)
2) Acute chest syndrome
- fever, cough, chest pain and pulmonary infiltrates, mostly seen in children (Streptococcus
pneumoniae, Mycoplasma pneumoniae)
- follows lung infection, pulmonary blood flow becomes sluggish, associated with
increased mortality
3) CNS
- seizures / strokes due to hypoxia
- catastrophic complication seen in children and young adults
- clinically: abrupt onset of hemiparesis/aphasia/seizures
- patients may make a complete recovery
4) Retinopathy
- loss of visual acuity or sometimes blindness 28

5) Severe Infections
- congestion and poor blood flow in spleen in children
- infarction and autosplenectomy in older children and young adults
Loss of splenic function:
- prone for infections
- pneumococci and Hemophilus influenzae
- sepsis and meningitis are common in children
- osteomyelitis caused by Salmonella paratyphi

6) Priapism
- sustained erection for more than 4 hours (in Sickle cell pts)
- medical emergency
7) Cholelithiasis
- pigmented gall stones
Sickle cell anemia (HbS): Complications
Sequestration crises
- massive sequestration of sickled cells in spleen
- rapid splenic enlargement, hypovolemia, and sometimes shock (dangerous)
- seen in children (first 2 years of life)
- occurs before spleen undergoes auto-infarction, and fibrosis
Aplastic crises
- transient bone marrow failure of erythropoiesis
- infection by parvovirus B19
- due to direct cytotoxicity of parvovirus to erythroid precursors (developing red cells in
bone marrow)
- worsening anemia and reduced reticulocyte count
Complications:
Cholelithiasis: hyperbilirubinemia
Chronic leg ulcers: (medial surface of tibia)
Renal: (renal papillary necrosis, loss of

concentration, dilution of urine)
Eye: sludging of blood in conjunctival vessels,

sickle retinopathy
Fat embolism: Leg ulcer in Sickle cell disease

G6PD deficiency
Introduction
- hereditary deficiency of G6PD, recessive X linked trait
- African Americans, Mediterranean variant (Italy/Greece)
- males are at risk for symptomatic disease
- mild hemolysis
- mostly intravascular hemolysis
- protection against plasmodium falciparum malaria

G6PD deficiency
Pathogenesis:
Functions of G6PD
- glucose 6 phosphate dehydrogenase reduces NADP to NADPH while oxidizing
glucose 6 phosphate
- NADPH then provides reducing equivalents needed for conversion of oxidized
glutathione to reduced glutathione
- which protects against oxidant injury by participating as a cofactor in reactions
that neutralize compounds such as H2O2
34
G6PD deficiency
Pathogenesis (cont’d):
- G6PD deficient red cells when exposed to oxidants, results in oxidation of reactive
sulfhydryl group on Hb chain
- which becomes denatured, forms a membrane bound inclusion: ‘Heinz body’
- Heinz bodies damages cell membrane
- results in intravascular hemolysis
extravascular hemolysis (less than intravascular): in spleen macrophages pluck out
Heinz bodies (bite cells)
35
G6PD deficiency
Factors which precipitate hemolysis:
- follows oxidant stress
- oxidant stress is noted with:
Drugs: anti malarials (primaquine, chloroquine, sulfonamides, nitrofurantoins)
Infections: viral hepatitis, pneumonia, typhoid fever
Fava bean ingestion (favism): mostly in children between 2 to 5 years
- Sardinia, Greece, Sicily
37
G6PD deficiency
Lab diagnosis:
Hb: decreased, mild to severe
Peripheral smear:
- normocytic anemia
- Heinz bodies, bite cells
G6PD levels: low
Hemogobinemia: intravascular hemolysis, pink/red plasma
Haptoglobin: reduced
Hemopexin: reduced (especially when hemolysis lasts longer)
Urine: hemoglobinuria
38
G6PD deficiency
Clinical features:
- features of acute hemolysis (2 to 3 days following oxidant stress & lasting about 7
days)
- associated with stress: drug administration (anti-malarial), infection, fava bean
ingestion
(decreased NADPH impairs microbial killing thru’ oxygen dependent pathway.
Therefore infections produces oxidant stress)
- transient splenomegaly
40
Pyruvate kinase deficiency
Introduction:
- hemolytic anemia due to glycolytic enzyme deficiency
- most cases reported in Europe
Pathogenesis:
- Pyruvate kinase deficiency results in impaired glucose utilization
- thereby decreased pyruvate kinase, and lactate production
- also glycolytic intermediates proximal to the block accumulate in the red cells
- the levels of 2,3 diphosphoglycerate may increase threefold (rightward shift with
increased delivery of oxygen to tissues)
- the major problem: diminished capacity to generate ATP
- in severe pyruvate kinase deficiency, the fall in ATP leads to irreversible cell injury
(failure of sodium potassium pump)
- these red cell lose their plasticity, and become rigid
- these rigid red cells are marked for premature destruction in spleen:
extravascular hemolysis
- these patients have higher number of circulating reticulocytes
Note the right shift of oxygen Hb curve, leading to
more unloading of oxygen as there is an increase in 2,3
DPG
Labs:
Hb: reduced
MCV: normal range
Peripheral blood smear: nucleated red
cells, acanthocytes with surface spicules
(spur cells)
S. Bilirubin: increased
Quantitative assay of PK: reduced
Clinical Features:
- may range from mild anemia to hydrops fetalis and neonatal jaundice
- beyond neonatal period anemia of varying degree, jaundice, splenomegaly
- cholelithiasis (bilirubin)
- aplastic crisis (parvovirus B19)
Treatment:
- during the early years, severe anemia is managed by red cell transfusions
“Whoever undertakes to set himself up as a judge of Truth and Knowledge is shipwrecked by the
laughter of the gods”
-Albert Einstein.
46
Autoimmune Hemolytic Anemia
Dr. Roy
Immune Hemolytic Anemia: Introduction
- immune hemolytic anemia is characterized by

- shortened red blood cell survival due to deposition of immunoglobulin and/or
complement on red cell surface
Autoimmune Hemolytic Anemia (AIHA)
- due to altered immune response, resulting in the production of antibody against
the host’s own erythrocytes
- some of the AIHA are mediated by antibodies with maximum binding affinity at
37℃ (Warm or Incomplete Ab)
- while other are mediated by antibodies with maximum binding affinity at 4℃
(Cold or Complete Ab)
Immune Hemolytic Anemia: Classification
Warm Antibody Type (IgG Ab active at 37℃, mostly Incomplete Ab)

Primary (idiopathic)
Secondary
- Autoimmune disorders (SLE)
- Drugs
- Lymphoid neoplasms (CLL)
Cold agglutinin type (IgM Ab active below 4℃, Complete Ab)
Acute (Mycoplasmal infection, Infectious Mononucleosis)
Chronic
- Idiopathic
- Lymphoid neoplasm (Lymphoplasmacytic lymphoma)
Cold Hemolysisn type (IgG Ab active below 37℃)
- rare, occurs in children following viral infections
An approach to AIHA
Points to remember
- a shortened red blood cell life span
characterized by:
1) low Hb
2) low Hematocrit
3) elevated bilirubin
4) elevated LDH (remember, LDH is not specific)
5) If hemolysis is Extravascular: spherocytes
6) If hemolysis is Intravascular: low hemopexin, hemoglobinuria, hemosiderinuria
7) DAT test (Direct Coombs test): positive
Coombs test
Direct Antiglobulin Test
Objective:
- to determine the presence of ‘anti- rbc antibodies’
- this test also determines the presence of ‘anti- complement antibodies’
Procedure:
- collect patient’s red cells
- wash the red cells (only firmly attached ‘antibodies’ would then remain)
- add Coombs reagent (Coombs reagent made by injecting a lab animal with
human red cells, and then obtaining the antibodies produced against human red
cells)
Coombs test
Direct Antiglobulin Test
- Coombs reagent would then contain Anti Human Globulin)
- if there are prefixed Ab on the patient’s red cell, adding
Coombs reagent would agglutinate these cells
- the test is interpreted as: positive
- DAT is a one stage test (in vivo)
Remember: Coombs in ‘Cold type’ tests for the
Clinical Importance presence of antibodies directed against complement
(C3b).
- Autoimmune Hemolytic anemia (AIHA)
- Drug induced AIHA
- Post transfusion immune mediated hemolysis
- Rh incompatibility
Coombs test
Indirect Antiglobulin Test (Indirect Coombs test)
Objective:
- to determine the presence of ‘anti- rbc antibodies’
- this test also determines the presence of ‘anti- complement antibodies’
- (the difference is that, in “Indirect Coombs”, the test is for detecting free
antibodies)
Procedure:
- collect patient’s red cells
- wash the red cells
- here, the test is done for: 1) Pre transfusion testing, 2) Prenatal screening testing
(Rh negative mother)
Coombs test
1) Pre transfusion test
- here, donor’s red cells are taken
- these cells are washed
- now, add recipient’s serum
- if antibodies (free) are seen in serum, they will react
- this can’t be detected on examination, for this to be seen Coombs reagent needs
to be added
- the resultant agglutination can be detected on microscopy
Coombs test
2) Pre natal screening test in Rh negative mother
- serum is taken from mother
- this is mixed with red cells which are commercially available (these red cells are
Rh positive)
- if the mother’s serum has Rh antibodies, there will a reaction
- to visualize the reaction on microscopy, Coombs reagent ahs to be added
- this test is ‘in vitro’
Immune Hemolytic Anemia
1) Warm Ab type
- IgG globulins having relatively high binding affinity for human RBCs at 37℃
- most common type
- may or may not fix complement
- typically leads to extravascular hemolysis (macrophages in spleen recognizes these
antibodies thru’ Fc portion)
Secondary causes include:
Autoimmune disorders (SLE). Most common.
Lymphoid neoplasms (CLL)
Drugs (Penicillin, α-methyldopa)
1) Warm Ab type (cont’d)
Pathogenesis:
- the patient’s RBCs typically are coated with IgG autoantibodies with or without
complement proteins
- autoantibody-coated RBCs are trapped by macrophages in spleen
- red cell hemolysis is extravascular
- a small amount of intravascular hemolysis may be seen
- IgG coated red cells bind to Fc receptors on macrophages
- which will remove red cell membrane during partial phagocytosis
- the red cell become spherocytic following loss of membrane
- these red cells are then broken down in spleen
2) Cold Ab type
Mycoplasma pneumoniae infection
- other infections: EBV, CMV, Influenza, HIV
Pathogenesis:
- IgM Ab binds to red cells and fixes complement at low temperatures. (Remember: IgM is a
large molecule, therefore it can span several red cells leading to agglutination)
- when the IgM/C3b-coated RBC circulates to warmer tissues, the IgM dissociates, leaving
complement C3b on the original RBC
- remember C3b is an excellent opsonin, which the leads to removal of affected red cells
by phagocytes in spleen, liver, and bone marrow
- in infections, this is usually transient and self-limited
- Raynaud’s phenomenon may be seen
Immune Hemolytic Anemia: Labs (Warm type)
Hb: reduced
Peripheral Blood Smear: spherocytic red cells,
polychromatophils, nucleated red cells
S. bilirubin: may be increased
Haptoglobin: may be reduced (remember: warm
type is extravascular)
DAT: positive
CLL: warm type. Note spherocytic rbc, n-rbc, and
“smudge” cells (Gumprecht shadows)
Warm Type:
CLL: note smudge cells
(Gumprecht shadows), and
spherocytes
Immune Hemolytic Anemia: Labs (Cold type)
Hb: reduced
Peripheral Blood Smear: schistocytes (fragmented red
cells), polychromatophils, nucleated red cells
S. bilirubin: may be increased
Hemopexin: reduced
Hemoglobinuria, Hemoglobinemia, Hemosiderinuria:
may be present
Cold type: schistocytes (fragmented rbc), & n-rbc
DAT: positive
Peripheral blood smear: note agglutination of red
cells. This is from a case of “cold type”
Microangiopathic hemolytic anemia
Introduction:
- are mechanical hemolytic anemias
- red cells are traumatized with contact with endothelium, fibrin, etc
- “intravascular” hemolysis
Causes:
1) Platelet thrombi
- hemolytic uremic syndrome (HUS)
- thrombotic thrombocytopenic purpura (TTP)
2) Fibrin thrombi
- disseminated intravascular coagulation (DIC)
- HELLP syndrome. (Hemolytic, Elevated transaminases, Low platelets, associated with preeclampsia)
3) Aortic stenosis
Labs:
- normocytic anemia
- reticulocytosis
Intravascular hemolysis
- decreased haptoglobin, and hemopexin
- hemoglobinuria, hemoglobinemia,
hemosiderinuria
- schistocytes in peripheral blood smear
Note: schistocytes, polychromatophils.

Investigations
Rh- incompatibility
Introduction:
- not common
Pathogenesis:
- mother is Rh (D antigen) negative, and the fetus is Rh positive
- this leads to immunization of the mother by the blood group antigens on the
fetal red blood cells, and
- the free passage of of antibodies from the mother through the placenta to the
fetus (as it is IgG)
- fetal red cells may reach the maternal circulation during the third trimester of
pregnancy
- when the cytotrophoblast is no longer present as a barrier, or during childbirth
itself (now mother becomes sensitized)
Rh- incompatibility
- initial exposure to Rh antigen leads to formation of IgM antibodies, that unlike
IgG do not cross the placenta
- therefore, Rh incompatibility is uncommon with first pregnancy
- exposure during a second pregnancy leads to a rapid IgG antibody response
Prevention:
- Rhesus immunoglobulin (RhIg) containing anti-D antibodies is administered at
28th week, and within 72 hours of delivery to Rh negative mother
- RhIg is also administered following abortions
Rh- incompatibility
Postnatal investigations:
if newborn has signs of hemolysis: Coomb Test
Rh incompatibility: positive
Clinical Features:
- Prematurity
- Neonatal anemia
- Neonatal jaundice (usually present at birth or within first 24 hrs). Remember:
kernicterus*
Rh- incompatibility
Prevention:
-to prevent maternal Rh-alloimmunization, Rh –ve mothers must be prevented from forming an immune
response against D antigen
-Anti-Rh (D) immunoglobulin is a polyclonal Ab product, composed of IgG anti- D antibodies collected from
pooled donor plasma
-it is routinely administered to Rh negative women at 28th week of gestation, and immediate post-partum
period
-once given, Anti-Rh (D) antibodies bind to Rh positive fetal red cells that enter maternal circulation
-therefore, preventing their interaction with maternal immune system via sequestration and elimination by
mother’s spleen
-Remember: administration of Anti D IgG antibodies during pregnancy does not cause significant
transplacental fetal hemolysis
-because the quantity of Rh (D) administered is very little compared to that produced in a typical
immunologic reaction
Rh- incompatibility
Clinical Features:
- hemolytic anemia
- unconjugated hyperbilirubinemia
- risk of developing kernicterus
- unconjugated bilirubin is normally bound to albumin
- liver function is impaired
- albumin synthesis may be reduced
- this increases the risk of kernicterus
Hemolytic disease of newborn:
ABO incompatibility
Etiopathogenesis:
- ABO hemolytic disease of the newborn (HDN) occurs almost exclusively in infants with
blood group A or B who are born to group O mothers (only those who have presence of
anti-A & anti-B IgG type antibodies)
- most anti-A and anti-B antibodies are of the IgM type,
- which is incapable of crossing the placenta (normal)
Remember:
- only a small proportion (1%) of group O individuals produce anti-A and anti-B of the IgG
antibodies type that may be transferred across the placenta
- hemolysis occurs by non-complement mediated phagocytosis of IgG coated red cells
“The only real voyage of discovery consists not in seeking new landscapes but in having new eyes”
Marcel Proust, novelist (1871-1922)
White Blood Cell
Disorders

Lecture Plan
General
Ø Bone marrow
Ø Peripheral Blood
Nomenclature
Disorders of WBC
Ø Non-Neoplastic
Ø Penias (low count)
Ø Philias or Cytosis (high counts)
Ø Lymph node (Lymphadenitis)
Ø Pre-malignant
Ø Myelo Dysplastic Syndromes (MDS)
Ø Myelo Proliferative Disorders (MPD)
Ø Malignant
Ø Lymphoid
Ø Myeloid
Lecture Plan
Disorders of
ØLymph Nodes
ØSpleen
ØThymus
Others
ØLangerhan cell Histiocytosis
PBLs

3
General

Bone Marrow

Normal Bone Marrow
***
Mast cells ?

7
Bone Marrow
Aspirate Trephine Biopsy

Bone marrow
• Marrow activity - ratio of hematopoietic

elements to fat cells (Cellularity)
ØNormal (Normocellular)- ratio is about 1:1
ØDecreased (Hypocellular)- - hypopalsia -
Aplastic anemia
ØIncreased (Hypercellular)- Hematopoiesis -
hemolytic Anemias, Leukemias

Bone marrow
Marrow Distribution
Ø 65% - Granulocytes and their precursors
Ø 25% - Erythroid precursors
Ø 10% - Lymphocytes and Monocytes & their
precursors
Myeloid : Erythroid ratio –3:1 (why?)
Ø Myeloid cells- Myelocytes, Metamyelocytes, &
Granulocytes
Ø Erythroid cells - Polychromatophilic &
Orthochromic normoblasts
Clinical significance of M:E ratio
Ø Increased: in Myeloid Leukemia
Ø Decreased: in Anemias, *** Polycythemia
Points need to know about
Bone Marrow
1. What is bone marrow ?
2. What are the types of bone
marrow?
3. Where is bone marrow located?
4. What is bone marrow ’s
composition?
5. How to obtain bone marrow?
6. Why is bone marrow testing done
(Indications for bone marrow
evaluation)?
Peripheral Blood

Peripheral Blood
Serum or plasma -55-60%
Buffy coat – 1%
Hematocrit ( PCV)
or
Red cell volume 39-44%

• Difference between serum and
plasma?
• What is Buffy coat made up of?
• Hematocrit or PCV ( packed Cell
Volume) ?
• How to know the functional status of
marrow by looking at peripheral
blood?
Peripheral Blood
Cells
Serum
RBC
Or
WBC
Plasma
Platelets

WBC
Myeloid Lymphoid
HSC
17
WBC- Normal
• Total Leukocyte count

• Differential Leukocyte
count
• Basic functions of
Leukocytes

WBC – CBC/ CBP

Lecture Plan
General
Ø Bone marrow
Ø Peripheral Blood
Nomenclature
Disorders of WBC
Ø Non-Neoplastic
Ø Pre-malignant
Ø Malignant
Ø Lymphoid
Ø Myeloid
Non – neoplastic Abnormalities
↓in # - Penias

WBC :Non – neoplastic Abnormalities
• Penias (Leukopenia)
– Neutropenia –Most Common (MC) and important
• low circulating neutrophil count
– <1500 cell/cu. mm – Neutropenia
– 1000 cells/ cu. mm – worrisome
– 500 cells/ cu. mm - serious infections
– 100 cells/ cu. mm - life threatening ( also called
Agranulocytosis)

• Penias (Neutropenia)
• Causes
– ↓ or ineffective production
» Act on Hemopoietic stem cell(HSC) àAplastic
anemia &infiltrative marrow disorders
» Act on committed granulocytic precursorsà
Drugs
» Disorders causing ineffective Hematopoiesis
àVit. deficiencies ( B12, Folate),
Myelodysplastic syndromes (MDS)
» Inherited disorder – kostmann syndrome
(defective genes of granulocyte differentiation)

• Penias (Neutropenia)
• Causes
– ↑ removal from circulation
» Immunological – Idiopathic, SLE, Drugs
» Hypersplenism
» ↑utilization – infections ( mainly Bacterial,
fungal, Rickttsial)
• Agranulocytosis
– MCC – Drug toxicity
• Can be dose related (used in cancer Rx.)
• Idiosyncratic (Cholamphenicol, Sulphonamides, Thiouracil etc.,)
– Idiopathic – Antibodies to Neutrophil specific antigen
– LGL (Large Granular Lymphocytic) Leukemia – Specific
CD8+ cells against Tumor cells cause suppression of
marrow granulocytic precursors
• Neutropenia
– Morphology = marrow changes depend on underlying
cause
• Hypercellular marrow if the cause is peripheral
destruction or ineffective hematopoiesis
• Hypocellular if it is due to Myelo- suppression
• Necrotizing , ulcerative lesions à oral cavity
(agranulocytic angina)
• High risk for à deep fungal infections (Candida,
Aspergillus)
• Sometimes there is exuberant growth of bacterial
colonies à Botryomycosis)

• Neutropenia
– Clinical course
• S &S (Signs & Symptoms) à related to infection
• Malaise, Chills, Fever
• Life threatening infections à die in hours to days (due
to Agranulocytosis)
– Treatment
• Broad spectrum antibiotics
• G-CSF à if Neutropenia is following immuno -
suppressive treatment.

• Lymphopenia - Less common
– Causes
• Congenital (Syndromes)
• Other causes
– HIV/ADIS
– Rx. Related à Corticosteroids. Cytotoxic drugs
– Auto – immune Disorders
– Malnutrition
– Severe Acute Viral infection
» Mechanism à ↑ type 1 interferons à T cell proliferation
à migration to Lymph nodes and Adhere to Endothelium
à Relative Lymphopenia

Lecture Plan
General
Ø Bone marrow
Ø Peripheral Blood
Nomenclature
Disorders of WBC
Ø Non-Neoplastic
Ø Pre-malignant
Ø Malignant
Ø Lymphoid
Ø Myeloid
Non – neoplastic Abnormalities
Philias or Cytosis (high counts)

Leukocytosis
• Leukocytosis:
• Increase in number of WBC
• Mostly due to infections/ Inflammatory conditions
• Pathogenesis
– WBC homeostasis
– Factors influencing homeostasis
• 1. Cell storage pool à bone marrow, Thymus,
circulation, peripheral tissues
• 2. Rate of release into circulation
• 3. Marginal pool à adherent to vascular endothelium
• 4. Rate of extravasation

Leukocytosis - causes
Neutrophilic Acute bacterial infections( pyogenic)
leukocytosis
Eosinophilic Allergic disorders such as asthma, hay fever, allergic
leukocytosis skin diseases parasitic infestations; drug
(Eosinophilia) reactions; certain malignancies (e.g., Hodgkin
disease and some non-Hodgkin lymphomas);
Basophilic leukocytosis Rare, indicative of CML
(Basophilia)
Monocytosis Chronic infections (TB, SLE) inflammatory bowel
diseases (ulcerative colitis)
Lymphocytosis Accompanies monocytosis in many disorders viral
infections (e.g., hepatitis A, cytomegalovirus,
Epstein-Barr virus); Bordetella pertussis infection

Leukocytosis
• Leukocytosis:
• Pathogenesis cont’d…
– WBC homeostasis
• Maintained by cytokines, Growth factors, Adhesion
molecules
• Maintained through à Commitment, proliferation,
differentiation, extravasation
• Mechanisms
– For Neutrophilia
• Immediate response à ***release from marrow
storage pool by IL-1, TNFα
• Prolonged à IL-1, TNFα act on macrophages, Marrow stromal
cells, T cells à G-CSF à Granulocytopoiesis
– For Eosinophilia - IL-5

Leukocytosis - Mechanisms

Leukocytosis - Neutrophilia

Leukocytosis
• Morphology
– In Severe infection/ Inflammations (Kawasaki’s) à
– Neutrophils
• Toxic granulations ( abnormal azuriphilic granules),
• Dohle bodies (Endoplasmic Reticulum) –like in disease)
– Lymphocytes à Reactive/Transformed lymphocytes
• Differential Diagnosis
– Leukemoid reaction (inflammation) from leukemia (CML)

Leukemia Vs. Leukemoid Reaction
Characteristic feature CML Leukemoid Reaction
Age >40 yrs Any age

Leukocytosis >100,000 30,000 – 50,000
Absolute Basophilia Present May not
Splenomegaly Prominent May not

Philadelphia Present Absent
Chromosome
LAP / NAP Very low / Absent High

Transformation to Yes No
Acute leukemia
Leukocytosis -Toxic Granulation:

Toxic Granulation

Normal Neutrophils 38
39
Leukocytosis -Lymphocytosis

Lecture Plan
General
Ø Bone marrow
Ø Peripheral Blood
Nomenclature
Disorders of WBC
Ø Non-Neoplastic
Ø Pre-malignant
Ø Malignant
Ø Lymphoid
Ø Myeloid
Lecture plan
Ø Lymph node disorders
ØNon – neoplastic or Inflammatory
ØAcute Nonspecific Lymphadenitis
ØChronic Nonspecific Lymphadenitis
ØHemophagocytic Lymphocyto Histiocytosis (HLH)

Diseases of
Lymph Node
Non malignant Malignant

(Lymphadenitis) (Lymphomas)
Acute Chronic
HLH Hodgkin’s NHL
(infections) (immunologic)

Acute Nonspecific Lymphadenitis
Ø Causes
Ø Microbes, Cell debris, Foreign material
Ø Sites
Ø Generalized – (esp. children) Viral, Bacteremia
Ø Localized –
ØCervical – infections in oral cavity
ØAxillary/ inguinal – infections in limbs
ØMesenteric – acute appendicitis
Ø Morphology
Ø Large lymphoid follicles with prominent germinal centers
Ø Tingible body macrophages
Ø Clinically
Ø Enlarged & Tender lymph nodes
Ø Clinical Course
Ø Heal with scar Dr.T.Krishna MD, www.mletips.com 44
Acute lymphadenitis
• Causes
– Microbes, Cell debris, Foreign material
• Sites
– Generalized – (esp. children) Viral, Bacteremia
– Localized –
• Cervical – infections in oral cavity
• axillary/ inguinal – infections in limbs
• Mesenteric – acute appendicitis
• Morphology
– Large lymphoid follicles with prominent germinal centers
– Tingible body macrophages
• Clinically
– Enlarged & Tender lymph nodes
• Clinical Course
– Heal with scar Dr.T.Krishna MD, www.mletips.com 45
Tingible Body Macrophages

Reactive Lymph node

Lymphoma

Reactive LN Vs. Lymphoma
Size of the follicles

Zonation
Dark and light areas
Capsular invasion

Chronic Nonspecific Lymphadenitis
ØMorphological types
ØFollicular hyperplasia
ØHumoral response – esp. RA, Toxoplasma, HIV
ØGerminal centers with secondary follicles
ØTingible body macrophages
ØMantle zone- surrounding the sec. follicles
ØGives dark and light zones
ØDifferential Diagnosis – Follicular Lymphoma
ØMarginal zone hyperplasia
ØSubtype, common in Toxoplasma, HIV
ØIt is outside the Mantle zone
ØMonocytoid B cells (memory cells)
Chronic
Chronic lymphadenitis
Nonspecific Lymphadenitis
– Para-cortical Hyperplasia
• Cellular response
• Dilantin Rx, Viral (IM), Viral vaccines
• Large T immunoblasts in Para cortical areas
– Sinus Histiocytosis
• Prominent lymphatic sinusoids
• Due to immune response to tumors or their products

ChronicLymphocyto
Hemophagocytic lymphadenitis
Histiocytosis
ØAKA
ØMacrophage Activation syndrome
ØReactive conditionà Macrophage + CD8+
CTL’s
ØTypes
Ø1. Familial – young age(infants)
Ø2. Sporadic à Any age

ChronicLymphocyto
Hemophagocytic lymphadenitis
Histiocytosis
Ø Pathogenesis
ØIn both forms à systemic activation of Macrophage & CTL’s
ØMacrophages à phagocytose both progenitor cells in marrow
and formed elements in the peripheryà Cytopenias
ØMacrophages & CTLs’
Ø Release large amounts of inflammatory mediators
Ø Interferon γ, TNFα, IL-6, IL-12, soluble IL-2 receptor (IL-2R)
Ø Effects
Ø Suppress Hematopoiesis à severe cytopenias
Ø Cytokine storm à SIRS
Ø Shock like picture
ØFamilial (also in sporadic) à mutations affect à formation/
deployment of toxic granules in CTL, NK cells
ØMC trigger à infection (EBV)

Hemophagocytic Lymphocyto Histiocytosis
Ø Clinical course (MC common presentation)
ØAcute febrile illness, Splenomegaly ,
Hepatomegaly
ØBone marrow à Hemophagocytosis (neither
essential nor sufficient in diagnosis)
ØLab data
ØAnemia ,Thrombocytopenia, ↑↑↑ plasma Ferritn
and IL-2R
ØAbnormal LFT and TGL à Hepatitis
ØAbnormal coagulation profile à DIC (?)

Ø Prognosis
ØIf untreated à die of shock /MOF in 2 months
(familial)
ØIn treated à develop
Ørenal failure (adults) ,
ØGrowth retardation , Mental retardation
(children)
Ø Treatment
ØHSC transplantation (familial) à 50% survive
ØImmunosuppressive Rx., mild chemotherapy


57
Thank you

Objectives
Ø Define – Neoplasm, Malignancy, neoplastic Hematologic
disorders
Ø Compare and contrast
Ø Myelodysplastic (MDS) vs. Myeloproliferative
disorders(MPD)
Ø Acute (AL) vs. Chronic Leukemias (CL)
Ø Names and classification of
Ø MDS
Ø MPD
Ø Leukemias
Ø Methods to Categorize Leukemias
Ø Lab findings
Lecture Plan
Ø White cell disorders
Ø Non – neoplastic Abnormalities
Ø ↓in # - Penias & ↑ - Philias
Ø Neoplastic disorders
Ø Introduction
Ø Etio – pathogenesis
Ø Classification
Ø Myeloid neoplasms
Ø Myelodysplastic Syndromes
Ø Histiocytoses
Ø Introduction to Leukemias
Ø Lymphoid neoplasms
White cell disorders
Ø Neoplastic disorders - Introduction
§ Morphology of tumor cell à
Ø Resembles one of the stages of developments of
normal counterparts
Ø Helps in diagnosis & Classicification
Neoplastic White cell disorders - Introduction cont’d…
Ø 1. Lymphoid neoplasms
Ø B – Cell
Ø T – Cell
Ø NK Cell
Ø 2. Myeloid neoplasm
Ø Acute Myeloid Leukemias – AML (immature/blasts)
Ø Myelodysplastic syndromes – MDS (dysplastic/disorganized)
§ Morphology
§ Marrow à ineffective Hematopoiesis
§ Peripheral blood à corresponding cytopenias
Ø Myeloproliferative disorders MPDs’ (proliferation of mature cells)

§ Morphology
§ Marrow à Hypercellular
§ Peripheral blood à Philias/Cytosis
Ø 3. Histiocytoses – group of disorders

§ Cell of origin à Histiocyte (tissue cell); immature dendritic cell
Lecture Plan
Ø Introduction
Ø Classification
Ø Histiocytoses
Ø Neoplastic disorders : Etio – pathogenesis
Ø I). Chromosomal
Ø A. Translocations àMC abnormality
Ø i. Genetic abnormalities/mutations lead to à
Ø Dominant negative (Loss of Function)
Ø Gain of Function
Ø B. Genomic aberrations àProduce oncoproteins
Ø Activate growth signalling pathways
Ø Inhibit apoptotic cell death
Ø Mechanisms
Ø ↑proliferation but arrest in differentiation (Eg: APML)à loss
of function mutations
Ø ↑self renewal of tumor cells (act like stem cells)à ↑Tyrosine
kinase activity à signal RASà *activation of PI3K/AkT &
MAPK à Warburg metabolism
Ø Neoplastic disorders : Etio – pathogenesis cont’d..
Ø C. Genetic aberrationsà Antigen receptor gene
rearrangements & diversification àActivation of
Oncogenes
Ø Ag àactivate B cellsà enter germinal cantersà ↑
expression of “AID” enzyme (Activation Inducted
cytosine Deaminase) – DNA modifying enzyme leads
to
Ø Ig gene modifications à cell switching in two
ways
Ø1. Intragenic recombinations
Ø2. Replacement of “C” gene segment of IgH
with different “C” gene segmentà Antibody
class switch (IgG1, IgG2 etc.)
Ø Examples cont’d…
Neoplastic White cell disorders Etio – pathogenesis cont’d..
Ø 3. Genetic aberrations cont’d…
Ø Examples
Ø Somatic Hypermutation à point mutations of Ig
genesà ↑ Ab affinity to Ag (Eg: Translocation of Ig
locus à MYC activation in germinal canter à B cell
Lymphomas)
Ø AID cause lesions in DNA à chromosomal breaks à
MYC/Ig translocations
Ø AID misdirected Bcl-6 gene breaks (transcription
factor) à activation of Bcl-6 à B-cell Lymphomas
Ø Aberrant recombinations of V(D)J recombinase
(normally cuts DNA at specific of Ig and T –cell
receptor loci and assembly of Ag receptor genes) à
abnormal joining of other genes to Ag receptorsà lead
to Precursor T- cell tumors
Ø II). Inherited Genetic Factors
Ø Genetic disease lead to Genomic instability
Ø Examples – Bloom’s Fanconi’s anemia, Ataxia
telengectasia
Ø Genetic disorders associated with ↑susceptability to
childhood Leukemias à Down’s, NF1
Ø III). Oncogenic Viruses
Ø HTLV-1 à Adult T Cell Leukemia/Lymphoma
Ø EBV à Burkitt’s, Hodgkin’s, NK cell Lymphomas
Ø KSHV/HHV8 à B cell lymphoma with malignant pleural
effusion
Ø HIV patientsà B cell lymphoma or any organ
Ø IV). Chronic Inflammation
Ø H. pylori à Gastric B cell lymphoma
Ø Gluten sensitive enteropathy à T cell Lymphoma
Ø Breast Implants à T cell lymphoma
Ø HIV patientsà B cell lymphoma or any organ
Ø T cell dysregulation leads toà systemic B cell
hyperplasia à Germinal canter B cell Lymphomas
(particularly EBV and KSHV/HHV8)
Ø V). Iatrogenic
Radiotherapy , Chemotherapy lead to à Mutagenic effects
Ø
on hematopoietic and lymphoid tissues à Myeloid and
Lymphoid tumors
Ø VI). Smoking (carcinogen – Benzene compounds)
Ø Risk of AML à ↑1.3 to 2 X
Lecture Plan
Ø Introduction
Ø Classification
Ø Histiocytoses
Naming & Classification
FAB & WHO
FAB & WHO
Lecture Plan
Ø Introduction
Ø Classification
Ø Histiocytoses
MDS
MDS
Ø Group of Clonal stem cell disorders
Ø Characterized by
Ø Maturation defect à Ineffective Hematopoiesis
Abnormal
Ø Multipotent stem cellsà differentiate
Ineffective
Ø Result à Peripheral blood cytopenias
Ø ↑risk of Acute Myeloid Leukemias (AML)
Ø Types
Ø Primary or Idiopathic = > 50yrs, Gradual in onset, risk of
AML ↑
Ø Rx ( RT or Drugs) related (t MDS) = after 2 -8 of RX,
complication of Rx, Higher risk of AML (↑ ↑ ↑)
MDS
Ø Pathogenesis
Ø Not clear
Ø Proposed MDS genomes – Three groups
Ø 1. Epigenetic factors à similar mutations of genes in
AML (DNA methylations, Histone modifications)
Ø 2. RNA splicing factors à Mutations of 3’end
Ø 3. Transcription factors (TF) à mutations of TFs of
normal myelopoiesis
Ø Cytogenetic abnormalities
Ø Chromosomal abnormalities
Ø Deletions (5q,7q,20q)
Ø Monosomy (5 & 7)
Ø Trisomy (8)
Ø Others à 10% of MDS
Ø Affect both types of MDS
Ø Loss of function mutations of TSG (p53)
MDS
Cytogenetic abnormalities cont’d…
Ø MYC oncogene mutations
Ø slight excess à Very common in neoplasms
Ø MYC oncoprotein (Chromosome – 8) stimulate cell

proliferation
Ø 5q deletion
Ø Loss of gene coding ribosomal protein à RPS14 à
ineffective erythropoiesis
MDS
Ø Morphology
Ø Marrow = usually Hypercellular, with abnormal
differentiation of all three cell lines
Ø Erythroid precursors
Ø ring sideroblasts (perinuclear iron laden mitochondria)
Ø megaloblastoid maturation
Ø budding nucleated cells
Ø Granulocytic precursors
Ø Neutrophils – Hypogranular (IIº), Toxic granules, Dohle
bodies
Ø Pseudo Pegler – Huet neutrophils (two nuclear segments) or
un-segmented nuclei
Ø Megakaryocytes- uni or multilobated (Pawn ball like)
Ø Myeloblasts <20% of marrow cellularity
Ø Peripheral Blood = Cytopenias ( Pancytopenia)
Ø Patients present with Refractory Anemia’s (not responding to
hematenics even after 6 months of Rx )
MDS
Ø Morphology
Ø Peripheral Blood
Ø Cytopenias ( Pancytopenia)
Ø Pseudo Pelger –Huet neutrophils
Ø Giant platelets
Ø Macrocytes
Ø Monocyotsis
Ø Myeloblasts (<10%)
Ø Clinical Features
Ø Common in - elderly (mean age -70 years)
Ø 50% - Asymptomatic
Ø In symptomatic – present with complications of
cytopenias (what are they?)
MDS
Ø Prognosis
Ø Primary MDS à 8 groups (WHO)
Ø Worst prognosis with à high blast count & multiple
chromosomal aberrations
Ø Median survival
Ø Best prognostic group à up to 5 years
Ø Worst group à 9 - 24 months
Ø Progression to AML à 10 -40% (associated with
additional mutations)
Ø tMDS(Secondary)
Ø Median survival à 4 - 8 months
Ø Progression to AML à ↑↑↑
Ø Severe cytopenias (commonly lead to death)
MDS
Ø Treatment
Ø Limited in elderly (symptomatic à Antibiotics and blood
transfusions)
Ø In young people à stem cell transplantation offers cure
Ø Some patients with isolated mutations (5q- )
Immunomodulator à Thalidomide
Ø DNA methylation inhibitors
budding nucleated ring sideroblasts

MDS (Pawn ball Megakaryocytes)
MDS (Pawn ball Megakaryocytes)
MDS
budding nucleated
Ring sideroblasts
MDS
Pseudo Pegler – Huet neutrophils
(two nuclear segments)
MDS
Pseudo Pegler – Huet neutrophils
(two nuclear segments)
Normal
Lecture Plan
Ø Introduction
Ø Classification
Ø Histiocytoses
Leukemias
Leukemia
Myeloid Lymphoid
Acute Chronic Acute Chronic

(AML- 8 types) (CML) (ALL- 3 types) (CLL)
Leukemias
Feature Acute Chronic
Rate of Growth Rapid Slow
Cells Immature Mature but
(Blasts) abnormal
Symptoms Yes (Bleeds, Usually
Anemia & Asymptomatic
Infections)
P/S Blasts (+ ) Blasts ( - )
BM Blasts ***≥20% <20%
Acute Myeloid Leukemias
Acute Lymphoid Leukemias
Blasts
Features Myeloblasts Lymphoblasts
Cytoplasm Mod/abundant Scant/Mod
Cytoplasmic Common Uncommon
Granules
Nucleus fine Coarse

(chromatin)
Nucleoli Prominent Variable

Auer rods Present Absent
Lecture Plan
Ø Introduction
Ø Classification
Ø Histiocytoses
MPD
MPDs’
n Normal pathway
n Growth factor/ligand binds with its receptor on
the cell
n Activation of Tyrosine Kinase (TK) by
dimerization and autophosphorylation
n Activated TK activate signal pathways (RAS,
MAP etc.,) by phosphorylation
n Finally act on hemopietic progenitor cells à
lead to proliferation and survival of cells
MPDs’
n Mutations of TK /Signal pathways
n Hemopoietic progenitor cells become growth
factor independent à lead to uncontrolled
proliferation of mature or differentiated cells.
n Proliferation of mainly granulocytic and
megakaryocytic elements (and lymphoid cells if
pleuripotent cells are affected as in case of
CML)
n Common features of MPDs
n Stages
¨ Proliferativephase à marrow –Hypercellular,
Peripheral blood à Philias/cytosis
¨ Phase of Extramedullary hematopoiesis à homing
of hemopoietic progenitor cells in secondary
hemopietic sites
¨ Spent /burnt-out phase à Marrow fibrosis with
cytopenias in peripheral blood
¨ Transformation phase à into Acute Leukemias
n Unique features of certain MPDs

¨ SpecificTK mutations (refer next slide) à help in
diagnosis and targeted therapy
¨ In Systemic Mastosis à KIT TK mutations
Tyrosine Kinase mutation in MPDs’
n Chronic Myelogenous/Myeloid Leukemia
(CML)
n Molecular Mechanisms
¨ In>90% of cases( MC) à reciprocal translocation of
genes in chromosomes 9:22;
n Forms Philadelphia chromosome à Chr. 22
n BCR (Ch-22)+ ABL (Ch-9) à fusion/chimeric gene;
Demonstrated by cytogenetics/karyotyping;
n BCR-ABL fusion gene (210 KDa) has TK activity
¨< 5%: fusion gene (BCR-ABL) formed by complex/
unknown mechanisms; diagnosed by à FISH, PCR
n Tumor Cell à PLEURIPOTENT STEM CELL
CML cont’d……(Chimeric gene – BCR-ABL)
CML cont’d…(Chimeric gene – BCR-ABL)
CML cont’d… (Karyotyping)
CML cont’d…
n Morphology cont’d…
¨ Bone marrow findings:
n Erythroid seriesà normal or ↓ in number
n Most important à Macrophages with green-blue

cytoplasm “Sea Blue Histiocytes”
n ↑ Reticulin (not to the extent of Myelofibrosis in
early stages)
CML cont’d…
n Peripheral blood findings
¨ Marked leukocytosis (TWBC:100,000 cells/cu.mm)
¨ ↑ number of Mature neutrophils and its precursors
(up to Myelocytes)
¨ ↑ number of Eosinophils and Basophils (Absolute
Basophilia – useful finding)
¨ Myeloblasts < 10%
¨ ↑ number of Platelets
CML cont’d…(Sea Blue Histiocytes)
CML cont’d…(Peripheral Blood- Leukocytosis)
CML cont’d…
n Organs
¨ Spleen à mild in early stages (due congestion)
n Marked splenomegaly (2.5-3.0 Kg, what is the
normal weight of spleen?); EMH; sub capsular
infarctsà medical emergency
¨ Liver à Hepatomegaly
¨ LNà also enlarged
¨ Normal weight of spleen – 150 grams

CML cont’d…(Splenomegaly)
(External surface)
(Cut section)
CML cont’d …(Splenomegaly)
No infarct Infarcts
CML cont’d…
n Clinical features
¨ Age: common in adults > 40 years (Juvenile CML
can be seen in teens)
¨ Onset: insidious
¨ Anemia: mild to moderate (present with easy
fatigability, weakness, weight loss etc.,)
¨ Dragging sensation of left side of abdomen à
marked splenomegaly
¨ Acute abdomen à due to Splenic infarcts
CML cont’d…
n Lab findings
¨ Karyotyping /PCR demonstrate BCR-ABL
translocations in >95%cases
n Course of disease
¨ Median survival 3 years
¨ 50% develop accelerated phase with peripheral
blood findings of worsening anemia, ↓Platelets, ↑
Basophilia
n Molecular- Trisomy 8, isochromosome 17, double
Philadelphia)
n In next 6-12 months progress to blast crisis {AML-
70%, ALL (Pre-B)-30%)}
¨ Other 50% à directly develop blast crisis
CML cont’d…
n IKAROS
¨ Transcription factor Which regulates differentiation
of hemopoietic progenitors
¨ Mutations of IKAROS noted in lymphoid blast crisis
(Acute Lymphoid Leukemia) developed in pts with
CML
¨ Similar mutations are seen in all ALL cases who
develop disease denovo
CML cont’d…
n How CML is different from other MPDs’
¨ In CML mutations lead to activation of TK {(Tyrosine
Kinase) top in the signal pathway)} which lead to
different, and stronger signals
¨ In other MPDs’ (PV, ET, and Myelofibrosis)à
mutations in JAK signals are different and relatively
weaker
CML cont’d…
n Treatment
n 1. BCR-ABL inhibitors (1st Generation)
¨ Remission is seen in >90% of cases; ↓BCR-ABL “+”
cells which leads to ↓risk of Accelerated CML and
Blast crisis: Limitations;- 1. Can’t eliminate CML
tumor /stem cell, 2) Drug resistance leads to Rx.
failure
¨ Drug resistance is an issue
n Mechanism of drug resistance
¨ 50% (half) of cases – mutations in BCR-ABL
¨ Remaining half – mutations in other kinases
¨ 2nd and 3rd generation BCR-ABL inhibitors overcome

these issues.
CML cont’d…
n 2. HSC transplantation
¨ Indications – young patients and early stage of
disease (offers cure in 75% of cases)
¨ Not effective in - Accelerated phase, blast crisis
and elderly patients
Myeloproliferative Disorders (MPDs)
Polycythemia Vera (PV)
n Introduction
¨ Tumor cell – Myeloid progenitor cell (in CML what
is the tumor cell?)
¨ Molecular mechanism – Point mutations in TK,
JAKà activating or gain of function mutations
¨ Mutations result in proliferation of all the three cell
lines but clinically significant is – Erythroid cell line
¨ Types – Primary PV needs to differentiated from
reactive/secondary PV by testing blood levels of
Erythropoietin (EPO)
Tumor cell in CML – Pleuripotent stem cell

PV cont’d…
n In Primary PV
¨ Blood EPO levels are low
n Clinical features are due to
¨ 1. High Hct à hyper viscosity
¨ 2. Abnormal PLTS function à Bleeding/Thrombi
PV cont’d…
n Pathogenesis (Molecular Mechanisms)
¨ In97% of cases à Point mutations in JAK2 at 617
(phylalanine in place of Valine)
¨ 3% have other mutations
n Experimental evidence à Murine models with

JAK2 mutations show PV like features
PV cont’d…
n 30% of cases (Special sub type)
¨ Two mutated copies of JAK2 (? Dose related)
¨ ↑↑ WBC population
¨ Marked splenomegaly
¨ Pruritus
¨ Rapid progression to spent/fibrotic phase
PV cont’d…
n Morphology
¨ Marrow à Hyper cellular;
n ↑number of all three lineages;
n 10% of cases show ↑concentration of reticulin

fibres;
n late stage produce Myelofibrosis and marrow will
be Hypocellular
¨ Peripheral blood à ↑ three cell line but ↑ in RBC
mass clinically important),
¨ Organomegalyà mild in early stages (due to
congestion) – later moderate ( due to EMH – Extra
Medullary Hematopoiesis)
PV cont’d…
n Clinical features
¨ Related to ↑RBC mass (clinically important),
n ↑ Hct, ↑ blood volume
n venous (low pressure venous side of circulation)

à distended vessels, Plethora, cyanosis
n Headaches, dizziness, HTN, GI symptoms
¨ Histamine from Basophils cause intense Pruritus ,

peptic ulcers
¨ Hyperurecemia with secondary gout is seen in 7-
10% cases (from ↑cell turnover)
PV cont’d…
n Clinical features cont’d…
¨ Bleeding/clotting episodes (25% of cases)
n Due to abnormal blood flow & Platelet dysfunction
lead to DVT, MI, Strokes, Budd-Chairi syndrome
(Hepatic Vein Thrombus, Abdominal pain, Ascitis)
n Bowel infarctions from portal and mesenteric
Vein Thrombi
n Early findings èEpistaxis and gum bleeds,
n Massive bleeds are seen in 5-10% cases

PV cont’d…
n Lab
¨ Hgb% - 14-28 gm/dL,
¨ Hct - >60%;
n Hct may be low following chronic bleeding (auto-
correction of disease)
n WBC – 12k to 50 k (normal: 4k to 11k)
n Platelets – 500k (normal: 150k to 450 k) with giant

Platelets (which are functionally abnormal)
PV cont’d…
n Clinical course
¨ If
Untreated èdie in months from bleeds, clots
¨ Only phlebotomy improves life by 10 years
¨ JAK2 inhibitors are in clinical trails
n Prognosis
¨ Spent/fibrotic phase develops despite Rx. in 15 –
20% of cases by 10 years
¨ AML transformation à in 2% of cases
n Tumor cells lack JAKs mutations
PV- summary of clinical course
RBC - episodic symptoms
Bleeding
(25% of cases)
thrombosis
Acute leukemia
( 2%)
Myeloproliferative Disorders (MPDs’)
Essential Thrombocytosis or Essential
Thrombocythemia (ET)
n Introduction
¨ Uncommon: 1-3/100,000 per year.
n Etio-pathogenesis
¨ 50% of cases have JAK 2 point mutations
(activating – gain of function)
¨ 5-10% of cases show MPL mutations (MPL –
receptor tyrosine Kinase activated by
Thrombopietin)
¨ Rest of them (around 40-45%) have Calreticulin
mutations (Calreticulin à protein of cytoplasm, ER
and has several functions)
¨ Important à JAK2 & Calreticulin mutations are
mutually exclusive
ET cont’d…
n Clinical
¨ ↑↑ Platelets (functionally abnormal)à bleeding
/Thrombotic ( like in PV) events
¨ Polycythemia and Myelofibrosis à Absent
¨ Need to rule out conditions that cause reactive
Thrombocytosis (CML, IDA)
¨ JAK2 mutations in PV and ET similar but show
different phenotypic picture
ET cont’d… (Molecular Mechanism)
JAK, MPL (Normal) JAK, MPL (mutated)

↓ ↓
Marrow MK Progenitors Marrow MK Progenitors
(Thrombopietin dependent) (Thrombopietin independent)
↓ ↓
Normal Platelet synthesis Abnormal ↑ Platelet synthesis
ET cont’d…
n Morphology
¨ Bone marrow shows Mild hyper cellularity
n ↑number of Megakaryocytes with
abnormal morphology
¨ Peripheral blood
n ↑ number of functionally abnormal
Platelets, Giant Platelets,
n Mild leukocytosis
¨ Organomegaly (due to EMH) seen in 50%

of cases
ET cont’d…
n Clinical course
¨ Age >60s’ (sometimes in young)
¨ Present with bleeding /clotting (more common)
complications (like PV)
¨ Spent phase and AML transformation à
uncommon
¨ Erythormelalgia – throbbing and burning hands and
feet
n Due to occlusion of small arterioles
n Also seen in PV
ET cont’d…
n Clinical course cont’d…
¨ Indolentcourse like PV with long term remissions
and episodes of clotting /bleeding complications
¨ Median survival:10 -15 years
¨ Severe Thrombotic events are seen in à Pts’ with
very high PLTS counts & Homozygous JAK2
mutations
n Rx. à Gentle Chemotherapy

Myeloproliferative Disorders (MPDs’)
Myelofibrosis (MF)
n Introduction
¨ Themost characteristic feature is obliterative bone
marrow fibrosis
n Molecular mechanisms
¨ JAK 2 mutations: 50-60% - Most Common
¨ MPL mutations: 1-5%
¨ Calreticulin mutations: remaining (40-45%)
MF cont’d… (pathogenic mechanism)
Neoplastic Megakaryocytes
↓
Fibrogenic growth factors
↓
(TGFβ, PDGF)
↓
Fibroblasts in the marrow (not neoplastic)
↓
↑ Deposits collagen
↓
Myelofibrosis
MF cont’d…
n Morphology
¨ Bone marrow
¨ Early phase à Hypercellular marrow with abnormal
Megakaryocytes;
n Megakaryocytes are large, dysplastic, clustered
n Other cell lines are structurally normal
¨ Progressive phase à Hypocellular marrow with increased

fibrosis
n Megakaryocytes show irregular nuclear pattern (cloud –
like)
¨ Late phase à marrow becomes bony (osteosclerosis) – like
spent phase of other MPDs’
n Homing of hemopoietic stem cells to EMH sites (Spleen,
Liver, and LNs’)
n RBC produced in EMH site are not normal à Anemia
MF cont’d…
n Peripheral blood
¨ Leukoerythroblastosis,
¨ Tear drop RBC (dacryocytes: damage during birth)
¨ Basophilia, (these features are also seen in other disorders)
¨ Large/giant Platelets
n Spleen
¨ markedly enlarged (4 kilograms in weight)
¨ EMH starts in sinusoids and later extends to cords,
¨ Cut section à looks like that of CML.
¨ Sub-capsular infarcts can lead to medical emergency
n Liverà enlarged ( due to EMH)

n LNs’ à show EMH, but not very significant
MF cont’d…
n Clinical features
¨ Age - >60 yrs.
¨ MF is less common than PV, ET
¨ Present with anemia and splenomegaly
¨ Other features à loss of weight, night sweats (due
to ↑metabolism by EMH)
¨ Hyperurecemia (due to ↑ cell turnover)lead to
secondary Gout
MF cont’d…
n Lab
¨ Moderate to Severe anemia
¨ Leukoerythroblastosis
¨ Leukocytosis in early stage, later Leukopenia or
normal counts
¨ PLTSà normal counts initially, laterà
Thrombocytopenia
Best and must investigation for diagnosis
↓
Bone marrow evaluation (both Bone marrow
Aspiration & biopsy)
MF cont’d…
n Rx.
¨ Primary Myelofibrosis: difficult to Rx., Median
survival is 3-5 yers
¨ HSC transplantation à in young and fit patients
¨ JAK2 inhibitors improve clinical condition
n Complications
¨ Infections
¨ Thrombotic/bleedingepisodes
¨ Transformation to AML -5-20% (highest of all
three MPDs’),
¨ AML can develop in EMH sites
Myelofibrosis
Myeloproliferative Disorders
Summary
n Disorders of multipotent progenitor cells ( myeloid &
Lymphoid precursor)
n Increased, Functionally abnormal cells.
n Extramedullary hemopoiesis - Organomegaly
n End stage
¨ Progress to Leukemia
¨ Myelofibrosis /spent phase
n Classification:
¨ Chronic Myeloid Leukemia (CML)
¨ Polycythemia rubra vera (PV)
¨ Essential Thrombocythemia (ET)
¨ Myelofibrosis (MF)
Organomegaly
Lecture Plan
Ø Introduction
Ø Classification
Ø Histiocytoses
Langerhans Cell Histiocytosis
n Group of disorders
n Characteristic feature - ***increased dendritic
cells/macrophages
n Spectrum of diseases range from rare
malignant Histiocytic lymphomas to benign
reactive Histiocytosis
Benign Intermediate Malignant
Reactive sinus Langerhan cell Histiocytic

Histiocytosis Histiocytosis Lymphoma
n Molecular mechanisms(pathogenesis)
¨ Oncogenic mutations affecting BRAF( in 600,
Valine-Glutamic acid) in 50-60% of cases
n Tumor Addiction ( refer chapter Neoplasia)
¨ BRAF- Serine/Threonine Kinase, immediately acts

in downstream of RAS
¨ Similar mutations are also seen in Hairy cell
leukemia
¨ TP53,RAS, TK MET-uncommon mutations
n MORPHOLOGY
n Langerhans cell
¨ 1) Abundant, vacuolated cytoplasm
¨ 2) Vesicular nuclei, nuclear grooving/nuclear folds
¨ 3)Birbeck granules; cytoplasmic tennis racket- like
and seen in electron microscopy, pentalaminar
tubules contain protein Langerin
4) Tumor cells express HLADR,S-100 and CD-1a
n CLINICOPATHOLOGICAL FORMS
n 1. Multifocal multisystem Langerhans cell
Histiocytosis also called Letterer-Siwe disease
¨ Age: less than 2 years
¨ Present with seborrhic cutaneous lesions (due to
LC infiltration) present on trunk(front and back) and
scalp
¨ Hepatosplenomegaly, lymphadenopathy, pulmonary
lesions
¨ Destructive osteolytic lesions(in the late stage)
n Letterer-Siwe disease Cont’d…
¨ Marrow - infiltrated with Langerhan Cells
¨ Peripheral blood - Anemia, thrombocytopenia, also
infections mainly otits media, mastoiditis
¨ Transform to Langerhan Cell sarcoma with
anaplastic tumor cells
¨ Clinical course-
n fatal if untreated
n Treatment-Intensive chemotherapy 5-year

survival-50%
n 2. Unifocal and Multifocal-
Unisystem(Eosinophilic Granuloma)
¨ 1)Langerhans cell with polymorphous populations of
Eosinophils(most prominent), lymphocytes, plasma
cells, neutrophils
¨ 2)location/site-Medullary cavities of calvarium, ribs,
femur
n Unisystem - any of the following(skin, lungs, stomach)
n Unifocal – affect Bones(older children and adults)
¨ asymptomatic or with pathological fractures
¨ insidious, spontaneous healing or cure by local excision

or irradiation)
n 3)Multifocal Unisystem
¨ Affect young children
¨ Multiple bone destructing masses expand into soft
tissues
¨ Effect pituitary stock of hypothalamus in 50% of
patients, present with diabetes insipidus
¨ Hand-Schiller-Christian disease Present with
diabetes Insipidus, bone defects(calvarial),
exophthalmos
¨ Prognosis- spontaneous remission or mild
chemotherapy
n 4)Pulmonary LCH - special type
¨ adult
smokers (regress after stop smoking)
¨ 60% are reactive proliferations
¨ 40% present with BRAF mutations
n How Langerhan Cells of normal cutaneous
distribution home into storage sites?
¨ Normal epidermal Langerhan cells express CCR-
6(chemokine receptors)
¨ Neoplastic Langerhan cells express CCR-6 and
CCR-7: Migrate to tissues which express these
chemokines
Ligand Receptor Tissue
CCL-20 CCR-6 Skin and bone
CCL-19,CCL-21 CCR-7 Lymphoid tissues

Vesicular nuclei
Abundant, Vacuolated cytoplasm

Birbeck
granules
Lecture Plan
ØNon – neoplastic Abnormalities
Ø↓in # - Penias & ↑ - Philias
ØPre-Neoplastic Conditions
ØMyelodysplastic Syndromes
ØMyelo Proliferative Disorders (MPD)
ØHistiocytoses
ØNeoplastic disorders
ØIntroduction
ØEtio – pathogenesis
ØClassification
ØMyeloid neoplasms
ØLymphoid neoplasms
Myeloid Neoplasms:
• Pathogenesis:
– Most common – t(8;21) disruption RUNX-1, inv(16)
disrupts CBF-B gene
– Deep DNA sequencing shows epigenetic alterations
like DNA methylation, post translational modifications
of Histone proteins, which are central and critical.
• Morphology
– Myeloblasts description
– Aleukemic Leukemia à, absence of blasts in the
peripheral blood
• Summary – Table (ref. Next slide)
Blasts
Cytoplasm Moderate/Abundant Scant/Moderate
Granules
Nucleus Fine, Lacy Coarse, condensed

(chromatin)

(inconspicuous)
Auer rods (MPO) Present Absent
Acute Myeloid Leukemias
AML - summary
1. Mo- undifferentiated, Ultra structurally Myeloblasts

2. M1- No maturation, =/>3%Blasts are MPO +ve
3. M2- MC(30-40%), Auer rods, t(8:21)
4. M3- APML, many Auer rods/cell, Younger age ( 35 yrs),
DIC, t(15:17), mutations of Retinoic receptor gene
5. M4- Myelomonocytic, NSE +ve, inv(16)
6. M5- Monoblastic (MPO -ve, NSE +ve), older pts,
Organomegaly, Lymphadenopathy, Tissue infiltration (
skin- Chloromas),
7. M6- Erythroleukemia , older age, 20% of Rx related
AMLs
8. M7- Megakaryocytic Leukemia- Myelofibrosis
Myeloid Neoplasms cont’d...
• Clinical Features:
– Pancytopenia
– APML or AML M3 presents with DIC
– Infections with fungi, pseudomonas, other
commensals (due to immune deficiency)
– Localised soft masses are also called
myeloblastoma, granulocytic sarcoma or
chloroma (inM5)
• Prognosis:
– Difficult to treat, except AML M3 or APML with
t(15;17) - ATRA (all trans retinoic acid)
Myeloblasts
Myeloblasts
Myeloblasts – Myeloperoxidase (MPS) positive
AML-M5 - Gum Hypertrophy
AML 5 – Non-specific Esterase
AML 5 Control
Lecture Plan
ØHistiocytoses
ØIntroduction
ØClassification
ØMyeloid neoplasms
Lymphoid Neoplasms
Definitions
1) Leukemia - Neoplasms involves bone marrow and
peripheral blood
2) Lymphoma involves-Tissue masses (lymph node /nodal
and or extra-nodal)
3) Leukemia/Lymphoma describes tissue distribution at the
time of diagnosis
4) Lymphomas-Hodgkin’s Vs NHL (Hodgkin's has special
pathological features and totally different from Non
Hodgkin's Lymphomas- NHL)
5) Plasma cell neoplasms mainly arise from bone marrow
6)Incidence-100,000 new cases per year in USA
Hodgkin’s Vs. NHL
HD vs. NHL
Feature Hodgkin’s NHL
Nature Similar to Similar to
infection malignancy
Spread Contiguous Discrete

Leukemic No yes
state
Extra nodular Un- common common

spread
Lymphoid Neoplasms
Classified into
i) Precursor B cell-Immature B cells
ii) Peripheral B cells-Mature B cell
iii) Precursor T cell - Immature T cell
iv) Peripheral T/NK- mature cells
v) Hodgkin’s lymphoma with RS cell(Reed

Sternberg)
1. Precursor B and T cell neoplasms
A. Acute Lymphoblastic leukemia/lymphoma(
ALL)
2. Peripheral B cell Neoplasms (8 important
subtypes)
3. Peripheral T cell, NK cell neoplasms (6
subtypes)
4. Hodgkin’s Lymphoma (5 subtypes)
1. Precursor B and T cell neoplasms
A. Acute Lymphoblastic leukemia/lymphoma(
ALL)
Lymphoid Neoplasms
2. Peripheral B cell Neoplasms
1. Follicular lymphoma
2. Diffuse large B cell lymphoma
3. Burkitt's lymphoma
4. Plasma cell neoplasms and related disorders
5. Mantle cell lymphoma
6. Marginal zone lymphoma
7. Hairy cell leukemia
8. Plasma Cell Neoplasms
Lymphoid Neoplasms cont’d….
3. Peripheral T cell, NK cell neoplasms
1. Peripheral T cell lymphoma, Unspecified
2. Anaplastic large cell lymphoma(ALK positive)
3. Adult T cell leukemia/lymphoma
4. Mycosis Fungoidosis / Sezary syndrome
5. Large granular lymphocytic leukemia
6. Extra nodal NK/T cell lymphoma
4. Hodgkin’s Lymphoma (subtypes)
1. Nodular Sclerosis
2. Mixed Cellularity
3. Lymphocyte rich
4. Lymphocyte depleted
5. Lymphocyte predominance
• GENERAL PROPERTIES
– Biopsy and histopathologic examination(HPE) is
a must
– Antigen receptor gene (AgR) rearrangements
• Takes place before neoplastic transformation of
lymphoid cells
• All the tumor cells will have same AgR gene
sequence(monoclonal) –
• Every AgR rearrangement leads to unique DNA
sequence
– monoclonal marker (helps to identify sub clones of residual
tumor cells after treatment)
• GENERAL PROPERTIES
– Biopsy and histopathologic examination(HPE) is
a must
– Antigen receptor gene (AgR) rearrangements
• Takes place before neoplastic transformation of
lymphoid cells
• All the tumor cells will have same AgR gene
sequence(monoclonal) –
• Every AgR rearrangement leads to unique DNA
sequence
– monoclonal marker (helps to identify sub clones of residual
tumor cells after treatment)
• Tumor cells resemble one of the stages of their normal
differentiated cells (B, T cells) à helps in identifying specific
features of Leukemias and lymphomas
• Lymphoid neoplasm can cause
– Loss of immunityà infections
– Loss self – tolerance à Autoimmune diseases
• People with inherited /acquired immunodeficiency disorders have
increased risk of lymphoid neoplasms (EBV induced B cell type)
• Neoplastic B/T cells home in to their normal anatomical sites (Ex
– follicular lymphomas (B cell) à germinal centres of lymph
nodes, Cutaneous (T cell) cell lymphomas to skin)
• Neoplastic cells (lymphoid) widely distributed by the time
diagnosis with Exceptions
– Hodgkin’s – contiguous spread
– Marginal zone Lymphomas à limited to site of
inflammation
Precursor B & T cell neoplasms
Acute Lymphoblastic leukaemia/lymphoma( ALL)
Lecture Plan
ØHistiocytoses
ØIntroduction
ØClassification
ØMyeloid neoplasms
Precursor B & T cell neoplasms
Acute Lymphoblastic leukaemia/lymphoma( ALL)
Acute Lymphoblastic leukemia/lymphoma( ALL)
Ø Neoplastic cell- Pre B / Pre T Lymphoblast
Ø 85% are B-ALL
Ø MC cancer of children (<15 yrs. age group)of USA
Ø In 70% of cases mutations of Transcription factors like
– GOF of NOTCH1 in T cell,
– LOF affect ETV 6,RUNX-1 in B cell
Feature B-ALL T-ALL
Age Young children-3 yrs. Adolescents(relates
(relates with predominance with thymus
of B cell population in young development
Frequency MC(85%) Less common
Sex Male Male
AKA Childhood acute leukemia Thymic lymphoma

ALL cont’d...
Ø Single gene mutations are not sufficient to develop
Leukemia (at least 10 mutations are needed)
Ø ***Hyperploidy / hypoploidy are only seen in B ALL and MC
chromosomal abnormality (90% of cases)
Morphology
Ø Marrow; Hypercellular, ↑Lymphoblasts, ↑mitotic rate
Ø Thymic mass - in > 50% of T ALL, (also have splenomegaly
and lymphadenopathy)
Ø Starry sky appearance (due to macrophages engulfing
apoptotic cells) also seen in Burkitt’s
Ø Staining (lymphoblast àMPO - ve, PAS +ve)
Ø Immunophenotyping
Ø 95% of Pre B, Pre T Lymphoblasts TdT Positive (Terminal
deoxynucleotide Transferase)
ALL cont’d...
Ø Clinical Features
Ø Bone marrow ↑ blasts which suppress hematopoiesis
Ø Peripheral blood à anemia, thrombocytopenia
Ø T ALL may have stormy onsetà mass effect (bone pains,
lymphadenopathy and Hepatosplenomegaly, testicular
enlargement, compress large vessels, Air ways of
mediastenum)
Ø CNS symptoms (due to meningial spread) àHeadache,
vomiting, nerve palsies
Ø Prognosis
Ø Paediatric ALL have Excellent prognosis with
chemotherapy (remissions in 95%, cure in 85% of cases)
Ø ALL is still leading cause of death in children – why?
Ø Adults with ALL have poor results (due to molecular
Pathogenesis and poor tolerance to treatment)
Acute Lymphoblastic leukemia/lymphoma( ALL)
Poor prognostic factors Good prognostic Factors
Age less than 2 years Age 2-10 years
MLL (Myeloid Lymphoid Low WBC count

Leukemia or Mixed Lineage
Leukemia)Gene translocations
Adolescents with ALL Hyperploidy
Peripheral blood blasts with Trisomy 4,7,10, t(12:21)

more than 100000(high
tumor burden
ALL
Scant cytoplasm
Inconspicuous Nucleoli
ALL
Markers of Pre B, T Lymphoblast Pan B-Cell markers

ALL
AML
Blasts
Cytoplasm Moderate /abundant Scant
Granules
Nucleus fine Coarse
(chromatin)

Auer rods Present Absent
Lymphoid Neoplasms
• Precursor B And T cell neoplasms
– Acute Lymphoblastic leukemia/lymphoma( ALL)
• Peripheral B cell Neoplasms
1. Chronic lymphocytic Leukemia (CLL)/
Small lymphocytic lymphoma (SLL)
Peripheral B cell Neoplasms
• Chronic lymphocytic Leukemia (CLL)/
– Difference between CLL and SLL is -number of
peripheral blood Lymphoblasts
– For CLL absolute lymphocytic count is >
5,000/microlitre (normal count?)
– MC leukemia in adults of western world
• Age more than 60 years and twice as common in
males
– SLL is 4% of all NHL
CLL and SLL cont’d….
Pathogenesis
• MC genetic abnormalities are
– deletion of chromosomes 13,11,17
– Trisomy 12q
• Tumor suppressor genes - miR15A miR161 are on
chromosome 13
• Tumor cell - Post germinal centre memory B cell
• B cell receptor (membrane bound Ig) stimulates
cascade of kinases - BrutonTK-BTK which
lead to CLL tumor cell growth and survival
– BTK inhibitors are used in treatment of CLL
– BTK defective in X linked Agammaglobulinemia
Morphology:
• Lymph Node- Architecture is effaced; tumour cells are 6-12
microns (mature normal Lymphocyte – 8 microns), small
activated lymphocytes
– Activated Lymphocytes are larger and form
proliferation centre, which are pathogenic of
CLL/SLL
• Peripheral blood shows smudge cells (disrupted nuclei of
tumour cells)
• Bone Marrow: Infiltrated with tumour cells
• Spleen: Tumour infiltrates are seen in red and white pulp
• Liver: Infiltration is mainly in Portal tract
Immunophenotype:
• Tumour cells express Pan B: CD-19, 20, 23, 5 and low level
expression of IgM and IgD
Clinical Features:
• Commonly asymptomatic
• If symptomaticà they present with easy fatigability, weight
loss, anorexia
• Generalised lymphadenopathy, Hepatosplenomegaly
• White cell count varies (In CLL- more than 200,000/
microlitre)
Clinical Features cont’d…
• Both CLL and SLL affect immune function result in
– Bacterial infection (from Hypogammaglobulinemia)
– Immune Hemolytic Anemia and Thrombocytopenia
Clinical Course:
– Median survival is 4-6 years
– Survival is more than 10 years in patients with minimal tumour
burden
Poor prognostic features:
– Deletions of 11q, 17q
– Absence of somatic Hypermutations
– ZAP-70 expression (leads to increased signals of Ig receptor)
– NOTCH- 1 mutations
– High tumour burden
Treatment:
• Asymptomatic patients: Only observation
• Symptomatic:
– Chemotherapy with Antibodies against CD-20
– HSC transplantation in young patients
– BTK inhibitors
• Most important factor: CLL/SLL transform into diffuse large
B-cell lymphoma in 5-10% of cases, called Richter syndrome,
– Richter syndrome
• rapidly growing mass in lymph node and spleen due to
additional mutations
• survival is < one year.
SLL cont’d….
CLL cont’d….
Smudge cell
CLL cont’d….
Lymphoid Neoplasms
5. Plasma cell neoplasms and related disorders
Follicular Lymphoma:
Introduction
• Mc form of indolent NHL in
– Common in middle age, and Male: Female – 1:1
Pathogenesis
• Tumor cell – Germinal center B cell
• Associated with translocations of BCL2 –t(14:18), BCL2 on 14
and IgH on 18
– BCL2 of Bcl2 family is anti-apoptotic à mutations leads
to survival of tumors cells
– Tumor grows by survival rather than proliferation of cells ,
– Low growth fraction (5%),
– less sensitive to chemotherapy and Radiotherapy
Follicular Lymphoma cont’d….
Morphology
• Nodular or nodulo - diffuse pattern,
• LN architecture is preserved.
• Two population cells are present
– Centrocytes – small with scant cytoplasm with nuclear cleaving
– Centroblasts – large cells, moderate cytoplasm, several nucleoli
• Centrocytes predominate
• Peripheral blood involvement is less common,
• BM infiltrates are seen in 85% of cases
• Tumour cells infiltrate
– Splenic white pulp
– hepatic portal tracts
Follicular Lymphoma cont’d….
Immunophenotype
• Tumor cell express CD 19, 20, 10, surface Ig, BCL 6 (resemble germinal
center B cells)
• Most important à CD5 is absent (differentiated from CLL/SLL, Mantle C)
• BCL2 is expressed in 90% of case (normal B cell are BCL2 negative)
Clinical features
• Lymph Nodes- painless, enlarged, Cut Section- uniform size follicles with
loss of Zonation,
• Extra-nodular – mass lesion any where , but common in waldeyer ring,
• Liver, Spleen may develop large destructive masses,
• Other extra-nodal sites – GIT, skin, bone brain
• Bone marrow involvement is late and uncommon
Treatment
• Remission is 70% and, cure in 40-50% with intensive CT
• Adjuvant Rx. with anti-CD20 antibodies
Lymphoid Neoplasms
Diffuse Large B Cell Lymphoma (DLBCL)
• MC NHL in USA, in Elderly males
• MC molecular abnormality is over-expression of deregulated
BCL6
Morphology – Large tumor cells, abundant cytoplasm, Vesicular
nuclei with prominent nucleoli
– Lymph node architecture is effaced
– Anaplastic cells- multinucleated with nucleolar
inclusions (like RS cells)
Immunophenotype – B cell markers (CD 19, 20, 10, and BCL6,
surface Ig)
• DLBCL cont’d…
Special types
1. Immunodeficiency type – seen in patients with
severe immunodeficiency (HIV-AIDS, BM
transplantation recipients) – EBV infected B cell
2. Primary Effusion Lymphoma – Advanced HIV cases,
elderly, present malignant pleural effusion all
tumors cells have KSHV/HHV8 viral genome and
lack B and T cell markers
Treatment
– Intensive Chemotherapy – 70% remission, 50%
cure)
Prognosis: Wide spread and MYC translocations carry poor
prognosis
DLBCL cont’d… Open Nucleolus Mitosis
chromatin
DLBCL cont’d…
Diffuse Large B Cell Lymphoma (solitary large)
Follicular Lymphoma
(multifocal)
Lymphoid Neoplasms
Burkitt Lymphoma
Feature African Sporadic With HIV
(endemic)
Molecular Break point in Class switch Both (in B cells)

IgH region – AID
EBV infection 100% 15-20% 25%
Age Children/young - Adults
Bone Marrow Uncommon Common COMMON

involvement
Extra nodal Mandible, Ileocecum, Any site
sites Abd.viscera peritoneum
Burkitt Lymphoma cont’d….
Pathogenesis
• MC molecular mechanism – MYC translocations (Ch.8) – MYC
– master transcription factor
T(8:14) –repositioning of MYC close to Ig promoter and
increased expression of MYC
↓
Increased expression of genes of aerobic glycolysis
↓
Warburg metabolism – increased synthesis of building blocks of cell –
nucleotide, lipids, proteins
↓
Increased growth and survival of tumor cells
Pathogenesis
Most rapidly growing tumor of humans

fastest growing tumor of all
Morphology
• Tumor cell –B cell with 10-25μ, (similar size as in diffuse type), high mitotic
index, increased number of apoptotic cells
• Macrophages with clear cytoplasm à starry sky
• If marrow is infiltrated à tumor cells show clumped nuclear chromatin with
2-5 nucleoli, Royal blue cytoplasm with clear cytoplasmic vacuoles
Immunophenotype
• Tumor cell express – IgM, CD 19, 20, 10, BCL6
• Most important – BCL2 is absent in 100% cases
Clinical
• MC in extra nodal site,
• Very aggressive in growth but excellent response to
Chemotherapy (cure in case of young people)
Starry sky
Abnormal mitosis
Lymphoid Neoplasms
Mantle CELL LYMPHOMA
Introduction:
• 2.5% of NHL (uncommon in USA)
• 5-6th decade, common in male
• Tumour cells resemble B-cells of mantle zone (Outside
germinal centre)
Pathogenesis:
• All tumour cells have t(11;14) which affects IgH locus of
ch. 14; lead to Cyclin D-1 over expression on ch.11
• 70% of cases diagnosed by karyotyping, 100% by FISH
• Cyclin D-1 acts at G1-S phase in the cell cycle, leads to
proliferation of tumour cells
Mantle CELL LYMPHOMA cont’d….
• Morphology:
– MC is generalised lymphadenopathy, 50% have spleen,
liver, marrow, gut involvement
– Polypoidal lesions in the intestine are called lymphoid
polyposis (unique for mantle cell lymphoma)
– Nodular appearance in low power and homogenous small
lymphocytes with nuclear cleaving in high power
– Centroblasts are absent (helps to differentiate from
Follicular lymphoma and CLL/SLL)
– Blastoid variant – intermediate size cells with open
chromatin and high mitosis
Immunophenotype:
Positive Negative
CD-19, 20, 5, surface Ig (M and D) CD-23
Clinical Features:
– Most common is painless lymphadenopathy
– Symptoms related to spleen and gut
– Media survival (3-4 years)
– Incurable
– Tumour cells infiltrate and lead to organ dysfunction
– Blastoid infiltrates have poor prognosis
Treatment:
– HSC transplantation
– Proteasome inhibitors
Mantle CELL LYMPHOMA cont’d…. Atrophic Germinal center
Homogenous small lymphoid cells

Lymphoid Neoplasms
MARGINAL ZONE LYMPHOMA
• Introduction
– Heterogeneous group with nodal and extranodal
expression
– Extranodal also known as MALTomas
– Somatic Hypermutations indicate the tumour cell is of
memory B-cell origin
– Three characteristics:
• Arise at sites of chronic infection or autoimmune background- -
Sjögren's (salivary gland), Hashimoto’s (Thyroid), MALToma
(Stomach)
• Tumour localised for long term
• Regress with eradicating underlying cause
MARGINAL ZONE LYMPHOMA cont’d….
• Pathogenesis:
– In extra nodal sites, lymphoma arises as continuation of
reactive hyperplasia to lymphoma, (unknown
mechanism)
– In early stages, B-cell proliferation is dependent on T-
helper cells and withdrawal of antigen leads to involution
of the tumour
• Best example is regression of MALToma by treating H. Pylori
– After additional mutations, like t(14;18); leads to antigen
independent tumour cells, which has a poor response to
treatment
MARGINAL ZONE LYMPHOMA cont’d….
Pathogenesis cont’d...
• Translocations up regulate BCL10/MALT-1
which activates NK-ƙB, which leads to
– 1. Clonal evolution, which spreads to distant
sites, transform to diffuse large B-cell lymphoma.
– 2. Similar molecular abnormalities are also seen
in EBV induced lymphomas
Lymphoid Neoplasms
Hairy Cell Leukemia:
Introduction:
• Rare, 2% of all Leukemias
• Middle age (55y), white, common in male (M:F = 5:1)
Pathogenesis:
• More than 90% of cases - BRAF activation point
mutations at 600, Valine to Glutamate
• BRAF is a Serine/Threonine Kinase, acts at
juxtaposition to RAS in MAPK pathway
• BRAF mutations are also seen in Langerhan cell
Histiocytosis, Melanomas
Hairy Cell Leukemia cont’d...
Morphology:
• Tumour cells show fine, hair-like processes, well demonstrated in
phace-contrast microscope
• Nuclei are round, oblong, reniform, with pale blue cytoplasm
having thread-like or bleb-like extension
Marrow:
• Show more hairy cells, enmeshed by ECM, which leads to
entrapment in the marrow
• Result in bone marrow aspirate which is dry tap
• Spleen white pulp is obliterated and looks red, beefy
• Liver – hairy cells infiltrate portal tract
Hairy Cell Leukemia cont’d...
PAN-B Markers Unique Markers

CD 19,20 Surface Ig CD11C, 25, 103, Annexin A1
Clinical Features:
• Most common is massive splenomegaly, liver less common, lymph
nodes rare
• Peripheral smear – Pancytopenia, unexplained Monocytopenia
Clinical course:
• 1/3rd of patients have atypical mycobacterial infections
• Indolent with excellent response with mild chemotherapy
• Relapses after 5 years are common, but still respond with chemotherapy
Hairy Cell Leukemia
Hairy cell
Lecture Plan
Ø Pre-Neoplastic Conditions
ØHistiocytoses
ØIntroduction
ØClassification
ØMyeloid neoplasms
Lymphoid Neoplasms
PLASMA CELL NEOPLASMS AND OTHER RELATED CONDITIONS
Introduction
• AKA – Plasma cell dyscrasias
• Monoclonal gammopathy, Dysproteinemia, Paraproteinemia
• Tumor cell
– Plasma cell
– secrete monoclonal Ig /Ig fragments (non-neoplastic
Polyclonal plasma cells secrete different Igs’)
– Monoclonal Ig/Ig fragments – are tumor markers,
important in pathogenesis
• 15% of deaths from lymphoid neoplasms are due to PLASMA
CELL NEOPLASMS
• MC and most dangerous subtype
“***Multiple myeloma”
PLASMA CELL NEOPLASMS cont’d….
• M component
– Monoclonal Ig,
– High mol. Wt. -160,000,
– Can’t filter through kidneys (not preset in urine)
• Light chains
– Toxic to tubular epithelium
– produced by tumor cells (ƙ>ƛ),
– Bence–Jones proteins (BJP) – ƙ light chains
• low mol. weight (present in urine)
– Amyloid – ƛ (ƛ4, ƛ6) light chains deposit in kidney
• Cause Amyloidosis (AL Amyloid)
Clinico-pathologic types
1. Plasmacytoma – solitary, in bone or soft tissue
2. Smoldering myeloma – high M component but asymptomatic
3. Waldenstrom’s macroglobulinemia – high levels of IgM,
hyperviscosity of blood,, associated with Lympho-plasmacytic
lymphoma (LPL)
4. Heavy Chain Disease - rare and present with LPL, small marginal
zone Lymphoma, in malnourished – Mediterranean L, secretes
heavy chain fragments
5. Primary/immunocyte associated Amyloidosis –ƛ chain
prominence, deposit as AL Amyloid (Amyloid-Light Chain)
6. MGUS – Monoclonal Gammopathy of Undetermined
Significance – asymptomatic, mild to moderate elevated M
component, older people, low but constant procession to MM
7. Multiple Myeloma/Plasma cell myeloma –
widely distribute in skeletal system, secretes IgG
• MM (Plasma cell neoplasm) à
– Elderly
– multiple osteolytic lesions (Low back pains)
– Hypercalcemia
– renal failure
– acquired immune defects
Pathogenesis
• Molecular mechanisms
– Translocations – IgH heavy chain (ch.14q32), results in
abnormalities of Cyclin D1, Cyclin D3,
– Deletion of Ch 17p affects p53 (poor prognosis)
– MYC rearrangements – in late stage, highly aggressive
tumours à can lead to plasma cell leukemia
Myeloma cell secrete
• Cytokine IL-6
– controls proliferation and survival of tumour cells
– produced by tumour cell and marrow stromal cells
– Increased serum IL-6 levels correlate with active disease
and poor prognosis
• MIP-1 alpha
– Upregulates RANKL (It is an activator of NF-ƙB ligand on
the marrow stromal cells – leads to activation of
osteoclasts)
• WNT pathway mediators
– inhibit osteoblast activity à Hypercalcemia
Immunophenotype: Tumour cells are positive
– CD 138 positive, (AKA syndecan 1)
– CD-56
Clinical Features are due to:
– Increased plasma cell tumour growth in bones
– Excess production of Immunoglobulins
– Bones – chronic pain (Lower back), pathological fractures,
– Hypercalcemia (confusion, weakness, lethargy,
constipation, polyuria) also renal dysfunction.
– Decreased immunity – due to decreased normal Ig levels,
leads to recurrent bacterial infections, ***CMI is normal
– Renal failure – is called myeloma kidney
• Causes – multifocal
– Most important is BJP (ƙ chains) àtoxic to
tubular epithelial cells.
– amyloidosis (ƛ6, ƛ3 chins)à Amyloid light -AL
type)
Lab:
1. MC (99% of cases) - ↑ serum Ig (M-component) and
BJP in urine
Cut off for diagnosis
– Ig in serum >6mg/dL (IgG >IgA)
– BJP in urine >3grams/dL
2. Immunofixation- shows M-spike (Serum Ig) in blood and
urine
• 70% of patients have free light chains and M-protein
• 20% have only free light chains
• 1% are non-secretory
3. Bone marrow plasma cell ≥30% (for diagnosis of
MM)
4. Peripheral blood - Normocytic Normochromic
Anemia, Leukopenia, Thrombocytopenia
• Median Survival – 4-7 years
Treatment:
• Proteasome inhibitors
• Immunomodulators – Thalidomide, Lenolidomide (activate
ubiquitin ligase, targeted proteolysis of proteins required by
myeloma
• Biphosphonates – inhibit bone resorption, decrease
pathologic fractures, inhibit hypercalcemia
• Stem cell transplantation – prolongs life
Poor prognostic factors: Good prognostic factors

Multiple bone lesions Smoldering myeloma
Deletion of 17p t(4;14) Cyclin D-1 translocations
Solitary Plasmacytoma
• Osseous or extra osseous site (lung, oronasophrynx, nasal
sinuses)
• local excision cures extra osseous upper respiratory tumors
Smoldering myeloma
• Lies between MM and MGUS
• progression MGUS>Smoldering myeloma > MM
• Marrow plasma cells; 10-30%
• Serum “M” component >3mg/dL
• Asymptomatic
• 75% progress to MM in 15 yrs.
MGUS – Monoclonal Gammopathy of Undetermined
Significance
• MC Plasma cell Dyscrasia
• Asymptomatic,
• 1% develop MM every year (constant conversion rate)
• Molecular abnormalities – translocations and deletions are
same as in MM – MGUS is early stage of MM
• Progression is unpredictable,
• Follow –up with Serum “M” protein, Urine BJP
Lympho Plasmacytic Lymphoma
• B cell neoplasm of 6-7th decades,
• Resembles CLL/SLL, but there is presence of plasma cells
• Monoclonal “M” from plasma cell à Hyperviscosity
syndrome (like Waldenstrom's Macroglobulinemia)
• Complications of light chains and Bone lesions – absent
Pathogenesis
• MYD88 mutations ( adapter protein important in activity of
NK-ƙB factor & downstream signals of B-cell receptor -Ig
complex) lead to growth and survival of tumor cells
Lympho Plasmacytic Lymphoma cont’d…
• Morphology
– Marrow – Lymphocytes, Plasma Cells,
Plasmacytoid cells, and mast cell hyperplasia
– Tumor plasma cells – PAS positive cytoplasmic
RUSSEL bodies, nuclear DUTCHER bodies
• Immunophenotype
– Lymphoid component – CD 20 +ve, surface Ig
+ve,
– Plasma cells – secrete IgM
• Clinical features
– Nonspecific – weakness, fatigue, weight loss,
– 50% with lymphadenopathy and splenomegaly
Lympho plasmacytic Lymphoma cont’d…
• Myelophthesic anemia and IHA (Cold Agglutinin IgM) in 10%
of cases
• Hyperviscosity syndrome - IgM secreting, present with visual
impairment
• Neurological: Headaches, Dizziness, Deafness, Stupor (Due to
sluggish blood flow),
• Bleeding – Formation of complexes between macroglobulins
and clotting factors, abnormal Platelets,
• Cryonglobulinemia – Precipitates of macroglobulins at low
temperatures lead to Reynaud phenomena and cold
urticaria.
Clinical Course:
• Incurable and progressive
• Most of the symptoms related to intravascular IgM –
(Plasmapheresis improves symptoms)
• Transforms to large cell lymphoma
• Median survival is 4 years
M protein Light chain
Serum Protein Electrophoresis

Punched-out osteolytic lesions

Neoplastic plasma cells,

Hodgkin’s Lymphoma
• Lymph nodal involvement/spread - contiguous
• Diagnostic cell – Reed – Sternberg cell (RS cell) – derived from
germinal or post-germinal center B-cell.
• Common in USA (8000) cases per year
• Age- 32 years
• First human cancer successfully treated with radiotherapy and
chemotherapy
• Classification – (Refer table in next slide)
• Molecular abnormality – most common is activation of NF-KB by
EBV,
– EBV expresses latent membrane protein 1 (LMP-1) which activates NF-
KB.
• How to differentiate HL from NHL (ref. Table –next slide)
• Hodgkin’s Lymphoma cont’d…
Hodgkin;s Vs. NHL

HD vs. NHL
Feature Hodgkin’s NHL
Nature Similar to Similar to
infection malignancy
Spread Contiguous Discrete

Leukemic No yes
state
Extra nodular Un- common common

spread
• RS cells
– secrete a variety of cytokines and attract reactive cells like
lymphocytes, plasma cells, Eosinophils and Monocytes, RS cells are
aneuploid, and
– have REL proto-oncogene changes, which result in activation of NF-
KB,
– infected with EBV in 90% of cases
• Subtypes- Refer Tables in slides 26
• Clinical staging – Refer Tables in slides 27,28
• Treatment
– Radiotherapy and Chemotherapy give excellent
results,
• late complications: 1. Secondary malignancy of lung,
breast, skin (Melanoma), and AML (mainly M6)
RS cell
Classic RS Mononuclear variant
Lacunar RS Popcorn like RS

• Subtypes
• Staging
HD Staging & Classification
Clinical Histological
• Nodular sclerosis
– Mixed cellularity
Lymphocyte Predominant
– Mixed cellularity
• Peripheral T cell, NK cell neoplasms
1. Peripheral T cell lymphoma, Unspecified
2. Anaplastic large cell lymphoma(ALK positive)
3. Adult T cell leukemia/lymphoma
4. Mycosis Fungoidosis / Sezary syndrome
5. Large granular lymphocytic leukemia
6. Extra nodal NK/T cell lymphoma
Peripheral T cell/NK cell Neoplasms
• Introduction – heterogeneous group, resemble mature T/NK
cells , T cell tumor à5-10% of NHL in USA and Europe, NK
tumors à rare,
• Both T/NK cells are common in Eastern countries
1) Peripheral T cell Lymphoma, unspecified
• Not easily categorised, diffuse type with loss of LN
architecture, Tumour cell- pleoromorphic T cell, along with
Eosinophils and macrophages, increased neo-angiogenesis
• Markers – CD 2, 3, 5, either ƛβ, or δ TCR, also 4, 8. Many
tumour cells look like mature cells – require DNA analysis
• Morphology – Generalised Lymphadenopathy, Eosinophilia,
Pruritus, fever, weight loss,
• Prognosis – worse than B cell tumors
Peripheral T cell Lymphoma
Mixed populated population of small, intermediate and large

Lymphoid cells
2) Anaplastic Large cell Lymphoma (ALK positive)
• Uncommon, ALK gene rearrangements(ch.2q23) lead to break in
ALK to form chimeric gene – results in activation of TK, RAS,
JAK/STAT (like CML but not BCR-ABL)
• Morphology – tumor cells – large , anaplastic , most abundant
cytoplasm, horseshoe like nuclei (HALLMARK cell), infiltrate LN
sinuses (resemble Mets.)
• ALK is expressed only Tumor cells – very important
• Clinical course & Prognosis – seen in children and young adults,
Mc in soft tissues very good prognosis (cure rate – 75%), older
people – poor prognosis
• Both ALK + and ALK – cells express CD30 (TNF receptor family)
horseshoe like nuclei
IHC with ALK protein

3) Adult T cell Leukemia/Lymphoma
• Introduction – CD4+ Tumor cells, only seen in adults with HTLV-1
infection, common in Japan, West Africa and Caribbean
• Clinical features – skin lesions , Generalised lymphadenopathy,
Hepatosplenomegaly, Hypercalcemia; Peripheral blood –
Lymphocytosis
• Morphology – Tumor cells have multilobated nuclei – “CLOVE LEAF
or FLOWER” cells, TC contain HTLV proviruses , Tax – viral encoded
protein activated NK-ƙ B
• Clinical Course – fatal, death within 1 yr, some cases with localised
skin lesions – indolent course,
– HTLV-1 causes non-neoplastic progressive demyelininating disease of CNS
(Robbin’s page 605 – CNS and spinal cord?)
“CLOVE LEAF or FLOWER” cells
• 4) Mycosis Fungoides (MF) /Seary syndrome
• MFà involves epidermis, upper dermis, tumor cells have cerebriform
nuclei (convolutions or folding)
• Markers – responsible homing of tumors cells to skin – CCR4, CCR10
• Clinical course – indolent with survival -8-9 yrs.
• Rarely develop T cell Leukemia/lymphoma
Feature MF Sezary
Skin lesions Three phases Generalised

(inflammatory, exfoliative lesions
Plaque, (Erythoderma)
Tumorification) Tumorification- rare
Leukemic phase Absent Present with Sezary
cells in blood
Extra cutaneous Lymph Node, NA
spread Marrow
Mycosis Fungoides
Cutaneous T cell Lymphoma

Mycosis Fungoides
CD4
Positive
5) Large Granular Lymphocytic Leukemia
• Rare T/NK cell variants
• 35% cases – STAT3 (TF) mutations lead to cytokine
independent STAT3 signalling.
• Morphology – Tumor cells – large lymphocytes with
abundant blue cytoplasm, few coarse azuriphilic cytoplasmic
granules, Marrow – Tumour cell infiltrates scatter (difficult to
identify), Spleen, Liver – show Tumour cell infiltrates
Feature T cell NK cell

Lymphocytosis, Present Absent
Splenomegaly
CD3, CD56 CD3+, CD56 “-” CD3-, CD56 “+”
Clinical course Indolent Aggressive
5) Large Granular Lymphocytic Leukemia cont’d...
– Clinical Features – Anemia and Neutropenia,
marrow has marked decrease in late precursors
of myeloid series, rarely – pure red cell aphasia
– Sometimes associated with rheumatoid
conditions “FELTY Syndrome” – RA,
Splenomegaly, Neutropenia (may progress to
Leukemia)
6) Extra nodal T/NK cell Neoplasms
• Rare in USA (common in Asia)
• MC present with destructive nasopharyngeal mass, also involve
testis, skin
• Tumor cells compress and invade vessels – Ischemic necrosis
• Large Azurophilic cytoplasmic granules are seen (like NK cells)
• Associated EBV and all tumor cells will have EBV episomes (100%
association) but EBV infected tumor cell lack CD21?
• Markers T cells – CD3, TCR, NK -16, 56
• Highly aggressive but respond well to RT (not CT)
ET
Marked Thrombocytosis
with Giant PLTS
Primary MF
Common Double
Lymphoid pool Negative
Pre B
Lymphoblast
Double
Positive
Naïve B cell
Plasma
cells
Mantle
Cell
Marginal
Zone
Spleen
Spleen
Billroth
Thymus
Thank you
Spleen
Anatomy
• Normal weight (in adults) – 150 grams
• End arterial supply (no collaterals) –
significance?
• No lymphatics
• Circulation – Two circuits
– Open circuit – circulation through capillaries to
cards of Billroth, RBC squeeze through and
examined by macrophages
– Closed circuit- through capillaries to veins
Spleen
Physiology (functions)
1.Phagocytosis
– of senescent cells (RBC, WBC etc.,), particulate
matter- like bacteria)
– Pitting – remove Heinz bodies and Howell-
Jolly bodies
2. Antibody production
– by dendritic cells in spleen
– Auto-Ab to self antigens (in Auto-immune disorders)
3. Hematopoiesis
– minor role in fetus
– EMH in Thalassemia, MPDs’
Spleen
Physiology (functions)
4. Sequestration of blood cells
– PLTS (up to 40% in normal and 90% in
Hypersplenism)
– also sequesters RBC, WBC
5. Part of Mononuclear phagocyte system
– involved in all forms infections, inflammations,
Haematological disorders, Metabolic disorders
– MC manifestation – Splenomegaly
Spleen
Pathology
1. Autosplenctomy or post splenctomy patients
– ↑risk of bacterial infections& septicaemia with
encapsulated bacteria (Strep. pneumoniae, H.
influenza, Meningococci – all three have vaccines)
2. Hypersplenism - Clinico-pathologic condition
– Massive splenomegaly,
– Peripheral blood cytopenias (due to marked
sequestration),
– Compensatory marrow hyperplasia‘s
– reversal of haematological features by splenctomy
Spleen
Pathology
3. Congestive splenomegaly - due to impaired
venous drainage
• Causes
– cardiac in RHF
– Hepatic in Cirrhosis,
– Spontaneous portal venous obstruction in
(intrahepatic obstruction or Portal venous
inflammation – Pylephlebitis)
– Splenic vein obstruction – in Ca. Pancreas, and Ca.
Stomach
Spleen
Pathology
4. Infarcts
– MC with occlusion of splenic artery or branches
– Spleen has no collaterals (end arterial supply) – infracts or
pale or white
– Thrombo - emboli MC originate from heart
5. Neoplasms
– rare
– MC are benign (MC benign neoplasms are –
Lymphangioma, Hemangioma)
• 6. Congenital anomalies
– Absence – rare and associated with Situs inversus
– Hypopalsia – more common
– Accessory spleens or Spenaculi ( clinically important in HS,
ITP)
Spleen
Pathology
7. Rupture
– Medical emergency (causes massive intraperitoneal
bleed à death
– Rx. à emergency splenctomy
• Causes
– - MC organ injured in blunt trauma of abdomen
– Spontaneous rupture – very fragile due to
• infections (IM, Malaria, Typhoid),
• Tumors (Lymphoid neoplasms, MPDs’) ,
– Chronical splenomegaly – rare to rupture (due to
extensive reactive fibrosis)
Thymus
• Role in cell-mediated immunity (CMI)
• Eembryologically derived from the third and
inconstantly, the fourth pair of pharyngeal
pouches
• Involution
– Physiologic-
• 10 to 35 gm at birth it weighs
• 20 to 50 gm at puberty
• 5 to 15 gm in the elderly
– Pathologic (in children and young adults)
• in severe illness and HIV infection
Thymus
• Anatomy
– two fused, well-encapsulated lobes, outer cortical
layer enclosing the central medulla.
• Histology
– Thymic epithelial cells and immature T lymphocytes
– Cortical (peripheral) epithelial cells - polygonal with
abundant cytoplasm with dendritic extensions
– Medullary (central) epithelial cells- densely packed,
spindle-shaped, scant cytoplasm, devoid of
interconnecting processes,
– Hassall corpuscles( characteristic keratinized cores)
present in medulla
Thymus
• Physiology
– CMI - progenitor cells of marrow origin migrate to
the thymus and mature (learn to differentiate self
from non-self antigens) into T cells
– Native cells - Macrophages, dendritic cells, a
minor population of B lymphocytes, rare
neutrophils and Eosinophils, and scattered myoid
(muscle-like) - ? role in the development of
myasthenia gravis
Thymus
Pathology
1. Developmental Disorders
• - Thymic hypopalsia or aphasia -seen in DiGeorge
syndrome ( severe defects CMI, abnormalities of
parathyroid development & hyperparathyroidism)
– DiGeorge syndrome may be part of the 22q11 deletion
syndrome
• Thymic cysts – incidental findings, maximum size up to
4 cm in diameter, lined by stratified to columnar
epithelium and have serous or mucinous fluid
• Cystic neoplastic thymic masses – important,
symptomatic, can be lymphoma or a Thymoma
• “ Asymptomatic cysts – simple / non-neoplastic ,
Symptomatic cysts - neoplastic and can be malignant
Thymus
Pathology
2. Thymic hyperplasia or thymic follicular
hyperplasia (B cell areas)
• Causes –
– chronic inflammatory and immunological states, MCC
- myasthenia gravis, ( 65% to 75% of cases),
– also seen in Graves’, SLE, scleroderma, RA, and other
autoimmune conditions
• Morphology – Thymic enlargement for age (MC
presentation),
• Clinical significance - mistaken radiologically as
Thymoma, (leading to unnecessary surgery)
Thymus
Pathology
3. Thymomas
• Tumors of epithelial cells
• Thymocytes (T cells) – non-neoplastic
• Common Features
– Age- 40 yrs.
– Sex- M: F:: 1:1
– Site- Anterior/ superior Mediastenum
(Hodgkin’s, NHL can also develop at same
site)
Thymus
Pathology
3. Thymomas
• Morphology
– Majority (75%) – encapsulated
– Secondary changes – cystic necrosis, calcification
• Clinical course:-
– Mostly due to local invasion (40%), MG (35%) and
paraneoplastic syndromes (Hypo-
gammaglobulinemia, Grave’s, Pure Red cell
aplasia, Cushing’s)
– Cortical or type-I tumors – autoimmune diseases
Thymus
Pathology
3. Thymomas
• Morphology
– Majority (75%) – encapsulated
– Secondary changes – cystic necrosis, calcification
• Clinical course:-
– Mostly due to local invasion (40%), MG (35%) and
paraneoplastic syndromes (Hypo-
gammaglobulinemia, Grave’s, Pure Red cell
aplasia, Cushing’s)
– Cortical or type-I tumors – autoimmune diseases
Thymomas

TYPE – I TYPE- II
Frequency 50% (mc) 25% 5%
Cell type Medullary or Cortical Thymic
mixed carcinomas(sq
(Medullary & uamous or
cortical Lympho-
epithelial epithelial type)
cells) Resemble
Nasophryngeal
ca.
T-cells Sparse Plenty Sparse
Hassall’s Rare Common --
corpuscles
Thymomas
TYPE – I TYPE- II
Hassall’s Rare Common --
corpuscles
Most important Benign cells Cytologically Cytologically
feature benign, malignant &
locally Anaplastic,
invasive, metastasis – lungs
metastasize
Survival - Minimally Bad
invasive –
90%
Extensive
invasion -
<50%
Virus association - - EBV in 50% pts.
CD5 marker on Absent Absent present
tumor cells
3 R’s of success:
Respect for self,

Respect for others and
Responsibility for all your actions.
Hematology:
Introduction to Bleeding Disorders
Dr. Roy
Introduction
Three major components of normal hemostatic mechanism:
Vessel wall
Platelets
Coagulation cascade
1) Vessel wall:
histologically comprised of 3 layers: intima, media, and
adventitia
intima lined by endothelial cells
endothelial cells rest basal lamina, and subendothelial collagen
2
3
Introduction
1) Vessel wall (cont’d):
Endothelial cell: Most dynamic cell, regulates vasomotor tone, controls cellular
trafficking, and maintains blood in a fluid state
- these cells express receptors for a wide variety of physiological mediators.
Examples: Thrombin, Angiotensin II, adhesion molecules, vWF, protein C
- these cells synthesize: PGI2, NO
- these cells express surface molecules glycosaminoglycans, which are important in
binding factors
4
Platelets
2) Platelets:
- platelets are formed in the bone marrow from megakaryocytes
- mature megakaryocytes are the largest cells in bone marrow
(30 to 90 microns), containing 14 to 16 nuclear lobes
- platelets appear to be formed through fragmentation
- platelets are small, discoid, anucleate cells
- platelets are seen in circulation for about 8 to 10 days
Receptors:
GpIb: receptor for vWF
GpIIb-IIIa: receptor for fibrinogen
5
Platelets
2) Platelets (cont’d):
α granules: contain Factor V, vWF, fibrinogen, PDGF
dense bodies: contain ADP (aggregating agent), calcium,
serotonin
Major Functions:
1)Platelet Adhesion: spreading adhesion involves 3 components-
adhesive proteins (vWF); Calcium; Platelet surface molecules
2)Platelet Aggregation: requires shape change, activation of
Gp2b/3a, bridging with fibrinogen
Normal values:
150,000 to 400,000 cells/mm3
remember: platelet counts tend to be a subject of error due to
clumping, and sometimes due to anti-coagulant used (EDTA)
6
Platelet tests: Bleeding time
- provides assessment of platelet count and function
- bleeding time is the time required for cessation of hemorrhage from a small
puncture wound of the skin made under standard condition. Common method:
Ivy
- Normal: 2 to 7 minutes
Examples for Hereditary causes of abnormal bleeding time:
- von Willebrand disease
- Glanzmann thrombasthenia
Acquired causes:
- aspirin (common)
- liver failure
7
Platelet tests
Platelet aggregation study
- harvest platelets from thrombocytopenic patients
- use aggregators: ADP, collagen, Ristocetin
Ristocetin cofactor activity
- Ristocetin produces an activated conformation of vWF such
that
- it will bind to platelets via the platelet GPIb receptor (this
mimics the in vivo sequence of events that allow interaction
between subendothelial tissue bound vWF & platelets)
- vWF:Ristocetin cofactor activity is the most sensitive and
specific assay for vWF
Abnormal test: seen in vWF disease, Bernard-Soulier dis.
8
Introduction
3) Coagulation system:
-main function of coagulation system is to produce thrombin, in
the event of injury
-which first aids activation of platelets in hemostasis
-secondly forms a stable fibrin network from circulating
fibrinogen
-thirdly by stimulating coagulation inactivating system, thus
limiting the the process to the area of injury
Coagulation cascade
Extrinsic pathway (Factor VII)
Intrinsic pathway (Factor XII, XI, XI, VIII)
9
10
11
12
Blood Coagulation & Tests
13
Partial Thromboplastin Time (PTT)
- PTT is a simple test of the intrinsic and common pathways of coagulation

- PTT is somewhat more sensitive to deficiencies of factors VIII and IX
- in the test, contact activation is provides by glass tube
- but addition of activators such as particulates of silica, or kaolin provides more
optimal contact activation
Normal: 35 to 45 seconds
- interpretation of this test must be in conjunction with other tests
14
Prothrombin time (PT)
- production of fibrin by means of extrinsic and common
pathway requires ‘tissue factor’ and Factor VII
- in addition to Factors X and V, prothrombin, and fibrin
- these pathways are measured by Prothrombin time (PT)
- of the five coagulation factors measured by PT (Factors V, VII, X
and fibrinogen), three (Prothrombin, Factors VII and X) are
vitamin K dependent
- so, these factors are decreased by anticoagulant drugs
(coumarin)
One stage PT: eliminates the role of factor V, by directly
activating factor X
- uses an enzyme present in venom of Russell viper
Normal: 11-13 seconds
15
INR (International Normalized Ratio)
- Prothrombin time has a high variation
- hence, a standardized test was then created where patient’s PT is
compared to a standardized PT
- this test is most beneficial for patients on long term anti-coagulants (like
warfarin)
- normal value for INR: 0.2 to 1.2
Anticoagulants
Heparin
- naturally occurring, its action requires the presence of cofactors:
antithrombin III, and heparin cofactor 2
- major site of action is against factor Xa (prevents conversion of
prothrombin to thrombin)
- this means, a small amount of heparin in the presence of antithmrombin III
prevent formation of thrombin
Anticoagulants
Heparin (cont’d)
- inhibition of factor Xa is instantaneous in the presence of heparin
- however, if thrombin has been formed, larger quantities of heparin is needed to
exert anticoagulant effect
Indications for heparin
- post-op or recumbency stasis thrombosis
- prophylaxis of deep vein thrombosis
- maintenance of anti-coagulation
- arterial embolization
Anticoagulants
Heparin (cont’d)
Administration
Unfractionated heparin
- these are large molecular weight proteins, and hence only one third of heparin
molecules administered to patients have anticoagulant function,
- and the anticoagulant profile and clearance of heparin are influenced by the
chain length of the molecules
- differential clearance results in accumulation of the lower-molecular-weight
species, which have a lower ratio of antithrombin III to anti-factor
- also, high mol. wt. heparin has a higher affinity to bind platelets
- monitoring Heparin: aPTT remains the most convenient despite its limitations
Anticoagulants
Heparin (cont’d)
Administration
Low Molecular Weight Heparin (LMWH)
- these are derived from depolymerization of standard heparin, which yields
fragments approximately one third the size of the parent compound
- anticoagulant effect by inhibiting factor Xa and augmenting tissue-factor-pathway
inhibitor but minimally affect thrombin, or factor Iia
- aPTT, a measure of antithrombin (anti-factor IIa) activity, is not used to measure
the activity of low-molecular-weight heparins, which requires instead a specific
anti-Xa assay
Fibrinolytic tests
Fibrinogen Degradation Product (FDP’s)
- FDP’s are protein fragments of varying sizes that result from the
proteolytic action of plasmin on fibrin
- plasma levels of these are increased in DIC
D-Dimer assay
- detects cross-linked insoluble fibrin monomers in a fibrin clot
- does not detect fibrinogen degradation products, as these are not
cross-linked
Uses of D-Dimer assay:
- deep vein thrombosis
- pulmonary thromboembolism
- DIC
20
21
Platelet Coagulation
Petechiae, Purpura Hematoma, Joint bl.

22
Petechiae: 1 to 2 mm
Purpura: more than 3 mm

23
Porphyrias
Introduction
- porphyrias arise from various inherited enzyme defect in heme biosynthesis
pathway
Classification
1) congenital erythropoietic porphyria
2) erythrohepatic protoporhyria
3) acute intermittent porhyria
4) porphyria cutanea tarda
5) mixed porphyria
24
25
acute intermittent porphyria (AIP)
Introduction
- disease is widespread but is especially common in Scandinavia
and Great Britain
Etiology
- autosomal dominant condition resulting from the half-normal
level of HMB synthase activity
Pathogenesis
- induction of the rate-limiting hepatic enzyme ALA synthase in
heterozygotes who have half-normal HMB synthase activity is
the probable cause for acute attacks
- this disorder remains latent (or asymptomatic) in the most who
are heterozygous for HMBS mutations
26
Pathogenesis
- when heme synthesis is increased in the liver, half-normal HMB
synthase activity may become limiting,
- and ALA, PBG, and other heme pathway intermediates may
accumulate and be excreted in the urine
- almost always latent before puberty suggests that adult levels
of steroid hormones are important for clinical expression
- more common in women, suggesting a role for estrogens or
progestins
- pre-menstrual attacks is likely due to endogenous
progesterone
- acute porphyrias are sometimes exacerbated by exogenous
steroids, including oral contraceptive preparations containing
progestins 27
28
Labs
CBC: normal
urine: porphobilinogen increased (darkens on
exposure to light)
porphobilinogen levels: elevated in urine or
plasma (specific for acute porphyria)
Dark Brown or Black urine:
-‘cola colored’: rhabdomyolysis
-dark brown: alkaptonuria, melanoma
-port wine: acute intermittent porphyria
29
Clinical Features
- typical presentation is that of patient (young woman) who is previously healthy
- who has had several days of severe fatigue and an inability to concentrate,
followed by progressively worsening abdominal pain, nausea, vomiting, and
subtle neurologic signs
- no abdominal findings on examination
30
31
“If you break your neck, if you have nothing to eat, if your house is on fire, then you've got a problem. Everything else is an
inconvenience. Life is inconvenient. Life is lumpy. A lump in the oatmeal, a lump in the throat, and a lump in the breast are
not the same kind of lump. One needs to learn the difference”
Robert Fulghum, author (b. Jun 1937)
Bleeding Disorders
Dr. Roy
Bleeding Disorders
- excessive bleeding can result from
i) increased fragility of the blood vessels
ii) platelet deficiency and dysfunction
iii) derangement of coagulation
iv) combination of these
2
Bleeding Disorders
Investigations:
1) Complete Blood Counts
- Total Platelets Count (normal: 150,000 to 400,000 cells/mm3)
2) Bleeding Time
- rate at which a stable platelet plug is formed following a superficial puncture
wound
- bleeding normally stops within 1-9 minutes
3) Platelet aggregation test
Bleeding Disorders (coagulation tests)
4) Prothrombin Time (PT)
- production of fibrin by means of the Extrinsic and Common pathway
- this requires Tissue Factor, Factors: VII, X, and V, Prothrombin, and Fibrinogen
Methods of estimating PT
- “Quicks PT”, its one-stage test based upon the time required for a fibrin clot to
form after the addition of Tissue Factor (Normal 11 to 15 seconds)
International Normalized Ratio (INR)
- used to standardize prothrombin time (PT) results for patients taking warfarin
(coumadin)
- INR does not have unit. Normal: 0.9 to 1.1
5) Activated Partial Thromboplastin Time (aPTT or PTT)
- time it takes to produce fibrin polymers after adding phospholipids
- normal: 25 to 40 seconds
Prolonged aPTT/PTT
Common Pathway: F X, cofactor V, Thrombin, Fibrinogen
Intrinsic Pathway: F XII, XI, IX, VIII
Prolonged in:
- Hemophilia, Vitamin K deficiency, DIC, vWF, SLE (if anticoagulant is present),
monitoring of patients on unfractionated heparin (UHF) therapy
6) Plasma Thrombin Time (TT)
- time it takes to produce fibrin polymers after adding phospholipids
- Tests Fibrinogen activity
Prolonged TT:
- Fibrinogen deficiency
- monitoring Heparin therapy, and Fibrinolytic therapy
Bleeding disorders related to vessel wall abnormalities
Henoch-Schönlein purpura
Incidence:
- seen in childhood and adolescence (90% below 10 years)
- frequently following upper respiratory tract infection
Pathogenesis:
- systemic hypersensitivity disease of unknown cause
- characterized by widespread inflammatory reaction of the capillaries and small
vessels
- deposition of immune complexes (IgA)
7
Lab Investigations:
CBC:
- Total WBC: mild increase
- Total Platelets: normal or mild increase
Hb: reduced
Bleeding Time: normal
Coagulation work-up: normal
Urine: proteinuria, microscopic hematuria, RBC casts
Skin biopsy: IgA, C3, fibrin deposits
Renal biopsy: IgA, C3 deposits, fibrin deposits
Lab Investigations:
BUN: increased
Serum creatinine: increased
Serum antibodies and complement:
- IgA: detected
- Complement levels: reduced
ESR and CRP: increased
Stool Guaiac test: positive
Skin biopsy: note upper dermal blood vessels with a
collar of inflammatory infiltrate. Arrow shows blood
vessels obliterated by inflammation & fibrinoid
necrosis
Clinical Features:
-immune disorder seen in children following upper respiratory infection
-characterized by vasculitis presenting as palpable purpura especially notes in
buttocks and lower limbs
-also noted are GI bleed, glomerulonephritis
-IgA nephropathy
-disease is self-limited, respond well to steroid therapy
11
IgA nephropathy
- most prevalent primary chronic glomerular disease
- diagnostic hallmark of IgA nephropathy is the predominance of IgA deposits,
either alone or with IgG, IgM, or both, in the glomerular mesangium
- complement C3 and properdin are almost always present
- C4 or C4d, mannose-binding lectin, and terminal complement complex (C5b–C9)
are frequently detected, whereas C1q is usually absent
- these findings suggest involvement of the alternative and lectin pathways of
complement activation
Light Microscopy: an increase in mesangial matrix and hypercellularity are
common; other glomerular lesions may include focal necrosis
12
IgA nephropathy
13
Bleeding related to reduced platelet number
Thrombocytopenia
Decreased production of platelets: Aplastic anemia, Leukemias
Decreased platelet survival: Immune Thrombocytopenia as in Immune

Thrombocytopenic Purpura (ITP), or Non-immunologic as in Disseminated
Intravascular Coagulation
Sequestration: hypersplenism
Dilutional: massive transfusions

14
Immune Thrombocytopenic Purpura
Types:
1) Primary Immune Thrombocytopenia
2) Secondary
Primary Immune Thrombocytopenia

Acute Immune Thrombocytopenia
Chronic Immune Thrombocytopenia
Secondary Immune Thrombocytopenia
Drug induced: most causing aplastic anemia
Leukemias
Bone marrow infiltration
Hypersplenism
SLE
Infections: HIV
15
Acute Immune Thrombocytopenic purpura
Incidence:
- seen in children, usually following viral infection (within 6 weeks). Varicella zoster
and EBV most common
Pathogenesis:
- formation of anti platelet antibodies (GpIIb/IIIa)
- self limited, resolves spontaneously within 6 months
- steroid therapy indicated, if, thrombocytopenia is severe
16
Acute Immune Thrombocytopenic purpura
Lab findings:
CBC: Thrombocytopenia
Bleeding time: prolonged
Coagulation study: normal
Clinical Features:
- petechial hemorrhages on skin, epistaxis, (mucocutaneous bleeding)
- 80% of children with untreated acute immune thrombocytopenic purpura (ITP) have a
spontaneous recovery
- with completely normal platelet counts in 2-8 weeks
17
Acute ITP: A) extensive petechiae and purpura on the legs of a child
with immune thrombocytopenic purpura. B) shows a conjunctival
hemorrhage
18
Chronic Immune Thrombocytopenic Purpura
Incidence:
- young to middle aged women
- vary from mild to severe
Pathogenesis:
- formation of ‘auto antibodies’
- these are directed against ‘platelet membrane glycoproteins’ (IgG antibodies
directed against GpIIb/IIIa)
- anti-platelet antibodies act as opsonins, that
- are recognized by IgG Fc receptors on phagocytes
- results in platelet destruction, mostly in spleen
19
Lab investigations:
CBC: Thrombocytopenia
Partial Thromboplastin Time (aPTT): normal
Bone marrow:
- bone marrow is not required as a routine
- but is indicated if clinical history/physical examination raises a possibility of
leukemia
- as leukemia is associated with thrombocytopenia
Findings in BM: slight increase in megakaryocytes
20
Clinical features:
- easy bruising, menorrhagia, melena, bleed from mucus membranes
- subconjunctival and retinal hemorrhages may be seen
- transplacental transfer of IgG may produce transient thrombocytopenia in infants
- most patients responds to corticosteroids
- splenectomy (in refractory cases)
- IVIG (intravenous immune globulin) for steroid resistant cases
21
Treatment strategies for Chronic
Immune Thrombocytopenia
22
Drug induced thrombocytopenia
Heparin-induced thrombocytopenia (HIT)
- is a complication of heparin therapy
Types:
Type 1 HIT
- presents within the first 5 days after treatment with heparin
- there is only moderate reduction of platelet (remains more than 100, 000
cells/mm3)
- and the platelet count normalizes with continued heparin therapy
- Type 1 HIT is a non-immune disorder that results from the direct effect of heparin
on platelet activation
23
Type II
- is an immune-mediated disorder that typically occurs 5-14 days after exposure to
heparin
- platelet count fall below 100,000 cells/mm3, clinically significant
- associated with life- and limb-threatening thrombotic complications
- Type II must be suspected when a patient who is receiving heparin (usually
unfractionated) has a decrease in the platelet count
- caused by antibodies that recognize complexes of heparin and platelet factor 4,
which is a normal component of platelet granules
24
Type II
Pathogenesis:
- induced by IgG antibodies recognizing
Platelet Factor 4 and Heparin complexes
- resulting immune complexes that
activates thrombin
- remember: increased thrombin, not
thrombocytopenia, causes the clinical
problems (paradox)
Lab:
- Serotonin release assay
25
Type II
Treatment:
- stop Heparin therapy immediately!
- ELISA for confirmation
- other test: functional assay for PF4 autoantibody
- start non-heparin anticoagulation. Example: direct Thrombin inhibitor-
Argatroban
- or, synthetic Heparin
- avoid Warfarin until platelet count has normalized to 150,000 cells/mm3
26
Hypersplenism
Introduction:
- enlarged spleen which causes rapid and premature destruction of blood cells. It is
also referred to as congestive splenomegaly
- by definition: one or more blood cytopenias
Types:
1) Primary hypersplenism: rare, affects women. Banti syndrome
2) Secondary hypersplenism: portal hypertension, spontaneous portal vein
thrombosis, hepatic vein thrombosis (Budd Chiari), CHF
3) Occult hypersplenism
27
1)TTP
2) HUS
3) HELLP
28
Thrombotic Thrombocytopenic Purpura (TTP)
Incidence:
- mostly women, more in African Americans
Pathogenesis:
- deficiency of enzyme ADAMTS 13 (vWF metalloprotease)
- normally degrades very high molecular weight multimers of vWF
- in its absence there would be excess of vWF (as it cannot be cleared)
- this increases platelet adhesion in areas of endothelial injury (high shear stress
conditions, drugs)
- therefore thrombus formation occurs with minimal endotheilal injury
- thrombocytopenia results from platelets used up in widespread thrombus formation
Lab Findings:
Hb: reduced
CBC: leukocytosis, thrombocytopenia
Peripheral blood smear: schistocytes, low platelets
Coagulation studies: normal (PTT is normal)
Intravascular hemolysis: hemogloblinemia, hemoglobinuria, may be seen
BUN, and S. creatinine: elevated
Urine: hematuria, proteinuria
ADAMTS 13 assay: not reliable
30
TTP Peripheral blood smear: Note: schistocytes (arrow), scant platelets,
polychromatophils
31
Clinical features:
Pentad
1)fever
2)thrombocytopenia
3)renal failure
4)microangiopathic hemolytic anemia
5)neurological symptoms
32
Hemolytic-Uremic syndrome (HUS)
Incidence:
- children (less than 10 years in most cases)
- epidemic , summer months, under cooked red meat, unpasteurized milk, contaminated water
- may be transmitted from person-person
- common cause for acute renal failure in children
Pathogenesis:
- infectious gastroenteritis caused by E.coli strain 0157:H7
- shiga-like toxin (STEC) is elaborated which may act both locally and systemically
- these shiga-like toxins binds and damages endothelial cells in gut mucosa (hemorrhagic colitis)
33
-shiga-like toxins are released into circulation and glycosphingolipid receptor for
Stx is globotriaosylceramide (Gb3) present in renal endothelial cells
-cytotoxicity of shiga-like toxins leads to endothelial cell swelling, abnormal
vascular permeability, cell death
-endothelial cell disruption exposes the underlying thrombogenic GBM
-triggering platelet activation and aggregation, and cytokine activation
-thrombi formation
34
Lab Findings:
Hb: reduced
CBC: thrombocytopenia
Peripheral blood smear: schistocytes (see next slide)
Haptogloblin: reduced
Intravascular hemolysis: hemoglobulinemia may be seen
Coagulation studies: PTT and PT are normal
Fibrin degradation product (FDP): may be increased
35
HUS: note schistocytes, and reduced number of platelets.
36
Lab Findings (cont’d):

BUN & Serum creatinine: raised
Urine analysis: hematuria, proteinuria
Stool examination:
- EHEC 0157:H7 detected by plating fresh feces on
sorbitol-MacConkey agar
- ELISA and PCR more sensitive
Note: fibrin thrombi in glomeruli

(bright red) 37
Clinical Features:
Triad:
1)acute renal failure
2)thrombocytopenia
3)microangiopathic hemolytic anemia
- infants and young children
- bloody diarrhea, vomiting, fever, hypertension
38
HELLP
Introduction:
Hemolysis Elevated Liver enzymes Low Platelets
-life-threatening condition that can potentially complicate pregnancy. Association
with Preeclampsia, increasing age, and multiparity may be risk factors
Preeclampsia: is characterized by a triad of hypertension, proteinuria, and edema
hypertension is defined as an elevation of 30 mm Hg (systolic) or 15 mm Hg
(diastolic) above the value in the first trimester or any value above 140/90 mm Hg
Eclampsia: is marked by seizures or coma in addition to the signs of preeclampsia
* most occur between the 27th and 36th week of gestation
39
HELLP
Labs:
CBC:
-thrombocytopenia
Hb: decreased
Peripheral blood smear: schistocytes, reduced number of platelets
Liver Function Tests:
-increased serum bilirubin
-abnormal liver enzyme levels
40
HELLP
Clinical Features:
- abdominal pain, and vomiting by 27th
week of gestation
- right-upper-quadrant or epigastric
pain, headache, visual disturbance,
weakness, and jaundice
- edema with secondary weight gain
Placenta: normal
41
Bleeding disorders related to defective platelet function
42
Bleeding Disorders: defective platelet function
Defective adhesion
Bernard-Soulier syndrome
- deficiency of platelet membrane glycoprotein complex
Defective aggregation
Glanzmann’s thromboasthenia
Defective platelet secretion

Storage pool defects
43
Introduction:
- rare autosomal recessive, consanguinity
- platelets have a quantitative, or qualitative abnormality of membrane GpIb complex
- therefore failure to bind von Willebrand factor
- characterized by mild thrombocytopenia, and giant platelets
Pathogenesis:
- severe bleeding tendency due to defective adhesion of platelets to subendothelium
Normally: a domain of the of the GpIb alpha binds directly to vWF mediating normal
platelet adhesion
44
CBC:
mild reduction in platelets
- giant platelets
Coagulation studies: normal
Ristocetin test: abnormal
45
Clinical Features:
- bleeding tendency seen early in life or early
childhood
- purpura, epistaxis, gingival hemorrhages,
menorrhagia
- severe bleed may be sometimes seen
- patients must avoid anti-platelet treatment (aspirin)
Bernard-Soulier: blood smear
46
Glanzmann Thrombasthenia
Introduction:
- rare, familial, autosomal recessive, consanguinity
- bleeding tendency due to quantitative or qualitative abnormalities of platelet
receptor GpIIb-IIIa
Pathogenesis:
Normally: platelet receptor GpIIb-IIIa binds to fibrinogen
Glanzmann Thrombasthenia: a genetic defect in GpIIb-IIIa prevent normal assembly
and processing of the functional receptor. This results in the lack of a fibrinogen
receptor and a defective fibrinogen binding after platelet activation
- therefore results in a problem with platelet aggregation
47
48
Glanzmann Thrombasthenia
Lab diagnosis:
CBC:
total platelet count: normal
Peripheral blood smear: normal looking platelets
Platelet aggregometry: abnormal study
Coagulation studies: normal
Clinical Features:
- bleeding tendency more in homozygous mutations
- purpura, epistaxis, gingival hemorrhages, menorrhagia
49
Bleeding disorders (Platelet function)
Drugs:
- aspirin causes platelet aggregation defect
- inhibits COX pathway, which blocks synthesis of thrombaxane A2 (TXA2)
Uremia:
- reduction in ADP induced platelet aggregation (please read footnotes)
Infections:
- in the setting of DIC
50
Bleeding Disorders (abnormalities in clotting factors)
von Willebrand disease
Hemophilia A
Hemophilia B
51
von Willebrand Disease
Incidence:
- inherited, autosomal dominant disease
- clinically resembles Hemophilia, (pseudo-Hemophilia)
- it is the most common inherited bleeding disorder
Pathogenesis:
characterized by: 1) defect of platelet function leading to prolonged bleeding
2) coagulation defect due to deficiency of factor VIII activity in plasma
52
-FVIII and vWF are encoded by separate genes and synthesized in different cells
FVIII and vWF:
-FVIII is made by sinusoidal endothelial cells and Kupffer cells (liver), tubular
epithelial cells (kidney)
-once FVIII reaches circulation it binds to vWF (made by endothelilal cells)
-vWF stabilizes FVIII
-circulating vWF exists as multimers (exceed 20 x 106 daltons)
-most important function of vWF is to promote adhesion of platelets to
subendothelial collagen (occurs through platelet GpIb, and matrix components)
-vWF may influence platelet aggregation
53
Lab investigations:
CBC:
Platelet count: normal
Bleeding time: prolonged, (this is due to a platelet adhesion defect)
PT (prothrombin time): normal
aPTT (Partial Thromboplastin Time): prolonged
vWF activity: reduced
Factor VIII activity: reduced
Ristocetin test: abnormal
54
Clinical Features:
- spontaneous bleed from mucous membrane (epistaxis), menorrhagia (important)
- bleed from trivial wounds, menorrhagia, dental procedures, minor surgeries
Remember: vWF disease has several variants, so the severity of symptoms range from
mild to severe
- a well-known, serious, and possibly life-threatening bleeding complication is
gastrointestinal bleeding from angiodysplasia
- intraarticular (joint) bleeding is a frequent complication in patients with hemophilia but
has not been reported as a major problem in patients with von Willebrand’s disease
Treatment: 1) Desmopressin. It increases vWF and Factor VIII levels, and helps endothelial
cells to release vWF. 2) Aminocaproic acid. It inhibits plasminogen activator
55
Hemophilia A
Introduction:
- mutation in Factor VIII
- X-linked recessive trait
- affects males (and homozygous females)
- females are asymptomatic carriers
- transmits the abnormal X chromosome to 50% of male offspring
- no family history in 30%
- severity depends on amount of Factor VIII activity seen
- typically presents with spontaneous hemorrhages
56
57
Hemophilia A
Lab investigations:
CBC:
Platelet count: normal
Bleeding time: normal
Prothrombin time: normal
aPTT (Partial Thromboplastin Time): prolonged
(remember: Factor VIII is seen in Intrinsic pathway, so it would be abnormal.
However, Prothrombin is normal as it is in the Extrinsic pathway)
Factor VIII levels: reduced
detection of female carriers 58

Coagulation Studies
59
Hemophilia A
Clinical features:
- activity below 1% is associated with severe disease
- easy bruising, hemorrhage following trauma or operative procedures
- ‘spontaneous’ hemorrhages to knee joints (hemarthroses)
- chronic hemophilic arthritis
(soft tissue hematomas & hemarthroses)
- hematuria
- bleed into CNS (most serious)
Remember:
- clinically Hemophilia A & B are indistinguishable
- petechiae are absent
60
Bleed into joints X-knee: note soft tissue shadow (edema)
61
Hemophilia B
- clinically similar to Hemophilia A
- decrease in Factor IX activity
- also called as Christmas disease
- X linked recessive
- Factor IX deficiency may be mild, moderate or severe
- differentiated from Factor VIII deficiency by Factor assay
62
63
Disseminated Intravascular Coagulation
Introduction:
-complex systemic thrombo-hemorrhagic disorder
-disseminated intravascular clotting causes a hemostatic defect
-resulting from utilization of the coagulation factors and platelets (consumption
coagulopathy)
Etiology:
-it is not a primary disorder
-secondary to various causes
64
Etiology (cont’d):
- obstetric complications (abruptio placentae, amniotic fluid embolism, abortion, intra uterine
death)
- infections (sepsis)
- neoplasms (mucin secreting adenoca)
- massive tissue injury
- snake bite (viper)
Pathogenesis:
- pathologic activation of the coagulation or
- in some cases impairment of clot inhibiting mechanism
Two major mechanisms that trigger:
1)release of tissue factor, or procoagulants
2)widespread injury to endothelial cells
65
- once triggered, there will be formation of thrombin, which converts fibrinogen to fibrin
- widespread deposition of fibrin in microcirculation results in ischemia (microcirculation)
- fibrin in microcirculation leading to microangiopathic hemolytic anemia (which produces
injury to red cells in circulation)
- also there will be ischemia, due to microthrombi
- with widespread microthrombi in circulation, there will be release of
- plasmin, which then cleaves fibrin
- remember plasmin also digests Factors V, VIII therefore depleting their concentration
- also fibrin degradation products from fibrinolysis will inhibit platelet aggregation, and
inhibit thrombin
66
67
Pathogenesis of Septic shock
68
69
Morphology:
thrombi seen in various organs
- Brain
- Heart
- Lungs
- Kidneys
DIC may be associated with:

1.Waterhouse-Friderichsen syndrome (Neisseria meningitidis sepsis)
2.Sheehan postpartum pituitary necrosis
70
DIC Blood smear: schistocyte (arrow). Note: reduced platelets.
71
Fibrin in glomerulus. DIC.
72
Lab diagnosis:
CBC:
Platelet count: decreased
PT (prothrombin time): prolonged
aPTT (partial thromboplastin time): prolonged
Fibrinogen levels: reduced
Peripheral blood smear: schistiocytes, reduced platelets, polychromatophils
Fibrinogen degradation products and D- Dimers: present
73
Treatment:
1) Packed RBC: for patients with active bleeding, or Hb below 7 gm%
2) Platelet Transfusion: active bleeding , or high risk of bleeding (platelet count
less than 50,000 cells/mm3)
3) Fresh Frozen Plasma: PT or PTT less than 1.5 times the normal
4) Cryoprecipitate: bleeding and Fibrinogen levels less than 150 mg/dl (N 200 to
400 mg/dl) despite Fresh Frozen Plasma (FFP), or when FFP is not available
5) Anti-fibrinolytic therapy:
6) Anticoagulation: prophylactic Heparin, therapeutic Heparin, and other
anticoagulants
74
Clinical Presentation of Cytokine Storm
“only mediocrity can be trusted to be always at its best. Genius must always have lapses proportionate to its
triumphs”
Max Beerbohm, essayist, parodist, and caricaturist (1872-1956)
77
CNS Pathology, Part 1
Cerebral Edema
Developmental Malformations
Spinal cord diseases
CNS Trauma
March 2021
Introduction
Two major divisions
- central nervous system (CNS). Consists of brain, spinal cord, optic nerve and
retina, and contains the majority of neuronal cell bodies
- peripheral nervous system (PNS). Consists of nervous tissue outside the CNS and
consists of the cranial and spinal nerves, the peripheral autonomic nervous
system (ANS) and the special senses (taste, olfaction, vision, hearing and balance)
- autonomic nervous system. Subdivided into sympathetic and parasympathetic
components.
Introduction: histology
CNS:
- neuronal cell bodies are grouped together and are, more or less, segregated from
axons
- the generic term for such collections of cell bodies is grey matter
- small aggregations of neuronal cell bodies, which usually share a common
functional role, are termed nuclei
- neuronal dendrites and synaptic interactions are mostly confined to grey matter
- axons tend to be grouped together to form white matter, so called because axons
are often ensheathed in myelin, which confers a paler colouration
Introduction: histology
Spinal cord:
- lies within the vertebral column, in the upper two-thirds of the vertebral canal,
and is continuous rostrally with the medulla oblongata of the brainstem
- spinal cord receives afferent input from, and controls the functions of, the trunk
and limbs
- afferent and efferent connections between the periphery and the spinal cord
travel in 31 pairs of segmentally arranged spinal nerves that attach to the cord as
a linear series of dorsal and ventral rootlets
- dorsal roots carry primary afferent nerve fibres from
neuronal cell bodies located in dorsal root ganglia,
- whereas ventral roots carry efferent fibres from
neuronal cell bodies located in the spinal grey
matter
Blood supply
Blood supply
Layers of the brain
Types of cells
Reactions of neurons to injury
1) Acute neuronal injury (“red neurons”)
- seen following acute CNS hypoxia/ischemia, severe
hypoglycemia, and other acute insults, and are the
earliest morphologic markers of neuronal cell death
- red neurons are evident by 6 to 12 hours after an
irreversible hypoxic/ischemic insult
- characteristic features include: a) shrinkage of the
cell body b) pyknosis of the nucleus c)
disappearance of the nucleolus d) loss of Nissl
substance e) intense cytoplasmic eosinophilia
Normal
Cerebral Edema
Cerebral Edema
Types:
Vasogenic: caused by
- this is extracellular fluid accumulation
- due to impaired capillary permeability secondary to breakdown of endothelila
tight junction
Causes: localized (inflammation, or neoplasms), or generalized
Cytotoxic: caused by
- intercellular accumulation of fluids
- due to: impaired sodium/potassium ATP
Cerebral edema
Cerebral Edema: Hydrocephalus
Introduction:
- abnormal enlargement of cerebral ventricles and/or subarachnoid spaces due to excess
CSF
Types:
1) Communication (due to failure in CSF absorption or increased CSF production)
2) Non communicating ( obstruction of CSF from ventricles to the subarachnoid spaces)
3) Normal Pressure Hydrocephalus:
- chronic form of communicating hydrocephalus, seen in old (60 yrs and above)
- results from decreased CSF absorption
- there is no increase in ICP (normal pressure), as there is effective compensation as CSF
accumulates through ventricular dilatation
4) Hydrocephalus ex vacuo
Pathophysiology Clinical Features Diagnosis
Communicating - Increased CSF production - headache, nausea, - USG in infants less than 6
- Decreased CSF absorption vomiting months
- Papilledema - MRI/CT in older infants
- Abducens nerve palsy and adults
- Abnormal gait
- Impaired consciousness
- Cushing Triad
Non communicating - Obstructed flow of CSF from - Similar features as above - Similar to above
ventricles to subarachnoid
space
Normal pressure - Decreased CSF absorption Classic Triad - MRI preferred as initial
- Wet: urinary test
incontinence - CSF analysis would be
- Wacky: dementia confirmatory
- Wobbly: frequent falls,
broad-based gait with
short steps (ataxic gait)
Hydrocephalus ex - Loss of brain tissue - Symptoms of underlying - Cortical atrophy on
vacuo disease: Alzheimer, Pick radioimaging
Hydrocephalus: clinical features
- excessive CSF within the ventricular system
- enlargement of ventricles
- when it happens before closure of cranial sutures (infancy)
- there is enlargement of head circumference
- after infancy, hydrocephalus produces enlargement of lateral ventricles and
raised intracranial pressure, without increase in head circumference
19
CSF Pathway
20
Setting sun sign
21
Raised intracranial pressure & herniation
Introduction:
- herniation is displacement of brain tissue past rigid dural folds (falx and tentorium) or
through the openings in the skull because of
- increased intracranial pressure
- as volume of the brain increases, CSF is displaced, and
- cause compression of blood vessels
- leading to increased intracranial tension (ICT)
Causes:
Generalized brain edema
Tumors, abscesses, hemorrhages
22
Cerebral Herniation Syndromes
Subfalcine
- cingulate gyrus of one hemisphere is compressed and herniates beneath falx
cerebri
- compression of:
1) contralateral hemisphere: obstructs foramen Munro leading to herniation and
produce hydrocephalus
2) Pericallosal arteries: hemiparesis (especially affecting lower limbs)
Uncal herniation
- medial temporal lobe (uncus)
Compression of
1) ipsilateral oculomotor nerve (fixed dilated pupil)
2) ipsilateral posterior cerebral artery (cortical blindness with contralateral
homonymous hemianopia)
3) contralateral cerebral peduncle (ipsilateral paralysis plus Kernohan
phenomenon)
Foramen magnum herniation
- structures of the posterior fossa
(cerebrellar tonsils, medulla)
- herniate through the foramen
magnum
- this produces impaired consciousness,
decerebrate posturing, apnea,
impaired circulation, death
Duret hemorrhages:
- hemorrhage of the brainstem caused by progressive
brain herniation
- this results in compression of brainstem and
subsequent damage to ts blood supply
- linear or flame shaped lesions
- midline or paramedian location
- due to distortion or tearing of
- penetrating veins and arteries supplying
Duret hemorrhage: visible here as a midline
- the upper brain stem linear focus of hemorrhage
26
Malformations & Developmental disease
- Neural tube defects
- Dandy-Walker malformation
- Arnold-Chiari malformation
- Syringomyelia
- Tuberous Sclerosis
27
Neurulation: formation of neural tube
- processes involved in the formation of the neural plate and neural folds
- and closure of the folds to form the neural tube constitute neurulation
- neurulation is completed by the end of the fourth week, when closure of the
caudal neuropore occurs
- as the notochord develops, it induces the overlying embryonic ectoderm, located
at or adjacent to the midline, to thicken and form an elongated neural plate of
thickened epithelial cells
at 19 to 21 days
transverse sections
Neural tube defects
Introduction:
- failure of a portion of the neural tube to close
Neurulation (overview)
- notochord induces the overlying ectoderm to form the neural plate
- by the end of 3rd week neural folds grow over midline and fuse to form neural tube
- neural tube run in cranial to caudal axis
derivatives:
- CNS is derived from wall of the neural tube
- ventricles & spinal cord are derived from the hollow lumen
- peripheral nervous system is derived from the neural crest
Remember:
- neural tube comprises of a bundle of nerve sheath which closes to form brain at the anterior end and spinal cord at
posterior end
- closure should occur at around the 28th day of conception failing which brain or spinal cord doesn't form properly31
Neural tube defects
Neural tube defects
Neural Tube Defects:

Anencephaly:
- malformation in the anterior end of neural tube
- cerebrum and cerebellum are reduced or absent,
- but the hindbrain is present
- associated with fetal loss, stillbirth, or neonatal
death
- polyhydramnios in mother due to absence of fetal
swallowing of amniotic fluid
34
Neural tube defects
Causes:
- folic acid deficiency (others: Methotrexate, Valproic acid)
- maternal folic acid levels must be adequate before pregnancy to prevent an open
NTD
- also, exposure to agents that interfere with normal folate metabolism during the
- critical period of neural tube development (up to 6 weeks after last menstrual
period) increases the risk of neural tube defect
Screening:
- maternal serum alpha-fetoprotein (MSAFP) screening during the second trimester
of pregnancy (open neuropore leaks CSF proteins like AFP into amniotic sac where
AFP crosses placenta into maternal blood)
- Acetyl cholinesterase can also be increased in amniotic fluid
35
Posterior Fossa anomalies: Dandy-Walker malformation
Introduction:
- partial or complete absence of cerebellar vermis
- this results in cystic dilatation of the 4th ventricle, with
absent cerebellum
- enlargement of the posterior fossa (contains the
brainstem and cerebellum) with a high attachment of
the tentorium cerebelli
- (triad: agenesis of cerebellar vermis, cystic dilatation of
4th ventricle, and enlarged posterior fossa)
Posterior Fossa anomalies: Dandy-Walker malformation
Pathogenesis:
- congenital: failure of 4th ventricle to open
- produces non communicating hydrocephalus
- intra-uterine infection: TORCH
Clinical Features:
- ataxic gait
Dandy-Walker syndrome: a cyst-like structure, representing the greatly dilated fourth
ventricle, expands in the midline, causing the occipital bone to bulge posteriorly and
displace the tentorium and torcula upward.
38
Arnold Chiari Malformation
Type I Type II Type III
(Chiari I malformation) (Arnold Chiari malformation) (Chiari III malformation)
Definition - Ectopic extension of cerebellar - Protrusion of cerebellar tonsils, - Herniation of parts of

tonsils thru’ foramen Magnum Vermis, and Medulla oblangata cerebellum and brainstem
thru’ foramen Magnum through an abnormal
opening in the back of the
skull ( encephalocele)
Clinical Features - Usually asymptomatic in - Usually asymptomatic in infancy - Mostly fatal in infancy
adolescence/young - Obstructive hydrocephalus (brain - If infants srvive
- Occipital headache, neck pain shifts into foramen Magnum a) Upper and lower neuron
- Radicular pain in shoulder and arms resulting in stenosis of cerebral palsies
- Cerebellar symptoms: ataxia, aqueduct) b) Cranial nerve disorders
dyarthria - Breathing difficulty c) Cerebellar symptoms
- Downbeat nystagmus: eyes tend to - Always associated with d) Epilepsy
drift upwards, followed by a rapid myelomeningocele (motor and e) Intellectual disability
downward movement sensory deficit below the lesion)
- Some develop Syringomyelia
Radioimaging of Brain - Caudal displacement of cerebellar - Features similar to Chiari I - Features similar to Chiari I
tonsils - In addition: caudal displacement - In addition: caudally
- Small posterior cranial fossa of beaked dorsal midbrain displaced cerebellum, and/or
medulla protrudes through
defect (encephalocele)
Syringomyelia
Introduction:
- presence of abnormal fluid filled cavity (syrinx) that develops within the central canal of
the spinal cord
- this results from disrupted CSF drainage, caused by for example: Chiari malformation
- age: 30 to 40 years 9spinal cord injury related)
Causes:
- Arnold-Chiari malformation
- post traumatic
- post inflammatory (transverse myelitis)
- post infectious Imenigitis)
- intramedullary tumors (ependymoma, hemangioblastoma)
- Others: scoliosis, congenital malformations
Syringomyelia
Pathogenesis:
- there will be obstruction to the central canal of the spinal cord, usually cervical
- this leads to impaired CSF drainage formation of fluid filled cavity (syrinx)
compression of the anterior white commissure with damage (reactive gliosis)
affecting crossing neural fibers of the lateral spino-thalamic tract (therefore
affects pain and temperature) bilateral dissociated sensory loss and dysesthetic
pain
- expansion of the syrinx may damage: lower motor neuron in the medial part of
the lateral corticospinal tract (unilateral/bilateral spastic paresis below the level
of the syrinx
Syringomyelia
- descending hypothalamic fibers in Th1 to Th4 cord segments (Horner syndrome)
- posterior column may be affected in advanced disease, loss of postion sense and
vibration in the feet
- medulla when involved: syringobulbia
Clinical Features:
- often asymptomatic, cape-like distribution
- dissociated sensory loss, dysesthetic pain
- autonomic disturbances (Horner synd)
- respiratory insufficiency
Others: Charcot joints, Syringobulbia
Tuberous Sclerosis
Introduction:
- Autosomal Dominant disorder
- associated with intellectual disability, hamartomas, benign tumors and
characteristic skin lesions
Pathological findings:
- hamartomas in CNS (cortical tubers, subendymal nodules, subendymal giant-cell
astrocytomas)
- renal angiolipomas
- retinal glial hamartomas
- pulmonary lymphangioleiomyomatosis
- cardiac rhabdomyomas
Tuberous Sclerosis
Genetic abnormality:
- tuber sclerosis locus: TSC1 in chromosome 1 (Ch 9)
encodes for ‘hamartin’
- tuberous sclerosis locus: TSC2 in chromosome 2 (Ch 16)
encodes for ‘tuberin’
- TSC2 mutation is more commonly mutated
- both TSC1 & TSC2 bind forming a complex which inhibits
mTOR (kinase)
- mTOR is a key regulator which control cell size
Tuberous Sclerosis
Morphology:
- cortical hamartomas of tuberous sclerosis are firm areas of the cortex that
- in contrast to the softer adjacent cortex, have been likened to potatoes, hence the
appellation “tubers”
Clinical Features:
- intellectual disability
- infantile spasms (children less than 3 yrs old, head bobbing, flexor spasms, seizures)
- tumors and tumor-like lesions
- skin: ‘shagreen patches’, ash leaf spots, adenoma sebaceum (angiofibroma)
- CNS: cortical hamartomas, giant cell astrocytoma
- Cardiac: rhabdomyoma
Angiofibromas Tuberous Sclerosis
Ash leaf
Shagreen patch in tuberous sclerosis complex is a

firm, bumpy plaque that is usually located on the
lower back
Spinal cord lesions
- Poliomyelitis
- Werdnig-Hoffman disease
49
Poliomyelitis
Introduction:
- causes an acute systemic viral infection leading to
- a range of manifestations which include paralysis of limb muscles, and respiratory
muscles
Pathogenesis:
- aspherical unencapsulated RNA virus (Picornaviridae). Polio type 1 causes ‘paralytic
polio’
- fecal-oral transmission, (rarely droplet transmission). Incubation period: 7 to 14 days
- first infects oropharynx, then secreted into saliva and swallowed
Poliomyelitis
- then multiplies in the intestinal mucosa,
and lymph nodes
- produces viremia, and fever
- virus gains access into cells using CD155
- in some cases, the virus invades CNS and
replicate in motor neurons of the spinal
cord (spinal poliomyelitis),or
- replicates in in brain (bulbar poliomyelitis)
Poliomyelitis
Morphology:
- mononuclear inflammation around blood vessels (perivascular cuffing)
- neuronophagia of the anterior horn cells of spinal cord
- if inflammation extends may produce cavitation
Clinical Features:
1) Poliomyelitis without CNS involvement (abortive poliomyelitis): non-specific
symptoms, complete recovery
2) Poliomyelitis with CNS involvement: a) Non paralytic, b) Paralytic
Paralytic Poliomyelitis
- asymmetric flaccid paralysis affecting most commonly leg muscles
- paralysis most severe in proximal muscles
- ascending paralysis with diaphragm involvement respiratory failure
52
Werdnig-Hoffman disease
Introduction:
- rare, type I spinal muscular atrophy (infantile)
- progressive muscular weakness starting during infancy, fatal within 2

years
- associated with a high mortality rate
Etiology and Pathogenesis:
- Autosomal recessive
- commonly a mutation is seen to involve SMN1 gene (survival motor
neuron 1), found in chromosome 5
- this leads to degeneration of the anterior horn cells
- loss of lower motor neuron: denervation atrophy of muscles, inability to

breathe
Werdnig Hoffmann: anterior horn showing
chromatolysis.
Werdnig Hoffmann: groups of muscle bundles

undergoing atrophy
Traumatic Vascular Injury
Epidural Hematoma
Subdural Hematoma
Epidural Hematoma
Introduction:
- arterial bleed in the brain creating a blood-filled
space between the inner surface of the skull and
dura
Pathogenesis:
- dura is fused with the periosteum on the internal
surface
- dural arteries; especially: middle meningeal artery
is vulnerable to with temporal fractures (in children
temporary displacement of skull)
Epidural Hematoma
- following tear of middle meningeal artery, blood extravasate under arterial
pressure into epidural space (potential space) leading to dural separation
- most occur in temporal/tempero-parietal region
- when blood accumulates slowly patient may show ‘lucid interval’, compression of
anterior temporal lobe
Radiology:
- typically biconvex in shape and can cause a mass effect with herniation
Epidural Hematoma
Clinical Features:
- initial loss of consciousness
- temporary recovery of consciousness with return to normal/ner-normal
neurological function: Lucid interval
- followed by decline in neurological function and onset of symptoms due to
expanding hematoma
Epidural Hematoma
Subdural Hematoma
Introduction:
- venous bleeding between the dura and the arachnoid membranes
Classification:
1) Acute Subdural Hematoma: onset of symptoms within 3 days following the
event
2) Subacute Subdural Hematoma: onset of symptoms 4 to 20 days following the
event
3) Chronic Subdural Hematoma: onset of symptoms 21 days or more following the
event
Subdural Hematoma
Etiology:
1) Acute Subdural Hematoma
- most commonly caused by blunt head trauma
- non-accidental: “shaken baby syndrome” (child abuse)
- acceleration-deceleration injury (rupture of bridging veins)
2) Chronic Subdural Hematoma
- in adults: trauma from falls, seen with old age and alcohol use
- non-traumatic: cerebral atrophy associated with advanced age
Subdural Hematoma
Pathogenesis:
- subdural space: lies between the inner
surface of the dura mater and the outer
arachnoid layer of leptomeninges
- ”bridging veins” travel from the
convexities of the cerebral hemispheres
thru’ the subdural space
- to empty into superior sagittal sinus
Subdural Hematoma
Morphology:
- typically venous bleed is self-limited
- therefore, breakdown and organization of hematoma happens
1) lysis of the clot: 1 week
2) fibroblast proliferation from dural surface into hematoma: 2 weeks
3) early development of hyalinized fibrous tissue: 1 to 3 months
-repeat bleed may happen, as there are thin-walled blood vessels during
granulation tissue formation
Subdural Hematoma
Radiology:
- frontoparietal convexities and the middle
cranial fossa (can cross cranial sutures)
- typically crescentic shaped, concave
- does not cross mid-line
Subdural Hematoma
Clinical Features:
- lucid interval between time of injury and symptoms may be seen
- impaired consciousness and confusion
- focal neurological deficits
- headache, vomiting
The normal brain in this section (i.e., without
subdural hemorrhage) has been mildly
retracted posteriorly during removal,
highlighting the bridging veins
dura has been reflected back (with a small portion

visible at the lower right) 66
“fear is a disease that eats away at logic and makes man inhuman”
Marian Anderson, singer (1897-1993)
CNS Pathology
Part 2
Dr. Roy
Cellular Pathology of CNS
Reactions of neurons to injury (SDL)
Acute process
- as a consequence of depletion of oxygen/glucose/trauma
Chronic process
- slower process as a consequence of accumulation of abnormal protein
aggregates (degenerative disorders)
- Neurons are maintained for life span of an individual

- so protein turnover have be well regulated to ensure cellular integrity
Cellular Pathology of CNS
Reactions of neurons to injury (SDL)
Morphology:
Acute neuronal injury (red neurons)
- following CNS hypoxia/ischemia changes occur in neurons
- the earliest evidence of injury is: “red neurons”
- these are seen by about 12 to 24 hours after a hypoxic/ischemia insult
- red neurons show shrinkage of cell body, pyknosis of nucleus, loss of Nissl
substance, with intense cytoplasmic eosinophilia
Red neurons
Gliosis
Gliosis:
- gliosis is the most important histopathologic indicator of CNS injury
Cerebrovascular disease (CVA)
Definition:
- abrupt onset of focal or global neurological symptoms
- due to ischemia or hemorrhage
- symptoms must continue for more than 24 hrs, to be defined as “stroke”
- if symptoms resolve within 24 hrs, this episode will be referred to as “TIA”

Transient Ischemic Attack: less than 24 hours. Risk of full-fledged stroke in the next 3 months following a TIA.
Amaurosis fugax: transient monocular blindness occurs from emboli from to central retinal artery of one eye
Incidence:
- incidence of cerebrovascular diseases increases with age
5
Ischemic stroke Intracerebral h’age Subarachnoid h’age
Incidence - About 85% - About 10% - About 5%
Causes - Embolism - Ruptured cerebral artery or micro- - Ruptured berry aneurysm

- Thrombosis aneurysm - AV malformation
- Small vessel occlusion - Trauma
(lipohyalinosis) - Reperfusion injury after ischemic
- Systemic hypoperfusison stroke
Risk Factors - Age above 65 years - Age above 65 years - Hypertension

- Hypertension - Hypertension - Smoking
- Diabetes mellitus - Vasculitis - Family history
- Atrial fibrillation - Malignancy
- Carotid artery stenosis - Ischemic stroke
Pathology - Pale infarct liquefactive necrosis glial - Hematoma surrounded by pale - Hematoma surrounded by pale
scarring infarct and edema infarct and edema
- Hemosiderin lined cavity with glial - Hemosiderin lined cavity with
scarring glial scarring
CVA: classification
1) Ischemia (insufficient cerebral blood flow)
a. Global cerebral ischemia
b. Focal cerebral ischemia: TIA, Stroke
2) Hemorrhage (cerebral infarction due to hemorrhage)

a. Intracerebral hemorrhage (bleed within brain parenchyma)
b. Subarachnoid hemorrhage (bleed into subarachnoid space)
7
Ischemia: Global cerebral ischemia
Introduction:
- generalized reduction of cerebral perfusion
Etiology:
- cardiac arrest
- hypovolemic shock
- hypoglycemia (example: Insulinoma)
- carbon monoxide poisoning (chronic)
Remember: in all of these causes, there is reduced perfusion/oxygenation
8
Pathogenesis:
- depends on the severity (mild, moderate, or severe)
Mild cases
- post-ischemic confusional state
- followed by complete recovery
- no irreversible tissue damage
Hierarchy of severity: Neurons are the most sensitive, the most sensitive neurons are those in the
pyramidal layer of the Hippocampus(Sommer sector). Then comes the cerebellar Purkinje cells, and
pyramidal cells in the cerebral cortex
9
Severe cases
- widespread neuronal death occurs
- irrespective of regional vulnerability
- vegetative state in those who survive, others go into “brain death”. (brain
death is characterized isoelectric or flat EEG)
Border zone infarcts (watershed areas):
- border zone between anterior and middle cerebral artery
- produces a sickle shape band of necrosis over the cerebral convexity
Remember: border zone infarcts are usually seen after hypotensive episodes
10
Classic watershed infarct with secondary hemorrhagic
transformation (arrow); boundary between anterior and
middle cerebral artery circulations
Ischemia: Focal cerebral ischemia
Introduction:
- reduction or complete cessation of blood flow to a localized area of the brain
- due to arterial occlusion or low perfusion
- examples: TIA, and Stroke
Etiology:
1) Embolization
- cardiac mural thrombi (myocardial infarct, valvular disease, and atrial fibrillation)
- thromboembolism originating from arteries (atheromatous plaques within the
carotid arteries)
- paradoxical emboli (moistly children with cardiac anomalies)
most common territory: middle cerebral artery
13
Etiology (cont’d):
2) Thrombotic occlusion
- atherosclerosis and plaque rupture (carotid bifurcation, the origin of the middle cerebral artery,
and either end of the basilar artery)
- Inflammation (infectious vasculitis of small and large vessels seen with syphilis and tuberculosis,
or immunosuppression leading to opportunistic infection e.g., aspergillosis or CMV encephalitis)
Pathogenesis:
- with sustained ischemia, infarction would be seen in the territory of the compromised vessel
- size, location, and shape of the infarct and the extent of tissue damage would depend on :
a)duration of the ischemia, b) adequacy of collateral flow
14
Morphology:
- infarcts can be hemorrhagic, or non-hemorrhagic
- usually infarcts begin with loss of blood supply, therefore it would initially non-
hemorrhagic
- secondary hemorrhage happens with ischemia-reperfusion injury, (due to
through collaterals or following dissolution or fragmentation of the intravascular
occlusive material)
- secondary hemorrhages are petechial in nature, and may be multiple or even
confluent
Remember: clinical management of patients with these two types of infarcts differs
greatly as thrombolytic therapy is contraindicated in hemorrhagic infarcts
15
Morphology:
Non-hemorrhagic
- no major changes noted during the first 6 hours of irreversible injury
- by 48 hours, tissue becomes pale, soft, and swollen, and
- the cortico-medullary junction distinction is lost
- between 2 to 10 days, brain becomes gelatinous and friable, and
- the previously ill-defined boundary between normal and infarcted tissue
becomes more distinct as edema resolves in the viable adjacent tissue
- from 10 days to 3 weeks, the tissue liquefies, eventually leaving a fluid-filled
cavity that continues to expand until all of the dead tissue is removed
16
Cerebral infarcts. (A) Acute ischemic injury causes diffuse eosinophilia of neurons, which are beginning to
shrink. (B) After about 10 days, the lesion is characterized by the presence of foamy macrophages (best
seen on the left) and adjacent reactive gliosis with neovascularization (on the right). (C) Remote small
infarct is seen as an area of tissue loss surrounded by residual gliosis.
Morphology:
Hemorrhagic
- these are similar to non-hemorrhagic type
- however, there is blood extravasation and resorption
- patients receiving anticoagulant treatment, hemorrhagic infarcts may be
associated with extensive intracerebral hematomas
- venous infarcts are often hemorrhagic as seen with thrombotic occlusion of the
superior sagittal sinus
- malignancies, localized infections, and other conditions leading to a
hypercoagulable state increase the risk for venous thrombosis
18
Hemorrhagic infarct
Non-hemorrhagic infarct
Clinical Features:
- neurologic deficits produced by infarction are determined by the anatomic
distribution of the damage, rather than the underlying cause
- neurologic symptoms referable to the area of injury often develop rapidly, over
minutes, and may continue to evolve over hours
- as strokes are frequently associated with cardiovascular disease, many of the
genetic and lifestyle risk factors are similar
20
Hypertensive cerebrovascular disease
Morphology:
- two most common types are 1) Lacunar infarcts, 2) Slit hemorrhages, and 3)
Hypertensive encephalopathy
Lacunar Infarcts:
- involves the deep penetrating arteries and arterioles that supply the basal ganglia
and
- hemispheric white matter as well as the brainstem
- these blood vessels show narrowing (small vessels: arteriolosclerosis)
- lacunar infarcts are seen in hypertension, and diabetes
Lacunar infarct
- small vessels
- arteriolosclerosis, lipohyalinosis
Morphology:
Lacunar infarct (cont’d):
- lacunar infarct appear as single or multiple, small, cavitary infarcts known as ‘lacunes’
(lake-like measuring less than 15 mm wide)
- characteristically seen in the lenticular nucleus, thalamus, internal capsule, deep white
matter, caudate nucleus, and pons
Microscopy:
- lipohyalinosis, tissue loss surrounded by gliosis
- état criblé: affected vessels may also be associated with widening of the perivascular
spaces without tissue infarction
Clinical Features:
- may remain lacunae silent or cause severe neurologic impairment
Slit hemorrhages:
- associated with rupture of the small-caliber penetrating vessels and the
development of small hemorrhages
- with time these hemorrhages resorb, leaving behind a slit-like cavity (slit
hemorrhage)
- surrounded by brownish discoloration
Microscopic examination:
- slit hemorrhages show focal tissue destruction, pigment-laden macrophages, and
gliosis
Hypertensive Encephalopathy:
- clinicopathologic syndrome seen in the setting of malignant hypertension
- characterized by diffuse cerebral dysfunction, including headaches, confusion,
vomiting, and convulsions, sometimes leading to coma
- autopsy examination show an edematous brain with or without transtentorial or
tonsillar herniation
Microscopy:
- show petechiae and fibrinoid necrosis of arterioles in the gray and white matter
Clinical correlation:
Vascular Multi-infarct dementia:
- patients may suffer multiple, bilateral, gray matter and white matter infarcts may
develop a distinctive clinical syndrome
- characterized by dementia, gait abnormalities, and pseudobulbar signs, often
with superimposed focal neurologic deficits
- prolonged and severe cerebral ischemia
- risk factors: advanced age, chronic HTN, DM, obesity, prior history of stroke
Clinical correlation:
Binswanger syndrome:
- when there is an injury which preferentially involves large areas of the subcortical
white matter with myelin and axon loss, the disorder is referred to as Binswanger
syndrome
- not well-defined
Hemorrhage
Types:
- intracerebral hemorrhage and subarachnoid hemorrhage
Intracerebral hemorrhage
Introduction:
- rupture of small intraparenchymal vessels results in a hemorrhage within the brain
- hemorrhages which is located in the basal ganglia and thalamus are referred to as “ganglionic”
hemorrhages
- hemorrhages which is located in the lobes of cerebral hemispheres are referred to as “lobar”
hemorrhages
Etiology:
- hypertension (most important cause)
- cerebral amyloid angiopathy (Alzheimer disease)
28
Hemorrhage: Intracerebral hemorrhage
Pathogenesis:
Hypertension:
- causes vessel wall abnormalities, such as
- accelerated atherosclerosis in larger arteries, and hyaline arteriolosclerosis in smaller arteries
- there is weakening of the wall of the affected blood vessels
- therefore, vulnerable to rupture
Charcot-Bouchard microaneurysms:
- penetrating branches of the middle cerebral artery (lenticulostriate branches). These measure less
than 300 um in diameter (basal ganglia mostly)
- pressure-induced rupture causes bleeding in the putamen, thalamus, adjacent white matter, pons,
and cerebellum
- associated with chronic hypertension
29
Hemorrhage: Intracerebral hemorrhage
Morphology:
- common sites: putamen, thalamus
- extravasation of blood with compression of adjacent parenchyma
- old hemorrhages show cavitary destruction of brain
- rimmed by brownish discoloration
- later, hemosiderin laden macrophages appear
- surrounding gliotic reaction noted
cerebral amyloid angiopathy
- associated with Alzheimer disease
- leptomeningeal and cerebral cortical arterioles and capillaries
- dense and uniform deposits of amyloid seen 30
A) Massive hypertensive ganglionic
hemorrhage rupturing into a lateral
ventricle.
B) Hyaline arteriolosclerosis (fibrosis
and thickening of the arteriolar
walls) develops in the basal ganglia
and subcortical white matter of
patients with long-standing
hypertension; it is a risk factor for
hypertensive hemorrhages as well as
lacunar infarcts.
C) Large lobar hemorrhage due to
cerebral amyloid angiopathy; it
focally dissects into the
subarachnoid space. (D) Amyloid
deposition in cortical arterioles in
cerebral amyloid angiopathy; inset,
immunohistochemical staining for
Aβ highlights the deposited material
in the vessel wall.
Hemorrhage: Subarachnoid hemorrhage
Subarachnoid hemorrhage (SAH)

Introduction:
- rupture of sacular (berry) aneurysm
- bleed occur into the subarachnoid space
- most berry aneurysms are seen near major arterial branch points in anterior circulation
- rupture is into the subarachnoid space in the basal cisterns and often into the
parenchyma of the adjacent brain
- structural anomaly noted in berry aneurysm
- there is absence smooth muscle and intimal elastic lamina
- this anomaly is mostly seen to occur along branching points
32
Etiology and Pathogenesis (cont’d):

- may be linked to genetic factors
- autosomal polycystic kidney disease (ADPKD)
- Ehlers-Danlos synd (type IV)
- Marfan syndrome
- neurofibromatosis (NF-1)
- predisposing factors: smoking, hypertension
Remember: although berry aneurysms are sometimes referred to as congenital, they are not
present at birth. But develop over time due to an underlying defect in the media of the blood
vessel
33
ADPKD Polycystic kidney.
34
Morphology:
Gross:
- when un-ruptured, berry aneurysm appears as an outpouching
- seen at branching points along the circle of Willis
- range from few millimeters to 2 to 3 cm, translucent wall
Microscopy:
- smooth muscle and intimal elastic lamina do not extend into the neck of the
aneurysm
35
Subarachnoid hemorrhage (Berry aneurysm):
Note the absence (thinning) of media.
Image below: special stain (Trichrome), to highlight the
thinning.
36
Clinical Features:
- “worst headache of my life”
- sudden loss of consciousness may be preceded by
a brief moment of excruciating headache
- headache is usually generalized, often with neck
stiffness, and vomiting is common
- Kernig sign, Brudzinski sign
- prodromal symptoms: seen in 30 to 50%, may
precede days to weeks prior
37
Clinical Features:
- 25% to 50% patients die with first rupture
- lysis of the red blood cells and subsequent conversion of hemoglobin to bilirubin
stains the spinal fluid yellow within 6–12 hrs
- this xanthochromic spinal fluid peaks in intensity at 48 h and lasts for 1–4 weeks,
depending on the amount of subarachnoid blood
38
Subarachnoid hemorrhage
39
Subarachnoid hemorrhage: noted on the base of
the brain
40
“till round the corner there may wait,/ a new road or a secret gate”
J.R.R. Tolkien
41
CNS Pathology, Part 3
Demyelinating disorders
Degenerative disorders
Dr. A. Roy
Leukodystrophies
- Krabbe disease
- Metachromatic leukodystrophy
- Adrenoleukodystrophy
Leukodystrophies
Introduction:
- are genetic metabolic disorders of the white matter that affect glial cells
- myelin may be absent or decreased, of an abnormal structure, and unstable
Examples:
Krabbe Disease
Metachromatic Leukodystrophy
Adrenoleukodystrophy
3
Krabbe disease
- autosomal recessive disorder
- deficiency of galactocerebroside β-galactosidase (galactosylceramidase), chromosome 14q31 mutations
- this enzyme is required for the catabolism of galactocerebroside to ceramide and galactose
- absence of enzyme activity leads to accumulation of galactosylsphingosine, which is cytotoxic
- brain shows loss of myelin and oligodendrocytes
Histology:
- presence of ‘globoid ‘ cells in brain
Clinical Features:
- rapidly progressive, predominant motor signs such as weakness
- normal at birth, develop irritability, spasticity around 3 to 6 months of age. Usually die in two or three years
4
Krabbe disease: globoid cells. Globoid cells are hisiocytes filled with
galactocerebroside.
5
Metachromatic Leukodystrophy
- autosomal recessive disorder
- deficiency of lysosomal enzyme: arylsulfatase A (chromosome 22q)
- this enzyme cleaves sulfate from sulfate containing lipids (sulfatides)
- deficiency leads to accumulation of sulfatides (cerebroside sulfate)
- sulfatides causes white matter injury
Histology:
- macrophages contain membrane bound vacuoles which shows crystalloid structures
- these show ‘metachromasia’ when stained with Toluidine blue
6
Adrenoleukodystrophy
Introduction:
- X-linked recessive disorder
- mutation in a member of the ATP-binding cassette transporter family of proteins:
ABCD1
- inability to catabolize very long chain fatty acids, leading to these to be elevated
in serum
- leading to progressive loss of myelin in CNS and peripheral nerves
- adrenal insufficiency associated with adrenal atrophy
Clinical Features:
- childhood type: muscle spasms, gait abnormality strabismus 7
Demyelinating Disease
- Multiple Sclerosis
- Gullian-Barre Syndrome
- Progressive Multifocal Leukoencephalopathy
- Charcot-Marie-Tooth disease
- Metachromatic Leukodystrophy
- Krabbe’s Disease
Multiple sclerosis
Introduction:
“autoimmune demyelinating disorder characterized by distinct episodes of
neurologic deficits , separated in time, attributable to white matter lesions
that are separated in space”
Incidence:
- highest reported: Orkney Islands (Scotland)
- more in temperate than tropical
- more in females
- 20 to 40 years
9
Multiple sclerosis: Pathogenesis
Genetic
- MS is caused by an autoimmune response directed against components of the
myelin sheath
- involves genetic and environmental factors
- has a 15-fold higher when the disease is present in a first-degree relative
- some cases show association with a DR haplotype of the major histocompatibility
complex
- defects with the IL-2 and IL-7 receptor genes are also reported, this leads to
immune dusregulation
10
Multiple sclerosis: Pathogenesis
Environmental
- geographic variation is noted, with most cases diagnosed away from the equator
- low level of vitamin D (immune system modulator) in people who are not
exposed to sunlight during winter months
Immune
- immune response directed against components of myelin sheaths
- initiated by TH1 and TH17 T-cells that react against myelin antigens and secrete
cytokines
- TH1 cells also secrete IFN-γ, which activates macrophages, and TH17 cells promote
the recruitment of leukocytes
11
Multiple sclerosis: Morphology
Gross:
- MS is a white matter disease that is best seen in sections of the brain and spinal cord
- MS lesions are firmer than the surrounding white matter (sclerosis) and
- appear as well circumscribed, somewhat depressed, glassy, gray-tan, irregularly
shaped plaques
- area of demyelination often has sharply defined borders, which is best seen when
stained with stains for myelin
- sites where plaques commonly occur: adjacent to the lateral ventricles, optic nerves
and chiasm, brainstem, ascending and descending fiber tracts, cerebellum, and spinal
cord
12
Multiple sclerosis: Morphology. Microscopy
Active plaques:
- shows ongoing myelin breakdown
associated with abundant
macrophages containing lipid-rich,
PAS-positive debris
- also seen are lymphocytes and
monocytes, mostly as perivascular
cuffs, especially at the outer edge of
the lesion
- within a plaque there is relative
preservation of axons
- and depletion of oligodendrocytes Active plaque: myelin stain
Multiple sclerosis: Morphology. Microscopy
Inactive plaques:
- with time lesions inflammatory cells slowly disappear
- within inactive plaques, little to no myelin is found, and
- there is a reduction in the number of oligodendrocyte nuclei
- astrocytic proliferation and gliosis are prominent
- axons in old gliotic plaques show severe demyelination and are
also greatly diminished in number
Shadow plaque: in some cases of MS, the border between

normal and affected white matter is not sharply circumscribed
Inactive plaque:
Multiple sclerosis: Clinical Features
- unilateral visual impairment due to involvement of the optic nerve
(optic neuritis, retrobulbar neuritis) is a frequent initial manifestation
of MS
- MS affecting brainstem produces cranial nerve signs, ataxia,
nystagmus, and internuclear ophthalmoplegia from interruption of
the fibers of the medial longitudinal fasciculus
- MS affecting spinal cord lesions give rise to motor and sensory
impairment of trunk and limbs, spasticity, and difficulties with the
voluntary control of bladder function
Multiple Sclerosis
Multiple sclerosis: Clinical Features
CSF Findings:
- MS patients shows a mildly elevated protein
level
- in one third of cases a moderate pleocytosis
(increased cell count in fluids)
- IgG levels in the CSF are increased and
oligoclonal IgG bands are usually observed
on immuno-electrophoresis
- (these are indicative of the presence of a
small number of activated B cell clones,
postulated to be self-reactive, in the CNS)
Multiple Sclerosis: internuclear opthalmoplegia 18
Progressive Multifocal Leukoencephalopathy
Introduction:
- type of encephalitis caused by JC polyoma virus
- virus preferentially infects oligodendrocytes
- clinically characterized by demyelination
- seen exclusively in immunosuppressed patients
Morphology:
- irregular, ill-defined white matter injury
- on microscopy, lesions reveal demyelination
Clinical Features:
- visual loss, weakness, high mortality
19
Central Pontine Myelinolysis
Introduction:
- acute disorder characterized by loss of myelin in basis pontis and portion of the pontine
tegmentum
- myelin loss appears to be symmetrical
Pathogenesis:
- usually follows 2 to 6 days after rapid correction of hyponatremia (osmotic demyelination)
- mechanism is not well established
Clinical Features:
- rapidly evolving quadriplegia
- “locked in” syndrome (only eye movement is preserved)
20
Degenerative Diseases
Dementia
- there is gradual decline in cognition
- level of consciousbess is preserved
- memory loss, impaired judgemnet, and
personality changes
- irreversible
- 3 important findings: Aphasia, Apraxia, Agnosia
Degenerative diseases
Definition:
“an acquired deterioration in cognitive abilities that impairs the successful performance of activities
of daily living”
- memory is the most commonest cognitive ability lost with dementia
Degenerative disease affecting the cerebral cortex
1) Alzheimer disease
Frontotemporal dementia (affecting language, and personality. Involves temporal and frontal)
- Frontotemporal dementia with Parkinsonism linked to tau mutations
- Pick disease
Degenerative disease of Basal ganglia & brainstem
1) Parkinson disease
2) Dementia with Lewy body 22
Disease Clinical pattern Inclusions Genetic cause
Alzheimer disease Dementia Aβ (plaques), Tau APP, PS1, PS2

(tangles)
Frontotemporal lobar Behavioral changes, Tau Tau
degeneration language disturbance
Parkinson disease Hypokinetic movement α-synuclein α-synuclein
disorder
Progressive supranuclear Parkinsonism with Tau Tau
palsy abnormal eye movements
Huntington disease (HD) Hyperkinetic movement Huntington Htt
disorder (polyglutamine)
Amyotrophic lateral Weakness with upper and SOD1 SOD1
sclerosis (ALS) lower motor neurons
signs
Alzheimer disease
Introduction:
- most common cause of dementia in older adults (others: African Americans, Family history,
obesity, diabetes, TBI)
Patterns:
Sporadic:
- with aging
- epsilon 4 allele
- which promotes Aβ generation
Early Forms:
a) Familial: associated with presenilin 1
b) Down’s syndrome: remember chromosome 21 houses the Aβ (there will be 3 copies of chromosome
21 in Down’s syndrome)
25
Pathogenesis
Alzheimer disease
Pathogenesis:
- major abnormality in Alzheimer is
- is the accumulation of two proteins (Aβ and tau) in specific brain regions,
- possibly due to excessive production and defective removal of Aβ peptides
Plaques:
- these are deposits of aggregated Aβ peptides in the neuropil
Tangles:
- these are aggregates of the microtubule binding protein tau, which develop
intracellularly and then persist extracellularly after neuronal death
27
Alzheimer disease
Pathogenesis:
- neural dysfunction are related to plaques and tangles, and
- also the mechanisms that lead to the accumulation of these abnormal
aggregates
Formation of Aβ
- Amyloid Precursor Protein (APP) is a cell surface protein with a single
transmembrane domain that may function as a receptor (possibly for prion
protein (PrPc) among other ligands)
- Aβ portion of the protein extends from the extracellular region into the
transmembrane domain
28
Alzheimer disease
Pathogenesis:
- processing of APP begins with cleavage in the extracellular domain
- followed by an intramembranous cleavage
- here are two potential pathways, determined by the type of initial
proteolytic event
1) if the first cut occurs at the α-secretase site within the Aβ sequence, then
Aβ is not generated (the non-amyloidogenic pathway)
2) if the surface APP endocytosed, it may undergo cleavage by β-secretase,
which cuts at the N-terminal region of the Aβ sequence (the
amyloidogenic pathway)
29
Alzheimer disease
Pathogenesis:
- after cleavage of APP at either of these sites
- the γ-secretase complex performs an intramembranous cleavage
- if paired with a first cut by α-secretase, it produces a soluble fragment,
- but if paired with β-secretase cleavage, it generates Aβ
- when formed, Aβ is highly prone to aggregation: first into small oligomers
(which may be the toxic form responsible for neuronal dysfunction), and
- eventually into large aggregates and fibrils
30
Alzheimer disease
Pathogenesis:
Familial Alzheimer Disease
- there are two loci associated with majority of early-onset familial AD encode the
two presenilins:
PS1 on chromosome 14
PS2 on chromosome 1
- these mutations lead to a gain of function, such that the γ-secretase complex
generates increased amounts of Aβ
31
Alzheimer disease
Pathogenesis:
Formation of Tau protein:
- Tau is a microtubule-associated protein which is present in axons
- with the development of tangles in Alzheimer Disease, it shifts to a somatic-
dendritic distribution
- becomes hyperphosphorylated, and loses the ability to bind to microtubules
- formation of tangles is an important component of Alzheimer Disease
- increased tangle burden in the brain over the course of the illness eventually
appears to become independent of the Aβ
32
Alzheimer disease
Pathogenesis:
Other genetic risk factors
- apolipoprotein E (ApoE) is a risk factor of the development of Alzheimer
- of all the alleles, ε4 allele increases the risk of Alzheimer, and lowers the age of
onset of the disease
Role of inflammation
- deposits of Aβ elicit an inflammatory response from microglia and astrocytes
- these responses help in the clearance of the aggregated peptide, but may also
stimulate the secretion of mediators that cause damage
- inflammation also produces free radicals (ROS)
33
Alzheimer disease
Morphology:
Gross:
- cortical atrophy
- widening of sulci (frontal, temporal, parietal)
- compensatory ventricular enlargement (hydrocephalus ex vacuo)
Micro:
Neuritic plaques (senile plaques):
- focal, spherical collection of dilated, tortuous neuritic processes
- seen around a central amyloid core (stain positive for Congo Red). Aβ Amyloid is seen as
extracellular material
34
Alzheimer disease
Morphology (micro cont’d):
Neurofibrillary tangles:
- twisted neurofilaments in neuronal cytoplasm
- best seen on silver stains
- contains tau protein (microtubule associated protein)
- in Alzheimer tau protein is hyperphosphorylated
- leading to neurofibrillary tangles
Cerebral amyloid angiopathy (CAA):
Alzheimer Disease: neurofibrillary tangles
- accumulation of Aβ amyloid in wall of cerebral
arterioles (Congo Red positive)
- this may lead to intracerebral hemorrhage
35
Alzheimer’s disease
Granulovacuolar
degeneration
Morphology (micro cont’d):
Granulovacuolar degeneration:
- clear intraneuronal cytoplasmic
vacuolation
Hirano bodies:
- elongated, glassy, eosinophilic bodies
found in hippocampal pyramidal cells
Loss of cholinergic neurons:
- nucleus basilis of Meynert and
hippocampus
Hirano bodies 36
Alzheimer Disease
Morphology; Gross
Alzheimer Disease
- neuritic plaque
Alzheimer Disease
- neurofibrillary tangles
Alzheimer disease: Clinical Features
- based on a point system
- more or equals 27 out of 30 is Normal
- oriented to time and space
- repeat 3 items, and remember them
- serials of 7, or spell WORLD backwards
- name an item pointed out
- repeat a phrase
- draw an object drawn
Alzheimer disease
Clinical features:
- initial symptoms are forgetfulness and other memory disturbances
- as disease progresses other symptoms occur: language deficits, loss of
mathematical skills, and loss of learned motor skills
- final stages of Alzheimer Disease, affected individuals may become incontinent,
mute, and unable to walk
- intercurrent infections, mostly pneumonia, is usually the terminal event
40
Dementia causes
- Alzheimer disease
- multi-infarct dementia (stroke)
- Lewy body dementia
- Pick’s disease
- Creutzfeldt-Jakob disease
- HIV
Parkinson disease
Introduction:
- progressive neurodegenerative disease
- cardinal features: rest tremor, rigidity, bradykinesia, and gait impairment (masked
facies)
- usually seen after the age of 60 years
Pathogenesis:
1. degeneration of dopaminergic neurons in the substantia nigra pars compacta,
2. reduced striatal dopamine, and
3. intracytoplasmic proteinaceous inclusions known as Lewy bodies
(MTTP: methyl phenyl tetrahydropyridine, a contaminant of opioid drugs may cause
loss of dopaminergic neurons)
42
Parkinson disease
Morphology:
Gross:
- pallor of substantia nigra and locus ceruleus
Micro:
- loss of pigmented neurons in substantia nigra
- associated with gliosis
- presence of Lewy body
- Lewy bodies: composed of fine filaments, densely
packed in the core (composed of α-synuclein) 43
Parkinson’s disease
Clinical features:
- Motor features: bradykinesia,
hypophonia, sialorrhea, rest tremor
(pill rolling), cogwheel rigidity, gait
disturbance
- Non-motor features: depression,
anxiety, cognitive impairment, sleep
disturbances, restless legs, and rapid
eye movements
44
Huntington disease
Introduction:
- autosomal dominant
- progressive movement disorder dementia
- histologically shows degeneration of striatal neurons (GABAnergic neurons)
Molecular genetics:
- expanded repeats of of trinucleotide repeats (CAG)
- gene HD, HTT located on chromosome 4 encodes for huntingtin
- normal HD genes contain 6 to 35 copies of the repeat
- when the repeats expand, disease sets in earlier (seen with spermatogenesis)
- therefore with each following generation the disease would manifest at an earlier age than
the preceding
45
Huntington disease
- loss of medium spiny striatal neurons
- causes dysregulation of basal ganglion circuitry which modulates motor
output
- this is related to neuronal loss (cerebral cortex)
- neuron’s normal function: dampen motor activity
- in disease: increased motor output, seen as choreoathetosis
46
Huntington disease
Morphology:
- brain is small, atrophy of caudate nucleus
- secondary atrophy of globus pallidus
- secondary dilation of the lateral ventricles
Micro:
- severe loss of striatal neurons
- loss of medium spiny striatal neurons
- causes dysregulation of basal ganglion
- normal function: dampen motor activity
- in disease: increased motor output,
choreoathetosis
Huntington disease
Clinical Features:
- manifests by 4th decade of life
- involuntary jerky movements
- chorea, aggression, depression, suicidal, dementia
Amyotrophic lateral sclerosis
Introduction:
- also referred to as motor neuron disease or Lou Gehrig’s disease. Onset between
40 to 60 years.
- degenerative disease characterized by loss of upper and lower motor neurons
- leading to muscle atrophy (amyotrophy)
- the loss of cortical motor neurons results in thinning of the corticospinal tracts
that travel via the internal capsule
- loss of fibers in the lateral columns and resulting fibrillary gliosis impart a
particular firmness (lateral sclerosis)
49
Pathogenesis:
- mutation in gene encoding for copper-zinc superoxide
dismutase (SOD1)
- possible mechanisms:
1) reduced capacity to detoxify free radicals
- leads to neuronal death
2) mutated SOD1 is a misfolded protein

- therefore triggers a misfolded protein response
50
Clinical Features:
- asymmetric weakness of hands
- spasticity of arms and legs
- involuntary contractions: fasciculation
- combined upper motor neuron and lower motor neuron deficits
- no oculomotor deficits
(Remember: there is no sensory impairment. This differentiates ALS from

Syringomyelia.)
51
Friedreich ataxia
Introduction:
- autosomal recessive, affects cerebellum and spinal cord
Pathogenesis:
- expansion of unstable trinucleotide repeats (GAA) in
- frataxin
- this leads to undetectable or extremely low levels of frataxin mRNA
- causing mitochondrial dysfunction, affecting iron regulation
- iron accumulation leads to free radical injury (apoptotic cell death)
- degeneration of dorsal root ganglia, posterior column
52
Friedreich ataxia
Morphology:
Spinal cord:
- loss of axons, gliosis of posterior columns, degeneration
Heart:
- frequently manifests as hypertrophic cardiomyopathy , and pericardial adhesions
Clinical Features:
- manifests in 1st decade, gait ataxia (staggering)
- nystagmus, dysarthria, pes cavus, hammer toes, hypertrophic cardiomyopathy
- kyphoscoliosis in childhood
53
Friedreich ataxia
Kyphoscoliosis
Pes cavus
Subacute Sclerosing Panencephalitis
Introduction:
- rare progressive clinical syndrome characterized by
- cognitive decline, spasticity of limbs, and seizures
Pathogenesis:
- seen months or years after early-age acute infection with measles (altered measles virus)
- myelin degeneration and gliosis
- viral inclusions positive for measles seen on microscopy
Clinical Features:
- often fatal
55
Central Pontine Myelinolysis
“restlessness and discontent are the first necessities of progress”
Thomas A. Edison
57
CNS Pathology. Part 4.
Dr. Roy
CNS Tumors
Incidence:
- bimodal age frequency
Children: examples- medulloblastoma, pilocyctic astrocytoma
Adults: examples- glioblastoma
Childhood tumors are usually seen located beneath the tentorium
Adult tumors are usually located above the tentorium
Etiology:
- radiation
- familial: NF-1, NF-2, VHL, Li-Fraumeni syndrome, Trucot, and Gardener’s syndrome
2
CNS Tumors: Gliomas
i) Astrocytoma
ii) Oligodendroglioma
iii) Ependymoma
Astrocytomas
Introduction:
- tumors which show astrocytic differentiation
types:
i) infiltrating (Glioblastoma multiforme)
ii) non-infiltrating (pilocystic)
3
CNS Tumors: Astrocytoma
Introduction:
- astrocytic tumors are divided into Infiltrating and non infiltrating
Infiltrating (diffuse)
- accounts for 80% of adult brain tumors
- 4 tiered grading system (I to IV), grade (remember grade I & II are low-grade, grade III
& IV are high-grade)
- high grade: Glioblastoma Multiforme
Age incidence: 4th to 6th decade

4
Glioblastoma Multiforme: high grade
astrocytoma
5
Glioblastoma Multiforme
Morphology:
Gross:
- poorly defined, and infiltrative
- areas of hemorrhage and necrosis may be seen
Micro:
- increased glial cellularity
- variable nuclear pleomorphism
- network of astrocytic processes
GFAP stain: Glioblastoma
- stain positive for GFAP
Clinical features:
- headache, seizures, and focal neurologic deficit
6
CT scan: Glioblastoma: a large tumor with
central areas representing necrosis
Glioblastoma: Large mass with areas of

h’age & necrosis. Tends to cross the midline.
7
Glioblastoma: Note: necrosis in, vascular proliferation,
& cellularity.
Marked cellular & nuclear pleomorphism.
8
Non-Infiltrating: Pilocytic astrocytoma
Introduction:
- childhood tumor
-seen in cerebellum, slow growing
-grade I/IV (indicates low-grade, benign)
Morphology:
- often cystic, may be well circumscribed
- on micro: bipolar cells, with hair-like processes, Rosenthal fibers, eosinophilic granular bodies
- GFAP positive
Clinical features:
- raised ICT
- recurrence free intervals of more than 20 yrs
9
Pilocytic Astrocytoma
Pilocytic astrocytoma: Note the cystic

space bordered by reddish-brown tumor.
Fibrillary meshwork, Rosenthal fibers,

eosinophilic bodies, GFAP positive 10
CNS Tumors: Oligodendroglioma
Introduction:
- low-grade glioma resembling oligodendrocyte
- less infiltrative as compared to astrocytoma
Incidence:
- mostly adults (40 to 60 yrs)
- mostly seen in cerebral hemispheres, with predilection for white matter
Morphology:
- well circumscribed
- gelatinous with cystic spaces
- calcification (important point)
- low proliferative index 11
CNS Tumors: Oligodendroglioma
Clinical features:
- seizures, slow growing, long post-
operative survival time
- overall survival rate is good
Oligodendroglioma: Note ‘chicken-wire’ capillary

pattern, “fried egg” cells.
CNS Tumors: Ependymoma
Introduction:
- slow growing glioma, composed of neoplastic ependymal cells
- typically originates from wall of cerebral ventricles (i.e., the cells lining the
ventricles of the brain and the central canal of the spinal cord)
- chromosome 22q, which contains the neurofibromatosis 2 (NF2) gene
- ependymomas that can be completely resected, therefore are potentially curable
- partially resected ependymomas will recur and require irradiation
Location:
Children: 4th ventricle
Adults: spinal cord (myxopapillary)
13
Morphology:
- 4th ventricle ependymoma: solid or
papillary (cauliflower-like growths)
- location close to vital pontine and
medullary nuclei
Micro:
- cells show round to oval nuclei
- long dendritic processes
- perivascular pseudo-rosettes
MRI shows that the mass arising in the
fourth ventricle
14
Clinical features:
- depends on the location of the neoplasm
- headache, nausea, vomiting, or vertigo
- secondary to increased ICP
- from obstruction of CSF flow through the
fourth ventricle
- well-differentiated, slow growing
- 5 year survival rate: 45 to 50%
Ependymoma: Note tumor cells with round –oval nuclei, and

peri-vascular pseudo-rosettes
15
Medulloblastoma
Introduction:
- embryonal malignant tumor, poorly
differentiated (grade IV)
- arise from granular cells (cerebellum)
Incidence:
- 3 to 10 years
- slightly more in male child
16
CNS Tumors: Medulloblastoma
Morphology:
- midline of cerebellum
- may occlude CSF flow
- often well-circumscribed
Micro:
- sheets of tumor cells
- Homer-Wright rosettes
- seeding of the CSF is common
Note: undifferentiated (primitive cells), &

Homer-Wright rosettes. 17
CNS Tumors: Medulloblastoma
Clinical features:
- raised ICT, epilepsy, focal
neurologic deficit, grade IV,
aggressive
- children less than 3 years with
metastases have poor prognosis
- 70% of patients have long-term
survival but usually at the cost of Homer Wright rosettes: tumor cells
significant neurocognitive impairment arranged around a central clearing
containing neurofibrillary material
18
:
tumor grows from midline cerebellum obstructing CSF
pathway.
Medulloblastoma: tumor arising from

cerebellum. 19
Meningiomas
Introduction:
- benign tumors (majority)
- arising from meningothelial cell of arachnoid
- commonly located along the falx, cortical convexity, and sphenoid bone
Incidence:
- common primary CNS tumor
- mostly in females (reproductive age group)
Risk factors:
- low dose radiation
Molecular genetics:
- association with NF-2 (Chromosome 22q contains the NF2) 20
Meningiomas
Morphology:
Gross:
- rounded, encapsulated masses, has a dural base,
compress underlying brain
- en plaque: tumor spreads thinly over dural surface
Micro:
- various histological types: syncytial, fibroblastic,
transitional, psammomatous
- Psammoma bodies may be seen
Meningioma
- low risk of recurrence
21
Meningiomas
Clinical features:
- slow growing
- parasagittal aspect of brain convexity,
dura over lateral convexity, wing of
sphenoid, olfactory groove, sella
turcica, foramen magnum
- seizures, visual changes, mental status
changes, hearing loss, muscle
weakness
- surgery is best option Meningioma:
- psammoma bodies, calcification
Meningioma: note the typical location,
circumscription, compression of underlying brain.
Psammoma bodies: spherules of calcification.

Name the other tumors in which psammoma bodies are
seen? 23
Primary CNS Lymphoma (PCNSL)
Introduction:
- aggressive malignancy arising exclusively in the CNS
- often multifocal
- extra-nodal non-Hodgkin Lymphoma
- incidence is increasing, particularly in immunocompromised individuals
- Epstein-Barr virus (EBV) frequently plays an important role in the pathogenesis of HIV-
related PCNSL
Incidence:
- accounts for 1 to 2% of intracranial malignancies
Risk factors:
- congenital or acquired immunodeficiencies 24
Morphology:
- mostly multiple, and deeply situated
- tend to be well defined, but as discrete as
metastases
- periventricular spread
Micro:
- diffuse large B cell lymphoma (DLBCL)
PCNSL: monotonous appearance of the
- B-cell markers are positive (CD20, CD19) cells and their perivascular location
- EBV markers positive, in setting of
immunosuppression
25
Diagnosis:
- CSF, (commonly shows elevated protein and malignant cells)
- tissue sampling
- CBC, HIV testing
- bone marrow
Clinical features:
- focal neurologic deficits
- visual disturbances
- neuropsychiatric
- seizures
- raised ICT
- B symptoms: fever, weight loss, night sweats,
- poor response to treatment PCNSL: immunohistochemistry using
B- cell markers (CD20) 26
Craniopharyngioma
Introduction:
- hypothalamic suprasellar tumor
- derived from vestigial remnants of Rathke pouch
- usually suprasellar, but occasionally intrasellar
- slow growing benign tumors, most seen in 5 to 15 age group and second age
group 65 years and older
- children present with growth retardation (pituitary deficiency may result, from
involvement of the hypothalamus)
27
Craniopharyngioma
Morphology:
- small 3 to 4 cm diameter
- encapsulated, solid, and cystic pattern
- encroach on optic chiasm/cranial nerves
Micro:
- children: adamantinomatous (enamel, calcification)
- adults: papillary craniopharyngioma (rarely calcify)
- cysts show cholesterol rich thick brownish-yellow fluid
28
Craniopharyngioma
Clinical course:
- symptoms result from compression of adjacent
structures,
- especially the optic chiasm
- bitemporal hemianopia
- diabetes insipidus
- risk of recurrence associated with larger tumors
Sagittal MRI showing contrast- enhanced

suprasellar craniopharyngioma 29
Metastatic Tumors
Common tumors giving rise to mets:
1. Lung
2. Breast
3. Skin (melanomas)
4. Kidney
5. Gastrointestinal tract
Morphology:
- sharply demarcated masses, junction of gray & white matter
- meningeal carcinomatosis: seen with carcinoma of lung & breast
30
Metastatic Tumors
Clinical course:
- present as mass lesions
- sometimes may the
presenting feature
Meningeal carcinomatosis: note

opacities in meninges.
31
Retinoblastoma
Incidence:
- 200 to 500 cases/yr. (USA)
- 90% of cases are below 5 years of age
Molecular genetics:
- familial in 40%
- sporadic in 60%
- association with RB gene
32
Retinoblastoma: Knudson’s hypothesis
- both normal alleles of the RB locus must be inactivated (2 hits) for

retinoblastoma
Familial cases: children are born with one defective (first hit: germ cells)
and one normal copy of RB gene
- normal copy is lost thru’ somatic mutation in retinoblasts (second hit:
somatic cells)
Sporadic cases: children are born with 2 normal copies of RB gene
- these copies are lost thru’ somatic mutation occurring in one of the
retinoblasts (both hits: somatic cells)
33
“Two-hit” tumor formation in both hereditary and nonhereditary cancers. A “one-hit”
clone is a precursor to the tumor in nonhereditary cancers, whereas all cells are one-hit
clones in hereditary cancer
34
Retinoblastoma
Morphology:
- undifferentiated & differentiated elements are seen
- undifferentiated: small round cells, rosettes…
Flexner-Wintersteiner
Clinical:
- leukocoria (white pupillary reflex)
- strabismus, ocular pain
- tumor extension into brain
- involving optic nerve
35
Retinoblastoma
Clinical features (cont’d):
- treatment of retinoblastoma now largely
avoids enucleation,
- unless there is very extensive ocular
disease or regional extraocular extension,
- local obliterative therapy, supplemented if
necessary by intravenous or
- possibly intra-arterial chemotherapy, and
- avoidance of external beam radiotherapy
36
Rosettes:
- rosettes are little round groupings of cells found in tumors
- they usually consist of cells in a spoke-wheel or
- halo arrangement surrounding a central, acellular region
Homer-Wright rosette
- typically seen in neuroblastomas, medulloblastomas, and primitive neuroectodermal tumors (PNETs)
- it consists of a halo of tumor cells surrounding a central region
- containing neuropil (hence its association with tumors of neuronal origin)
Flexner-Wintersteiner rosette
- characteristic of retinoblastomas,
- it consists of tumor cells surrounding a central lumen
- that contains cytoplasmic extensions from the tumor cells
Perivascular pseudorosette
- consists of tumor cells collected around a blood vessel
- it’s called a pseudorosette because the central structure isn’t part of the tumor
- these rosettes are common in ependymomas,
- but also seen in medulloblastoma, PNET, central neurocytomas, and glioblastomas
“truth does not change according to our ability to stomach it”
Flannery O'Connor, writer (1925-1964)
4/19/21
Blood Vessel Pathology. Part 1.

Hypertension, Atherosclerosis
Dr. Roy
Introduction
- cardiovascular system (CVS) consists of the heart, major arteries,
arterioles, capillaries, venules, and veins
- these form a closed circulatory system that carries blood
- there are two major circuits that distribute blood to the body:
Systemic circulation, and Pulmonary circulation
- both these system depend on the heart as a pump
Systemic circulation
- carries blood from the heart to all the organs
Pulmonary circulation
- carries blood from the heart to the lungs for gaseous exchange
1
4/19/21
Introduction
Functions:
- gaseous exchange
- temperature control
- transport of oxygen, carbon dioxide, nutrients, hormones, metabolic
products
Types of Arteries
1. Elastic arteries: largest vessels (aorta, and pulmonary trunk). The
wall of these vessels are primarily made of elastic fibers
2. Muscular arteries: These are medium sized, and are the most
numerous. Their wall contain smooth muscle fibers
3. Arterioles: These are the smallest branches. Capillaries connect
arterioles to the smallest veins or venules
2
4/19/21
Structure of arteries
- wall of arteries contain 3 concentric layers (tunics)
- inner most facing the lumen: tunica intima, lined by simple squamous
epithelium referred to as endothelium. A small layer beneath this is
called as subendothelial connective tissue
- middle layer: tunica media, composed of primarily smooth muscle
fibers, along with elastic and reticular fibers. Also seen are collagen
fibers.
- outermost layer: tunica adventitia, composed of mostly type I
collagen fibers
* walls of some muscular arteries show two thin wavy bands of elastic
fibers. The internal elastic lamina, located between tunica intima and
media. And, the external elastic lamina, located between tunica media
and adventitia
Structure of Artery
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Structure of veins
- capillaries unite to form larger blood vessels called as: venules
- venules usually accompany arterioles
- initially blood flows through smaller “postcapillary venules” and then
into veins
- veins can be classified as: small, medium, and large
- compared to arteries, veins are more numerous, and have thinner
walls, larger diameter, and greater structural variations
- pressure within the venous system is low
- small, and medium sized veins of limbs (arms and legs), and those
that convey blood against gravity contains valves
- veins too have tunica intima, tunica media, and tunica adventitia
Structure of blood vessels

Vasa vasorum
- walls of medium, and large
arteries and veins are too thick
to provide nourishment by direct
diffusion from the lumen
- these walls are supplied by their
own smallest blood vessels from
adjacent small arteries
- these small arteries are called
as: vasa vasorum
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Diseases of Blood Vessels
1)Hypertension
2)Arteriosclerosis (Arteriolosclerosis, Monckeberg medial
sclerosis, and Atherosclerosis)
3)Aneurysm
4)Aortic dissection
Hypertensive Vascular Disease

Introduction:
- Hypertension is a leading risk factor for death and
disability, including
1) Stroke
2) Accelerated coronary and systemic atherosclerosis
3) Heart failure
4) Chronic kidney disease
5) Death from cardiovascular causes
10
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Hypertensive Vascular Disease

Guidelines:
Normal BP:
- should be below 120/80 mm Hg and
- elevated blood pressure as 120 to 129 mm Hg systolic with a diastolic
pressure below 80 mm Hg
Stage 1 hypertension:
- is defined as 130 to 139 mm Hg systolic or 80 to 89 mm Hg diastolic
Stage 2 hypertension:
- 140/90 mm Hg or higher
11
Hypertensive Vascular Disease: Guidelines
12
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Hypertensive Vascular Disease: Pathophysiology
Blood Pressure regulation:

- it is a function of cardiac output and peripheral vascular resistance
(Cardiac Output X Total Peripheral Resistance)
Cardiac output
- is: stroke volume X heart rate
- most important determinant of stroke volume is the filling pressure,
- which is regulated through sodium homeostasis and its effect on
blood volume
13

Blood Pressure regulation:
Peripheral resistance
- is regulated predominantly at the level of the arterioles by neural and
hormonal inputs
- vascular tone reflects a balance between
1) vasoconstrictors: Angiotensin II, catecholamines, and endothelin
2) vasodilators: Kinins, prostaglandins, and nitric oxide
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Renin:
- an enzyme produced by renal juxtaglomerular cells
- renin is released in response to
1) low blood pressure in afferent arterioles
2) elevated levels of circulating catecholamines
3) low sodium levels in the distal convoluted renal tubules
- (glomerular filtration rate falls when cardiac output is low leading to
increased sodium resorption by the proximal tubules)
15
- renin cleaves plasma angiotensinogen to angiotensin I,

- which in turn is converted to angiotensin II by angiotensin-converting
enzyme (ACE)
Angiotensin II raises blood pressure by
1) inducing vascular contraction (increasing peripheral resistance)
2) stimulating aldosterone secretion by the adrenal gland increasing
tubular sodium resorption
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Renin-Angiotensin-Adosterone (RAA Axis)
17
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Myocardial Natriuretic Peptides:

- these are released from atrial and ventricular myocardium in
response to volume expansion
- these inhibit sodium resorption in the distal renal tubules, thus
leading to sodium excretion and diuresis
- they also induce systemic vasodilation
19

Blood Pressure regulation
20
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Types and Causes of Hypertension

Essential Hypertension
- accounts for 90% to 95% of all cases
Secondary Hypertension
Renal
- Acute Glomerulonephritis
- Chronic Renal Disease
- Polycystic disease
- Renal artery stenosis
Endocrine
- Adrenocortical hyperfunction: (Cushing Synd, Primary Aldosteronism)
- Exogenous hormones (glucocorticoids, estrogen and oral contraceptives
- Phaeochromocytoma
- Acromegaly
- Hypothyroidism (myxedema)
- Hyperthyroidism (thyrotoxicosis)
- Pregnancy induced
CVS
- Coarctation of aorta
- Polyarteritis nodosa (PAN)
21
Hypertensive Vascular Disease: Pathogenesis
Pathogenesis of Hypertension:
- almost 90% to 95% are unknown (essential)
Pathogenesis of “essential hypertension”
1) Genetic Factors
2) Reduced renal sodium excretion (most important)
3) Vasoconstrictive influences
4) Environmental (stress, obesity, smoking, physical inactivity, and salt
consumption)
22
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Pathogenesis of Hypertension:
Reduced renal sodium excretion
- in presence of normal arterial pressure
- this leads to increase in fluid volume, increased cardiac output, and
peripheral vasoconstriction
- net result being elevated BP
- with elevated BP, additional sodium will be excreted by the kidneys to
equal intake and prevent further fluid retention
- now, there will be an altered but steady state of sodium excretion
- but this results in increase in BP
23

Pathogenesis of Secondary Hypertension
- best example is: ‘renovascular hypertension’
- caused by renal artery stenosis (narrowing), leading to decreased
glomerular blood flow, and decreased pressure within afferent
arteriole
- this leads to: a) increased Renin secretion which activates RAA axis
resulting in vasoconstriction, and increased peripheral resistance
b) increased sodium resorption which leads to increased blood volume
thru’ the action of aldosterone
24
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25
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Pathogenesis of Secondary Hypertension
Renal artery stenosis

- hypertension secondary to renal artery
stenosis is caused by increased production of
renin from the ischemic kidney
Causes:
1) Narrowing at the origin of the renal artery
by an atheromatous plaque (70%)
Fibromuscular dysplasia: note “string of beads”
2) Fibromuscular dysplasia. Characterized by appearance on renal angiogram. Typically noted in
fibrous or fibromuscular thickening that may middle-to-distal portion.
involve the intima, the media, or the Remember: atherosclerotic narrowing is typically noted
at the origin.
adventitia. More common in women and tend
to occur in younger age groups (i.e., in the
third and fourth decades).
27
Hypertensive Vascular Disease: Morphology

Hyaline Arteriolosclerosis
- arterioles show homogeneous, pink hyaline
thickening with associated luminal narrowing
- this happens due to plasma protein leakage across
injured endothelial cells, and
Hyaline Arteriolosclerosis.
- due to increased smooth muscle cell matrix note: luminal narrowing,
subsequent ischemia
synthesis following chronic hemodynamic stress
- Example: renal biopsy in hypertension, diabetes
mellitus
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30
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Hyaline arteriolosclerosis: Benign Nephrosclerosis

Morphology of the kidney
Gross:
- kidneys are either normal or moderately reduced in
size
- there is loss of mass is due mainly to cortical scarring
and shrinking
Micro:
-narrowing of the lumens of arterioles and small
arteries,
due to thickening and hyalinization of the walls (hyaline
arteriolosclerosis)
32
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Hyaline Arteriolosclerosis:
- is associated with Benign nephrosclerosis
- this is a renal condition seen with hypertension
- renal biopsy: hyaline arteriolosclerosis
Clinical Course:
- causes atrophy of the tubules and sclerosis of the
glomeruli
- progresses to renal failure
- retinal changes
Hyaline arteriolosclerosis
33
Hypertensive Vascular Disease: Morphology

Hyperplastic Arteriolosclerosis
- this is classically seen in severe hypertension (malignant
hypertension)
- where vessels show concentric, laminated (“onion-skin”) thickening of
the walls with luminal narrowing
- these laminations are composed of smooth muscle cells with
thickened, reduplicated basement membrane
- example: renal biopsy in hypertension
34
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Hyperplastic arteriolosclerosis: Malignant

Nephrosclerosis
Morphology of the kidney:
Gross:
- tiny, pinpoint petechial hemorrhages may
appear on the cortical surface from rupture of
arterioles or glomerular capillaries,
- which makes the kidney appear as “flea-bitten”
Micro:
- show fibrinoid necrosis of arterioles
35
Hyperplastic arteriolosclerosis
- associated with Malignant Hypertension (Malignant Nephrosclerosis)
- rapid increase in blood pressure accompanied by renal failure and cerebral
edema
Clinical Course:
- renal: on gross shows- “flea bitten” appearance
- acute renal failure, retinal changes, CNS complications
36
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Retinal changes in Hypertension

(cross reference: ICM)
Grade I: focal narrowing of the arterioles, mild arteriovenous nicking

Grade II: arteriole narrowing, copper wiring present, arteriovenous nicking more accentuated
Grade III: arteriole narrowing, silver wiring present, hemorrhages, soft and hard exudates, disappearance of
the vein under the arteriole, disk normal
Grade IV: arterioles are fine fibrous cords; same as grade III except papilledema is present
37
complications of Hypertension
CVS:
- LVH
- Acute Myocardial Infarction
CNS:
- Intracerebral hematoma
- Berry Aneurysm
- Lacunar infarcts
Renal
- Benign nephrosclerosis
- Malignant hypertension
Eyes
- Hypertensive retinopathy
38
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Arteriosclerosis
Definition:
- arteriosclerosis means hardening of the arteries
- a term given arterial wall thickening and loss of elasticity
Types:
1) Arteriolosclerosis
- this affects small arteries and arterioles, and may cause downstream
ischemic injury
- there are two anatomic variants: Hyaline arteriolosclerosis, and
Hyperplastic arteriolosclerosis (discussed earlier)
39
Arteriosclerosis
Types:
2) Mönckeberg medial sclerosis
- show calcification of the walls of
muscular arteries, typically
involving the internal elastic
membrane
- seen after the age of 50 years
- important to remember that the
calcifications do not encroach on
the vessel lumen and
- therefore are usually not clinically
significant
40
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Arteriosclerosis
Types:
3) Atherosclerosis
- derived from Greek word for
“gruel” and “hardening,”
- this is the most frequent and
clinically important pattern
41
Atherosclerosis
- Definition
- Epidemiology and Risk Factors
- Pathogenesis
- Morphology
- Clinicopathological
- Complications
42
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Atherosclerosis
Definition:
- characterized by intimal lesions called as ‘atheromas’
(atheromatous/atherosclerotic plaques)
- atheroma consists of a raised lesion with a soft, yellow, grumous core
of lipids
- lipids (mostly are cholesterol, and cholesterol esters)
- the lesion is capped by a fibrous lining
Problems: Ischemic Heart Disease (Myocardial Infarction), Stroke,
Aneurysm, Ischemic Bowel Disease, Peripheral Artery Disease
(gangrene)
43
Epidemiology and Risk Factors
44
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Atherosclerosis: Epidemiology
- highest incidence in developed countries
Risk Factors:
Constitutional risk factors: age, gender, genetics
Age:
- typically progressive, clinical manifestations occur between 40 to 60
years
Gender:
- premenopausal women are protected (estrogen)
- risk increases after menopause
Genetics:
- family history. Familial Hyercholesterolemia
45
Risk Factors:
Modifiable risk factors: hyperlipidemia, hypertension, cigarette smoking, and
diabetes mellitus
Hyperlipidemia:
- hypercholesterolemia
- major component of serum cholesterol associated with increased risk is: LDL
(low density lipoprotein ‘bad cholesterol’)
- LDL is the main cholesterol that is delivered to the tissues
- HDL (high density lipoprotein ‘good cholesterol’) mobilizes cholesterol from
tissues and transport it to the liver for excretion into bile
46
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Hyperlipidemia (cont’d):
- high dietary intake of cholesterol and dietary fat raises plasma
cholesterol level
- diets low in cholesterol lowers plasma cholesterol
- omega-3 fatty acids present in fish is considered healthy
- trans-unsaturated fat (margarine) is unhealthy
- exercise and “moderate” consumption of ethanol is cardioprotective
- as these increases HDL
- smoking increases cholesterol
47
Lipid metabolism
(Biochem review)
48
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Hypertension:
- increases in both systolic and diastolic pressure is a risk factor
Cigarette smoking:
- prolonged years of smoking increases the risk
- cessation of smoking is beneficial
Diabetes mellitus:
- induces hypercholesterolemia
49
Atherosclerosis: Pathogenesis (overview)

1) Endothelial injury: in presence of high lipid diet, endothelium can be injured
2) Accumulation of lipoproteins: LDL, and its oxidized form deposits in the vessel
wall
3) Monocyte adhesion: monocytes adhere on the injured endothelium, and later
migrate into intima. Monocytes then transforms to become macrophages
4) Platelet adhesion: platelets also adheres onto injured endothelium
5) Factor release: factors released from activated macrophages, and platelets. These
factors recruit other cells to this location
6) Lipid accumulation: both within, and outside the cells (intracellular and
extracellular)
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51
Atherosclerosis
52
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Atherosclerosis: Pathogenesis (detailed)

Endothelial Injury
- causes for endothelial cell injury: mechanical denudation,
hemodynamic forces, immune complex deposition, irradiation, or
chemical
- sometimes early lesions begin at sites of morphologically intact
endothelium. This is referred to as, endothelial dysfunction (non-
denudation)
Endothelial dysfunction causes: 1) increased endothelial permeability
2) increased leukocyte adhesion, and 3) altered gene expression
53
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55
56
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Endothelial Injury (cont’d)
- two most important causes of endothelial
injury are
1) hemodynamic disturbances: as in
hypertension
2) hypercholesterolemia: increased LDL,
decreased HDL, increased levels of the
abnormal lipoprotein (a)
57

- with hypercholesterolemia, lipoproteins
accumulate within the intima
- here they may aggregate or be converted to
oxidized LDL by free radicals produced by
inflammatory cells
- these modified LDL is taken by the
macrophages
- as modified lipoproteins cannot be completely
degraded, they ends up within macrophages
as lipid-filled macrophages called foam cells
58
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hypercholesterolemia (cont’d):
- smooth muscle cells migrate from the media, and then transform
- into lipid-laden foam cells by ingesting of the modified lipids
- the binding and uptake of modified lipids stimulates;
1) release of growth factors
2) cytokines, and chemokines
that create a vicious cycle of monocyte recruitment and activation
59

Inflammation
- adherent platelets on endothelium release PDGF,
- macrophages in subendothelium also releases PDGF
- smooth muscle cells too produce PDGF
- many of the factors stimulate smooth muscle cells to synthesize collagen
Stable atherosclerotic plaques: collagen produced will stabilizes
atherosclerotic plaques (absent inflammatory activity)
Unstable atherosclerotic plaques: if there are activated inflammatory cells in
atheromas, it leads to the breakdown of extracellular matrix components
(MMPs), resulting in unstable plaques
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Atherosclerosis: Endothelial Injury
61
Pathogenesis
62
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Atherosclerosis: Morphology
Fatty streaks:
- are composed of lipid-filled foamy macrophages
- they begin multiple minute flat yellow spots, which fuse to form
elongated streaks (1 cm long or longer)
- remember: fatty streaks are not sufficiently raised to cause any
significant flow disturbances
- fatty streaks can eventually form plaques, however not all end up
becoming advanced lesions
63
Atherosclerosis: Fatty streaks
64
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Atherosclerotic plaque:
- plaques results from intimal thickening and lipid accumulation
- plaques appear as: white-yellow and encroach on the lumen of the
artery
- when ulcerated plaques are superimposed with thrombus: red-brown
- these lesions are patchy, produce eccentric protrusions, and rarely
they are circumferential
65
Changes in Atherosclerotic Plaques:

- plaques are susceptible for
Rupture, ulceration, or erosion
- this exposes underlying thrombogenic substances
- triggering coagulation pathway
- ending in occlusive THROMBOSIS
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Atherosclerotic plaque
67
68
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Atherosclerotic plaque
Changes in Atherosclerotic Plaques

- Fibrous cap undergoes continuous remodeling that can stabilize the plaque, or conversely, render it more
susceptible to rupture
At molecular level:
- collagen is the major structural component of the fibrous cap, and accounts for its mechanical strength
and stability
- collagen turnover is controlled by MMPs, enzymes elaborated largely by macrophages and smooth
muscle cells within the atheromatous plaque; conversely, tissue inhibitors of metalloproteinases (TIMPs)
produced by endothelial cells, smooth muscle cells, and macrophages modulate MMP activity
69
History and morphologic features:
70
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Atherosclerotic plaques: Vessels commonly involved:
1) lower abdominal aorta
2) coronary arteries
3) popliteal arteries
4) internal carotid arteries
5) vessels of the circle of Willis
71
Components of Atherosclerotic plaques:
- there are 3 important components
1) superficial fibrous cap containing smooth muscle cells and collagen
2) beneath and to the sides (shoulder): smooth muscle cells,
macrophages, and T-cells
3) deep to the fibrous cap: necrotic core- intracellular and extracellular
lipid, necrotic material, lipid laden macrophages
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Atherosclerosis: Clinicopathological
1) Critical stenosis:
- a stage at which the occlusion is sufficiently severe to produce tissue
ischemia
- in the coronary circulations, critical stenosis occurs when the occlusion
produces a 70% decrease in luminal cross-sectional area
2) Acute plaque change:
- Rupture/fissuring: which exposes underlying highly thrombogenic plaque
constituents
- Erosion/ulceration: exposing the thrombogenic subendothelial basement
membrane to blood
- Hemorrhage into the atheroma: expanding its volume
73
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Coronary Angiogram:
critical stenosis
75
Atherosclerosis: Complications
1) Vessel weakness:
- this may lead to “aneurysms” (abdominal aorta)
2) Thrombosis:
- ruptures plaque exposes thrombogenic subendothelial collagen, which leads to platelet-fibrin
thrombus
- example for thrombus: acute MI, Stroke, Small bowel infarction
3) Hypertension:
- reduction of renal blood flow due to atherosclerotic narrowing
- Renin-Angiotensin-Aldosterone activation
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4) Cerebral atrophy:
- blood flow reduction in CNS following atherosclerotic narrowing of blood
vessels (circle of Willis, carotid artery)
5) Atherosclerotic embolization:
- leads to peripheral vessel blockage
6) Peripheral Arterial Disease (PAD):

- very important clinical complication
77
6) Peripheral Arterial Disease (PAD) cont’d:
- associated risk factors include smoking, diabetes, hypertension
- clinically produce:
Claudication- unilateral, gradual, and consistent cramping pain in the
buttock, thigh, and calf that may be associated with weakness and
numbness. It is due to decreased arterial blood flow to the affected leg.
Pain is relieved by resting
Ulcers- characteristically seen in lower limbs, may present as gangrene
(dry/wet). Also, may present as ‘non-healing’ ulcer.
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79
Atherosclerosis: Laboratory studies

Markers of cardiac injury:
1) Troponins:
- in contrast to other cardiac markers, cTnT and cTnI are nearly absent
from normal serum (are detected within 3 to 12 hrs following acute
MI, disappear by 7 to 10 days)
- therefore are important markers for cardiac ischemia
2) CK-MB:
- detected by 3 to 8 hrs, disappear by 1 to 3 days
80
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Atherosclerosis: Laboratory studies

Markers of coronary risk:
1) C-Reactive Protein (CRP):
- it is an “Acute Phase Protein”
- produced during inflammation during atherosclerosis
- and seen with unstable plaque
2) Coronary-artery calcium (CAC) scan:

- a CAC score below 100 indicates low risk, and a score above 300
indicates high risk
81
CAC scan: origin of the left coronary artery, show calcification of the left
main and proximal left anterior descending coronary arteries (Panel A) and
(Panel B).
82
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“Throughout history, it has been the inaction of those who could have acted, the indifference of those who should have
known better, the silence of the voice of justice when it mattered most, that has made it possible for evil to triumph”
Haile Selassie, Emperor of Ethiopia
83
Blood Vessel Pathology. Part 2.

Aneurysms, Aortic Dissection
Dr. Roy
84
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Aneurysm
Definition:
An aneurysm is a localized abnormal dilation of a blood vessel or the heart
that may be congenital or acquired
True aneurysm: involves an attenuated but intact arterial wall (all layers) or
thinned ventricular wall of the heart
Example: 1) atherosclerotic, 2) syphilitic, 3) congenital vascular aneurysms, 4)
ventricular aneurysms that follow transmural myocardial infarctions
False aneurysm (pseudo aneurysm): defect in the vascular wall leading to an
extravascular hematoma that freely communicates with the intravascular
space (pulsating hematoma)
Example: 1) ventricular rupture after myocardial infarction
85
Aneurysm
86
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Aneurysm: Types
Types:
1) Saccular aneurysms:
- spherical outpouchings involving only a
portion of the vessel wall
- they vary from 5 to 20 cm in diameter and
often contain thrombus
2) Fusiform aneurysms:
- diffuse, circumferential dilations of a long
vascular segment
- they vary in diameter (up to 20 cm) and in
length and can involve extensive portions of
the aortic arch, abdominal aorta, or even the
iliacs
87
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89
Aneurysm: Pathogenesis
1) Intrinsic quality of the vascular wall connective tissue is poor:
- Marfan syndrome (defective synthesis of fibrillin)
- Loeys-Dietz syndrome (aberrant TGF-β activity and weakening of
elastic tissue)
- Ehlers-Danlos syndrome (defective type III collagen synthesis)
2) Balance of collagen degradation and synthesis is altered by

inflammation and associated proteases:
- Atherosclerotic plaque (increased matrix metalloprotease (MMP)
expression, especially by macrophages)
- Vasculitis
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Aneurysm: Pathogenesis
3) Vascular wall is weakened through loss of smooth muscle cells or the
synthesis of non-collagenous or non-elastic extracellular matrix:
Systemic hypertension cause significant narrowing of arterioles of the
vasa vasorum (ischemia of the inner media)
- this medial ischemia produce “degenerative changes” of the aorta
- leading to loss of smooth muscle cell
- eventually scarring occurs (and loss of elastic fibers), inadequate
extracellular matrix synthesis, and production of increasing amounts
of amorphous ground substance (glycosaminoglycan)
Tertiary Syphilis (obliterative endarteritis)
91
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Syphilitic Aortitis (Tertiary Syphilis):

- focal ischemic necrosis, secondary to endarteritis obliterans
- leads to destruction of vaso vasorum of aorta
- thereby producing ischemic necrosis of aortic media
- dilatation of the thoracic aorta, and aortic valve ring
- endarteritis obliterans on histology reveals lymphocytes, and plasma
cells
93
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Aneurysm: Causes
Most important causes of Aneurysms are:
Atherosclerosis: Abdominal Aortic Aneurysm
Hypertension: Ascending Aortic Aneurysm
Others:
- trauma, vasculitis, congenital defects (fibromuscular dysplasia and berry
aneurysms), and infections (mycotic aneurysms)
Mycotic aneurysms:
- from embolization of a septic embolus, usually as a complication of infective
endocarditis, or fungal (Mucormycosis)
- as an extension of an adjacent suppurative process
- by circulating organisms directly infecting the arterial wall
95
Abdominal Aortic Aneurysm (AAA)

Definition:
- permanent pathologic dilation of the aorta with a diameter >1.5
times the expected anteroposterior (AP) diameter of that segment,
given the patient's gender and body size
Incidence:
- patients at greatest risk for are men who are older than 50 years with
history of smoking, and have peripheral atherosclerotic vascular
disease
Causes:
- Atherosclerosis, inflammatory, and infectious
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Abdominal Aortic Aneurysm (AAA)

Pathogenesis:
- atherosclerosis involving
abdominal aorta leads to
progressive weakening of the
wall
- produces thinning of the
underlying aortic media
- which predisposes to aneurysm
formation
97
Abdominal Aortic Aneurysm (AAA): Morphology
- typically positioned beneath the renal arteries and above the

bifurcation of the aorta
- can be saccular or fusiform, up to 15 cm in diameter, and up to 25 cm
in length
- there is often severe complicated atherosclerosis with destruction
and thinning of the underlying aortic media
- aneurysm frequently contains a bland, laminated, poorly organized
mural thrombus (Virchow’s triad…..turbulence, stagnation)
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Abdominal Aortic Aneurysm
99
Abdominal Aortic Aneurysm (Inflammatory)

Inflammatory Abdominal Aortic Aneurysm:
- account for 5% to 10%
- typically occur in younger patients
- present with back pain and elevated inflammatory markers (e.g.,
elevation of C-reactive protein)
- on histology: lymphocytes, plasma cells, and macrophages. periaortic
scarring is often noted
- cause: localized immune response to the abdominal aortic wall
100
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Abdominal Aortic Aneurysm (Inflammatory)

Other settings of Inflammatory Abdominal Aortic Aneurysm
Immunoglobulin G4 (IgG4)-related disease:
- there is high plasma levels of IgG4 and tissue fibrosis associated with frequent
infiltrating IgG4-expressing plasma cells
- responds well to steroid therapy
Mycotic Abdominal Aortic Aneurysm:

- due to lesions that have become infected by the lodging of circulating microorganisms
in the wall
- leads to suppuration which destroys the media, with the risk of rapid dilation and
rupture
101
Abdominal Aortic Aneurysm: Clinical Features

1) Asymptomatic:
- noted incidentally on physical exam as an abdominal mass
- may present as a pulsatile mass,
- bruit (harsh sound) may be heard on auscultation if renal artery stenosis
and/or arterial stenosis involving the orifices of the mesenteric arteries are
present along with the abdominal aortic aneurysm
- fragmented atherosclerotic plaques may embolize to give rise to “blue toe
syndrome”
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2) Rupture into the peritoneal cavity or retroperitoneal space
3) Obstruction of a vessel branching off from the aorta. This can result
in ischemic injury to the supplied tissue. Example: iliac (leg), renal
(kidney), mesenteric (gastrointestinal tract), or vertebral arteries (spinal
cord)
4) Compression on an neighboring structure. Example: compression of

a ureter or erosion of vertebrae
103

- when do you suspect a rupture?
Rupture triad:
1) sudden onset of severe flank pain (bleed is initially retroperitoneal)
2) followed by hypotension from blood loss into the retroperitoneum
and
3) presence of a pulsatile mass on physical examination
Investigations:
- Ultrasound study
- CT scan
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Aortic Dissection
105
Aortic dissection
Definition:
- aortic dissection occurs when blood separates the laminar planes of the media to
form a blood-filled channel within the aortic wall (weakening of media)
- this can become catastrophic if the dissection then ruptures through the
adventitia and hemorrhages into adjacent spaces
Incidence:
- occurs in two groups:
1) men aged 40 to 60 years with antecedent hypertension (more than 90% of
cases)
2) younger adults with systemic or localized abnormalities of connective tissue
affecting the aorta (e.g., Marfan syndrome)
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Classification
107
Aortic dissection: Pathogenesis

1) Hypertension:
- is the major risk factor for aortic dissection
- in hypertension aorta reveals medial hypertrophy of vasa vasorum
- associated with degenerative changes: loss of medial smooth muscle
cells, disorganized extracellular matrix
- these changes results from reduced blood flow through the vasa
vasorum
2) Marfan Syndrome, Ehlers-Danlos Syndrome:
- abnormal elastic/collagen synthesis
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Aortic dissection: Pathogenesis

Important to remember:
- that medial damage is not necessary for aortic dissection
- but, in the presence of medial weakness, once there is an intimal tear
blood flows under systemic pressure dissecting through the media
Cystic medial degeneration:
- may promote dissection
- medial degeneration happens due to ischemia (narrowed vasa
vasorum as in HTN, or defective ECM synthesis with connective tissue
disorder)
- in medial degeneration: loss of smooth muscle, deficient synthesis of
ECM, and increase in ground substance- proteoglycans)
109
Aortic dissection: Morphology
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Aortic dissection begins with an intimal tear
- in most cases tear is noted in the ascending aorta
- usually within the first 10 cm from the aortic valve
- tears are typically transverse with sharp edges
- remember that the dissection can extend retrograde (ie towards the
heart), as well as distally
- the dissection may extend distally into the iliac, or femoral
- hematoma is seen characteristically between the middle and outer
third (see image in previous slide)
111

- dissecting hematoma often
ruptures out thru’ the adventitia
- this causes massive hemorrhage:
into thoracic cavity, abdominal
cavity, or produce cardiac
tamponade
- another situation may arise, when
the dissecting hematoma may re-
enter into the aortic lumen by
producing an intimal tear
- this is seen as a “double-barreled
aorta”
112
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113
Aortic dissection: Clinical Features

- characterized by sudden onset of excruciating pain,
- usually beginning in the anterior chest, radiating to the back between
the scapulae, and moving downward as the dissection progresses
differential diagnosis: the pain can be confused with that of myocardial
infarction
- aortic regurgitation, compression of SVC lead to loss of pulses in
upper extremity
- most common cause of death: rupture of the dissection into the
pericardial, pleural, or peritoneal cavities
114
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“the dangerous man is the one who has only one idea, because then he'll fight and die for it”
Francis Crick, physicist, biologist, neuroscientist, Nobel laureate (1916-2004)
115
CVS: Vasculitis, Part 1
Dr. Roy
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Vasculitis
Definition:
- general term for vessel wall inflammation
- vessels of any type in virtually any organ can be affected,
- but most vasculitides affect small vessels ranging in size from
arterioles to capillaries to venules
Classification:
1) based on type of blood vessels
2) based on pathogenesis
117
Classification: Based on blood vessels
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119
Vasculitis: Classification
Based on Pathogenesis:
1) Non-infectious vasculitis:
- vasculitis caused by local or systemic immune response
- immunologic injury in non-infectious vasculitis may be caused by:
a) Immune complex deposition
b) Anti-neutrophil cytoplasmic antibodies
c) Anti-endothelial cell antibodies
d) Autoreactive T-cells
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Vasculitis: Non-infectious vasculitis

SLE:
- in SLE, there is autoantibody production and formation of immune
complexes that deposit in vessels
Other examples of Immune complex deposition include:
Drug hypersensitivity vasculitis:
- examples of drugs: penicillin, streptokinase
Vasculitis secondary to infections:
- Ab to microbial constituents can form immune complexes that
circulate and deposit in vascular lesions
121

Mechanism:
immune complex deposits endothelial injury inflammation
and complement activation formation of C5a amplification
of inflammation coagulation system activation Fibrinoid
necrosis
Example: Henoch-Schonlein purpura (Type III hypersensitivity)
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123

b) Antineutrophil cytoplasmic antibodies
- these patients have circulating antibodies that react with neutrophil cytoplasmic
antigens, referred to as ‘antineutrophil cytoplasmic antibodies’ (ANCAs)
ANCAs are autoantibodies directed against constituents (mainly enzymes)
- of neutrophil primary granules
- monocyte lysosomes
- endothelial cells
Importance of ANCAs
- they can help in diagnosis: their titers levels reflect clinical severity, and a rise in
titers after periods of quiescence is predictive of disease recurrence
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b) Antineutrophil cytoplasmic antibodies (ANCAs) cont’d
- there are 2 important ANCAs
1) Anti-proteinase-3 (PR3-ANCA)
- older terminology: c-ANCA
- PR3 is a neutrophil azurophilic granule constituent that shares
homology with numerous microbial peptides
- therefore generation of PR3-ANCAs is triggered by certain infections
- Example: PR3-ANCAs are associated with Granulomatosis with
Polyangiitis
125
b) Antineutrophil cytoplasmic antibodies (ANCAs) cont’d

2) Anti-myeloperoxidase (MPO-ANCA)
- older terminology: p-ANCA
- MPO is a lysosomal granule constituent involved in oxygen free
radical generation
- MPO-ANCAs are generated by several therapeutic agents, example:
propylthiouracil
- Example: microscopic polyangiitis, Churg-Strauss syndrome
126
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b)Antineutrophil cytoplasmic antibodies (ANCAs) cont’d
Mechanism:
- ANCA autoantibodies are directed against cellular constituents and do not
form circulating immune complexes
- the vascular lesions do not typically contain demonstrable antibody and
complement
- ANCA-associated vasculitides are often described as “pauci-immune”
- ANCA directed against proteins other than PR3 and MPO are often seen in
patients with non-vasculitic inflammatory disorders, such as inflammatory
bowel disease, sclerosing cholangitis, and rheumatoid arthritis
127

c) Antiendothelial cell antibodies
- antibodies to endothelial cells, may be generated by defects in
immune regulation
- example: Kawasaki disease
128
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Large Vessel Vasculitis

- Giant Cell Arteritis
- Takayasu’s Arteritis
129
130
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Giant cell arteritis

Introduction:
- most common form of vasculitis among older individuals (more than
50 years) in the United States and Europe
- chronic inflammatory disorder affecting typically large-sized, but also
seen to involve other arteries
- characteristically affects arteries in the head (mostly temporal
arteries, but also the vertebral and ophthalmic arteries)
- involvement of ophthalmic artery has a risk for blindness, and
therefore is a medical emergency
131

Pathogenesis:
- results from a T-cell mediated immune response against
- vessel wall antigens which generate proinflammatory cytokines
(particularly TNF)
- anti-endothelial cell and anti-smooth muscle cell antibodies can also be
demonstrated in roughly two thirds of patients
- cellular immune etiology is supported by the characteristic granulomatous
response
- prompt therapeutic response to steroids
- predilection for a single vascular site (temporal artery) remains unexplained
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Giant cell arteritis: Morphology

- arterial segments develop intimal thickening
- that reduces the luminal diameter
- characteristically lesions show granulomas in media
- lesions are located on the internal elastic lamina leading to elastic
lamina fragmentation
- inflammation is composed of T cells (CD4+ > CD8+) and macrophages
- although multinucleated giant cells are seen in approximately 75% of
adequately biopsied specimens, they maybe absent
- inflammatory lesions are only focally distributed along the vessel and
long segments of relatively normal artery may be interposed
133
134
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Giant cell arteritis: immunopathogenesis
135
Giant cell arteritis: Clinical Features

- rare before age 50, symptoms may be only vague and constitutional:
fever, fatigue, weight loss
- other clinical features:
- facial pain, headache, most intense along the course of the superficial
temporal artery, which can be painful to palpation
- jaw claudication (pain when chewing)
- ocular symptoms (ophthalmic artery) appear abruptly in about 50% of
patients
Diagnosis: depends on biopsy and histologic confirmation. Elevated ESR.
Treatment: Corticosteroids or anti-TNF therapies are typically effective
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Takayasu arteritis
Introduction:
- is a granulomatous vasculitis of medium and larger arteries (less than 50 years)
- characterized principally by ocular disturbances and marked weakening of the
pulses in the upper extremities
- therefore, referred to as: “pulseless disease”
Incidence:
- Asian (Japanese) population mostly, younger than 50 years (if older than 50 years-
giant cell arteritis). Women are affected more.
Pathogenesis:
- autoimmune
137
Takayasu arteritis: Morphology

- typically involves the aortic arch
- other blood vessels: may affect the remainder of the aorta and its
branches, with pulmonary artery involvement, coronary and renal arteries
may be similarly affected
Histology:
aortic arch and great vessels shows:
- irregular thickening of the vessel wall with intimal hyperplasia
- transmural fibrous thickening of the aorta
- with severe luminal narrowing of the major branch vessels
remember: histological features are similar to giant cell arteritis
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Takayasu arteritis
139
Takayasu arteritis: Clinical Features

- initially nonspecific: fatigue, weight loss, and fever
- as disease progresses: reduced blood pressure and weak pulses in the
carotids and the upper extremities
- discrepancy in BP in right and left side (more than 10 mmHg)
- involvement of ocular: visual defects, retinal hemorrhages, and total
blindness
- involvement of vessels distal to aorta: claudication of the legs
- involvement of pulmonary artery: pulmonary hypertension
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Takayasu arteritis: Clinical Features

- involvement of coronary: narrowing of the coronary ostia may lead to
myocardial infarction
- involvement of the renal arteries: systemic hypertension in roughly
half of patients
Clinical course
- variable, in some there is rapid progression
- while others enter a quiescent stage after 1 to 2 years, permitting
long-term survival, albeit with visual or neurologic deficits
- corticosteroid therapy
141
Medium-sized Vessel Vasculitis

- Polyarteritis Nodosa (PAN)
- Kawasaki Disease
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Polyarteritis Nodosa
Introduction:
- Polyarteritis nodosa (PAN) is a systemic vasculitis of small- or medium-sized
muscular arteries
Pathogenesis:
- typically involves renal and visceral vessels
- but sparing the pulmonary circulation
- no association with ANCA
- about 30% of patients with PAN have chronic hepatitis B and deposits containing
HBsAg-HBsAb complexes in affected vessels (immune complex mediated etiology
in these patients) subset
- cause remains unknown in the remaining cases
143
Polyarteritis Nodosa: Morphology
- characterized by segmental transmural necrotizing inflammation of

small- to medium-sized arteries
Vessels involved in descending order:

- kidneys
- heart
- liver
- gastrointestinal tract
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Polyarteritis Nodosa: Morphology cont’d

- inflammatory process weakens the arterial wall (leading to aneurysm: nodosa)
- impaired perfusion with ulcerations, thrombosis, infarcts, ischemic atrophy,
or hemorrhages may be the first sign of disease
Histology
Acute phase: transmural inflammation of the arterial wall with infiltrates of
neutrophils, eosinophils, and mononuclear cells
frequently accompanied by fibrinoid necrosis
Chronic phase: acute inflammation is replaced by fibrous thickening of the
blood vessel wall
145
Polyarteritis Nodosa
146
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Polyarteritis Nodosa: Clinical Features

- most patients are young adults, however may occur in pediatric and
older populations
- clinical features mostly result from ischemia and infarction of affected
tissues and organs
- Fever of Unknown Origin
Renal: Hypertension
GIT: Bowel infarction
- clinical course is remitting and episodic, with long symptom-free
intervals
- renal involvement is important as it is a major cause of mortality
147
Kawasaki disease
Introduction:
- an acute febrile, usually self-limited illness of infancy and childhood
- most patients (80% are 4 years old or younger)
- it is associated with an arteritis affecting large to medium-sized, and
even small vessels
- first described in Japan, but has worldwide incidence
Most important: coronary artery involvement that can cause
aneurysms that may rupture or develop thrombus
- resulting in acute myocardial infarction
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Kawasaki disease: Pathogenesis
- pathogenesis is unknown…. (? viral)

- possibly a delayed type of hypersensitivity reaction involving T-cells
- to an unknown antigen
- this results in cytokine production and macrophage activation
- and also B-cell activation, resulting in formation of auto-antibodies
against endothelial cells and smooth muscle cells
149
Kawasaki disease: Morphology
- dense transmural inflammatory infiltrate similar to PAN

- fibrinoid necrosis is less prominent compared to PAN
- the acute disease may disappear or in response to treatment, but
aneurysm formation due to vessel wall may happen
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Kawasaki disease: Clinical Features
Classical presentation:
1) conjunctival (conjunctivitis) and
oral erythema and blistering
(strawberry-appearing tongue)
2) edema of the hands and feet
3) erythema of the palms and soles,
a desquamative rash
4) cervical lymph node enlargement
(tender)
151
Kawasaki disease: Clinical Features

Complications:
- around 20% of untreated patients develop cardiovascular
complications:
1) asymptomatic coronary arteritis
2) giant coronary artery aneurysms (7 to 8 mm) leading to rupture or
thrombosis, acute myocardial infarction, and sudden death
Investigations:
- ECG findings suggestive of Acute Myocardial Infarction may be seen
Treatment: Immunoglobulin therapy, and Aspirin
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“there are a thousand hacking at the branches of evil to one who is striking at the root”
Henry David Thoreau, naturalist and author (1817-1862)
153
CVS: Vasculitis, Part 2
Dr. Roy
154
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Small-sized Vessel Vasculitis

- Microscopic Polyangiitis
- Churg-Strauss syndrome
- Behcet syndrome
- Granulomatosis with Polyangiitis
- Thromboangiitis obliterans
155
Microscopic Polyangiitis
Introduction:
- necrotizing vasculitis that generally affects capillaries, small arterioles
and venules
- other names: hypersensitivity vasculitis, leukocytoclastic vasculitis
how is it different from PAN?
- microscopic polyangiitis affects smaller blood vessels
- unlike PAN, in microscopic polyangiitis all lesions tend to be of the
same age in any given patient and are distributed more widely
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Sites of involvement:
1) skin (palpable purpura), and mucous membranes
2) lungs
3) brain
4) heart
5) gastrointestinal tract
6) kidneys (necrotizing glomerulonephritis (90% of patients)
7) muscle can all be involved
8) pulmonary capillaritis (common)
157
Microscopic Polyangiitis: Pathogenesis
- possible settings:
1) antibody responses to antigens such as drugs (e.g., penicillin)
2) microorganisms (e.g., streptococci)
- either one of these can lead to immune complex deposition or
trigger secondary immune responses
- like ANCAs
- most cases are associated with MPO-ANCA
- (this is due to recruitment and activation of neutrophils)
158
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159
Microscopic Polyangiitis: Morphology

typically shows:
- segmental fibrinoid necrosis of the media and
- focal transmural necrotizing lesions
- however, there is no granuloma formation
remember: may resemble PAN but spare medium-sized and larger
arteries therefore consequently, infarcts are not present
- leukocytoclastic vasculitis (seen mostly in post-capillary venules):
- presence of neutrophils, many undergoing apoptosis, karyorrhexis
160
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161
Diagnosis:
- skin biopsy along with clinical history/examination
162
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- high magnification
163
Microscopic Polyangiitis: Clinical Features

- hemoptysis
- hematuria
- proteinuria
- abdominal pain (bowel bleeding)
- palpable cutaneous purpura
Treatment:
- cyclophosphamide, and steroids (these are not given fro patients who
have widespread renal or brain involvement)
164
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Churg-Strauss syndrome
Introduction:
- is a rare disease with small-vessel necrotizing vasculitis
- classically associated with:
1) asthma
2) allergic rhinitis
3) lung infiltrates
4) peripheral hypereosinophilia
5) extravascular necrotizing granuloma
165
Churg-Strauss syndrome: Pathogenesis and Morphology

Possible mechanism:
- may be a consequence of hyperresponsiveness to an allergic stimulus
- as in patients with asthma
- leukotriene receptor antagonists have been reported to be a trigger
Morphology:
- vascular lesions can be histologically similar to polyarteritis nodosa or
microscopic polyangiitis
- but characteristically reveal granulomas (palisading), and tissue eosinophils
- in less than 50% of cases ANCAs (mostly MPO-ANCAs) are present
166
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Churg-Strauss syndrome: Clinical Features

- patients present with adult onset asthma (4th decade)
- allergic rhinitis, nasal polyps, and sinusitis
also noted are:
- multisystem diseases with cutaneous involvement (palpable purpura)
- gastrointestinal tract bleeding
- renal disease (focal and segmental glomerulosclerosis)
- myocardial involvement may give rise to cardiomyopathy
- remember: heart is involved in 60% of patients and accounts for almost
half of the deaths in the syndrome
167
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Behcet syndrome
Introduction:
- is a small- to medium-vessel neutrophilic vasculitis
- common in Middle East, and Far East (most cases reported in Turkey, along
the Silk route)
- that classically presents as a clinical triad
1) recurrent oral aphthous ulcers
2) genital ulcers
3) uveitis
- disease mortality is related to severe neurologic involvement or rupture of
vascular aneurysms
169
Behcet syndrome: Pathogenesis (hypothetical)

- there may be an association with certain HLA haplotypes
- such as HLA-B51
- and a cross-reactive immune response to certain microorganisms
(example: H. pylori) is implicated
- therefore, an abnormal immune response is triggered resulting in
formation of numerous auto-antibodies (example: anti-endothelial
cell antibody)
- T-cells also activates release of cytokines
- all of these result in inflammation
170
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Behcet syndrome: Clinical course

Clinical Course:
- recurrent oral ulcers (aphthous ulcers)
- genital ulcers which healing with scarring
- bilateral uveitis
- differential diagnosis: Reiter syndrome
Treatment:
- Immunosuppression with steroids or TNF-antagonist
171
Granulomatosis with Polyangiitis

Introduction:
- classic triad:
1) Necrotizing granulomas of the upper respiratory tract
2) Necrotizing or granulomatous vasculitis affecting small- to medium-
sized vessels (mostly in Lungs)
3) Focal necrotizing, often crescentic, glomerulonephritis
- older terminology: Wegener granulomatosis (no longer used)
172
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Granulomatosis with Polyangiitis: Pathogenesis

- it is a T-cell mediated hypersensitivity response
- to normally “innocent” inhaled microbial or other environmental
agents
- which then leads to granuloma formation
- PR3-ANCAs are also present in up to 95% of cases
- therefore, a useful marker of disease
remember: following immunosuppression, a rising PR3-ANCA titer is
often a harbinger of relapse
- most patients in remission have a negative test or falling titers
173
Granulomatosis with Polyangiitis: Morphology

Upper respiratory tract lesions:
- lesions are surrounded by granulomas with geographic patterns of
central necrosis and accompanying vasculitis
- around the necrotizing granulomas there is a zone of proliferating
fibroblasts associated with giant cells and leukocytic infiltrate
- numerous granulomas can coalesce to produce nodules that can
cavitate
Remember: Caseating granulomas in TB may look very similar to
granulomas found in Granulomatosis with polyangiitis
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Granulomatosis with Polyangiitis: Morphology
Upper respiratory tract lesions (cont’d):

- late stage disease may be marked by extensive necrotizing
granulomatous involvement of the parenchyma
- and alveolar hemorrhage may be prominent
Renal Lesions:
Early: glomeruli exhibit only focal necrosis with isolated capillary loop
thrombosis (focal and segmental necrotizing glomerulonephritis)
Advanced: parietal cell proliferation resulting in crescent formation:
Crescentic glomerulonephritis
175
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Granulomatosis with Polyangiitis
177
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Glomerulus: normal histology
179
Glomerulus;
Normal vs. Focal segmental sclerosis
180
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Crescentic GN
181
Granulomatosis with Polyangiitis: Clinical Features
- affects males more than females

- average age around 40 years
Classic features include:
1) persistent pneumonitis with bilateral nodular and cavitary infiltrates
(95%)
2) chronic sinusitis (90%)
3) mucosal ulcerations of the nasopharynx (75%)
4) renal disease (80%).
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Granulomatosis with Polyangiitis: Clinical Features
- if the patient is untreated

- disease is usually rapidly fatal, with 80% mortality within 1 year
Treatment:
- corticosteroids, cyclophosphamide, and more recently TNF
antagonists
183
Thromboangiitis obliterans
Introduction:
- is characterized by segmental, thrombosing, acute and chronic
inflammation of medium-sized and small arteries
- commonly affects tibial and radial arteries
- also referred to as Buerger disease
- Asian population
Incidence:
- seen exclusively in heavy cigarette smokers, usually before age 35
- more in males
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Thromboangiitis obliterans: Pathogenesis and

Morphology
- strongly linked to cigarette smoking
- possible direct endothelial cell toxicity due to some component of
tobacco
- or, an immune response to components of tobacco smoke that
modify host vascular wall proteins
Morphology:
- focal acute and chronic vasculitis of small- and medium-sized arteries
- affecting mostly the extremities
- on microscopy: acute and chronic inflammation, accompanied by
luminal thrombosis
185
TAO Histology
186
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Thromboangiitis obliterans: Clinical Features

Early features:
- cold-induced Raynaud phenomenon
- leg pain induced by exercise that is relieved on rest (intermittent
claudication)
- instep foot pain induced by exercise (instep claudication)
- superficial nodular phlebitis (venous inflammation)
Chronic features:
- extremity ulcerations develop, progressing over time (occasionally
precipitously) to frank gangrene
187
Vasculitis: Classification
Based on Pathogenesis:
1) Non-infectious vasculitis
2) Infectious
- Bacteria (Pseudomonas)
- Fung (Aspergillus and Mucor)
- Hematogenous spread of microorganisms during septicemia
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“the most exhausting thing in life is being insincere”

Anne Morrow Lindbergh, writer (1906-2001)
189
CVS: Vascular tumors.

Veins and Lymphatics
Dr. Roy
190
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Classification of Tumors
191
Benign and Malignant Vascular tumors

- the difference between Benign and Malignant vascular tumors is that:
Benign tumors most often produce obvious vascular channels filled with
blood cells, lined by a monolayer of normal-appearing endothelial cells
Malignant tumors tend to be more cellular and more proliferative, and

exhibit features of malignancy
- sometimes, in malignancy distinct vascular lumina need to be confirmed by
immunohistochemistry: endothelial cell-specific markers (CD31 or von
Willebrand factor)
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Benign Tumors and Tumor-like conditions

Vascular Ectasias
- meaning of “ectasia”, and “telangietasia”:
Ectasia: means any local dilation of a structure
Telangiectasia: means a permanent dilation of preexisting small vessels

(capillaries, venules, and arterioles) that form discrete red lesion
Example of Ectasia: Nevus flammeus
193
Vascular Ectasias
Nevus flammeus
- seen as a “birthmark”, very common
- presents as light pink to deep purple flat lesion on the head or neck
composed of dilated vessels
- most of these regress spontaneously
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Port wine stain

Vascular Ectasias
Special types
1) “port wine stain”
-these lesions tend to grow during
childhood, thicken the skin surface,
and do not fade with time
195
Vascular Ectasias
Nevus flammeus
Special types
2) Sturge-Weber syndrome
- not common, it is congenital
associated with
- facial port wine nevi
- ipsilateral venous angiomas in the
cortical leptomeninges
- mental retardation, seizures,
hemiplegia, and skull radio-opacities
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Telangiectasias
Spider telangiectasias
- these are non-neoplastic vascular lesions, which on clinical examination
appear spider-like. These arise from pre-existing blood vessels
- with radial, and often pulsatile arrays of dilated subcutaneous arteries or
arterioles (spider legs),
- around a central core (spider’s body)
- spider telangiectasias blanch with pressure
- most are seen on the face, neck, or upper chest
- frequently associated with hyperestrogenic states, such as pregnancy or
liver cirrhosis
197
Telengiectasias
Hereditary hemorrhagic telangiectasia
- also referred to as: Osler-Weber-Rendu disease
- rare, autosomal dominant, mutations involving TGF-β signaling pathway
Clinical Features:
- telangiectasias (malformations composed of dilated capillaries and veins)
- which are present at birth
- involving skin and oral mucous membranes, respiratory, gastrointestinal,
and urinary tracts
- major complications: spontaneous rupture causing epistaxis,
gastrointestinal bleeding, or hematuria
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Telangiectasia: Spider nevi
Telangiectasia: Hereditary
Hemorrhagic Telangiectasia (Osler
Weber Rendu)
An elderly Miss Muffett

Decided to rough it
On Whiskey and Gin
Red Palms and little spiders
Developed outside her
Such are the wages of sin
(Bailey & Love Surgery)
199
Benign Tumors and Tumor-like conditions

Hemangiomas
Incidence:
- common tumors characterized by increased
numbers of normal or abnormal vessels filled
with blood
- most are present from birth and initially
increase in size, but many eventually regress
spontaneously
Clinical Features:
- most are localized lesions seen in the head
and neck,
- sometimes more extensive: angiomatosis,
and can occur internally
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Hemangiomas
Types based on histology:
1) Capillary hemangiomas
- are the most common type,
- seen in the skin, subcutaneous tissues,
and mucous membranes of the oral
cavities and lips
- also see to occur in the liver, spleen, and
Capillary Hemangioma
kidneys
Histology: unencapsulated, composed of
thin-walled capillaries with scant stroma
201
Hemangiomas: based on histology

2) Cavernous hemangiomas
Histology:
- unencapsulated, show ill-defined borders
- composed of large, cavernous blood-filled vascular
spaces separated by connective tissue stroma
Clinical Features:
- composed of large, dilated vascular channels
- when compared to capillary hemangiomas,
cavernous hemangiomas are more infiltrative,
frequently involve deep structures, and do not
spontaneously regress
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Hemangiomas
3) Juvenile hemangiomas
- also referred to as: “strawberry type”
hemangiomas
- seen in newborn, extremely common, and can
be multiple
Clinical Features:
- seen in the skin, grow rapidly for a few months
- but then fade by 1 to 3 years of age and
- completely regress by age 7 in the vast
majority of cases
203
Lymphangiomas
- are considered to be benign lymphatic counterparts of hemangiomas

Cystic hygromas
- sometimes referred to as: cavernous lymphangiomas
- classically seen in the neck or axilla of children
- may sometimes be enormous (up to 15 cm in diameter) and may fill
the axilla or produce gross deformities about the neck
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Lymphangiomas
Cystic Hygroma (cont’d)
Clinical Importance: cavernous lymphangiomas of the neck are
common in Turner syndrome
Histology:
- massively dilated lymphatic spaces lined by endothelial cells
- separated by intervening connective tissue stroma containing
lymphoid aggregates
- margins are indistinct and unencapsulated, making definitive
resection difficult
205
Cystic Hygroma
206
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Glomus tumors
Introduction:
- rare benign tumors, arises from neuro-arterial structure called a
“glomus body”
- normal glomus body is located in the stratum reticulare throughout
the body, but is more concentrated in the digits (fingers)
- probably plays a role in thermoregulation
- age group: young adults
Pathogenesis:
- glomus tumors arise from smooth muscle cells rather than endothelial
cells
207
Glomus tumors
Histology:
- organoid proliferation of small
vascular channels
Clinical Features:
- typically subungal, very painful
- complete excision is curative
208
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Bacillary Angiomatosis
Introduction:
- vascular proliferation seen in immunocompromised (AIDS)
Pathogenesis:
- caused by Bartonella family (opportunistic gram-negative bacilli)
- Bartonella henselae: causative organism for cat-scratch disease
- Bartonella quintana: causative organism for of Trench fever
- formerly known as Rochalimaea quintana
209
Bacillary Angiomatosis
Histology:
- capillary proliferation with
prominent epithelioid endothelial
cells exhibiting nuclear atypia and
mitoses
- heavy infiltrate of neutrophils
Clinical Features:
- presents as multiple red papules
Warthin-Starry stain: to demonstrates bacilli
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Intermediate-grade (borderline) tumors:

Kaposi Sarcoma
- vascular neoplasm caused by human herpesvirus 8 (HHV8)
- which is highly associated with AIDS
- mostly sexually transmitted, and by poorly understood non-sexual
routes (? oral secretions)
1) Classic KS: uncommon in the United States
- most associated with malignancy or altered immunity,
- but is not associated with HIV infection
2) Endemic African KS: typically occurs in HIV-seronegative individuals
younger than age 40
211
211
Kaposi Sarcoma
3) Transplant-associated KS: occurs in solid organ transplant recipients
- in the setting of T-cell immunosuppression
4) AIDS-associated (epidemic) KS:
- is an AIDS-defining illness
- represents the most common HIV-related malignancy
- AIDS-associated KS often involves lymph nodes and disseminates
widely to viscera early in its course
- most patients eventually die of opportunistic infections rather KS
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Kaposi Sarcoma: Morphology

- classic KS, or sometimes in other variants
- the cutaneous lesions progress through three stages:
Patches are red-purple macules typically confined to the distal lower
extremities
Raised plaques composed of dermal accumulations of dilated, jagged
vascular channels lined and surrounded by plump spindle cells
Nodular and more distinctly neoplastic
- these lesions are composed of sheets of plump, proliferating spindle cells,
mostly in the dermis or subcutaneous tissues
- encompassing small vessels and slit-like spaces containing red cells 213
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Kaposi Sarcoma
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Malignant Tumors
Angiosarcoma
Incidence:
- it is a malignant endothelial neoplasm which
mostly affects older adults
- most often involves skin, soft tissue, breast, and
liver
Etiopathogenesis:
1) Hepatic angiosarcoma
- exposure to chemical carcinogens: arsenic
(pesticides), Thorotrast (radioactive contrast)
polyvinyl chloride (plastic)
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Malignant Tumors: Angiosarcoma

Etiopathogenesis:
2) Lymphangiosarcoma
- seen in the setting of lymphedema
- following radical mastectomy (with en-bloc
axillary lymph node resection for breast cancer)
- malignancy arise from lymphatic channels
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Morphology:
- begin as multiple deceptively small and asymptomatic red papules or
nodules
- on histology, full range of differentiation notes: from plump, atypical
endothelial cells forming vascular channels
- to undifferentiated tumors with a solid spindled appearance and no
blood vessel formation
- endothelial origin can be demonstrated by CD31 or von Willebrand
factor positivity
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Clinical Features:
Hemangiosarcoma
- of liver is an occupational related disease
- uncommon
Lymphangiosarcoma
- seen with long standing lymphedema
- lymphedema following post-mastectomy in the most common setting
- aggressive malignancy
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Varicose Veins
Thrombophebitis and Phlebothrombosis
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Veins: Varicose veins

Definition:
- abnormally distended and often tortuous veins that arise underneath the
skin or mucosal surface
Etiology:
- the superficial veins of the lower leg are commonly involved
- because venous pressures in these sites can be markedly elevated (up to 10
times normal) by prolonged dependent posture (standing)
- other risk factors include: obesity and pregnancy
Esophageal Varices
- liver cirrhosis (portosystemic shunts- left gastric vein anastomoses with the
esophageal vein)
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Varicose Veins
Varicose Veins: surgical treatment
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Porto-systemic shunts: Alcoholic Cirrhosis
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Thrombophlebitis and Phelbothrombosis

Introduction:
- both these terms are interchangeable, means venous thrombosis and
inflammation
Sites:
- seen mostly (90%) in deep leg veins
in males: periprostatic venous plexus
in females: pelvic venous plexus in females
Etiology:
- prolonged immobilization resulting in venous stasis
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Thrombophlebitis and Phelbothrombosis

Clinical Features:
Phlegmasia alba dolens
- painful white leg (milk leg)
- triggered by ileo-femoral venous thrombosis
- seen in the setting of peri-partum, due to pressure of gravid uterus
on pelvic veins leading to venous stasis, and increased
hypercoagulability
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Phlegmasia alba dolens: remember initially the

affected leg is pale, but later it reddens.
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“true religion is the life we lead, not the creed we profess”

Louis Nizer, lawyer (1902-1994)
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Immunopathology
Dr. Roy
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Introduction
- immune reactions against ‘self-antigens’
- there is loss of self-tolerance
What are self-antigens?
- refers to lack of responsiveness to an individual’s own antigens, therefore our
immune system will not attack our own antigens
What are considered as self-antigens?
- these include: Class I and Class II MHC antigens, nuclear antigens, and cytoplasmic
antigens
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Introduction
How does the body manage Immune Tolerance?
- mechanisms include 1) Central Tolerance, and 2) Peripheral tolerance
How does ‘Central Tolerance’ works?
- immature self-reactive T, and B lymphocyte clones that recognize self-antigens
are removed at their maturation sites.
- T-cells at Thymus, and B-cells at bone marrow
More detailed overview of Central Tolerance
- in T-cells , somatic gene rearrangements generate diverse TCRs, when these
immature lymphocytes encounter the antigens in the thymus, many of the cells
die by apoptosis
- this process, called negative selection or deletion
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Introduction
More detailed overview of Central Tolerance

- in B-cells, if they strongly recognize self antigens in the bone marrow,
- many of the cells reactivate the machinery of antigen receptor gene
rearrangement and begin to express new antigen receptors, not specific for self
antigens
- this process is called receptor editing
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Introduction
What is Peripheral Tolerance?
- similar mechanisms exist in the peripheral tissues
- as the Central Tolerance may not be perfect
What are the ways in which Peripheral Tolerance works?
Anergy:
- neutralizing self-reactive lymphocytes
T-cells requires two signals: one; recognition of peptide antigen in association with
self MHC molecules on the surface of APCs and a set of costimulatory signals
(“second signals”) from antigen presenting cells. If these cells do not express
c0stimulatory signals, they will be “anergic”
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Introduction
What are the ways in which Peripheral Tolerance works?

Anergy:
- similar mechanism works in B-cells too
- if B-cells encounter self antigen in peripheral tissues, especially in the absence of
specific helper T cells,
- these B-cells become unable to respond to subsequent antigenic stimulation and
may be excluded from lymphoid follicles, resulting in their death
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Introduction
Are there any other mechanisms which play a role in Peripheral Tolerance?
- yes, for the T-cells there are additional mechanisms
1) Suppression by regulatory T-cells
- population of T cells called regulatory T cells functions to prevent immune
reactions against self antigens
- these regulatory T-cells develop in Thymus in response to recognizing self-
antigens
- and these cells can be induced in the ‘peripheral’ tissues too
- for example: CD4+ cells
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Introduction
Are there any other mechanisms which play a role in Peripheral Tolerance?
2) Deletion by apoptosis
- T- cells that recognize self antigens may receive signals that promote their death
by apoptosis
- Two mechanisms of deletion: if T-cells recognize self antigens, they may express
a pro-apoptotic member of the Bcl family, called Bim, without antiapoptotic
members of the family, unopposed Bim triggers apoptosis by the mitochondrial
pathway
- second mechanism: activation-induced death of CD4+ T cells and B cells involves
the Fas-Fas ligand system
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Understanding ”autoimmune disorders”

- autoimmunity arises from a combination of the inheritance of
susceptibility genes, which may contribute to the breakdown of self-
tolerance, and environmental triggers, such as infections and tissue
damage, which promote the activation of self-reactive lymphocytes
Defective tolerance or regulation
- failure of the mechanisms that maintain self-tolerance
Abnormal display of self antigens
- abnormalities may include increased expression and persistence of
self antigens that are normally cleared, or structural changes in these
antigens resulting from enzymatic modifications or from cellular
stress or injury
- if these changes lead to the display of antigenic epitopes that are not
expressed normally, the immune system may not be tolerant to these
epitopes, thus allowing anti-self responses to develop
Inflammation or an initial innate immune response
- microbes or cell injury may elicit local inflammatory reactions
resembling innate immune responses, and these may be critical
inducers of the autoimmune disease
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Immunofluorescence
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Autoimmune Diseases
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Systemic Lupus Erythematosus (SLE)
Introduction:
- multisystem disease of autoimmune origin
- presence of numerous ‘autoantibodies’ (anti nuclear antibodies: ANA)
- injury mostly due to deposition of immune complexes
- acute in onset, thereafter it becomes a chronic, remitting and relapsing disease
- targets: skin, joints, kidney, and serosal membranes
Incidence:
- predominantly women (9:1; female to male)
- 2 to 3 fold higher in Hispanics, and African-Americans
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SLE: Etiology
Etiology:
- fundamental defect in SLE is a failure of mechanism that maintain self-tolerance
- factors that play:
1. Genetic Factors:
- evidences to support a genetic association
a) higher incidence among family members
b) higher incidence among monozygotic twins
c) HLA association: HLA-DQ locus associated with production of
- anti-double stranded DNA (ds)
- anti- Smith (Sm)
- antiphospholipid antibodies
d) in some cases inherited deficiencies of early complement factors (C2, C4, or C1q). Lack of complement
impair removal of circulating immune complexes
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SLE: Etiology
2. Immunological Factors
- this leads to persistence and uncontrolled activation of self-reactive lymphocytes
Failure of self-tolerance of B-cells
- due to defective elimination of self-reactive B-cells in the bone marrow or defects in
peripheral tolerance mechanisms
Failure of CD4+ helper T-cells
- specific for nucleosomal antigens also escape tolerance and contribute to the production
of high-affinity pathogenic autoantibodies
- these autoantibodies in SLE show characteristics of T cell-dependent antibodies
produced in germinal centers, and increased numbers of follicular helper T cells have
been detected in the blood of SLE patients
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SLE: Etiology
2. Immunological Factors (cont’d)
TLR engagement by nuclear DNA and RNA
- these contained in immune complexes may activate B-lymphocytes
- TLRs function normally to sense microbial products, including nucleic acids
- therefore, B-cells specific for nuclear antigens may get second signals from TLRs
and may be activated, resulting in increased production of antinuclear
autoantibodies
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SLE: Etiology
2. Immunological Factors (cont’d)
Type-I interferons:
- these play a role in lymphocyte activation in SLE
- high levels of circulating type I interferons is noted in SLE patients and correlates
with disease severity
- Type-I interferons are anti-viral cytokines that are normally produced during
innate immune responses to viruses
- self-nucleic acids mimic their microbial counterparts
- molecular mimicry
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SLE: Etiology
3. Environmental Factors
a) Exposure to UV rays:
- cells exposed to UV rays undergo apoptosis
- during the apoptotic process, these cells become immunogenic
- thereby inducing an immune response
- UV rays modulate immune responses, by activating keratinocytes to produce IL-1
- IL-1 will then promote inflammation
b) Estrogen:
- may stimulate B cells to produce antibodies directed against DNA
- pregnancy may aggravate SLE
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SLE: mechanisms of disease process
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SLE: Autoantibodies
1) Antinuclear antibody (ANA)
- Ag-Ab complexes found in circulation
- classically produce Type III hypersensitivity reaction
- these Ag-Ab complexes also activate complement system ( C3, and C4 levels are
decreased)
- these are positive in other diseases, and are seen in 5% of normal population
- test is sensitive but not specific
- if the test is negative, SLE is then unlikely
- test is reported in titers (1:160 is considered positive)
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SLE: Autoantibodies
2) Anti double stranded DNA (anti-dsDNA)
- this is specific for SLE, and is associated with disease activity (as in flare up)
- associated with renal involvement
3) Anti-Smith antibody
- this test is also specific for SLE
- antibodies are directed against small nuclear ribonucleoprotein (snRNP)
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SLE: Autoantibodies
4) Autoantibodies specific for rbc, white cells, and platelets
- antibody directed against red blood cells: hemolytic anemia
- antibody directed against leukocytes: leukopenia
- antibody directed against platelets: thrombocytopenia
5) Antiphospholipid antibody
- antibody directed against proteins present in phospholipids
- patients would have increased risk for thrombotic events (DVT)
- lupus anticoagulant, leads to increased PTT (falsely elevated)
- false positive syphilis (RPR, VDRL)
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Criteria for diagnosis of SLE. 4 out 11 must be present

1) Malar Rash
2) Discoid rash
3) Photosensitivity
4) Oral ulcer
5) Arthritis
6) Serositis
7) Cerebritis
8) Cytopenias (anemia, leukopenia, thrombocytopenia)
9) Renal disease
10) Anti-ds DNA
11) Anti-Sm
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SLE: Drug Induced SLE

- SLE (lupus)-like features associated with
drug administration
- examples: INH, Hydralazine,
Procainamide
- patients present with rash classically
seen in forearm (discoid), arthritis,
cytopenias, positive for ANA
- diagnosis: clinical features, drug history,
and presence of anti histone antibodies
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Antibody Features
Anti RNP MCTD, SLE, Systemic sclerosis
Anti Smith Specific for SLE
Anti SS-B (La) Sjorgen syndrome, SLE, Systemic sclerosis
Scl-70 Specific for Systemic sclerosis

Anti-Jo1 Dermatomyositis
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LE cells represent phagocytosis by neutrophils of

apoptotic bodies, most likely induced by antinuclear
antibodies
-damaged cells in tissue expose their nuclei to ANA (anti
nuclear antibodies)
-LE bodies or hematoxylin bodies: damaged nuclei
-LE cell is any phagocytic leukocyte that has engulfed
denatured nucleus of an injured cell
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SLE: Morphology
1) Blood vessels
- acute necrotizing vasculitis involving
capillaries, small arteries and arterioles
- these may be present in any tissue.
- characterized by fibrinoid necrosis
- during chronic stages, vessels undergo fibrous
thickening with luminal narrowing
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SLE: Morphology
2) Kidney
- nearly 50% of SLE patients have renal
involvement
- these are due to deposition of
immune complexes present in the
mesangium or along the entire
basement membrane
- both: in situ formation, and
deposition of preformed circulating
immune complexes may be noted
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SLE: Morphology
2) Kidney (cont’d):
Morphological classification: Class I to Class VI
Class I: minimal measngial lupus nephritis
Class II: mesangial proliferative lupus nephritis
Class III: focal lupus nephritis
Class IV: diffuse lupus nephritis
Class V: membranous lupus nephritis
Class VI: advanced sclerosing lupus nephritis SLE (Class IV): deposits thickening capillary
walls.
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Diffuse Lupus Nephritis (Class IV)

compared to Normal
Normal 260
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SLE: Morphology
3) Skin:
- erythema: butterfly rash (malar) in 50%
- exposure to sunlight accentuates the
erythema
Histology:
- vacuolar degeneration of basal cells in
epidermis
- dermis show edema, perivascular
inflammation
- vasculitis with fibrinoid necrosis
SLE: skin biopsy
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Fibrinoid necrosis
Skin: immunofluorescence. Lupus band
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SLE: Morphology
4) Joints:
- produces a non-erosive synovitis with little deformity, this is different from
rheumatoid arthritis
5) CNS:
- neuropsychiatric symptoms are noted in patients with SLE
- may be associated with acute vasculitis (cerebritis)
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SLE: Morphology
6) Cardiovascular system:
Pericarditis- acute, subacute, or chronic forms are see. During the acute phases, the
mesothelial surfaces are sometimes covered with fibrinous exudate
Heart- any layer may be involved
valvular (Libman-Sacks) endocarditis:
- characterized by non-bacterial verrucous endocarditis
- single or multiple 1- to 3-mm warty deposits on any heart valve, distinctively on
both surfaces of the leaflets
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RHD: Rheumatic Heart Disease (small warty vegetations, along the lines of
closure of valve leaflets)
IE: Infectious Endocarditis (large irregular masses on the valve cusps that can
extend into the chordae)
NBTE: Non Bacterial Thrombotic Endocarditis (small bland vegetations usually

attached to the lines of closure
LSE: Libman Sacks Endocarditis (small or medium-sized vegetations on either

or both sides of the valve leaflets)
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SLE: Diagnosis
Labs:
1) Serum ANA: high titers are generally more specific for SLE (>1 : 160) than other
autoimmune diseases. (sensitivity 98%)
2) Anti-dsDNA antibodies: most often used to confirm the diagnosis of SLE. (95%
specificity, and sensitivity 70%–98%)
3) Anti-Smith antibodies: used to confirm the diagnosis of SLE (99% specificity; rare false
positives)
4) Anti-Ro (SS-A) antibodies and anti-La (SS-B) antibodies: these have a low sensitivity
(26%–50% and 5%–15%, respectively)
5) APL antibodies (sensitivity 30%–40%)
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SLE: Diagnosis
Labs:
6) Anti-histone antibodies: especially for Procainamide (anti-histone antibodies
shows 96% sensitivity)
7) Serum complement: usually decreased because of activation of the complement
system by immune complexes
8) Erythrocyte sedimentation rate (ESR) is increased in active SLE
9) Immunofluorescence testing: skin, and renal biopsy
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SLE: malar rash (butterfly)
SLE: Class IV changes: narrowing of

glomerular capillaries by mesangial
and endocapillary proliferation.
Wire-loop deposits and hyaline
thrombi are segmentally
distributed
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Lupus erythematosus (L-E) cell. Pleural fluid. A neutrophil with an ingested large
nucleus (nucleophagocytosis) (arrow) compressing the nucleus of the neutrophil
(asterisk). This appearance is characteristic of the L-E cell found in the blood,
marrow, or serous effusions in patients with lupus erythematosus. Search for such
cells as diagnostic clues has been replaced by other immunological tests (e.g.
antinuclear antibody).
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Joints (introduction)
- joints allow movement while providing mechanical stability
- classification: solid (non synovial, and cavitated (synovial)
- solid joints lack joint space (cranial sutures)
synovial joints
- have joint space, situated between ends of bone formed via enchondral
ossification
- boundaries of joint space consists of the synovial membrane, which is firmly
anchored to the underlying capsule, and
- does not cover the articular surface
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Joints (introduction)
synovial joints
- synovial membranes are lined by two types of cells that are arranged 1 to 4 layers deep
Type A synoviocytes: are specialized macrophages with phagocytic activity.
Type B synoviocytes: similar to fibroblasts and synthesize hyaluronic acid and various
proteins
- synovial membrane lacks a basement membrane, this allows efficient exchange of
nutrients, waste and gasses between blood and synovial fluid
Synovial fluid:
- is a plasma filtrate containing hyaluronic acid that act as a viscous lubricant and provides
nutrition for articular hyaline cartilage
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Rheumatoid arthritis
Introduction:
- chronic inflammatory disease, characterized by symmetric, peripheral
polyarthritis
- inflammatory synovitis resulting in destruction of articular cartilage and ankylosis
(fusion) of joints
- increases between 25 and 55 years, more in females
- small joints of hands and feet (peripheral joints)
hands: symmetric involvement of proximal interphalangeal (PIP) and
metacarpophalangeal (MCP) joints
remember: distal interphalangeal joint (DIP) is usually spared (unlike osteoarthritis)
extra articular lesions may involve skin, heart, blood vessels, lungs
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Rheumatoid Arthritis: Pathogenesis
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Rheumatoid arthritis: Pathogenesis
1. Immune factors
- CD4+ T helper (TH) cells may initiate autoimmune response by reacting with an
‘arthritogenic’ agent
- arthritogenic agent: ?microbial, ?self-antigen
- T cells then produce cytokines
- these stimulate other inflammatory cells to effect tissue injury
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1. Immune factors (cont’d)

most important ones are:
IFN-ɣ: from TH1 cells activate macrophages and resident synovial cells
IL-17: from TH17 cells recruit neutrophils and macrophages
TNF and IL-1: from macrophages stimulate resident synovial cells to secrete
proteases that destroy hyaline cartilage
RANKL: expressed on activated T cells stimulate bone absorption
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Autoantibodies
- directed against citrullinated peptides (CCPs)
- antibodies directed against CCPs are diagnostic
- 80% of RA patients have serum IgM, or IgA autoantibodies (rheumatoid factor)
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2. Genetic
- 50% of the risk for RA, is genetic
- HLA-DRB1 alleles are linked to RA
- PTPN22 (encodes for a protein tyrosine phosphatase that inhibit T-cell activation)
3. Environmental
- smoking may promote citrullination of self-antigens
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- the pathologic hallmarks of RA

are synovial inflammation
(typically symmetric) and
proliferation, focal bone
erosions, and thinning of
articular cartilage
- chronic inflammation leads to
synovial lining hyperplasia and
the formation of pannus
Note: synovial thickening, & chronic

inflammation
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Pannus: is a thickened cellular membrane of granulation–reactive fibrovascular
tissue that grows over the articular cartilage and causes erosion
- once articular cartilage is destroyed, Pannus bridges opposing bones to form
‘fibrous ankylosis’
Bone damage:
1. structural damage to the mineralized cartilage and subchondral bone is
mediated by the osteoclast
2. another form of bone loss is peri-articular osteopenia that occurs in joints with
active inflammation
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Rheumatoid arthritis: Morphology

Joints
Gross: edematous synovium, which becomes thickened, and hyperplastic
Histology:
- synovial cell hyperplasia and proliferation
- dense inflammatory infiltrates (with lymphoid follicles)
- increased vascularity due to angiogenesis
- fibrinopurulent exudate on the synovial and joint surfaces
- osteoclastic activity in underlying bone (this allows synovium to penetrate into
the bone and cause peri-articular erosions and subchondral cysts)
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Morphology:
Skin:
Rheumatoid nodules: sites of pressure- ulnar aspect of the forearm, elbows, occiput,
lumbosacral area
- firm, non-tender round to oval skin lesions
Histology: central zone of fibrinoid necrosis, surrounded by epithelioid cells & numerous
lymphocytes and plasma cells
Blood vessels:
Rheumatoid vasculitis:
- affects medium to small sized blood vessels (kidney not involved)
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Rheumatoid nodule: note fibrinoid necrosis, surrounded by epithelioid cells.
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Clinical features:
- small joints of hands and feet
- early morning stiffness (more than 1 hour after arising)
Wrist:
- metacarpophalangeal (MCP), and proximal interphalangeal (PIP) joints stand out
as the most frequently involved joints
- flexor tendon tenosynovitis is a frequent hallmark of RA and leads to decreased
range of motion, reduced grip strength, and “trigger” fingers
- ulnar deviation results from subluxation of the MCP joints, with subluxation of
the proximal phalanx to the volar side of the hand
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Hyperextension of the PIP joint with flexion of the DIP joint (“swan-neck deformity”)
Flexion of the PIP joint with hyperextension of the DIP joint (“boutonnière deformity”)
Swan neck deformity boutonnière deformity
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Baker cyst:
- popliteal (Baker) cyst develops posteriorly and inferiorly to the knee as a distention of a local bursa
- diagnosis may be made by aspiration of mucinous fluid
- when rupture produces pain (D/D: DVT)
Rheumatoid nodule
- seen as nodules over the extensor surface of the fingers, forearms, and pressure points (Achilles tendon,
olecranon process) and in the lungs
- on microscopy show necrobiosis, and fibrinoid necrosis
- often associated with patients who are RF positive
Hematologic findings:
- Anemia of chronic disease (ACD)
Carpal tunnel syndrome:
- entrapment of median nerve under the transverse carpal ligament
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Rheumatoid Arthritis:
Baker’s cyst
Rheumatoid nodule
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Atlantoaxial joint
- cervical spine is commonly
involved
- cervical instability is the most
serious and potentially lethal
manifestation
- the most common problem:
subluxation at C1-C2
- muscle wasting, decreased range of
motion, and compressive
neuropathy
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- Subluxation of cervical spine in
rheumatoid arthritis.
A) Flexion, showing widening of the space
(arrow) between the odontoid peg of the
axis (behind) and the anterior arch of the
atlas (in front)
B) Extension, showing reduction in this
space
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Labs:
- Rheumatoid Factor (RF): most common is IgM
- Normal to increased serum C3
- antibodies to cyclic citrullinated peptides (ACAPA)
- increased C-reactive protein (CRP), and increased ESR
- association with HLA-DR4
Synovial fluid analysis:

- presence of white cells (tend to be neutrophils)
- remember: synovium contains chronic inflammatory cells
- also note: C3 levels are low in synovial fluid
- RF can be detected in synovial fluid
Radiology: Plain X-ray, USG, MRI

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antibodies to cyclic citrullinated

peptides (ACAPA)
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Felty Syndrome
Introduction:
- triad: splenomegaly, neutropenia, and
RA
- seen in patients who have severe,
prolonged RA
- repeated infections due to decreased
neutrophils
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Complications
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Sjögren Syndrome
Introduction:
- chronic, slowly progressive autoimmune disease
- characterized by lymphocytic infiltration of exocrine glands
- resulting in dry mouth (xerostomia), and dry eyes (keratoconjunctivitis)
Incidence:
- middle aged women
- Female:Male is 9:1
- increased association with: Rheumatoid arthritis, Systemic lupus erythematosus, Scleroderma, Mixed
connective tissue disease, Primary biliary cirrhosis, Vasculitis, Chronic active hepatitis
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Sjögren Syndrome
Pathogenesis:
- characteristic feature: decrease in tears and saliva (sicca syndrome)
- due to lymphocytic infiltrate and fibrosis of lacrimal and salivary gland
- CD4+ helper T-cells predominate the inflammation, along with B cells, and plasma cells
- lymphocytic sialadenitis (Type IV Hypersensitivity reaction)
- also antinuclear antibodies such as rheumatoid factor , and other are present
- most important are
1) SS-A (Ro)
2) SS-B (La)
- these two are seen in over 90% of the patients
- HLA association: HLA-B8, HLA-DR3
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Sjögren Syndrome
Morphology:
Salivary glands:
- periductal and perivascular lymphocytic infiltration
- lymphoid follicles with germinal centers may appear
- ductal lining cells show hyperplasia, this leads to obstruction
- later there is acinar atrophy, fibrosis, and hyalinization
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Salivary gland: lymphocytic infiltrates, interstitial fibrosis, and acinar

atrophy
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Sjogren synd: Note the asymmetric swelling of the

left parotid gland
Note: dense lymphocytic inflammation, fibrosis305
305
Salivary gland: normal and in Sjorgen

syndrome
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Sjögren Syndrome
Clinical features:
- dryness of mouth/eye
- dental caries
- keratoconjunctivitis sicca: dry eyes
- Xerostomia: dry mouth
- histological diagnosis: lip biopsy/salivary gland biopsy
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Sjögren Syndrome
Labs:
antibodies in serum:
- SS-A(Ro), SS-B(La), positive RF, positive ANA (not
specific)
Schrimer test:
- amount of wetting measured in mm over 5 minutes
Salivary gland scintigraphy:
- low uptake of radionucleotide
Whole sialometry:
- measures volume and weight of saliva
Complication:
- risk for low grade B-cell lymphoma (Extra nodal marginal
zone lymphoma)
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Diagnosis of Sjogren Syndrome: 4 out of 6
1) Eye symptoms
2) Ocular symptoms
3) Ocular signs: Schrimer test
4) Salivary testing
5) Biopsy: Lymphocytic sialadenitis
6) Anti-SSa or Anti-SSb
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Systemic Sclerosis (scleroderma)

Introduction:
- characterized by
1) chronic inflammation (autoimmune),
2) widespread damage to small blood vessels, and
3) progressive interstitial and perivascular fibrosis in skin and multiple organs
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Etiology:
Autoimmunity: CD4+ T cells responding to unknown antigen in skin
- this activates inflammatory cells and fibroblasts
- cytokines from T-cells (TGF-β, IL-13)
Vascular damage:
- intimal proliferation, endothelial activation, and injury
Fibrosis:
- activated macrophages, fibrogenic cytokines (PDGF, TGF-β)
- fibroblast activation leading to increasing collagen deposition
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311

Morphology:
Skin:
- diffuse sclerotic atrophy, begins with fingers and distal regions of upper extremities
- later involve neck and face
- histology: edema, perivascular infiltrates (CD4+ T cells)
- swelling and degeneration of collagen (eosinophilic staining)
- capillaries and small vessels show thickened basal lamina, endothelial damage, and
partial occlusion
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Morphology:
Skin (cont’d):
- increasing fibrosis of dermis, tightly bound to subcutaneous structures
- increase in compact collagen, and thinning of epidermis, loss of rete pegs
CREST syndrome (limited type of Systemic Sclerosis)
C: calcinosis cutis
R: Raynaud’s phenomenon
E: esophageal dysfuntion
S: sclerodactly (tapered fingers, claw-like)
T: telangiectasia
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Note: stretched
parchment-like
skin, claw hand
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Note skin induration on the fingers, and fixed flexion contractures at the proximal
interphalangeal joints in a patient with progressive systemic sclerosis
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perivascular mononuclear cell infiltrate at the early stages of disease. Perivascular

changes are shown at high power in the left panel.
Later stage disease show skin sclerosis, a low density of blood vessels, and absence of
inflammatory cells. At this stage, there may be associated epidermal changes with
thickening and loss of secondary skin structures, including hair follicles and sweat
glands. Absence of the rete ridges is also characteristic at the later stages 317
317

Raynaud's phenomenon
- is an episodic vasoconstriction in the fingers and toes that occurs in virtually every
patient with scleroderma
- vasoconstriction may also affect the tip of the nose and earlobes
- attacks are triggered by exposure to cold, a decrease in temperature, emotional stress,
and vibration
- typical attacks start with pallor, followed by cyanosis of variable duration
- eventually erythema develops spontaneously or with rewarming of the digit
- progression of the three color phases reflects the underlying pathogenic mechanisms of
vasoconstriction, ischemia, and reperfusion
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Sharply demarcated necrosis of the fingertip in a patient with systemic

sclerosis associated with severe Raynaud's phenomenon
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319

Morphology:
GIT:
- progressive atrophy and collagenous fibrous replacement of the muscularis may
develop
- at any level of the gut but are most severe in the esophagus
- lower two thirds of the esophagus often develops a rubber-hose like inflexibility
- lower esophageal sphincter (LES) hypotonia gives rise to gastro-esophageal reflux
and its complications, including Barrett esophagitis (metaplasia)
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Morphology:
Musculoskeletal system:
- synovium show inflammation, hypertrophy & hyperplasia
- however unlike RA, no joint destruction
Kidney:
- vascular lesions (interlobular arteries show intimal thickening with mucinous or
collagenous material)
- concentric proliferation of intimal cells, similar to those seen in hypertension
- hypertension seen in 30% (in some complicated by malignant hypertension)
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323

Morphology:
Lungs:
- involved in 50% of cases
- pulmonary hypertension, & interstitial fibrosis
- pulmonary hypertension may progresses to right-sided heart failure
Heart:
- pericarditis with effusion, myocardial fibrosis
- thickening of intra-myocardial arterioles
- conduction disturbances (bundle branch blocks)
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Clinical Features:
- females, peak incidence- 50 to 60 years
- cutaneous changes- skin thickening
- Raynaud phenomenon
- dysphagia
- abdominal pain, malabsorption, anemia
- respiratory- features of right-sided heart failure, myocardial fibrosis, arrhythmia,
- hypertension
Lab:
- presence of ANA in all patients
1) Anti topoisomerase I (anti-Scl 70)
2) Anticentromere antibody (seen in 20 to 30% of patients who tend to have CREST syndrome)
3) Anti-RNA polymerase III antibody
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Dermatomyositis
Introduction:
- heterogeneous group of genetically determined autoimmune disorders that predominately target
- the skeletal musculature and/or skin and typically result in
- symptomatic skeletal muscle weakness and/or cutaneous inflammatory disease
Pathogenesis:
- immunologic disease in which there is damage to small blood vessels
- contributing to muscle injury
- vascular findings are present as telangiectasia (dilated capillary loops)
- seen in nail folds, eyelids, and gums
- biopsy of muscle/skin show deposits of complement: C5b-9
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Dermatomyositis
- antibodies associated include:
Anti-Mi2 antibodies: associated with Gottron papules
Anti-Jo1 antibodies: associated with interstitial lung disease, non-erosive arthritis,

skin rash
Anti-P155/P140 antibodies: associated with paraneoplastic & juvenile cases of

dermatomyositis
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Dermatomyositis
Morphology:
Muscle biopsy:
- mono-nuclear inflammatory cells, mostly around perimysial connective tissue and
around blood vessels
- perifasicular atrophy of muscle
Immunohistochemistry: inflammatory cells stain positive for CD4+ T-helper cells,
and
- deposition of C5b-9 in capillary vessels
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328
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Dermatomyositis
Note atrophic changes in muscle bundles in the marked out area
329
Skeletal muscle: normal and in

Dermatomyositis
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Dermatomyositis
Clinical Features:
- muscle weakness, slow onset
- often accompanied by myalgias
- getting up from chair, climbing stairs become difficult
- elevated serum creatinine kinase levels
- lilac colored discoloration of upper eyelids (heliotrope rash)
- scaling erythematous eruption or dusky red patches over knuckles/elbows/knee:
Gottron papules
- dysphagia
- interstitial lung disease (10%) 331
331
Note the confluent macular violaceous erythema, most

Dermatomyositis: heliotrope rash pronounced over the metacarpophalangeal/interphalangeal
joints, extending in a linear array overlying the extensor
tendons of the hand and fingers. These changes, referred to
as Gottron sign, are a hallmark cutaneous feature of DM
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“Conscience is thoroughly well-bred and soon leaves off talking to those who do
not wish to hear it”
-Samuel Butler, writer (1835-1902)
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167

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Cell injury- Part 1

Physiologic hypertrophy of the uterus during pregnancy. A,

Urinary bladder: note, thickening of the bladder wall

Remember: in myocardial growth

Mechanisms of hypertrophy (cont’d):

Increase in cell size, and nuclear size

vEtiology: 50% of cases

ii) Compensatory hyperplasia- regeneration of liver following partial

Examples: Endometrial Hyperplasia

- This is associated with anovulatory cycles

Increase in GnRH pulse Insulin resistance

Theca cells- androstenediones & Testosterone Arrest in antral follicle development

Adipose tissue (aromatase) Estrones & estradiols Endometrial hyperplasia

Prostate: Benign Prostatic

iii) Endocrine glands:

Thyroid Hyperplasia: Grave’s disease 29

v. Loss of endocrine stimulation:

Tracheo-bronchial: Normal histo Metaplasia: squamous 38

i) columnar to squamous (cont’d)

Endoscopy: shows tongues of reddened mucosa, indicative of Barrett’s

iii) intestinal metaplasia

Warthin Stany silver stain

Gastric mucosa: Intestinal metaplasia 47

Urinary bladder: note, squamous metaplasia of the mucosa. Note, presence of

Urinary bladder: squamous metaplasia 50

Myositis ossificans: note – extra skeletal calcification 51

• Metaplasia- one adult cell replaced by other

Buerger disease (Thromboangiitis Obliterans

Tissues susceptible to hypoxia:

ii) Subendocardial tissue (left ventricle in

iii) Renal cortex and medulla

iv) Purkinje cells in cerebellum 9

-(hypoxia, metabolic injury best

Sloughing of tubular cells

Renal tubules: cell swelling 14

Irreversible cell injury:

amorphous densities in mitochondria –

- cytosolic calcium is kept

Normal Pyknosis Karyorrhexis Karyolysis

Ø Mitochondria can be damaged by: increases in

Examples of free radical injury

Conversion to toxic metabolites:

It is called situs inversus totalis when

Up to 20% of patients with situs inversus

Light Cytoplam: eosinophilic

Electron Myelin figures, amorphous densities in

• Death of a tissue (many cells)

Case: elderly, history of MI

localized area of ischemia: infarct

Cell outline is preserved

coronary occlusion by thrombi - pale area of infarct

Locate the scar!

Walling- off the abscess

-Acute Pancreatitis (Enzymatic fat necrosis)

Shadowy necrotic fat cells

-Presents as acute abdomen (medical

Grey Turner’s sign

Hyper plastic arteriolosclerosis Fibrinoid necrosis :

• Single cells death.