2

SY 2021 -
CLINICAL
m HEMATOLOGY 1
1st
2022
SEMESTER
BS-MLS
MR. NICKSON CAMMAYO, RMT

TOPIC TITLE (EXAMPLE: ENZYMES)

OUTLINE
I HEMOGLOBIN & IRON KINETICS and RBC ABNORMALITIES
A. HEMOGLOBIN FUNCTION AND STRUCTURE & IRON KINETICS
B. RBC ABNORMALITIES
i. Subsub Topic 1
ii. SubSub Topic 2
a. pag meron pa
b. pag meron pa
c. pag meron pa

HEMOGLOBIN

HEMOGLOBIN FUNCTION AND STRUCTURE

Hemoglobin structure
 red globular protein, which have a molecular weight of
about 64,000
 compromise almost one third of the weight of a red
cell
 Each red cell contains approximately 640 million Hb
molecules
 composed of four subunits. Each subunit contains 1
One gram of hemoglobin can carry 1.34 mL of Oxygen
heme and 1 globin
o 1 Heme:1 moles of Oxygen 1 Hb: 4 moles of One gram of hemoglobin can carry a constant 3.47 mg of Iron
of Oxygen
1. HEME REFERENCE RANGE
 iron-chelated porphyrin ring which functions as a AGE GROUP CONVENTIONAL S.I UNITS
prosthetic group (nonamino acid group) Children (8 to 13 12 to 15 g/dl 120 to 150 g/L
o PROTOPORPHYRIN IX – tetrapyrrole ring y.o)
with a ferrous iron inserted into the center Adult (male) 14 to 18 g/dl 140 to 180 g/L
o 1 heme can carry 1 molecule of oxygen Adult (female) 12 to 15 g/dl 120 to 150 g/L
bound to the central ferrous iron
o 1 hemoglobin = 4 molecules of oxygen STRUCTURE
2. GLOBIN 1. Four identical heme groups, each consisting of a
 2 types of chains: protoporphyrin ring and ferrous (Fe 2+) iron
Alphalike Alpha, Zeta 2. Four globin (polypeptide) chains
Nonalpha Epsilon, Beta, Delta, 3. A tetramer of four globin polypeptide chains, with a heme
Gamma molecule attached to each chain
4. The hemoglobin molecule can be described by its primary,
 Responsible for different functional and physical secondary, tertiary, and quaternary protein structures.
properties of hemoglobin. Primary structure of Refers to the amino acid
hemoglobin sequence of the polypeptide
GREEK GREEK NAME # OF AMINO chains
DESIGNATION ACIDS Secondary structure of Refers to chain
α Alpha 141 hemoglobin arrangements in helices and
β Beta 146 nonhelices.
δ Delta 146 Tertiary structure of Refers to the arrangement
hemoglobin of the helices into a pretzel-
γ Gamma 146
like configuration
ε Epsilon 146
Quarternary structure The complete hemoglobin
ζ Zeta 141
molecule structure
5. The quaternary structure of hemoglobin, also called a
tetramer, describes the complete hemoglobin molecule

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Heme Heme consists of a ring of carbon, hydrogen, and

nitrogen atoms called protoporphyrin IX, with a
central atom of divalent ferrous iron (Fe2+)
Hemoglobin = HEME + TWO DIMERS OF GLOBIN
CHAINS
1 RBC = consists of 4 HEME
1 RBC = CAN CARRY 4 MOLECULES OF O2
1 HEME MOLECULE= 4 PYRROLE RINGS
1 Heme molecule = 1 ferrous iron

IRON KINETICS

Iron
 most abundant transition metal in the body
 Most iron in the body is in Hemoglobin and must be in
the ferrous state to be used. Ferrous iron binds to
oxygen for transport to lungs and body tissues. Ferric
iron (Fe 3+) is not able to bind to hemoglobin, but
does bind to transferrin
Hemoglobin function  an essential mineral and is not produced in the body
 to transport oxygen from the lungs to tissues and
transport carbon dioxide from the tissues to the lungs IRON KINETICS
for exhalation - Normal daily diet contains about 15mg iron and only
 an oxygen transporting protein contained within 1-2mg of iron is absorbed in the duodenum and upper
erythrocytes jejunum
o heme portion of hemoglobin - In the duodenum, dietary free iron is reduced to
 gives erythrocytes their ferrous iron and taken up from the intestinal lumen
characteristic red color into the enterocytes by the iron transport protein
Divalent monotransporter 1 (DMT)
- Once absorbed, iron may be stored as ferritin in the
HEMOGLOBIN SYNTHESIS/PRODUCTION
enterocytes or exported into the circulation by another
1. 65 % hemoglobin synthesis occur in immature nRBCs iron transport protein,
2. 35 % hemoglobin synthesis occur in reticuloytes
3. Heme synthesis occurs in the mitochondria of normoblasts FERROPORTIN (Fpn1)
and is dependent on glycine, succinyl coenzyme A,  In the plasma, ferric iron binds to transferrin which is
Aminolevulinic acid synthethase, and vitamin B6 (pyridoxine) delivered into cells by binding to transmembrane
4. Heme leaves the mitochondria to combine with a globin glycoprotein the transferrin receptors (TfR)
chain in the cytoplasm
5. Globin synthesis occurs in the ribosome, and it is controlled Hepcidin
on chromosome 16 for alpha and zeta chains and  a liver-produced peptide hormone
chromosome 11 for all other chains  is the master regulatory hormone of systemic iron
6. Globin chain are released from the polyribosomes and metabolism.
combine with heme molecules released from the mitochondria.  The interaction of hepcidin with the plasma iron
7. Rubricyte = first stage of hemoglobin synthesis transporter, ferroportin, coordinates iron acquisition
8. Reticulocytes = last stage capable of hemoglobin synthesis with iron utilization and storage.
 Hepcidin deficiency
o causes common iron overload syndromes
but overexpression of hepcidin is responsible
for microcytic anemia (anemia of chronic
inflammation).
 It regulates the transport of iron from enterocyte into
the circulation by binding through ferroportin.

Two storage forms of iron.

A. Ferritin = Major storage form, water soluble, considered as
an acute phase reactants.
B. Hemosiderin = Second storage form of iron, water insoluble.

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 Oxygen affinity of hemoglobin is usually expressed as

the PO2 at which 50% of hemoglobin is saturated with
oxygen (P50)
o In humans, P50 is normally about 26 torr
o Higher P50 = lower binding affinity of
hemoglobin
o Lower P50 = higher binding affinity of
hemoglobin
 NORMAL SHAPE = SIGMOID (S-Shaped)
o Indicates that more than one molecule of
oxygen is binding to a molecule of
hemoglobin and that binding facilitates the
binding of additional oxygen (heme-heme
interaction)
 P50 values: defined in terms of
the amount of oxygen needed to
saturate 50% of hemoglobin
 Arterial blood: 95% - 95 mmHg
 Venous blood: 70% - 40 mmHg
 p50: 26.6 mmHg or 27mmHg

Bohr effect Demonstration the relationship between blood

pH and oxygen affinity of Hb A shift in the curve
because of a change in pH (or hydrogen ion
concentration)
Shit to the Oxygen dissociation curve in cases of hypoxic
right conditions such as altitude adaptation (in high
altitude) or anemia
Shift to the Oxygen dissociation curve in the Presence of
left increase myoglobin and fetal hemoglobin

Shift to the left Shift to the right

pH Increase Decrease
PCO2 Decrease Increase
2, 3- DPG Decrease Increase
Temperature Decrease Increase
Hb affinity for O2 Increase Decrease

SHIFT TO INCREASED DECREASED MORE O2

THE RIGHT P50 O2 AFFINITY RELEASED
TO THE
TISSUES
SHIFT TO DECREASED INCREASED LESS O2
THE LEFT P50 O2 AFFINITY RELEASED
TO THE
TISSUES
OXYGEN DISSOCIATION CURVE
 The curve produced when the 2 variables (partial HEMOGLOBIN DERIVATIVES
pressure of oxygen and affinity of hemoglobin for
oxygen) are plotted on a graph (percent saturation of Oxyhemoglobin • Hemoglobin with Ferrous + oxygen
hemoglobin versus the partial pressure of oxygen) • Seen in arterial circulation
o Y-AXIS: percent (RELAXED STATE)
saturation of hemoglobin Deoxyhemoglobin • Hemoglobin with Ferrous but no
o X-AXIS: pO2 Oxygen
• Seen in venous circulation
(TENSED STATE)

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Carboxyhemoglobin • Hemoglobin with ferrous and carbon • It can also combine to Carbon
monoxide CO; hemoglobin has monoxide to form
200x/240x carboxysulfhemoglobin
more affinity for CO than O2 • CANNOT BE REDUCED BACK TO
• Carbon monoxide will bind with Hb HEMOGLOBIN and it remains in the
even if its concentration in the air is cell until break down
extremely • Quantitated by spectrophotometry
low (eg 0.02 – 0.04 %)
• Cannot bind and carry oxygen RBC ABNORMALITIES
• Increasing concentration of HbCO will - high needed to correlate what is the possible
shift the ODC (Oxygen dissociation diagnosis of the patient
curve) to ANISOCYTOSIS
the left, thus adding to anoxia - alteration or change of the size of the red blood cell
• Light sensitive and imparts a typical
brilliant CHERRY RED COLOR to the
blood
• Chief sources of the gas are gasoline
motors, illuminating gas, gas heaters,
defective
stoves, and smoking of tobacco
• Quantitated by differential
spectrophotometry or by gas
chromatography
Methemoglobin/ • Hemoglobin with Fe 3+ (ferric);
Hemiglobin cannot transport oxygen
• Cause chocolate brown discoloration
of blood
• Causes Cyanosis and functional
anemia if present in high enough MEAN CELL VOLUME – indicator to know whether the RBC is
concentration microcytic, macrocytic or normochromic
• Sources: Chemical or drugs such as - reference range is 80-100
chlorate, nitrate, and nitrite - less than 80-100 (microcytic)
• Quantitated by spectrophotometry - beyond 80-100 (macrocytic)
• →An abnormal hemoglobin (Hb M) - between 80-100 (normochromic)
may also be responsible for
methemoglobinemia noted at birth or NORMAL RED BLOOD CELL
first months  Biconcave disc shape
Sulfhemoglobin • Hemoglobin with S; cannot transport  Salmon pink (color from Hb)
Oxygen  7.2 um (7 – 8 um)
• During oxidation of hemoglobin,  Has an increased life span due to enzyme
sulfur (from some source, which may o Hexose monophosphate pathway
vary) is incorporated into  Pathway that helps in the increase
heme rings of hemoglobin, resulting in lifespan of RBC
a green hemochrome  A nucleated
• Blood is mauve-lavender in  Is not rigid (must be deformable/flexible) and has no
sulfhemoglobinemia inclusions (no fragments in the RBC or in the
• Usually reported in following peripheral of RBC).
situations:
• Patients under prolonged treatment
with sulfonamide or aromatic amine
compounds ex.
Phenacitin
• Patients with severe constipation
• In cases of bacteremia caused by
Clostridium perfringens
• In condition known as enterogenous
cyanosis

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ANISOCYTOSIS: VARIATION IN SIZE o As red blood cells mature in erythropoiesis,

 A normal erythrocyte has an average diameter of 7.2 N/C ratio decreases, one the RBC is fully
mm with a usual variation of 6.8 to 7.5 mm. matured, the nucleus removed but the
 The extreme size limits are generally considered to be cytoplasm isn’t yet done on its
6.2 (normal) to 8.2 (normal) mm. This normal size is maturation/complete maturation. Resulting
referred to as normocytic. into having small RBCs/microcytes.
 Erythrocytes may be either larger than normal o How is the maturation being not
(macrocytic) or smaller than normal (microcytic). concurrent/sabay – because of increase in
The size of RBC depends on the protocol of the erythropoiesis (body demands more RBCs
laboratory. that would possibly lead to produce
 Macrocytic erythrocytes exceed the 8.2-mm diameter microcytes)
limit, whereas microcytic erythrocytes are smaller  Size of RBC: 5um or less
than the average 6.2- mm diameter.  MCV < 8o fL
 The general term used in hematology to denote an o Hypochromic microcyte
increased variation in cell size is anisocytosis – o Microspherocyte
(more than 3 RBC in diff size, either big or small).  Causes: Increased erythropoiesis, deprivation of Hb
This term is nonspecific because it does not indicate synthesis, defective globin synthesis
the type of variation that is present. Anisocytosis is o Hb contains iron. If the patient has Iron
prominent in severe anemias. deficiency, there will no Hb produced/formed
that will lead to having microcytic RBCs.
VARIATIONS IN RED BLOOD CELL MORPHOLOGY (not o Defective in globin: 2 types
normal)  Hemoglobinopathies – qualitative
In many disorders or disease states, erythrocytes may disorder of Hb/globin chains. There
demonstrate variations in appearance or morphology as the is a substitution of globin chains
result of pathological conditions. happening forming different Hb (Hb
Note: Some morphological appearance of RBC can be cause SC – not normal Hb, substitutions in
by the process in smear prep. Different morphology of RBC amino acid/globin chains)
shows pathologic - Hb E, S, C
 Thalassemia – quantitative (lack of
The variations from normal can be classified as amount in globin/amino acid chain
1. Variation in size (anisocytosis) that is being formed)
2. Variation in shape (poikilocytosis) - no substitution happens.
3. Alteration in color (hypochromia/hyperchromia)  Conditions: IDA, ANEMIA OF CHRONIC DISEASE,
4. Inclusions in the erythrocyte SIDEROBLASTIC ANEMIA, THALASSEMIA (ATIS)
5. Alterations in the erythrocyte distribution on a peripheral
blood smear MACROCYTES
 Nuclear maturation logs cytoplasmic maturation
Anisocytosis: Variation in size o the cytoplasm is bigger and the nucleus isn’t
 Red blood cell Diameter Width (RDW) – numerical yet done on its full maturation, therefore the
expression that correlates with the degree of larger size of cytoplasm comes out in the
anisocytosis. peripheral.
o Not similar with MCV – (measure what is the o Nucleus is not proportional in the maturation
volume capacity of RBC in the cytoplasm) of the cytoplasm, leading to the production of
o Measures what/how is the degree of the macrocytes/large RBCs
variation in the sizes of the RBC  Size of RBC: >9 um
o Present in CBC (automated method – EDTA  Marker - MCV >100 fL/femtoLiters
tube inserted, machine perform the specific A. THIN MACROCYTES
tests ordered) o MCV is normal & often hypochromic -caused by
 Normal Value: 11.5-14.5% increased membrane cholesterol & lecithin (part
 Normocytic (MCV 80-100 (femtoLiters/fL) of red cell membrane, should not be
 Microcytic (MCV <80 fL) increase/decrease)
 Macrocytic (MCV >100 fL) B. OVAL MACROCYTES
o Deficiency of vitamin B12 as folate
MICROCYTES C. ROUND MACROCYTES
o Seen in hepatic diseases, increased
 Cytoplasmic maturation logs behind nuclear
reticulocytosis, acute alcoholism & aplastic
maturation. (logs=retarded/slow development)
anemia causes decreased erythropoiesis,
o slow development of cytoplasm
impaired DNA synthesis, young rbc7s w/c
undergo early loss of nucleus.

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 Conditions: Pernicious anemia, Folic acid deficiency,

Post splenectomy, after chemotherapy, Obstructive
jaundice

MACROCYTOSIS (HUBBARD)
1+ SLIGHT If there are approximately 25% How is spike seen in the periphery?
macrocytic rbcs present per high- The content of RBCs has sphingomyelin and lecithin, the most
power field abundant in RBCs is lecithin but in acanthocytes, the most
2+–3+ MODERATE If there are 25% to 50% abundant is sphingomyelin so there is an imbalance. The lipid
macrocytic rbcs present per high- content is not balance. The sphingomyelin should go to the
power field plasma but it cannot be absorbing in the small intestine. The
4+ MARKED If there is >50% macrocytic rbcs lipids supposedly in the small intestine going to the plasma but
per high-power field it does not happen. So there is an incapacity to absorb excess
lipids, the sphingomyelin goes to RBCs resulting to thorny or
NORMOCYTIC spurr-like projections of the red cell membrane.
 Seen if there is no problem in the size of the RBC,  Diseases associated:
maybe other causes occur and no relation in o Abetapoproteinemia (there is an in flacs of
morphology of RBC. lipid)
 Size of RBC: 6-9 um o Liver disease (lipids cannot be metabolizing)
 MCV: 80-100 fL o McLeod blood group phenotype (abnormality
 Conditions: in membrane of RBC)
o Acute Hemorrhage – sudden bleeding (no o Post-splenectomy (spleen is removed,
relation with RBC morphology) nothing is responsible to filter RBC)
o Hemolytic Anemias – depends on the type of
hemolysis (external//antibodies/drugs)  SPHEROCYTES
o Aplastic Anemia – anemia causes decreased o RBCs lacks the biconcave shape and
blood cell lineage formation (low levels of becomes more spherical, no central pallor
blood cells, there is a problem in the bone is present with increased hemoglobin
marrow) content. No halo
POIKILOCYTOSIS o are found in hereditary spherocytosis
 Variation in shape (increased amount of spherocytes),
hemolytic anemia (all is associated with
spherocytes) and post transfusion
o lacks of protein - SPECTRIN
o Why having spherocytes? The defects are
in the spectrin. Lacks of spectrin resulting to
a shape of spherical RBC. It can be easily
undergoing lysis. RBC should have spectrin
so it become deformable and cannot easily
lyse.
o When there is no problem in the spleen,
possible problem in the erythropoiesis had
happen.
o Bronze Cell

POIKILOCYTOSIS – VARIATION IN SHAPE

 Poikilocytes are damaged and/or rigid red cells.
 Why poikilocytes present in the blood smear, this  OVALOCYTES
occur in certain diseases, spleen cannot breakdown o Oval shaped RBCs.
all the blood cells. Spleen doesn’t accommodate all o are found in Thalassemia major, hereditary
the blood cells. ovalocytosis, sickle cell anemia.
 ACANTHOCYTES/SPURR CELL/THORN CELL o Elongated, oval cells which tend to become more
o have multiple thorny, spike-like projections elongated as the cell ages.
that are irregularly distributed around the o Elongation may be due to redistribution of
cellular membrane and may vary in size. hemoglobin or cholesterol to ends.
o RBCs with irregularly spaced projections. o Ovalocytes occurs when the cholesterol content
Projections vary in width but usually contain of the RBC is reduced/decreased.
a rounded end. o Decreased cholesterol amount.

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 ELLIPTOCYTES/CIGAR SHAPED RBC

o The RBCs are oval or elliptical in shape, long
axis is twice the short axis.
o are found in hereditary elliptocytosis,
megaloblastic anemia, IDA, thalassemia,
myelofibrosis.  STOMATOCYTES/SLIT/STOMA
o Cigar shaped appearance is due to a o Red cells with a central linear slit or stoma. Seen
problem/defect in the cytoskeleton particularly in as mouth shaped form in peripheral smear.
the membrane protein band 4.1. o Why having slit in the middle? There is an
o Looks like pencil imbalance on the osmotic pressure/osmotic
change. Occurs usually due to imbalances in
sodium and potassium. (cation imbalance)
o are found in excess alcoholism, alcoholic liver
disease, hereditary stomatocytosis.
o Mouth or kiss cells
 TARGET CELLS/CODOCYTES
o Red cells have an area of increased staining
which appears in the area of central pallor.
o are found in obstructive liver disease, severe
IDA, thalassemia, Hemoglobinopathies, post
splenectomy.  KERATOCYTES
o Target cells (codocytes) o These are half-moon shaped cells with two or
o Mexican Hat Cell – the target cells look like a hat more spicules.
o The Hb is concentrated in the middle, shows the o Keratocytosis is seen in G6PD deficiency (prone
dot. This happens in increased surface to volume to oxidation, resulting to keratocytes production),
ratio. (in target cells, excessive amount of pulmonary embolism, disseminated intravascular
cholesterol in the membrane or an imbalance coagulation.
between the Hb concentration.  Oxidation products (keratocytes
o Can also be found as an artifact – pressure in and Heinz bodies)
smear forms target cells.

 TEARDROP CELLS/CRYOCYTES  ECHINOCYTES/BURR CELLS/SEA

o RBCs having the shape like teardrop or pear. URCHIN/CRENATED RBC
These are usually microcytic and often o Not similar with acanthocytes. DIFFERENCE:
hypochromic. evenly/uniformed distributed projections outside
o are found in myelofibrosis (infiltration of bone Acanthocytes – not even/uniformed projections
marrow – most common), megaloblastic outside
anemia, IDA, thalassemia. o Red cells with uniformly spaced pointed
o Dacryocytes, racquet forms projections on their surface.
o are found in hemolytic anemia, uremia,
megaloblastic anemia.
o Osmotic imbalance causing projections in the
side of the red cell membrane.

 SCHISTOCYTES
o These are helmet or triangular shaped,
fragmented or greatly distorted RBCs smaller
than normal size.
o are seen in Thalassemia, microangiopathic  SICKLE CELLS
hemolytic anemia, mechanical hemolytic anemia, o These are sickle-shaped Red Blood Cells.
uremia, artificial heart valves. o are seen in Hb-S disease/ sickle cell anemia.
o Fragmentation of red blood cells. o Hb SS, Hb SC (washington monument), Hb SD,
o Schistocyte/Schizocyte/Keratocyte/Helmet/Bite Hb S-beta Thalassemia
Cell

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Stomacyte RBC with slit- Hereditary helliptocytosis

KERATOCYTE like are of Rh deficiency syndrome
 horn-like projections, are erythrocytes that are central pallor Acquired stomatocytosis (liver
partially deformed but not cut. disease, alcoholism)
 The spicules, resembling two horns, result from a Artifact
ruptured vacuole. Sickle cell Thin, dense, Sickle cell anemia
 Usually the cell appears like a half-moon or spindle. elongated Sickle cell-B-thalassemia
 These cells are seen in conditions such as RBC pointed
disseminated (diffuse) intravascular coagulation at each end;
(DIC). may be
BLISTER CELL curved
 eccentric nucleus, are erythrocytes containing one or Hb C crystal Hexagonal Hb C disease
more vacuoles that resemble a blister on the skin. crystal of
 This cell has a significantly thinned area at the dense
periphery or outer border of the cell membrane. The hemoglobin
vacuoles may rupture. formed within
the RBC
KRIZOCYTE OR PINCH CELL membrane
 triangular with 2 pallor edges resemble a pinched Hb SC crystal Fingerlike or Hb SC disease
bottle. quartz-like
 This abnormality is associated with hemolytic crystal of
anemias, including hereditary spherocytosis dense
hemoglobin
TABLE 19-2 Description of Red Blood Cell (RBC) protruding
Abnormalities and Commonly Associated Disease States from the RBC
RBC Cell Commonly Associated membrane
Abnormality Description Disease States Target cell RBC with Liver disease
Anisocytosis Abnormal Hemolytic, megaloblastic, iron (codocyte) hemoglobin Hemoglobinopathies
variation in deficiency anemia concentrated Thalassemia
RBC volume in the center
or diameter and around
Macrocyte Large RBC Megaloblastic anemia the periphery
(>8 um in Myelodysplastic syndrome resembling a
diameter), Chronic liver disease target
MCV >1 100 Bone marrow failure Schistocyte Fragmented Microangiopathic hemolytic
fL Reticulocytosis (schizocyte) RBC due to anemia (along with
rupture in the microspherocytes)
Oval Large oval Megaloblastic anemia periphery Macroangiopathic hemolytic
macrocyte RBC circulation anemia
Microcyte Small RBC Iron deficiency anemia Extensive burns (along with
9>6 um in Anemia of chronic microspherocytes)
diameter), inflammation Helmet cell RBC fragment Same as schistocyte
MCV <80 fL Sideroblastic anemia (keratocyte) in shape of a
Thalassemia/Hb E disease helmet
and trait Folded cell RBC with Hb C disease
Poikilocytosis Abnormal Severe anemia; certain membrane Hb SC disease
variation in shapes helpful diagnostically folded over
RBC shape Acanthocyte Small, dense Severe liver disease (spur
Spherocyte Small, round, Hereditary spherocytosis (spur cell) RBC with few cell anemia)
dense RBC Immune hemolytic anemia irregularly Neuroacanthocyrtosis
with no Extensive burns (along with spaced (abetalipoproteinemia,
central pallor schistocytes) projections of McLeod syndrome)
Elliptocyte, Elliptical Hereditary helliptocytosis or varying length
ovalcyte (cigar- ovalocytosis Burr cell RBC with Uremia
shaped), oval Iron deficiency anemia (echinocyte) blunt or Pyruvate kinase deficiency
(egg-shaped), Thalassemia major pointed, short
RBC Myelophthisic anemias projections
that are
usually evenly

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spaced over
the surface of HYPOCHROMIA
cell; present  indicates the amount of
in all fields of hemoglobin in the erythrocyte
blood film but is decreased and the central
in variable pallor area is greater than
numbers per 1/3 the red cell diameter; the
field. MCHC may be <32.0%.
Teardrop cell RBC with a Primary myelofibrosis  The Hb and the central area of pallor is inversely
(dacryocyte) single pointed Myelophthisic anemia proportional – high amt of Hb content, the central
extension Thalassemia pallor is smaller. Low amt of Hb, central pallor is
resembling a Megaloblastic anemia bigger.
teardrop or  Decrease Hb content, increase area of pallor.
pear. HYPOCHROMIA GRADING
1+ Area of central pallor is one half of cell diameter
2+ Area of pallor is two-thirds of cell diameter
3+ Area of pallor is three-quarters of cell diameter
4+ Thin rim of hemoglobin

HYPERCHROMIC / HYPERCHROMIA / HYPERCHROMASIA

 These are seen in RBC with
seemingly increased Hb content
since the cell is thicker than
normal but the hemoglobin
content is actually within the
normal limit.
ANISOCHROMIA  Hb is in high amount but in normal limit, there is an
 Alteration in color (chromia-RBC vary in color
imbalance between the Hb content. The surface area
depending on the amt/conc. of Hb)
volumetric ratio is decreased causing spherocytes
 variation in hemoglobin content (can be seen RBC because of the damage in the spectrin
with pale color and RBC too dark color)
 2 most common (hypochromic and normochromic) POLYCHROMASIA/POLYCHROMATOPHILIA
1. Hypochromic cell - associated with microcytic  Residual RNA component of the RBCs. RNA is
2. Hyperchromic cell – usually not seen in some diseases, removed in the mature RBC. When there is still RNA
associated with spherocytes, cold agglutinins, error in testing. in the RBC, it means that the RBC in the peripheral
3. Normochromic cell – usually associated with macrocytic and blood smear is from the early precursors like
normocytic polychromatophilic normoblast.
o Normochromic (MCHC=31-36%)  Large red cells (8-10u) which display diffuse
o Hyperchromic (MCHC= >36%) basophilia (blue tinge) due to residual RNA when
o Greater than 36 is not associated with disease but stained with Wright's stain. The amount of
associated with cold agglutinins, samples that are polychromasia correlates with the number of
refrigerated and not taken back at room temp, circulating reticulocytes - (has residual RNA)
spherocytes.

ANISOCHROMIA – VARIATION IN HEMOGLOBIN

CONTENT
NORMOCHROMIC
 A normal erythrocyte has a moderately pinkish-red
appearance with a lighter-colored center (central area
of pallor) when stained with a conventional blood
stain. POLYCHROMASIA GRADING
 A normal erythrocyte contains an amount of GRADING POLYCHROMATOPHILIC
hemoglobin so only 1/3 (cental pallor) of its diameter RBCs
is empty in appearance (normochromia) SLIGHT 1%
1+ 3%
2+ 5%
3+ 10%
4+ >11%

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INCLUSION BODIES

INCLUSIONS SEEN IN THE ERYTHROCYTE

 NORMAL RBC – NO INCLUSIONS
 The presence of inclusions decreases red cell
deformability and shortens red cell survival.
 Normally, RBCs that have inclusions passes through Basophilic stippling
the spleen but it will later be phagocytized by the  These are considerable amount of small basophilic
macrophages. inclusions in red cells which represent precipitated
RNA.
HOWELL-JOLLY BODIES  Distributed in the entire red blood cell
 Small round cytoplasmic red cell inclusion with same membrane/structure
staining characteristics as nuclei. These are  Usually seen in lead poisoning
fragments of DNA.  Seen in: thalassemia, MBA, hemolytic anemia, liver
 Usually seen in Post splenectomy (removed spleen – damage, heavy metal poisoning, Lead Poisoning
nothing to filter inclusion), megaloblastic anemia
(hypersegmented, triple H), hemolytic anemia,
hyposplenism, aplenism, severe hemolytic anemia.

Cabot ring
 Reddish, purple, thread-like rings in RBCs of severe
anemias; FIGURE OF EIGHT IN RBC using Wright
Stain.
Heinz bodies  These are the remnants of mitotic spindle,
 These represent denatured hemoglobin microtubules.
(methemoglobin) within a cell. With a supravital stain  Seen in patient with Megaloblastic anemia,
like crystal violet (seen in supravital stain) myelodysplastic syndrome and lead poisoning.
 From the precipitation of oxidized methemoglobin
 appear as round blue precipitates or may be a
product of abnormal globin synthesis and composed
of unused, precipitated globin chains.
 Seen in G6PD deficiency, splenectomy (removed
spleen so it cannot remove inclusions), oxidant drugs
(produced oxidation that would lead to precipitation of
heinz bodies), favism and unstable hemoglobin (Hb
should be stable, because once the Hb is unstable, it
will be prone to oxidation then it will result to
precipitation of heinz bodies)

Pappenheimer bodies
 These represent iron deposits which as dense blue,
irregular granules in wright stain.
 are found in RBCs with hemolytic anemia
(hemolyzed RBC, iron in the heme deposits in the
Feulgen – presence of DNA (seen in Howell jolly body only)
RBC), splenectomy (np removal of inclusion),
Supravital – RNA/DNA (BS, HJb, Ret, Pb, Hb)
sideroblastic anemia (iron can be deposited as
Heinz bodies – usually negative in wright’s stain.
sidero granules or sideroblasts), post-splenectomy,
Reticulocytes used supravital stain or methylene blue.
thalassemia (quantitative defect of Hb synthesis, lack
of globin and excess in heme resulting to iron
Malarial inclusions
deposits in the RBC.
 The inclusions may be ringed, segmented or massive
 Distributed only in the periphery
forms depending on the maturity of the parasite.

AGTARAP, RICA 1
 LECTURE TITLE

 Associated with infestation of: Howell Dark blue- Dark blue- DNA Hyposplenism
o Plasmodium vivax (Schuffner's granules) -jolly purple purple (nuclear Postsplenecto
o Plasmodium falciparum (Mauer's dots) body dense, dense, fragmen my
o Plasmodium malariae (zeimanns) round round t) Megaloblastic
o Plasmodium ovale (james dots) granule; granule; anemia
usually usually one Hemolytic
one per per cell; anemia
OTHER PROTOZOAN INCLUSIONS cell; occasionally Thalassemia
1. Babesia microti – ringlike structures resembling occasional multiple Myelodysplasti
malarial parasite; Maltese cross ly multiple c syndrome
- confuse with the ring of P. falciparum Heinz Round, Not visible Denatur Glucose-6-
2. Bartonella – comma-shaped organisms near body dark blue- ed phosphate
Membrane. Categorized as BACTERIA purple hemogl dehydrogenas
granule obin e deficiency
Hemoglobin C Crystals Hemoglobin SC Crystals attached Unstable
Characteristics: Condensed Characteristics: Bizarre to inner hemoglobins
rod or bar-shaped crystals shaped crystal; denser than RBC Oxidant
with blunt ends found in sickle cells with fingerlike membrane drugs/chemical
RBC’s. Tetragonal, projections. “Washington
rectangular rod shaped (bar Monument” shape Pappenhei Irregular Irregular Iron Sideroblastic
of gold, clam shell) Associated with: Inherited mer bodies clusters clusters anemia
Associated with: Inherited presence of Hgb S and Hgb of small, of small, Hemoglobinop
presence of hemoglobin C C (Hgb SC). light to light to athies
dark blue dark blue Thalassemias
CRYSTALS granules, granules, Megaloblastic
HEMOGLOBIN C HEMOGLOBIN SC often often anemia
CRYSTALS CRYSTALS near near Myelodysplasti
periphery periphery c syndrome
of cell of cell Hyposplenism
Post-
splenectomy
Cabot ring Rings of Blue Remnan Megaloblastic
figure- rings or t of anemia
eights figure- mitotic Myelodysplasti
Inclusi Appearan Appearance Inclusio Associated eights spindle c syndrome
on ce in in Wright n Diseases/Con Hb H Fine, Not Precipit Hb H disease
Supravita Stain Compo ditions evenly visible ate of B-
l Stain sed of disperse globin
Diffuse Dark blue Bluish tinge RNA Hemolytic d, dark chains
basop granules throughout anemia blue of
hilia and cytoplasm; After treatment granules; hemogl
filaments also called for iron, imparts obin
in polychromas vitamin B12, or “golf ball”
cytoplasm ia (seen in folate appearan
(seen in polychromati deficiency ce to
reticulocyt c RBCs
es) erythrocytes)
Basop Dark blue- Dark blue- Precipit Lead poisoning RED CELL DISTRIBTUTION
hilic purple, purple, fine ated Thalassemia
stipplin fine or or coarse RNA Hemoglobinop NORMAL
g coarse punctate athies  even distribution of RBCs in the thin portion adjacent
punctate granules Thalassemia to the feather end of the film
granules distributed Hemoglobinop  Red cells should be slightly separated from one
distributed throughout athies another or barely touching without overlapping
throughout cytoplasm Megaloblastic  Thin area should represent at least one-third of the
cytoplasm anemia entire film.
Myelodysplasti
c syndrome

AGTARAP, RICA 1
 LECTURE TITLE

ROULEAUX FORMATION  It is important that we prepare and make a very good

 Stacks of RBCs resembling a stack of coins. smear to avoid confusion.
 Does not form in the presence of antibodies
 Due to increased plasma globulins BLOOD SMEAR
 Results to increased ESR
 These are found in hyperglobulinemia,
hyperfibrinogenemia.

AUTOAGGLUTINATION
 RBCs aggregate into random clusters or masses
when exposed to red cell antibodies
 May cause anticoagulated blood to appear somewhat
“grainy” or granular as tubes is being rotated at room
temperature.
 Seen in Cold Agglutinin Disease and Autoimmune
hemolytic disease

REPORTING OF ABNORMALITIES

GRADING CHARTS IN HEMATOLOGY

VARIATIONS IN RED BLOOD CELL MORPHOLOGY GRADED AS

PATTERN/DISTRIBUTION Polychromatophilia 1+ = 1 to 5/field
Helmet cells 2+ = 6 to 10/ field
Red cell agglutinate Teardrop RBC 3+ = >10/field
 These are irregular clumps of RBCs. Acanthocytes
 These are found in cold agglutinations, warm Schistocytes
autoimmune hemolysis. Spherocytes
 Clumping of RBC's with no pattern. May see visible Poikilocytes 1+ = 3 to 10/ field
agglutination when the blood smear is prepared Ovalocytes 2+ = 11 to 20 / field
(grainy appearance). Elliptocyte 3+ = >20 / field
Burr cells
Bizarre- shaped RBC
Target cells
Stomatocytes
Rouleaux 1+ = aggregates of 3 to 4 RBC
2+ = aggregates of 5 to 10 RBC
REMEMBER:
3+ = numerous aggregates with
 A well-made, properly stained smear is essential to only few free RBC
prevent erroneous morphologic evaluation of the Sickle cells Grade as positive only
RBCs. Basophilic stippling
 Often times, types of specific poikilocytosis may form Pappenheimer bodies
as artifact......this may be due to an improperly made Howell- Jolly bodies
smear or improper/excess anticoagulant may alter the
cells. Precipitated stain or water artifact can mimic
RBC inclusions.
 If the abnormality is only present on the edge of the
slide or not found throughout the smear, it is probably
artifact.

AGTARAP, RICA 1
 LECTURE TITLE

SAMPLE CRITERIA FOR ERYTHROCYTIC MORPHOLOGY

EVALUATION

Morphologic WNL 1+ 2+ 3+ 4+
Characteristics
Macrocytes 0-5 5-10 10-20 20-50 >50
Microcytes 0-5 5-10 10-20 20-50 >50
Microcytes 0-2 3-10 10-50 50-75 >75
Poikilocytosis 0-2 3-10 10-20 20-50 >50
Burr cells 0-2 3-10 10-20 20-50 >50
Acanthocytes <1 2-5 5-10 10-20 >20
Schistocytes <1 2-5 5-10 10-20 >20
Dacryocytes 0-2 2-5 5-10 10-20 >20
Codocytes 0-2 2-10 10-20 20-50 >50
Spherocytes 0-2 2-10 10-20 20-50 >50
Ovalocytes 0-2 2-10 10-20 20-50 >50
Ovalocytes 0-2 2-10 10-20 20-50 >50
Sickle cells Abse Report as +1 to indicate
nt presence, do not quantitate
Polychromaatophilia <1 2-5 5-10 10-20 >20
Adult 1-6 7-15 15-20 20-50 >50
Newborn
Basophilic stippling 0-1 1-5 5-10 10-20 >20
Howell- Jolly abs 1-2 3-5 5-10 >10
bodies
Pappenheimer abs 1-2 3-5 5-10 >10
bodies

GRADING SCALE FOR RED CELL MORPHOLOGY

ANISOCYTOSIS / POIKILOCYTOSIS

NORMAL 5%
SLIGHT 5-10%
1+ 10-25%
2+ 25-50%
3+ 50-75%
4+ >75%

GOODLUCK AND GODBLESS!

AGTARAP, RICA 1
 2
SY 2021 -
CLINICAL
m HEMATOLOGY 1
1st
2022
SEMESTER
BS-MLS
MR. NICKSON CAMMAYO, RMT

TOPIC TITLE (EXAMPLE: ENZYMES)

OUTLINE
I ANEMIA
A. RBC ABNORMALITIES
B. IRON KINETICS
i.
ii.
a.
b.
c.

LMS MODULE
ANEMIA o “Absolute” – true change in RCM
 reduced capacity to carry oxygen to the body tissues.
The red blood cells (erythrocytes) have the important c. Chemical and Physical Response:
function of carrying oxygen to the tissues.  Anemia results to hypoxia
 can be caused by several factors. The red blood cells o shifting the oxyhemoglobin dissociation curve
can be reduced due to a series bleeding episode. TO THE RIGHT
 can occur because the red cells are not being o selective redistribution of blood
produced in normal quantities. o Cardiac output is increased
 can also occur if the amount of hemoglobin produced
is insufficient to fill the red cells. d. Hematologic response:
 result when the destruction of red cells exceeds  erythropoietic marrow stimulation “Shift reticulocytes” =
production. RPI
 A healthy body can lose 20% of its blood volume  Erythropoietin from the kidney.
without loss of body function (blood donations cause
no problems for the healthy). If the blood loss is 30% BASIC CLASSIFICATION OF ANEMIAS:
to 40% of the total blood volume, then shock and
circulatory collapse can be seen. If the loss reaches 1. Hypoproliferative
50%, death can be imminent.  Decreased RBC precursors
 caused by loss of blood is first seen as “being tired.”  Hypocellular BM
o As the loss increases, muscle fatigue or  INADEQUATE ERYTHROPOIESIS
Weakness develops, possibly accompanied 2. Maturation disorders
by a persistent headache. Fainting may also  Normal RBC precursor
be common. With continued loss, the patient  Decreased raw materials
goes into coma because the brain is not  DEFECTIVE ERYTHROPOIESIS
receiving enough oxygen. 3. Hemolytic disorders
4. Blood loss
RBC ABNORMALITIES
a. Anemia CAUSES OF ANEMIA:
 decreased red cells;
 best defined in reference to a decreased Hb level,  Acute blood loss
 the physiologic consequences and symptoms are the  Accelerated RBC destruction (Immune or non-immune)
direct result of the decreased oxygen carrying capacity  Nutritional deficiency (iron, folate or B12)
of the blood  Bone marrow replacement (e.g. cancer)
 Infection
b. Erythrocytosis and Polycythemia  Toxicity
 denote too many red cells; best defined in relation to  Hematopoietic stem cell arrest or damage
hematocrit levels above the reference range.  Hereditary or acquired defect
 Primary consequences are hypervolimia and
hyperviscosity. Diagnosis of Anemia:
1. Clinical history
o “Relative” – change in RCM secondary to a 2. Physical signs such as pallor, fatigue, weakness and
change in the plasma volume shortness of breath
 Relative anemia - plasma volume 3. LABORATORY TESTS
increases, diluting the RCM
- pregnancy
- hyperproteinemia
 Relative erythrocytosis - plasma
volume decreases
- dehydration (e.g., burns)

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 LECTURE TITLE

MORPHOLOGIC CLASSIFICATION OF ANEMIA o It is absorbed in jejunum and released

from enterocytes as N5-methyl folate.
1. MACROCYTIC ANEMIA After transport in blood stream, it is
converted to THF by the B12-dependent
A. Megaloblastic Anemia methyltransferase.
 (RBC's are abnormal in the bone marrow and the o The active form of folate,
circulating peripheral blood.) tetrahydrofolate(THF), acts as a cofactor
 They are group of disorders characterized by in methyl transfer reactions (conversion of
defective nuclear maturation caused by impaired dUMP to dTMP for use in DNA Synthesis)
DNA synthesis. o Most common cause is a dietary
 RBC’s are fragile, lifespan is shortened and may deficiency
die in the bone marrow which causes increase o Inadequate dietary intake
LDH. o Malabsorption
1. Poverty
 Granulocytes and thrombocytes are affected as
2. Old age
well.
3. Alcoholism
 Nuclear replication is slowed down resulting in
4. Chronic disease
maturation delays:
1. Macrocytic-normochromic red cells (many
o Increased requirement
macro-ovalocytes)
1. Pregnancy. There is increased
2. Granulocytes are hypersegmented
demand during pregnancy and should be
(macropolycytes)
supplemented prior to and during
3. Megakaryocytes are abnormal resulting in
pregnancy. Deficiency during pregnancy
thrombocytopenia
can cause neural tube defects in utero.
 Causes: 2. Infancy
a. Vit B12 (cyanocobalamin) Deficiency –
Pernicious anemia o Drugs
o Vitamin B12 is found in red meat, fish, 1. Methotrexate (chemotherapy drug that
poultry, eggs, dairy products is folate antagonist)
o It is a cofactor for methyltransferase 2. Alcohol, oral contraceptives, etc.
enzymes necessary for the conversion of
circulating form of folate to active form
Chemistries Folate Increased:
(tetrahydrofolate)
deficiency LDH
o It is also a cofactor for methylmalonyl
Indirect bilirubin
CoA mutase which converts
Urinary FIGLU (foriminoglutamic
methylmalonyl CoA to succinyl CoA.
Acid)
o B12 is ingested in animal products
(mostly) Decreased: Serum and RBC
o Intrinsic factor (IF)-bound B12 is
folate
absorbed in the ileum bound to
B12 deficiency Increased:
transcobalamin I & II (90%) in
LDH
enterocytes and exported to the blood
Indirect bilirubin
stream.
Urinary methylmalomic acid
o Inadequate dietary intake
o Malabsorption
Decreased:
1. Pernicious anemia or Addisonian’s
Serum B12
disease
RBC folate (2/3 of cases
 Caused by gastric parietal cell
Schilling’s test)
atrophy which causes decreased
secretion of Intrinsic factor (IF) which
c. Inherited disorders of DNA synthesis
is necessary for B12 absorption.
d. Drug induced disorders of DNA synthesis
 Onset is usually after age 50.
 Neurologic problems
LABORATORY EVALUATION OF MEGALOBLASTIC
2. Gastrectomy
ANEMIA
3. Blind loop syndrome (bacteria use
up the B12)  Blood Indices:
4. Fish tape worm (D. latum) – o MCV: Increased
competes for B12 o MCHC: Usually normal
o Drugs (alcohol, Nitrous oxide, anti-  WBC and platelet counts: Decreased
tubercular drug)  Bone Marrow Biopsy and Aspiration:
o Megaloblasts (large nRBCs with a webby
b. Folic acid (folate) deficiency – nutritional chromatin pattern) are present. The cells
megaloblastic show asychrony in that the cytoplasmmatures
faster than the nucleus.
o Folic Acid is found in green leafy o Erythroid hyperplasia with left shift
vegetables, fruits, dairy products, cereals, Treatment of Megaloblastic Anemia:
liver and kidney.  B12 deficiency
1. Vitamin therapy

2
 LECTURE TITLE

2. Intramuscular or subcutaneous injections for o Hereditary condition caused by increased iron

pernicious anemia to bypass absorption through the gut absorption which deposits in vital organs such
as liver, spleen and pancreas which then
2. HYPOCHROMIC MICROCYTIC ANEMIA become fibrotic.
a. Iron deficiency anemia o People with this take on a bronze color.
b. Thalassemia o Therapy consists of iron removal by
c. Sideroblastic anemia therapeutic phlebotomy or chelation
d. Anemia of chronic disorders  Sideroblastic anemia
 Lead poisoning
Note: We can also include Lead poisoning, knowing that lead o Lead interferes with iron storage in the
inhibits heme synthesis anything that will hinder cytoplasmic mitochondria
maturity (hinder hemoglobin synthesis) will cause microcytic o It damages the activity of enzymes used for
hypochromic anemias heme synthesis (basophilic stippling)

Heme Synthesis:

 Porphyria - excessive production of porphyrins in the

BM (or liver), decreased or defective production of Hb
(Heme portion)
o 1. Rare disease caused by accumulation of
porphyrins in the developing RBC’s
o 2. Characterized by dermal photosensitivity
and rash caused by the sun. The original
werewolf was probably a person with
erythropoietic porphyria.

b. Thalassemia
 hereditary disorder
 abnormal gene causing a decreased rate of synthesis
of certain polypeptide chains
 hemolytic anemia

Take note! All other hemolytic anemia are normocytic,

Iron Metabolism: normochromic
 Primary function of iron is oxygen transport
 Iron absorption and storage is influenced by: Laboratory Evaluation:
1. The amount and type of available iron in the diet  Peripheral Blood Smear:
2. Incomplete absorption due to GI tract problems o Micro hypo with marked anisocytosis and
3. Current iron stores poikilocytosis (i.e. target cells, teardrop cells,
4. Increased demand (Pregnancy, growth years) schistocytes, red cell inclusions), moderate
5. Excessive loss due to acute or chronic hemorrahage polychromasia. NRBCs usually present.
(menstrual period for women, GI bleeding for men)  WBC Count: often slightly elevated
 Platelets: often increased
a. Iron Deficiency Anemia  Bone Marrow: erythroid hyperplasia
 most commonly occurring microcytic, hypochromic  Osmotic Fragility: decreased
anemia (90%)  Heinz Bodies: seen with correct stain - HbF and HbA2:
 chronic blood loss: major cause of iron deficiency increased
anemia
 IDA occurs when iron stores in the body are c. Sideroblastic Anemia
inadequate to preserve hemeostasis  characterized by iron overload or increase in total body
iron
Anemia due to iron depletion may be caused by:  does not always occur as micro/hypo
1. Increased physiologic demand for iron during rapid  Dimorphic blood picture (one cell population is
growth, pregnancy and increased hematopoiesis micro/hypo, the other is normocytic, normochromic)
2. Increased loss of iron due to blood loss, hemolytic
anemia or cancer Laboratory Evaluation:
3. Malabsorption of iron due to gastrointestinal disorders
 Peripheral blood smear
4. Dietary deficiency. It is possible but not common in
o usually normaldimorphic picture of microcytic,
develop countries since the body efficiently recycles iron.
hypochromic RBCs and normocytic,
normohromic RBCs Siderocytes maybe seen.
Iron excess and Sideroblastic States:
Pappenheimer bodies may be seen.
 Hemosiderosis
 Bone Marrow
o accumulation of excess iron in tissue
o Erythroid hyperplasia, "Ringed" sideroblasts
macrophages which could be due to organ
(nucleated RBCs in which iron is deposited in
bleeding
a ring around the nucleus).
 Hemochromatosis
 Serum Iron: increased
 Storage Iron: increased

3
 LECTURE TITLE

d. Anemia of chronic disorders 3. NORMOCHROMIC NORMOCYTIC ANEMIA

 starts normocytic normochromic and gradually  Seen during:
becomes microcytic, hyporchromic. a. Recent blood loss
 Associated with: Chronic urinary tract infections, b. Overexpansion of plasma volume
rheumatoid arthritis, chronic leukemia. c. Hemolytic diseases
d. Hypoplastic bone marrow – e.g. aplastic anemia
Iron Def. Sideroblasti Anemia of
Anemia c Anemia Chronic
e. Infiltrated bone marrow – e.g. leukemia
Disorder f. Endocrine abnormality – e.g. hypothyroidism
g. Chronic disorders
Ferritin decreased Increased Normal to h. Renal diseases
Increased i. Liver diseases

Serum Iron Severely Increased Decreased A. Hemolytic Anemia

Decreased  results from an increase rate of RBC destruction and
Normal to Normal to Decreased the inability of the BM to respond to the stimulus of
Total Iron Increased Increased anemia which may be due to:
Binding a. intrinsic disorder: defect in the red cell itself
Capacity(TIB b. extrinsic disorder: a hemolytic factor
C) outside the red cell is acting on it.
Increased Increased Increased  hemolysis results when there is increased RBC
Free destruction and shortened cell survival.
Erythrocyte  If the hemolysis is extravascular, no hemoglobinemia,
protophorphi hemoglobinuria and hemosidenuria is present.
rin Hemolysis is detected by measuring an increase in one
Percent iron Decreased Severely Decreased of the products of heme catabolism.
saturation Increased
General laboratory findings following:
a. Intravascular hemolysis (catabolism)
Mar Ser Marro Seru Total Trans Free 1. RBC survival studies: Decreased
row um w m Iron ferrin Erythroc 2. Haptoglobin: Immediate decrease
Iron Fer Sidero Iron Bindi Satur yte 3. Methalbumin: Increased for several days
ritin blasts ng ation
Protopo 4. Urine hemosiderin: Increased for several days
Cap
acity rphyrin 5. Plasma hemoglobin: Immediate increase
(TIB (FEP) 6. Urine hemoglobin: Positive
C) 7. LDH & Bilirubin: Increased
Referenc Nor 30- 25%- 50- 250- 20%- <50 8. Presence of schistocytes
e Range mal 250 30% 150 450 55% ug/dL
ug/ ug/d ug/d RBC b. Extravascular hemolysis(catabolism)
L L L 1. RBC survival studies: Decreased
(var (vari 2. Serum unconjugated bilirubin: Increased
ies es
w/ w/
3. Urine & fecal urobilinogen: Increased
age age, 4. Normal to decreased haptoglobin
, sex) 5. Increased LDH
sex 6. Presence of microspherocytes
)
IRON Abs Dec 0 Dec. Inc. Dec. Inc. Category of Hemolytic Anemia:
DEFICIE ent . A. Intracorpuscular – intrinsic defect of the red cell
NCY 1. Hereditary
ANEMIA a. Membrane defects
ANEMIA N/In Inc. 0-few Dec. Dec. N/Dec Inc. b. Metabolic defects
OF c. . c. Hemoglobinopathies
CHRONI
d. Thalassemia syndromes
C
DISORDE 2. Acquired
RS a. Paroxysmal Nocturnal Hemoglobinuria (PNH)
SIDEROB Inc. Inc. Inc. Inc. Dec. Inc. Mixed
LASTIC (ringed /N B. Extracorpuscular – extrinsic defect of the red cell
ANEMIA ) 1. Immune hemolytic anemia
LEAD N N Inc. N to N Inc. Markedl 2. Infections
POISONI (may Inc. y Inc. 3. Chemical agents and toxins
NG be (adu 4. Physical agents
ringed lts) 5. Microangiopathic and macroangiopathic hemolytic
) N to anemia
dec.
6. Splenic sequestration (hypersplenism)
(child
ren) 7. General systemic disorders (w/o dominant hemolysis)
THALASS Inc./ Inc. N (may Inc./ N Inc./N N
EMIAS N /N be N
ringed)

4
 LECTURE TITLE

1. Intrinsic Disorders o NB: Normal RBC can withstand up to 49C

A. Defects of the RBC Membrane (Cytoskeletal D/O) heat.
a. Hereditary spherocytosis
b. Hereditary elliptocytosis (ovalocytosis) 2. Spherocytic type
c. Hereditary acanthocytosis o Results from double heterozygosity for HS and HE
d. Hereditary stomatocytosis
B. Metabolic defects 3. Stomatocytic type
a. G6PD Deficiency o A.K.A. Southeast Asian Ovalocytosis (SAO)
b. Pyruvate Kinase Deficiency o Common in Malaysia
C. Hemoglobin Defects o Due to a band 3 CHON defect and confers against
a. Sickle Cell Disease (sickle cells) P. vivax malaria.
b. Hemoglobin C Disease (Hgb C crystals)
c. Hemoglobin SC Disease (Hgb SC crystals) B. Additional Laboratory evaluation:
1. Hgb and Hct: Normal to severly decreased
2. Extrinsic Disorders 2. PBS: Elliptocytosis
a. chemical agents (lead) 3. Retic count: Slightly elevated
b. vegetable and animal poisons (fava beans) 4. Osmotic fragility: Variable
c. infectious agents (malarial organisms) 5. Autohemolysis test: Variable
d. physical agents (burn patients)
e. immunological agents (ABO-Rh incompatibility)
3. Hereditary Acanthocytosis
CONGENITAL DEFECTS OF THE RBC MEMBRANE A. Cause: decreased plasma and membrane lipids
CAUSING H.A. resulting in irregularly formed RBCs.
1. Hereditary Spherocytosis B. Additional laboratory evaluation
A. Also known as: hereditary hemolytic jaundice 1. Triglycerides and cholesterol: Decreased
B. Cause: 2. PBS: Marked acanthocytosis
1. autosomal dominant disorder 3. Retic count: Normal to increased
2. caused by a defect in cytoskeletal CHONs:
band 3, spectrin , ankyrin 4. Hereditary Stomatocytosis
3. This abnormally permeable cell membrane A. Cause:
results from a reduction of phospholipids and o The exact membrane defect is unknown, but
cholesterol that allows the passive influx of sodium studies suggest that there is an absence of a
ions into the cell resulting to interference with RBC membrane CHON located in the band 7
bi-concavity and deformity region called stomatin.
B. Associated with abnormal Na/K permeability
C. Features: chronic hemolysis, jaundice and C. It is also a feature of Rh null red cell phenotype.
splenomegaly
Main types:
D. Additional laboratory evaluation: 1. Hydrocytosis (overhydrated)
a. Hgb and Hct: Normal to moderately decreased o more severe
b. MCHC: Usually increased (the only condition in o Red cells take on extra water
which MCHC can be truly increased) o Additional Laboratory Evaluation:
c. PBS: Spherocytes and anisocytosis 1. MCV: Increased
d. Osmotic fragility: Increased 2. MCHC: Decreased
e. Autohemolysis test: Increased 3. PBS: Stomatocytes (elongated,
f. LDH and Bilirubin: elevated (extravascular mouth-like central pallor)
hemolysis) 4. Osmotic fragility: Increased
2. Xerocytosis
E. Treatment: Splenectomy o less severe
- NB: Mild anemia o allows water to exit the RBC
o Additional Laboratory Evaluation:
2. Hereditary Elliptocytosis/ Ovalocytosis 1. MCHC: Increased
A. Cause: 2. PBS: Normocytic red cells with
a. autosomal dominant disorder Codocytes (Target cells), mild
b. due to defective formation of spectrin tetramers. stomatocytosis and spiculated
 Associated with severe hemolytic anemia in infants. dessicocytes.
 Of all patients with HE, 90% are non-anemic. 3. Osmotic fragility: Decreased

Three types: 5. Hereditary Pyropoikilocytosis

1. Common type A. Cause: Defective spectrin results in variable RBC
o Found in African- American (heterozygotes shapes and size
have mild or no hemolysis while the B. Additional Laboratory Evaluation
homozygotes have moderate to severe 1. MCV: Decreased
anemia) 2. Osmotic fragility test: Increased
o Hereditary pyropoikilocytosis (HPP) is a 3. PBS: RBC budding, fragments,
variant of common HE which is characterized microspherocytes and elliptocytes
by unusual sensitivity of red cells to heat (45 4. Heat sensitivity: RBC fragmentation when
C) warmed to 45 oC

5
 LECTURE TITLE

6. Glucose-6-phosphate dehydrogenase (G6PD) appear in large number only after splecnectomy

A. Inheritance pattern: Sex-linked 3. Incubated osmotic fragility: Often increased
B. Cause: 4. Fluorescent spot test for pyruvate kinase: decreased
o A decrease of G6PD in the HMP shunt 5. Quantitative PK assay: Decreased
prevents the production of NADPH = 6. Autohemolysis: positive but does not correct with
synthesizes reduce glutathione. addition of glucose (unlike in HS)
o Reduce glutathione is required to prevent
hydrogen peroxide buildup in the RBC during 8. Methemoglobin Reductase Deficiency
oxidant stress due to infections and certain A.Inheritance pattern: Autosomal recessive
chemical exposure. B. Cause
o Increased levels of H202 irreversibly o A decrease of methemoglobin reductase in the EM
denatures hemoglobin resulting in Heinz glycolytic pathway allows Hgb to remain in the
bodies. Ferric (Fe3+) state. The oxidized iron molecule
o As RBCs with Heinz bodies traverse the cannot carry oxygen to the tissues.
microcirculation, hemolysis results.
C. Additional laboratory Evaluation:
 Usually associated with sensitivity to certain drugs and 1. Methemoglobin assay: Increased
to ingest in or even inhalation of the pollen of the 2. NADH methemoglobin reductase activity:
common European broad bean (Vicia faba). Thus the Decreased
disorder is called favism
 Favism – a subgroup of G6PD-deficient 9. Paroxysmal Nocturnal Hemoglobinuria (PNH)
patients having severe life-threatening o an acquired intravascular hemolytic anemia
hemolysis that occurs within hours after characterized by intermittent (paroxysmal) sleep-
eating fava beans associated (nocturnal) blood in the urine
 Anemia caused is found in all races, but the highest is (hemoglobinuria)
among African and Mediterranean cultures. o Onset is insidous occuring mostly in middle-aged
 there are 2 genetic isoenzyme variations of G6PD have adults of either gender.
been identified: (depending on electrophoretic mobility) o Cause: Red cell membrane defect. At least 9 cell
1. Type A – individuals have clinically mild conditions surface proteins that regulate complement are
but can be affected by oxidants to produce a hemolytic missing. As a result, the RBC membrane is
crisis. markedly sensitive to complement.
2. Type B – patient have an enzyme activity of only 1% = Complement is a series of CHONs circulating in
and are more susceptible to severe oxidant hemolysis the blood which, when activated, can cause
than type A. disruption of cell membrane (RBC and
 G6PD is normally highest in young RBCs and microorganisms)
decreases as cell ages.
 Oxidized hgb denatures and precipitates intracellularly o known as Marchiafava- Micheli Syndrome
as Heinz bodies, which adhere to the membrane and o Clinical features:
cause rigidity, a tendency to lyse, and splenic trapping. 1. Hemoglobinuria in first morning specimen:
 A patient’s susceptibility to hemolytic crisis can greatly hemosidenuria
increase with an illness or exposure to drugs that have 2. Chronic anemia (hgb: normal to <6g/dL),
oxidant properties. leukopenia and thrombocytopenia
3. Infections
C. Additional laboratory Evaluation: 4. Thrombosis
1. Retic count: increased during hemolytic 5. 5-10% of cases will convert to acute
episodes myelogenous leukemia; 25% will convert to
2. Supravital stains for Heinz bodies: Positive aplastic anemia
3. Flourescent spot test for G6PD: No o Additional Laboratory Evaluation
fluorescence 1. Retic count: Increased
4. Ascorbate cyanide test: Positive 2. RBC acetylcholinesterase: decreased
5. Quantitative G6PD assay: Decreased 3. WBC, RBC and platelet count: Decreased
4. Urine hemoglobin: positive in first morning
7. Pyruvate Kinase Deficiency urine
A. Inheritance pattern: Autosomal recessive 5. Screening test: Sugar water test (Sucrose
B. Cause hemolysis test: Positive
o A decrease of pyruvate kinase in the Embden 6. Confirmatory test: Acidified serum lysis
Meyerhof Glycolytic pathway prevents synthesis test: (Ham’s test) Positive
of levels of ATP needed for adequate RBC 7. BM biopsy and aspirate: Erythroid
function. Hyperplasia
o A by-product of the glycolytic pathway is the o PNH Therapy
conversion of NADH to NAD  Therapy is directed at the complications that
o PK deficiency leads to: arise from infections, anemia and
 ATP depletion impaired ion pumps red thromboses.
1. Anemia – iron therapy or transfusion if
cell dehydration and finally hemolysis
severe enough
C. Additional laboratory Evaluation: 2. Hemolytic episodes – corticosteroid
1. Osmotic fragility: Normal therapy to stimulate erythropoiesis
3. Thromboses – anticoagulant therapy
2. PBS: presence of echinocytes (dessicocytes) which

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 LECTURE TITLE

B. Aplastic Anemia ANEMIA PDF

• a disorder characterized by aplasia of the bone marrow
or its destruction by chemical agents or physical RED BLOOD CELL DISORDERS
factors. All cell lines are affected. (Pancytopenia) ANEMIA
• Also known as panmyelophthisis or panmyelopathy • One of the most common disease worldwide
• derived from the Greek word anaimia, meaning
A. Acquired “without blood.” (or there is a small amount of RBC)
1. Idiopathic or primary with no clear cause • A decrease in the number of RBCs, or the amount of
2. Secondary due to: hemoglobin in the RBCs, results in decreased oxygen
a. Chemical agents such as benzene, insecticides, delivery and subsequent tissue hypoxia.
weed killers
b. Drugs such as chloramphenicol(antibiotic), • Most anemic patients have low oxygen saturation/low
phenylbutazone (anti-inflammatory) and Dilantin O2 level due to decreased level of RBC or Hb
(anti- convulsants) • considered to be present if the hemoglobin
c. Ionizing radiation from nuclear fallout, x-rays or concentration of the red blood cells (RBCs) or the
adiation therapy, radium packed cell volume of RBCs (hematocrit) is below the
d. Infections such as hepatitis, Epstein-Bbarr lower limit of the 95% reference interval for the
Virus, CMV, HIV e. Miscellaneous – pregnancy, individual’s age, gender, and geographical location
malnutrition, imunologic dysfunction (black Asians resistant to malaria).
• A functional definition of anemia is a decrease in the
NB: "Approximately 70% of acquired aplastic oxygen carrying capacity of the blood. It can arise if
anemia cases are idiopathic. (Rodak, 2012)" there is insufficient hemoglobin (low Hb count) or the
hemoglobin is nonfunctional (rare,
B. Congenital: Fanconi’s anemia hemoglobinopathies, Hb S, Hb C, Hb SC. Common:
o It is one of many inherited d/o that may lead to thalassemia). (either quality or quantity effect. E.g Hb
aplastic anemia can’t function because of lack in amino acid or there is
o Diagnosis is confirmed with cytogenetics studies no supporting Hb to transport O2). The former is the
displaying an increased propensity for more frequent cause.
spontaneous chromosomal breakage. • defined operationally as a reduction, from the baseline
value, in the total number of RBCs, amount of
Clinical Manifestation of Aplastic Anemia: circulating hemoglobin, and RBC mass for a particular
1. Progressive fatigue patient.
2. Dyspnea • Low value of RBC and Hb including the red blood cell
3. Palpitations mass.
4. Bleeding (petechiae, purpura, ecchymoses and mucosal The causes of anemia fall into three major
bleeding) pathophysiological categories:
5. Infection Laboratory Findings in Aplastic Anemia: 1. Blood loss (acute blood loss – certain individual
1. RBC, hgb (RBC, hgb (<7), hct (<20), retic (absolute and encountered stab wound, continuous external bleeding
corrected): decreased resulting to decrease in blood volume. The RBC is
2. WBC (<1.5), absolute neutrophil count (<0.5): decreased associated with the blood, continuous bleeding will lead to
3. Platelets (<20,000-60,000): decreased anemia)
2. Impaired red cell production (defect in the development
Treatment: of RBC or factors a decrease erythropoiesis, can be drugs
o Bone marrow transplantation or chemotherapy)
 If this is not an option, immunotherapy is 3. Accelerated red cell destruction (hemolysis in excess of
given. Spontaneous recovery may occur if the the ability of the marrow to replace these losses)
offending agent is removed. (hemolyzed RBC due to external factors in the external
body that will lead to hemolysis/abnormality or the RBC
Pure Red Cell Aplasia morphology is abnormal which triggers the macrophages to
• Hyporegenerative anemia lyse the RBC ------hemolysis ------- ANEMIA)
• involves a reduced production of RBCs which cannot
be remedied by any of the known hematopoietic factors Is anemia a disease or just a symptom of another disease?
• Rare condition in which only the red cell precursor in o Anemia may be a sign of an underlying disorder. Or
the bone marrow are decreased. anemia maybe a disease (The patient has illness but
• Causes: the anemia is not the major copndition. Anemia is
1. Acquired pure red cell aplasia: thymoma and caused by the first disorder (alcoholism). Dilutional
infection with parvovirus B19 anemia with normal or increased total red cell mass
2. Congenital pure red cell aplasia: The Blackfan- may occur with pregnancy, macroglobulinemia, and
Diamond syndrome splenomegaly.
 Dilutional anemia – the proportion of the
Congenital Dyserythropoietic Anemia plasma and RBC is not equal. (Many plasma)
o Some anemias have more than one pathogenetic
• HEMPAS – hereditary erythroblast multinuclearity with mechanism and go through more than one
a positive acidified serum test. morphological state, such as blood loss anemia. In the
case of accelerated red cell destruction, hemolysis in
excess of the ability of the marrow to replace these
losses occurs.

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 LECTURE TITLE

QUANTITATIVE (amount/count) CHANGES IN RBCs 2. INSUFFICIENT ERYTHROPOIESIS– decrease in

ANEMIA ERYTHROCYTOSIS or the number of erythroid precursor cells. Quantity of
POLYCYTHEMIA (excess the red blood cells. If the number of RBC is correct in
or in flacs of RBCs) one maturation series. (A normal pronormoblast
Decreased RBC’s (low level ERYTHROCYTOSIS – produces 16 matured RBCs, once the pronormoblast
RBC count in the automated increase in RBC count just produce 10 RBCs, there is a problem in the correct
machine) POLYCYTHEMIA – appear production of RBC)
RBC count because the
Decreased oxygen carrying amount of the plasma is not INEFFECTIVE ERYTHROPOIESIS – QUALITY
capacity of blood (due to proportional to the • refers to the production of erythroid progenitor cells
decreased Hb. RBC without hematocrit that are defective. These defective progenitors are
Hb cannot carry O2)
often destroyed in the bone marrow before their
−Too many RBCs in the
maturation and release into the peripheral circulation.
circulation
−Increased PCV above ref • Ex. Megaloblastic anemia (DNA synthesis – portion for
range nuclear maturation, defect in the quality), Thalassemia
−Hypervolemia (increase (decrease amount of globin chains that would lead to
blood volume. ineffective erythropoeisis), Sideroblastic anemia
Usual blood volume is 5-7 • The peripheral blood hemoglobin is low despite an
Liters) increase in RBC precursors in the bone marrow (even
−Hyperviscosity there are too many RBC precursors but has low level
(increased nblood formed of Hb, that would be non-functional. Hb should be
elements, the blood incorporated in the RBC because it functions to carry
becomes more viscous) O2)
INSUFFICIENT ERYTHROPOIESIS – QUANTITY
• refers to a decrease in the number of erythroid
precursors in the bone marrow, resulting in decreased
RBC production and anemia.
1. Iron Deficiency Anemia
Iron – major portion where heme is poroduce,
without heme – no Hb, without Hb – no RBC.
2. Malabsorption – without nutrients coming from the
small intestine, or when the nutrients are not absorbing
back on the organs, there is a lack in nutritional
development.
3. Excessive loss from chronic bleeding – removed
blood loss/decrease blood volume.
4. Deficiency of erythropoietin
Erythropoietin – indicator on erythropoiesis
5. Loss of the erythroid precursors due to an
autoimmune reaction – happens for an instance when
Don’t just rely on signs and symptoms. Clinical signs and
the polychromatophilic normoblast attacked by an
symptoms are non-specific. There is a need for further
antibody, the polychromatophilic needs to mature in
diagnosis through lab tests.
the next stage but when it is being destroy or killed,
Blood smear to see the size or morphology of the RBC
there is no RBC that will going to form. Thus, it results
Microcytic hypochromic - problem in RBC maturation
to a decrease in RBC count.
Microcytic cytoplasmic – maturation logs behind nuclear
6. Infection (parvovirus B19) – attaches in the RBC that
maturation. Small cytoplasm
will lead to hemolysis. When the RBC is hemolyzed,
Macrocytic – nuclear maturation logs behind cytoplasmic
the number of RBC circulating in the bloodstream is low
maturation. Larger cytoplasm
7. Suppression of the erythroid precursors due to
Normal – hypoproliferation (different blood cell proliferate)
infiltration of the bone marrow with granulomas
Iron Deficiency Anemia can be associated in normocytic and
(sarcoidosis) or malignant cells (acute leukemia)
normochromic
Anemia is the underlying signs
CATEGORIES OF ANEMIA
MECHANISM OF ANEMIA
1. BLOOD LOSS
ERYTHROPOIESIS
• Acute
• 120 days’ life span, 1% removed and produced, with • Chronic
nutritional requirements, healthy BM 2. IMPAIRED PRODUCTION
• Senescent RBC/aged RBC – RBC which already • Aplastic
fulfilled its life span. This is ready to be lyse. • Iron deficiency
• It is the term used for marrow erythroid proliferative • Sideroblastic anemia
activity • Anemia of chronic disease
1. INEFFECTIVE ERYTHROPOIESIS – defective • Megaloblastic
progenitor cells. Quality of the erythropoiesis. Defect 3. HEMOLYTIC
on the development or the process of RBCs. (Eg: • Inherited defects
polychromatophilic normoblast is release on the BM, • Acquired disorders
bone marrow cannot function well) 4. HEMOLYTIC-HEMOGLOBIN DISORDERS

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CLASSIFICATION OF ANEMIA • Red Cell Morphology Examination – peripheral blood

• MORPHOLOGIC CLASSIFICATION smear
o Size of red blood cells (MCV) • Reticulocyte Count
o Hgb concentration (MCHC) • White blood cell count/PLT count
MCV – 80-100 fL femtoLiters • Bilirubin Measurement
MCHC – 31-37 g/dl • Estimation of serum haptoglobin (hemolysis, the Hb
bind to haptoglobin)
• ETIOLOGIC CLASSIFICATION (pathway/mechanism) • Free Hgb in the blood
o Causes: pathophysiological mechanism responsible • Osmotic Fragility Test (OFT) – test for spherocytes. It
for the RBC deficit tests the fragility or integrity of RBCs
o Can be due to insufficient erythropoiesis that will result • Haptoglobin
to decrease erythrocytes production. Can also be due
to hemolysis.
• Lactic Dehydrogenase test (marker for HA)
o increased in hemolytic anemia
o Decreased erythrocyte production
o Increased erythrocyte loss • Serum ferritin
o reflects the body’s tissue iron stores
MORPHOLOGICAL CLASSIFICATION o measured through immunoassays
o Male: 12-300ng/mL Female: 12-150ng/mL
• Normocytic normochromic anemia
o Bleeding • Total Iron Binding Capacity
o Hypoproliferation of hematopoietic stem cells o Test the availability where the iron binds. Too
many iron can just bind to other spaces.
• Microcytic hypochromic anemia o Iron deficiency anemia – lack of iron. There is
o IDA
available spaced for the freely iron to bind to
o Sideroblastic anemia
specific receptor.
• Macrocytic normochromic anemia o Unique marker for iron deficiency anemia.
o Megaloblastic anemia Increase TIBC.
o body’s ability to transport iron
APPROACH TO EVALUATING ANEMIAS o measured to identify how much iron is being
• Complete history taking (done by Physician) carried in the blood.
• Physical examination o Serum iron: 60-170 mcg/dl, TIBC: 240-450
• Hgb and Hct, Red Cell Count (medtech) mcg/dl Transferrin saturation: 20-50%
• Reticulocyte count and Anemia Classification • Fecal urobilinogen
o Check the effectivity or the sufficiency of the o measures the total excretion of the
erythropoiesis. breakdown products of heme
• MCV and Anemia Classification • Free Erythrocyte Protoporphyrin (FEP) test or Zinc
o Red blood cell indices (MCV, MCHC) Protoporphyrin (ZPP)
o determine the amount of protoporphyrin not
Signs and Symptoms of Anemia used for Hgb synthesis
Signs/Symptoms Yes or Signs/Symptoms Yes or o Measured by direct fluorescence-
No No hematofluorometer (15-18 ug/dL of RBCs)
• Plasma iron turnover
1. Dizziness Yes 8. Sensitivity to Yes o Not usually done in the laboratory
cold o radioactive iron is injected IV
2. Menstrual Yes 9. Paleness Yes o special test done in other countries
irregularity o Rate of disappearance from the blood (N is 2-
3 hrs)
3. Tachycardia Yes 10. Prone to NO
o 70-80% of the injected Fe reappears within 10
Infection
days
4. Faintness Yes 11. Dyspnea Yes
• Red cell life span
5. Feeling of Yes 12. Orthopnea Yes
o Chromium substance inject inside the body,
fatigue
the chromium attach in the RBC, blood
6. Nausea Yes 13.Flatulence Yes sample will be get and in the blood sample of
7. Anorexia Yes 14. Ruby NO the patient, we can see red blood cells that
complexion produce radioactivity. When RBC suddenly
disappear while seeing the radioactivity, that
LABORATORY PROCEDURES USED TO DIAGNOSE is a short life span meaning the RBC cannot
ANEMIA handle the chromium attached to it.
• Red Cell Count o Not routinely used in the laboratory
o Radioactive chromium
• Hemoglobin o Shorter life span = faster disappearance
• Hematocrit
• Red Cell Indices (MCV) (MCH) (MCHC) Increased RBC Concentration
o The most important of these indices is the • Hct: higher than 0.52 L/L – male
MCV, a measure of the average RBC volume • Hct: higher than 0.50 L/L – female
in femtoliters (fL)

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RETICULOCYTE COUNT
• serves as an important tool to assess the bone
marrow’s ability to increase RBC production in
response to an anemia.
• Reticulocytes are young RBCs that lack a nucleus but
still contain residual ribonucleic acid (RNA).
• Normally, they circulate peripherally for only 1 day
while completing their development.
• The adult reference range for the reticulocyte count is • Mean corpuscular hemoglobin (MCH)
0.5% to 1.5% expressed as a percentage of the total o The average weight of hemoglobin of the
number of RBCs.13 The newborn reference range is individual red cell.
1.5% to 5.8%, but these values change to
approximately those of an adult within a few weeks
after birth

PERIPHERAL BLOOD FILM EXAMINATION

• An important component in the evaluation of an anemia
is examination of the peripheral blood film, with
particular attention to RBC diameter, shape, color, and
inclusion.
• The peripheral blood film also serves as a quality
control to verify the results produced by automated
analyzers.
• Mean corpuscular hemoglobin concentration (MCHC)
• Certain shape abnormalities of diagnostic value (such o The percent of hemoglobin in the average red
as sickle cells, spherocytes, schistocytes, and oval cell.
macrocytes) and RBC inclusions (such as malarial
parasites, basophilic stippling, and Howell-Jolly
bodies) can bedetected only by studying the RBCs on
a peripheral blood film
• Finally, a review of the WBCs and platelets may help
show that a more generalized bone marrow problem is
leading to the anemia.

BONE MARROW EXAMINATION

• sternum, iliac crest, spinous processes of the
vertebrae, tibia, lumbar spinous processes determine
the M: E ratio
MEAN CELL VOLUME AND ANEMIA
CLASSIFICATION
• Microcytic anemia is characterized by an MCV of less
than 80 fL with small RBCs (less than 6 μm in
diameter). Hypochromia, characterized by an MCHC of
less than 32 g/dL and increased central pallor in the
RBCs, may accompany the microcytosis.
• iron deficiency, inability to utilize iron (chronic
inflammatory states)
• Bone marrow samples can be obtained by aspiration • globin synthesis
and trephine biopsy. Sometimes, a bone marrow
examination will include both an aspirate and a biopsy. • defect (thalassemia)
• The aspirate yields semi-liquid bone marrow, which • heme synthesis defect (sideroblastic anemia, lead
can be examined by a pathologist under a light poisoning)
microscope as well as analyzed by flow cytometry, • Macrocytic anemia is characterized by an MCV
chromosome analysis, or PCR. greater than
• Frequently, a trephine biopsy is also obtained, which • 100 fL with large RBCs (greater than 8 μm in diameter).
yields a narrow, cylindrically shaped solid piece of • Macrocytic anemias may be megaloblastic or non-
bone marrow which is examined microscopically megaloblastic.
(sometimes with the aid of immunohistochemistry) for • Megaloblastic anemias are caused by conditions that
cellularity and infiltrative processes. impair synthesis of deoxyribonucleic acid (DNA), such
as vitamin B12 and folate deficiency or
RED CELL INDICES myelodysplasia.
• Mean corpuscular volume (MCV)
o The average volume of the individual red
blood cell. Femtoliter (fl) or 10-15 liter = 1 fl.

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 LECTURE TITLE

MACROCYTIC - NORMOCYTIC, MICROCYTIC -

NORMOCHROMIC NORMOCHROMIC HYPOCHROMIC
a. Vitamin B12 a. Defective formation of Iron deficiency
deficiency – Folic the red blood cells or the anemia
acid deficiency presence of tumor cells in Thalassemia
Pernicious anemia the bone marrow Sideroblastic
Sprue Aplastic Anemia Leukemia anemia Chronic
Following Hodgkin’s disease Multiple blood loss
gastrectomy myeloma Anemia of
Ditary Leukoerythroblastosis chronic
Abnormal intrinsic Metastatic Cancer Anemia inflammation
factor associated with renal and Lead poisoning
Tapeworm endocrine disease
infection (D.latum)
b. Abnormal hemoglobin,
Red Blood Cell Distribution Width b. Disease of the increased destruction of
• The RDW can help determine the cause of an anemia liver red blood cells
when used in conjunction with the MCV Certain acquired hemolytic
• Each of the three MCV categories mentioned anemia PNH
previously (normocytic, microcytic, macrocytic) can Sickle cell anemia HDN
also be subclassified by the RDW as homogeneous Anemia of chronic renal
(normal RDW) or heterogeneous (increased or high insufficiency
RDW)

TABLE 18-4 Classification of Anemia Based on Red Blood

Cell Mean Volume (MCV) and Red Blood cell Distribution
Width (RDW)
MCV LOW MCV NORMAL MCV HIGH
RDW RDW RDW RDW RDW RDW High
Normal High Normal High Normal MICROCYTIC / HYPOCHROMIC ANEMIA
Heterozyg Iron Anemia of Early Aplastic Folate or
ous deficienc chronic iron, anemia vitamin B12 • Due to defective hemoglobin synthesis resulting to
Thalasse y inflammati folate, Chronic deficiency cytoplasmic maturation defect
mia Sickle on or liver Myelodyspla
Anemia of cell-B- Anemia of vitamin disease stic
chronic thalasse renal B12 alcoholi syndrome A. DEFECTIVE HEME SYNTHESIS
inflammati mia disease deficien sm Chemothera o Abnormalities of iron homeostasis (deficiency
on Acute cy py
Hb E hemorrah Mixed Cold
of metabolism)
disease/tr ge deficien agglutinin  SIDEROPENIC ANEMIA – commonly
ait Hereditary cy of disease known as the IDA
spherocyt
osis
iron +
B12 or
Chronic liver
disease
 SIDEROACHRESTIC ANEMIA
folate – adequate or excess iron but
Sickle defective utilization
cell - Anemia of Chronic Inflammation
anemia
Hb SC o Defective porphyrin metabolism
disease  SIDEROBLASTIC ANEMIA

B. DEFECTIVE GLOBIN SYNTHESIS

o results to Thalassemia •
o Deletion or mutation of globin genes

1.IRON DEFICIENCY ANEMIA

• Most common form of anemia •
• Prevalent in infants and children, pregnancy, excessive
menstrual flow, elderly with poor diets, malabsorption
syndromes, chronic blood loss
• ETIOLOGY
a. Inadequate intake of iron
b. Increase demand
c. Impaired iron absorption
d. Chronic blood loss
• Clinical signs and symptoms
o Fatigue, weakness, and shortness of breath,
especially with exertion, sore tongue
(glossitis),
o inflamed cracks at the corners of the mouth
(angular cheilosis), Koilonychia (spooning of

1
 LECTURE TITLE

the fingernails) may be seen if the deficiency siderocytes; inclusions are siderotic granules
is long-standing. (Pappenheimer bodies on Wright’s stained smears)
o Patients also may experience cravings for • Siderocytes are best demonstrated using Perl’s
nonfood items, called pica. Prussian blue stain
• PICA = The cravings may be for things such as dirt, • TYPES
clay, laundry starch. 1. PRIMARY – irreversible; causes of block is unknown
STAGES OF IDA  Two RBC populations (Dimorphic)
STA DESCRIPT HEMOGL SERUM TIBC FERRIT  One of myelodysplastic syndromes-
GE ION OBIN IRON IN Refractory anemia with ringed sideroblasts
(RARS)
Stag Storage Normal Normal Normal Decrea 2. SECONDARY- reversible; causes include alcohol,
e1 depletion sed anti-TB drugs, Chloramphenicol

4. ANEMIA DUE TO LEAD POISONING

Stag Transport Normal Decrea Increas Decrea  Anemia, when present in lead poisoning, is most often
e2 depletion sed ed sed normocytic and normochromic; however, with chronic
exposure to lead, a microcytic, hypochromic clinical
picture may be seen
Stag Functional Decreased Decrea Increas Decrea
o Multiple blocks in the protoporphyrin pathway
e3 depletion sed ed sed that affect heme synthesis
o Presence of many coarse Basophilic Stippling
o lead poisoning will lead to acquired
Parasites associated with IDA sideroblastic anemia and acquired porphyria
• Hookworms o It inhibits ferrochelatase and D-ALA synthase
enzyme in Heme/Protoporphyrin pathway
• Trichuris trichiura
• Schistosoma mansoni
• Schistosoma haematobium
IRON AND RELATED STUDIES
TRANSFERRIN -Beta-globulin responsible for binding
iron and its transport in the blood stream.
-1 gram can bind 1.25 mg of iron
TIBC -Refers to the total amount of iron that
transferrin can carry
-Functional measurement of the
capacity of transferrin to bind iron. MACROCYTIC / NORMOCHROMIC ANEMIA
% SATURATION An index of iron storage Calculated
value: 100 x serum iron/ TIBC • It is characterized by an MCV greater than 100fl w/
FERRITIN Storage form of iron Rough estimate of large RBC (greater than 8 um diameter). Macrocytic
body iron content anemias maybe megaloblastic or non-megaloblastic.
• The most common causes of megaloblastic anemia
2. ANEMIA OF CHRONIC DISEASE/INFLAMMATION are acquired, although congenital forms exist.
• Second most common type of anemia Deficiencies in cobalamin, folate, or both account for
the majority of cases
• Commonly associated with systemic diseases,
including chronic inflammatory conditions such as
• Red blood cells in megaloblastic anemias have an
rheumatoid arthritis, chronic infections such as abnormal nuclear maturation and imbalance between
tuberculosis or human immunodeficiency virus nuclear and cytoplasmic maturation.
infection, and malignancies • Refers to the abnormal marrow erythrocyte precursor
• ETIOLOGY seen in processes, such as pernicious anemia,
a. Due to inability to use available iron for hemoglobin associated with altered DNA synthesis.
production
b. Impaired release of storage iron associated with
increased Acute phase reactants such as Hepcidin
(decreases release of iron from stores), Ferritin, and
Lactoferrin

3. SIDEROBLASTIC ANEMIA
• Caused by blocks in the protoporphyrin pathway
resulting in defective hemoglobin synthesis and iron
overload.
• Excess iron accumulates in the mitochondrial region of
the immature RBC in the BM and encircles the nucleus;
cells are called ringed sideroblasts.
• Excess iron accumulates in the mitochondrial region of
the mature RBC in circulation; cells are called ringed

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 LECTURE TITLE

NON- MEGALOBLASTIC ANEMIA o Functional Aspects

MEGALOBLASTIC/NORMOBLAS  Regulates the folate synthesis
TIC ANEMIA (DNA)
-Disruption of the cholesterol-to- -Impaired DNA synthesis
phospholipid ratio Causes of Vitamin B12 Deficiency
-No impairment of DNA synthesis • Dietary intake (rare) – total vegetarianism
-Mostly round red blood cells -Oval macrocytes, • Intestinal malabsorption
-No megaloblastic in the bone hypersegmented PMNs in the −Lack of Intrinsic Factor (Pernicious Anemia)
marrow -No hypersegmented blood −Jejunal bacterial growth (stagnant loop
-Megaloblasts in the bone syndrome)
marrow −Fish tapeworm infestation (D. latum)
-Normal newborn status -Vitamin B12 deficiency −Ileal dysfunction
-Reticulocytosis -Folic acid deficiency
-Liver disease -Chronic alcoholism -Acute erythroleukemia b. FOLIC ACID DEFICIENCY
-Bone marrow failure -Myelodysplasia • Associated with poor diet, pregnancy or
-Congenital Dyserythropoietic chemotherapeutic anti folic drugs such as
Anemia Type 1 and 3 METHOTREXATE.
1. MEGALOBLASTIC ANEMIAS
• Folic acid has low body stores
• Defective DNA synthesis causes abnormal nuclear • Clinical symptoms: same with vitamin B12 deficiency,
except NO CNS involvement
maturation; RNA synthesis is normal, so the cytoplasm
is not affected. The nucleus matures slower than the
Folic Acid (Pteroylmonoglutamic acid)
cytoplasm (Asynchronism)
o Dietary Aspects
• It is an example of ineffective erythropoiesis  Green leafy vegetables (spinach,
• Caused by either vitamin B12 or folic acid deficiency lettuce, broccoli, asparagus)
• Laboratory picture of Pancytopenia, Oval macrocyte,  Liver, kidneys, yeast and mushroom
Howell- jolly bodies, Hypersegmented neutrophil o Metabolic Aspects
 Absorb at the jejunum (from food –
SCREENING TEST USED TO DIAGNOSE MEGALOBLASTIC polyglutamate, intestine –
ANEMIA monoglutamate)
o Functional Aspects
Five tests used to screen for megaloblastic anemia are the  Acts as co-enzyme in the formation
1. Complete blood count (CBC) - check for oval of thymidylate synthetase
macrocytes and pancytopenia  Thymidylate synthetase – needed to
2. Reticulocyte count- decrease retic count produce thymidine (one of the
3. White blood cell (WBC) manual differential- check pyrimidine bases of DNA)
for hypersegmented cells
4. Serum bilirubin- increase indirect bilirubin and total Causes of Folic Acid deficiency
bilirubin
5. Lactate dehydrogenase- elevated LD (flipped LD)
• Decreased intake
o Low reserves
o Excessive cooking destroys FA
A. VITAMIN B12 (COBALAMIN) DEFICIENCY
o Liver impairment −Malabsorption (tropical
• Pernicious anemia – caused by deficiency of intrinsic sprue, celiac disease, intestinal lymphoma,
factor, Antibody to intrinsic factor or antibodies to resection of the jejunum)
parietal cells
• Increased requirements
• Other causes: Malabsorption, total gastrectomy, total o Pregnancy
vegetarian diet, D. latum infection, Intestinal blind o Growing children
loops, and sprue
• Hyperactive hematopoiesis (excessive
• Vit. B12 deficiency takes 3-6 years to develop because bleeding)
of high body stores
• Celiac disease
• Clinical symptoms: Jaundice, weakness, glossitis, GI o is a condition that
bleeding, numbness and CNS PROBLEMS damages the lining of
the small intestine and
Vitamin B12 prevents it from
o Chemical Aspects absorbing parts of food
 Cobalamins that are important for
o Dietary Aspects staying healthy.
 Foods of animal origin (liver, organ
meats, seafood, dairy products) • Tropical sprue
o Metabolic Aspects o is a malabsorption disease commonly found
 Vitamin B12 is attached to the protein in the tropical regions, marked with abnormal
complexes in food. flattening of the villi and inflammation of the
 IF (intrinsic factor) is needed for its lining of the small intestine.
proper absorption in the ileum,
 IF is produced in the parietal cells in the
wall of the stomach.

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2. NON- MEGALOBLASTIC ANEMIA

• Include Alcoholism, Liver disease, and conditions
causes accelerated erythropoiesis
• RBC are round and not oval as seen in the
megaloblastic anemias

IMPORTANT FINDINGS IN MEGALOBLASTIC ANEMIA

• HYPERSEGMENTED NEUTROPHILS
• INEFFECTIVE ERYTHROPOIESIS AND
DECREASED RETICS, WBC, PLATELETS
• MACROOVALOCYTES • HOWELL-JOLLY BODIES

GOODLUCK AND GODBLESS!