Egypt J Pediatr Allergy Immunol 2014;12(2):49-61.

Review article
Pathophysiology of immune thrombocytopenic purpura: a bird's-eye view.
Amira Abdel Moneam Adly
Pediatrics Department, Faculty of Medicine, Ain Shams University, Cairo, Egypt
ABSTRACT and B lymphocytes (including T-helper, T-
Immune thrombocytopenic purpura (ITP) is a cytotoxic, and T-regulatory lymphocytes) 6.
common autoimmune disorder resulting in isolated The triggering event for ITP is unknown7, but
thrombocytopenia. It is a bleeding disorder continued research is providing new insights into
characterized by low platelet counts due to decreased the underlying immunopathogenic processes as well
platelet production as well as increased platelet as the cellular and molecular mechanisms involved
destruction by autoimmune mechanisms. ITP can in megakaryocytopoiesis and platelet turnover.
present either alone (primary) or in the setting of other Although historically ITP-associated thrombo-
conditions (secondary) such as infections or altered cytopenia was attributed solely to increased rates of
immune states. ITP is associated with a loss of destruction of antibody- coated platelets, it has
tolerance to platelet antigens and a phenotype of become evident that suboptimal platelet production
accelerated platelet destruction and impaired platelet also plays a role 8.
production. Although the etiology of ITP remains Bleeding is due to decreased platelet production
unknown, complex dysregulation of the immune as well as accelerated platelet destruction mediated
system is observed in ITP patients. Antiplatelet in part by autoantibody-based destruction
antibodies mediate accelerated clearance from the mechanisms9. Most autoantibodies in ITP are
circulation in large part via the reticuloendothelial isotype switched and harbor somatic mutations10,
(monocytic phagocytic) system. In addition, cellular and as such a role for CD4+ helper T cells in
immunity is perturbed and T-cell and cytokine profiles disease pathogenesis has been invoked. Consistent
are significantly shifted toward a type 1 and Th17 with this, ITP patients have activated platelet-
proinflammatory immune response with impaired autoreactive T cells with increasing cytokine
regulatory compartment, including Tregs and Bregs, imbalance toward IL-2 and IFN-γ, indicating a shift
have been reported, suggesting a generalized immune toward Th1 cells11. More recently, increased Th17
dysregulation in ITP. Understanding how cells or IL-17 cytokine were reported in ITP
Th1/Th17/Treg differentiation and expansion are patients12, implicating a possible role for Th17 cells
controlled is central to uncovering how autoimmunity in ITP immunopathology.
may be sustained in ITP. Moreover, a role for cytotoxic T cells in direct
lysis of platelets and megakaryocytes in the bone
INTRODUCTION marrow has been proposed 13. In addition to an
Immune thrombocytopenia (ITP) is recognized as increase in the effector T cell arm of the immune
an immune mediated disorder in which platelets are response (Th1, Th17 and CD8 cells); a decrease in
opsonized by autoantibodies and prematurely the regulatory immune compartment of patients
destroyed by reticuloendothelial system1. It is a with ITP has been described. Specifically, a
hematologic disorder affecting children with an deficiency in generation and/or defective functions
incidence of four to five cases per 100,000 children of ITP regulatory T cell (Treg)14 and regulatory B
per year2.It is characterized by immune-mediated cells (Bregs)15.
accelerated platelet destruction and suppressed This brief review will highlights the
platelet production. Although the etiology of ITP is mechanisms, and their elements, underlying the
not yet known, and the diagnosis continues to be pathogenesis and cellular kinetics of ITP and
one of exclusion.3 A number of studies have discusses the aspects of current understanding of
provided evidence of disturbed innate and adaptive immune dysregulation. Also it addresses the recent
immune responses in patients with ITP.4 The findings on the state of the Breg and Treg
pathophysiology of ITP increasingly is understood compartments by which this information may guide
better5. Not surprisingly, it is complex with therapy in ITP patients in the future.
involvement of many players in the human immune
orchestra, including antibodies, cytokines, antigen-
presenting cells, costimulatory molecules, and T
49
Adly

Implicating agents in ITP of ITP often occurs seemingly rapidly and between
ITP is a heterogeneous group of disorders with monitoring time points19.
potentially differing etiologies, natural histories and Despite these limitations, data from numerous
response to therapy3. Some ITP patients appear to studies in recent years are beginning to come
have fundamental disturbances in their innate and together to form a picture of an unbalanced immune
adaptive immunity, while in others a specific often response. Not surprisingly, the immune changes
times self-liming inciting agent may be involved. observed during ITP are complex. The long-held
Several potential inciting agents have been dogma of platelet-bound Abs leading to
identified or proposed. Fc_receptor (Fc_R)–mediated clearance of platelets
In the case of childhood ITP, many patients give by phagocytes residing in the spleen (and liver)
a history of a recent infection. In adults, infection continues to be a central theme in our current
with human immune deficiency virus, Helicobacter understanding of ITP. In addition to this, the
pylori and hepatitis C have been implicated in a evidence supports a wide array of immune shifts
high percentage of cases of ITP in endemic areas; involving all components of the immune system,
the causal relationship is strongest in settings where resulting in both shortened platelet survival and
microbial elimination leads to resolution of the inhibition of the production of platelets20.
associated ITP16. Acute ITP can also occur after
vaccination. The evidence for vaccine associated The role of the spleen
ITP is most compelling in those immunized with In 1916, Kaznelson, while a medical student in
measles-mumps-rubella vaccine (~1:40,000). There Vienna, prevailed upon the attending surgeon to
have also been case reports of ITP in children perform a splenectomy in a patient with ITP. The
following other vaccinations against hepatitis B, splenectomy was successful in normalizing the
diphtheria-tetanus-pertussis, and hepatitis A, but the platelet count and, with other cases, first established
relationship is less compelling17. the critical role of the spleen in ITP21. However, the
It has been hypothesized that ITP results from cause of thrombocytopenia remained unclear. Was
molecular mimicry engrafted on a normal immune the spleen destroying the platelets or did it secrete a
repertoire, such that the immune thrombocytopenia suppressive substance that inhibited platelet
resolves once the inciting antigen has been production and/ or release into the circulation?
eliminated. Although microbial antigens may play a Doan et al.22 examined a number of spleens from
role in the induction of vaccine associated ITP, patients with ITP. They demonstrated sea blue
many vaccines also contain adjuvants (e.g., alum) (lipid laden) histiocytes in the spleen, suggesting it
that might potentiate the immune response. was the platelet ‘destroyer’. What directed the
Recent work from Shoenfeld and coworkers spleen to prematurely destroy platelets, however,
suggests that some autoimmune syndromes and remained unclear23. The spleen, with its unique
inflammatory states might be induced by these microvascular architecture, is well suited for
immune adjuvants themselves18. The name they immunologic surveillance of the body platelet mass
have given to this activity is as it has the capacity to store a large number of
Autoimmune/Inflammatory Syndrome Induced by platelets in an exchangeable pool in close proximity
Adjuvants (ASIA). The role that ASIA might play to cells of the immune system23. The intrasplenic
in the initiation of ITP requires further study, but platelet transit time has been estimated to be around
the observation that ITP has been seen after the 10 min and is independent of spleen size. The
administration of several different vaccines demonstration that splenic tissue from patients with
provides an interest in this possibility. ITP produces antiplatelet IgG suggests that the
Identification of factors that precipitate ITP is spleen has capacity of forming these autoantibodies
extremely difficult due to the likely transient nature and that this mechanism is active in the disease24. In
of the provoking event, the inherent difficulty in addition, the splenic macrophages have been shown
diagnosing ITP early in its course, and the to avidly phagocytose platelets sensitized by serum
prolonged time period over which monitoring from ITP patients25. The spleen therefore, both
would need to occur to capture the onset of ITP. produces antiplatelet antibodies and sequesters
Even in subjects known to be at high risk for ITP opsonized platelets. The central role of the spleen in
due to a personal history of ITP, a strong family ITP is further illustrated by the fact that 70% of
history of ITP (rare), or comorbidity with a chronic ITP patients enter a durable remission after
condition predisposing to secondary ITP, the onset splenectomy26.

50
 Pathophysiology of ITP

Figure 1. Model of relationship of contributing factors in ITP.

(Johnsen J. Pathogenesis in immune thrombocytopenia: new insights. Hematology Am Soc Hematol Educ Program.
2012;2012:306-12).

Antigen presenting cells represents another mechanism by which platelet

The role of antigen-presenting cells (APCs) for the autoantibodies may lead to immune-mediated
loss of tolerance in ITP remains unclear, but a platelet destruction. Several studies have
model has been advanced in which APCs are demonstrated increased platelet-associated C3 and
crucial in generating a number of new or cryptic C4 on ITP platelets but these are thought to be
epitopes from platelet glycoproteins. In this model, secondary in importance to platelet IgG and/or the
APCs expressing these novel peptides, along with result of antiplatelet IgM29. Furthermore, there is an
costimulatory molecules, induce the activation of T- association of (especially) C4 deficiency and ITP
cells that recognize these additional platelet that has not been well studied and is of additional
antigens. Thus, this acquired recognition of new interest because the C4 genes are in the midst of the
self-determinants, or epitope spreading, may play HLA region on chromosome 630.
an important role in the initiation and perpetuation
of ITP. T-cell clones that react with cryptic epitopes Role of T cells
may escape the negative selection in the thymus T-cell abnormalities in patients with ITP
when self-determinants are present at a sub- A number of T-cell abnormalities have been
threshold concentration5. demonstrated in patients with ITP (summarized in
Fig 2) and it is likely that there are three main
Complement system mechanisms by which T cells could be involved in
In general, Abs specifically bound to cell-surface the thrombocytopenia in patients with ITP. First, a
antigens not only mediate clearance from number of studies suggest a Th1 bias, compared
circulation by Fc_Rs, but also can serve to fix with Th2, in adults with chronic ITP. For example,
complement on cells. Recently, plasma from ITP increased numbers of HLA-DR+ T cells, increased
patients was shown by 2 groups to be capable of soluble IL-2 receptors, and a cytokine profile
fixing complement to platelets in vitro27. suggesting the activation of precursor helper T and
Furthermore, platelets from ITP patients also type 1 helper T cells have been described31.
exhibit detectable complement, and the ability to fix Reduced levels of IL-10 have also been described
complement was correlated with the presence of in patients with active disease when compared with
detectable antiplatelet Abs28. Therefore, those in remission or healthy controls32 but
complement-mediated immunity, either by conversely, raised IL-10 levels have been described
enhanced clearance or direct cell destruction, in children with chronic ITP33. Further evidence of
51
Adly

Th1 involvement in the pathology of ITP is DNA microarray screening, Olsson et al.37 found
illustrated by an increase in the Th1 cytokines, IL-2 increased expression of several cytotoxic genes,
and IFN-c, in patients with ITP when compared such as granzyme A, granzyme B and perforin, as
with controls34. Interestingly, this increase was well as increased expression of genes involved in
more marked in patients in remission than in those the Th1 cell response, such as INFc and IL-2
with active disease. receptor-b in a small number of patients with ITP
Overall, they describe significantly increased when compared with controls. As apparent
Th1/Th2 ratios in patients with both active and compensation for this increased cytotoxicity, they
quiescent disease when compared with controls. also found increased expression of the killer cell
These findings may be related to ongoing immune immunoglobulin-like receptor (KIR) family on
activation as part of autoimmunity. The activity of CD3+ T cells in patients with ITP in remission
regulatory T cells and the potential for in vivo T- when compared with controls and to those with
cell exhaustion because of prolonged in vivo active ITP. KIRs downregulate cytotoxic T
activation has not been well studied in ITP30. lymphocytes (CTL) and natural killer cell (NK)
A second method of potential T-cell responses by binding to MHC class 1 molecules,
involvement is the release of cytokines that preventing lysis of target cells. These findings
interfere with megakaryocyte maturation and/or suggest that CTLs may be involved in ITP. In a
platelet release. Transforming growth factor (TGF)- direct assay similar to that measuring NK cell
b1 level has been inversely correlated with disease activity by using radiolabelled K562 targets, these
activity32. The role of TGF-b1 in ITP is thought to investigators assessed platelet destruction in vitro
be as a potent inhibitor of megakaryocyte by T cells. They found that six of eight patients
maturation. Two studies have shown increased with active ITP showed platelet lysis by T cells
granulocyte-macrophage colony stimulating factor whereas none of the patients in remission did. The
(GM-CSF) levels, and one increased macrophage effector cells were found to be CD3+CD8+ T
(M) CSF levels, suggesting that cells37.
monocyte/macrophage activation is associated with This expanded role of cytotoxic T cells may
ITP35. Circulating cytokines may also alter the explain why not all of the patients originally
response of HLA class II presentation, and/or described by Harrington et al.38 had a fall in their
influence the interaction between B and T platelet count following the infusion of plasma from
lymphocytes causing pre-existing B cells to patients with ITP, and may also explain a
proliferate and produce high-affinity percentage of patients without measurable
autoantibodies36. Finally, there is evidence to antiplatelet antibodies, and again points to the
suggests a direct cytotoxic effect of T cells, by heterogeneity of this disease.

Figure 2. Summary of the variety of T-cell abnormalities found in adults with ITP.
PBMC, peripheral blood mononuclear cells; MLR, mixed lymphocyte reaction; ITP, immune thrombocytopaenic
purpura (Cooper N, Bussel J. The pathogenesis of immune thrombocytopaenic purpura. Br J Haematol. 2006
May;133(4):364-74.)

52
 Pathophysiology of ITP

T-cell phenotypes in ITP (CD8+) cells42. Moreover, the increase in Tc17 cells
From studies of the immune system in animals, is correlated with skewing of the CD4:CD8 ratio in
immune homeostasis is thought to be maintained ITP42.
via a balance of type 1 (IFNγ, IL-2, TNFα, and An additional discrete T-cell subset, Th22
TNFβ1 related) and type 2 (IL-4, IL-5, IL-6, IL-10, (CD4+IFNγ_IL-17_IL-22+), has recently been
and IL-13 related) responses. Although the identified and found to be up-regulated in several
distinctions between type 1 and type 2 T cells and autoimmune diseases. Th22 cells are also
cytokine profiles in humans are less clear, these significantly increased in ITP patients, and this
categories are helpful in broadly placing common increase is correlated with the observed increased
immune patterns into context. Type 1 reactions are numbers of Th1 and Th17 cells43. Another cytokine,
classically thought to be involved in response to IL-21, is produced by some CD4+ T cells and
intracellular pathogens, and a type 1 response is one natural killer T cells and is capable of up-regulating
which generally promotes pro-inflammatory, cell- both Th17 cells and B cells. In ITP, IL-21
mediated, complement fixing phenotypes. On the expression on T cells is elevated in untreated newly
other hand, type 2 immunity is thought to function diagnosed ITP patients, although circulating IL-21
in the fight against extracellular pathogens, and a is unchanged44.
type 2 response typically elicits an immediate-type Simplistically, the evidence supports a type 1 T-
hypersensitivity response. During an inflammatory cell response with an up-regulated Th17 response in
event, dominance of either a type 1 or 2 profile is ITP. Interestingly, similar patterns of immune
characteristic, because the prevailing dominant type dysregulation can also be seen in other autoimmune
both expands and engages in negative feedback disorders43. These types of responses,
loops to suppress the “other” T-cell types. stereotypically proinflammatory, cell-mediated,
Resolution of an immune inflammatory event is complement-fixing phenotypes, would be expected
then characterized by suppression of the dominant to propagate and enhance the ongoing autoantibody
type phenotype and restoration of the type 1/type 2 mediated platelet immune process. Moreover, the
balance19. increase in Tc17 cells is correlated with skewing of
Therefore, the shifting of the balance of type 1 the CD4:CD8 ratio in ITP44.
and type 2 permits tailoring of the immune response
to the perceived threat. In ITP, type 1/type 2 ratios
are unbalanced (Figure 3). In primary ITP, adult
chronic primary ITP patients have high Th1/Th2
(“helper” CD4+ cells) ratios and high Tc1/Tc2
(“cytotoxic” CD8+cells) ratios39. Furthermore, the
Th1/Th2 ratio imbalance is inversely correlated
with disease severity40, meaning the higher the
Th1/Th2 ratio, the lower the platelet count. ITP
patients also exhibit decreased numbers of
CD4+CD25+ T-regulatory cells (Tregs), which Figure 3. Schematic of shifts in the T-cell balance
function to down-regulate T-cell responses20. Not in ITP.
surprisingly, the degree of decrease in numbers of (Cines DB, Bussel JB, Liebman HA, Luning Prak ET.
Tregs is associated with more severe disease in ITP The ITP syndrome: pathogenic and clinical diversity.
41
. In addition to these type 1/2–specific changes, Blood. 2009; 113(26):6511-6521).
the total CD4:CD8 ratio is also observed to be
diminished in ITP42 and improves with disease T cell tolerance failure
remission39. An autoantigenic stimulus may occur in the
More recently, other subsets of T cells distinct extrathymic periphery that could derive from the
from type 1 and type 2 have also been in implicated host’s tissues in the form of an altered self-antigen
in autoimmune diseases, including ITP. Similar to or from a cross-reactive environmental stimulus
type 1 and type 2 T cells, these T-cell subsets are that could mimic self-antigens. Alternatively high-
defined by their cytokine secretion profiles. T cells, affinity atuoreactive CD4+ Th cells may have
which secrete IL-17, are pro-inflammatory and of escaped thymic selection during T-cell ontogeny
interest in ITP in part due to a large body of and may stimulate the anti-platelet autoantibody
evidence implicating IL-17 in autoimmunity. response45. T cells recognizing peptides generated
Within IL-17–secreting T-cell subsets, Th17 from native GPIIb/IIIa by normal processing
(CD4+) cells are increased in ITP, as are Tc17 pathways are hypothesized to be deleted in the
53
Adly

thymus (negative selection) since GPIIb/IIIa has Reduced Treg activity may also be due to
been shown to be expressed abundantly as early as increased resistance of effector T cells to
the 16th week of intrauterine life on epithelial cells suppression, although we specifically demonstrated
of thymic stoma46. However, some autoreactive T that effector T cells from ITP patient and healthy
cells directed against membrane antigens present on controls were equally inhibited by Tregs from
bone marrow-derived cells and also expressed in healthy controls, arguing against the refractoriness
the thymus may not be eliminated by intrathymic of ITP effector cells to suppression53. Indeed, Tregs
deletion47. They can be detected in the peripheral, from ITP patients were less effective that Tregs
even though they may not cause autoimmune from healthy controls in inhibiting effector T cell
response. Some of these T cells to native GPIIb/IIIa proliferation from either patients or healthy
that escape thymic deletion and exist in periphery, controls, suggesting that reduced Treg activity is
are inactivated by a post-thymic mechanism of due to an intrinsic defect in ITP Tregs53. Failure to
peripheral tolerance48 Thus, even though the maintain immune suppression by Tregs may be
process of autoreactive T cells deletion or rendered responsible for the reported platelet autoantigen-
anergic is not totally efficient, there are several specific T cell proliferative responses and the
peripheral tolerance mechanisms that could proinflammatory phenotype in ITP patients53.
potentially suppress autoreactive T and B cells from Nevertheless, the observed polyclonal Treg
becoming activated (Fig. 4). These include dysregulation fails to explain why immune
activation-induced cell death through expression of autoreactivity in ITP is directed toward platelets
Fas and FasL, inhibitory action of membrane rather than other cell types. Detailed identification
molecules like CTLA-4, presence of a group of and characterization of platelet antigen-specific
suppressor T cells and some cytokines that help Tregs in ITP may help clarify why loss of tolerance
inhibit the activation and proliferation of is toward platelets rather than other tissues54.
autoreactive T cells, such as CD4+CD25+ T cells49.
All these tolerance mechanisms contribute to the Role of B cells
inhibition of autoreactive T cells that exist in Autoreactive B lymphocytes secrete antiplatelet
peripheral tissue and it is their failure that in various antibodies
ways leads the autoreactive T cells to mount an The most commonly occurring autoantibodies
attack against self-antigen. In clinics, immuno- (~75%) in patients with ITP are directed against the
suppressants such as azathioprine, cyclo- platelet surface glycoprotein (gp) complexes gpIIb–
phosphamide and cyclosporine have been used to IIIa and gpIb–IX55.
suppress T cells. Therefore, investigations of these Antibodies against other glycoproteins (Ia–IIa,
aspects in the T cells tolerance not only help IV, and V) have been identified, and multiple
understand the disorder or imbalance of immune platelet antigen specificities can be found in most
system in ITP, but may also help development of patients Although antibodies are primarily of the
novel clinical treatment of ITP by targeting these IgG subtype, IgM and IgA may be found56.
mechanisms50. Platelets are targeted by the attachment of
autoantibodies to their surface gp antigens, bound
Defective Treg compartment in ITP to Fcγ receptors expressed on tissue macrophages
Tregs, as characterized by high level expression of of the reticuloendothelial system and cleared from
the CD25 surface marker and of the transcription the circulation. Complement- induced lysis
factor forkhead box protein 3 (Foxp3) on CD4+ following antibody binding may also play a role57.
cells, suppress proliferation of many immune cell After platelet internalization and degradation,
types including T and B cells, either directly macrophages express platelet epitopes on their
through cell contact or indirectly through secretion surface and secrete cytokines that stimulate
of cytokines, thereby dampening inappropriate initiating CD4+ T-cell clones and clones with
immune activation and autoreactivity51. Possible additional specificities5.
reason for decreased Treg numbers can be due to Unique to patients with ITP, autoreactive CD4+
impaired development, proliferation, survival T cells recognize several distinct epitopes on gpIIb–
and/or stability of Tregs whereas defective Treg IIIa, leading to autoimmune response expansion and
function may be explained by failed cell contact accelerated platelet destruction. The trigger for the
dependent suppression or reduced secretion of initiating autoantibody response is unknown,
cytokines that mediate suppression including IL-10, although autoreactive T helper (Th) cells that
TGF-β or IL-3552. interact with antibody-producing B cells are
required7. Platelet-associated autoantibodies are
54
 Pathophysiology of ITP

detected in 50%– 70% of patients with ITP58, production. It is increasingly clear that cellular
emphasizing the limitations of the currently immune mechanisms play a pivotal role in ITP5.
available assays and/or suggesting that other or The production of antiplatelet antibodies by B
additional mechanisms are involved. cells requires antigen-specific, CD4-positive, T-cell
Assays for antibodies targeting gpIIb–IIIa, gpIb– help (Fig 5). It also is possible that in some ITP
IX, and gpIIa–IIIa may be more specific59, but have cases, cytotoxic T cells play a role in the
limited sensitivity, and the diagnosis remains destruction of platelets. A possible sequence of
dependent on clinical presentation for the most events in ITP is as follows. A trigger, possibly an
part.9 infection or toxin, leads to the formation of
antibodies/ immune complexes that attach to
Altered Bregs in ITP platelets. Antibody-coated platelets then bind to
Similar to the T regulatory compartment, Bregs antigen-presenting cells (macrophages or dendritic
inhibit T cell and monocyte activation and they do cells) through low-affinity Fcg receptors (Fcg
so in part through secretion of anti-inflammatory RIIA/Fcg RIIIA) and are internalized and degraded.
IL-1060, which in turn regulates Th polarization, Activated antigen-presenting cells then expose
pro-inflammatory differentiation of other antigen novel peptides on the cell surface and with
presenting cells (APCs) and autoimmune costimulatory help facilitate the proliferation of
responses61. The alteration that we have detected in platelet antigen-specific, CD4- positive, T-cell
ITP patients is both at the phenotypic and clones. These T-cell clones drive autoantibody
functionality of B cells14. Specifically, frequency of production by platelet antigen-specific B-cell
previously described as Breg population62, clones5. As part of the platelet destructive process
characterized as CD19+CD24hiCD38hi cells, was in ITP, cryptic epitopes from platelet antigens are
decreased in nonsplenectomized ITP patients off exposed, leading to the formation of secondary
treatment. platelet antigen-specific T-cell clones, with
ITP B cells have impaired IL-10 response after stimulation of new platelet antigen-specific B-cell
stimulation and a reduced ability to dampen of clones and broadening of the immune response. The
monocyte activation15. In mouse models, IL-10 autoantibody profile of individual patients who
secreting regulatory B cells promote differentiation have ITP reflects activity of polyclonal autoreactive
of Tregs63 or their recruitment64, indicating that B-cell clones derived by antigen-driven affinity
these two immunoregulatory cell types interact with selection and somatic mutation6.
each other.
Although the ability of human Bregs to control B cell tolerance failure
Treg differentiation has not yet been demonstrated, Besides T cells, B cell tolerance are also important
the possibility remains that altered Bregs in ITP in the pathogenesis of ITP, because they are the
patients contributes to compromised Treg ultimate producer of autoantibodies. Genetic
compartment14. analysis of autoantibodies in idiopathic
This also brings up the question as to how these thrombocytopenic purpura reveals evidence of
two immunoregulatory cell types interact with each clonal expansion and somatic mutation66. In the
other and that there may be a hierarchy amongst germinal-center B cells, the degree of down
these regulatory compartments with Bregs regulation is negatively correlated to the up
controlling Tregs. Moreover, as with the data in regulation of autoantibody IgG response. Central
Tregs, altered Breg activity identified in ITP and for tolerance suppresses the unwanted expansion of
that matter in other disease states in humans62 does autoreactive B cell population. If some B cells
not explain the antigen-specific autoreactivity, but escape this suppression or deletion, peripheral
rather is consistent with a perturbed immune mechanism may also be launched to maintain
reactive state. tolerance and one of the pivotal elements for
maintaining peripheral tolerance is Fc_RIIB.
Autoantibodies Suppress Megakaryopoiesis ITAM-containing FcRs, on the other hand can
Chang and colleagues65 showed that plasma from prime autoimmune diseases and the impairment of
patients with ITP containing autoantibodies against the functional balance between activating and
gp1b and gpIIb–IIIa significantly suppressed inhibitory FcRs leads directly to immune complex-
megakaryopoiesis in vitro. They proposed that mediated autoimmune diseases67. Possible
platelet autoantibodies may affect megakaryocyte mechanisms that result in the activation of
maturation or survival, leading to decreased platelet GPIIb/IIIa-reactive T cells are expression of cryptic
determinants of GPIIb/IIIa and generation of cross-
55
Adly

reactive B cells. The mechanism is thought to be mechanism. In ITP, these antibodies produced by B
the generation of cross-reactive B cells, initially cells recognize self-antigens on platelet and cause
primed by foreign protein serving as a molecular in platelet phagocytosis via the reticuloendothelial
mimic that then bind, process, and resent self- system. In clinical therapy, patients may be
protein. Then B cells would have the ability to responsive to splenectomy70, but treatment with an
efficiently concentrate and present to T cells small immunosuppressant that inhibits T- and B-cell
quantities of determinants that are typically function and cooperation, including azathioprine,
sequestered. Cross-reactive B cells can cyclophosphamide, cyclosporine, mycophenolate
subsequently prime naive autoreactive T cells if mofetil or anti-CD20, may be required. Monoclonal
they express the appropriate costimulatory anti- CD20 antibody causes B-cell depletion, which
molecules68. Autoantibodies can then be abundantly may inhibit T-cell–B-cell interactions50.
produced by B cells after this T–B interactivation.
In principle, autoimmune disorders arise because of Impaired Platelet Production
the failure to eliminate or deactivate self-reactive Although increased platelet destruction clearly
lymphocytes, which is reflected in a deficiency of plays a key role in the pathogenesis of ITP, it is
central and/or peripheral tolerance induction now recognized that impaired platelet production
mechanism69. Central tolerance selection in the also is important in many cases. In adults, as many
thymus may be faulty and allows the release of high as 40% of ITP cases may have reduced platelet
affinity autoreactive T cells. Alternatively, an turnover, reflecting the inhibitory effect of platelet
environmental agent can mimic a self-antigen that autoantibodies on megakaryopoiesis71.
leads to the breakdown of peripheral tolerance

Figure 4. Mechanisms of central and peripheral T cell tolerance.

(1) Fas/FasL pathway induces the activated T cells to apoptosis, (2) inhibitory cell surface molecules help T cell anergy
and (3) regulatory T cells secrete a profile of suppressive cytokines (Zhou B, Zhao H, Yang RC, Han ZC. Multi-
dysfunctional pathophysiology in ITP. Crit Rev Oncol Hematol. 2005; 54(2):107-16)

Studies of platelet kinetics in children who have Summary of multi-dysfunctional

ITP are limited but it is possible that a similar pathophysiology in ITP
situation exists. There also is evidence that platelet The increase in serum cytokine levels and activated
autoantibodies may induce thrombocytopenia by T cells, the alteration of cell communication and the
inhibiting proplatelet formation72. Circulating impaired megakaryocytopoiesis in patients with
thrombopoietin (TPO) levels in patients who have chronic ITP are related to a continual self-antigen-
ITP typically are normal or increased only slightly, stimulated autoimmune response that is caused by
reflecting the normal or only slightly reduced TPO tolerance failure. It is still unclear what causes the
receptor mass in this acquired platelet disorder. In breakdown of central and/or peripheral tolerance to
contrast, TPO levels are high in inherited platelet trigger autoreactive lymphocytes’ response, but
production Disorders73. both environmental and genetic factors are thought

56
 Pathophysiology of ITP

to be crucial. It is apparent that dysfunction in step (e.g., induction of mimic antigen by foreign
multi-steps, particularly in T of cellular immunity, antigen) is not necessarily accompanied by the
play a central role in the final outcome of development of ITP because of the absence of
megakaryocytopoietic suppression and platelet dysfunction of the latter steps. The immune system
destruction. These steps are tightly connected and in human body is extremely delicate and
should not be viewed in isolation or mutually complicated. An error in one step can sometimes be
exclusive from each other. Dysfunction in one step compensated or be restored by other remedy
may be a result of concurrent dysfunction of mechanism. ITP occurs only if multi-step
another. For example, Th1 cells increase in the dysfunctions exist. Circulating antibodies to the
circulation promotes an increase in cytokines IL-2, GPIIb/IIIa complex may represent only part of the
IL-10 and INF-γ, which result in feedback relevant antibody pool therefore reducing platelet
activation and proliferation of Th1 cells and APCs. antigen specific antibody is clearly downstream of
Fig. 6 also shows that dysfunction of one item in a treatment and too insensitive to predict relapse50.

Figure 5. Pathogenesis of epitope spread in ITP.

The factors that initiate autoantibody production are unknown. Most patients have antibodies against several platelet-
surface glycoproteins at the time the disease becomes clinically evident. Here, glycoprotein IIb/IIIa is recognized by
autoantibody (orange, inset), whereas antibodies that recognize the glycoprotein Ib/IX complex have not been generated
at this stage (1). Antibody-coated platelets bind to antigen-presenting cells (macrophages or dendritic cells) through Fcg
receptors and then are internalized and degraded (2). Antigen-presenting cells not only degrade glycoprotein IIb/IIIa
(light blue oval), thereby amplifying the initial immune response, but also may generate cryptic epitopes from other
platelet glycoproteins (light blue cylinder) (3). Activated antigen-presenting cells (4) express these novel peptides on
the cell surface along with costimulatory help (represented in part by the interaction between CD154 and CD40) and the
relevant cytokines that facilitate the proliferation of the initiating CD4-positive T-cell clones (T-cell clone 1) and those
with additional specificities (T-cell clone 2) (5). B-cell immunoglobulin receptors that recognize additional platelet
antigens (B-cell clone 2) thereby also are induced to proliferate and synthesize antiglycoprotein Ib/IX antibodies (green)
in addition to amplifying the production of anti-glycoprotein IIb/IIIa antibodies (orange) by B-cell clone 1 (6) (Cines
DB, Blanchette VS. Immune thrombocytopenic purpura. N Engl J Med 2002;346:995–1008)

57
Adly

Figure 6. Multi-step disorders in ITP.

(1) Self-antigen generation, (2) T, B central and peripheral tolerance failure, (3) activation of T–B cells and (4) effector
phase that lead to platelet clearance. (#) Single dysfunction does not always lead to clinical symptom, for the final stage
of antibody production usually needs 1, 2 and 3 (Zhou B, Zhao H, Yang RC, Han ZC. Multi-dysfunctional
pathophysiology in ITP. Crit Rev Oncol Hematol. 2005; 54(2):107-16).

Conclusion and future perspectives 5. Cines DB, Blanchette VS. Immune

ITP is a complex, chronic, often cell-specific, thrombocytopenic purpura. N Engl J Med
autoimmune disease that is still not fully 2002;346:995–1008.
understood. The improved understanding of the 6. Blanchette V, Bolton-Maggs P. Childhood
innate and adaptive immune systems however is immune thrombocytopenic purpura: diagnosis and
allowing us to understand and appreciate some of management. Hematol Oncol Clin North Am. 2010;
the complex interactions between platelets, the 24(1):249-73.
immune system, and the development of ITP. 7. Kuwana M, Ikeda Y. The role of autoreactive T-cells
Immune-mediated platelet destruction in ITP in the pathogenesis of idiopathic thrombocytopenic
occurs by a complex process involving multiple purpura. Int J Hematol 2005;81:106 –112.
components of the immune system. The initiating 8. Gernsheimer T, Stratton J, Ballem PJ,
event for the dysregulation remains unclear, Slichter SJ. Mechanisms of response to treatment
although recent evidence helps to explain the in autoimmune thrombocytopenic purpura. N Engl J
Med 1989; 320:974 –980.
processes by which the disorder may perpetuate
itself. 9. Gernsheimer T. Chronic idiopathic
thrombocytopenic purpura: mechanisms of
Rational therapy available for ITP awaits a more
pathogenesis. Oncologist. 2009; 14:12–21 .
thorough understanding of the relative contribution
of each item in the different steps in the 10. Roark JH, Bussel JB, Cines DB, Siegel DL.
Genetic analysis of autoantibodies in idiopathic
maintenance of normal immune status and how the
thrombocytopenic purpura reveals evidence of clonal
relative contribution of dysfunction of each could expansion and somatic mutation. Blood. 2002;
be accurately quantified in the treatment of ITP. 100:1388–1398 .
11. Ogawara H, Handa H, Morita K, Hayakawa M,
REFERENCES Kojima J, Amagai H, et al. High Th1/Th2 ratio in
1. George JN, Woolf SH, Raskob GE. ITP: Practice patients with chronic idiopathic thrombocytopenic
guideline developed by explicit methods for the purpura. Eur J Haematol. 2003; 71:283–288 .
American Society of Hematology. Blood 1996; 88:3– 12. Wang JD, Chang TK, Lin HK, Huang FL, Wang
40. CJ, Lee HJ. Reduced expression of transforming
2. Buchanan GR, Adix L. Outcome measures and growth factor-β1 and correlated elevation of
treatment end points other than platelet count in interleukin-17 and interferon-γ in pediatric patients
childhood ITP. Semin Thromb Hemost 2001;27:277– with chronic primary immune thrombocytopenia
285. (ITP). Pediatr Blood Cancer. 2011 Oct;57(4):636-40.
3. Cines DB, Bussel JB, Liebman HA, Luning Prak 13. Li S, Wang L, Zhao C, Li L, Peng J, Hou M. CD8+
ET. The ITP syndrome: pathogenic and clinical T cells suppress autologous megakaryocyte apoptosis
diversity. Blood. 2009; 113(26):6511-6521. in idiopathic thrombocytopenic purpura. Br J
4. Lazarusa AH, Semplea JW, Cines DG. Innate and Haematol. 2007; 139(4):605-11.
adaptive immunity in ITP. Semin Hematol. 2013
January; 50(1): S68–S70.
58
 Pathophysiology of ITP

14. Audia S, Samson M, Guy J, Janikashvili N, 27. Peerschke EIB, Andemariam B, Yin W, Bussel
Fraszczak J, Trad M, et al. Immunologic effects JB. Complement activation on platelets correlates
of rituximab on the human spleen in immune with a decrease in circulating immature platelets in
thrombocytopenia. Blood. 2011 Oct patients with immune thrombocytopenic purpura. Br
20;118(16):4394-400. J Haematol. 2010;148(4):638-645.
15. Li X, Zhong H, Bao W, Boulad N, Evangelista J, 28. Najaoui A1, Bakchoul T, Stoy J, Bein G,
Haider MA, et al. Defective regulatory B-cell Rummel MJ, Santoso S, Sachs UJ. Autoantibody-
compartment in patients with immune mediated complement activation on platelets is a
thrombocytopenia. Blood. 2012 Oct common finding in patients with immune
18;120(16):3318-25. thrombocytopenic purpura (ITP). Eur J Haematol.
16. Cines DB, Liebman H, Stasi R. Pathobiology of 2012 Feb;88(2):167-74.
secondary immune thrombocytopenia. Semin 29. Hed J. Role of complement in immune or idiopathic
Hematol. Jan; 2009 46(1 Suppl 2):S2–14. thrombocytopenic purpura. Acta Paediatrica
17. O’Leary ST, Glanz JM, McClure DL, Akhtar A, Supplementum, 1998; 424:37–40.
Daley MF, Nakasato C, et al. The risk of immune 30. Cooper N, Bussel J. The pathogenesis of immune
thrombocytopenic purpura after vaccination in thrombocytopaenic purpura. Br J Haematol. 2006;
children and adolescents. Pediatrics. Feb; 2012 133(4):364-74.
129(2):248–55 . 31. Andersson PO, Wadenvik H. Chronic idiopathic
18. Shoenfeld Y, Agmon-Levin N. ‘ASIA’ - thrombocytopenic purpura (ITP): molecular
autoimmune/inflammatory syndrome induced by mechanisms and implications for therapy. Expert
adjuvants. J Autoimmun. Feb; 2011 36(1):4–8 . Reviews in Molecular Medicine, 2004; 6: 1–17.
19. Johnsen J. Pathogenesis in immune 32. Andersson PO, Olsson A, Wadenvik H. Reduced
thrombocytopenia: new insights. Hematology Am transforming growth factor-beta1 production by
Soc Hematol Educ Program. 2012;2012:306-12. mononuclear cells from patients with active chronic
20. Neunert C, Lim W, Crowther M. The American idiopathic thrombocytopenic purpura. British Journal
Society of Hematology 2011 evidence-based practice of Haematology 2002; 116:862–867.
guideline for immune thrombocytopenia. Blood. 33. Mouzaki A, Theodoropoulou M, Gianakopoulos
2011;117(16):4190-4207. I, Vlaha V, Kyrtsonis MC, Maniatis A. Expression
21. Kaznelson P. Verschwinden die Hamorrhagischen patterns of Th1 and Th2 cytokine genes in childhood
Diathese beieinem Falle von essentielle idiopathic thrombocytopenic purpura (ITP) at
Thrombopenie (Frank) nach Milz extirpation. Wiener presentation and their modulation by intravenous
Klinische Wochenschrift 1916; 29: 1451–1454. immunoglobulin G (IVIg) treatment: their role in
prognosis. Blood. 2002; 100(5):1774-9.
22. Doan CA, Bouroncle BA & Wiseman BK.
Idiopathic and secondary thrombocytopenic purpura: 34. Panitsas FP, Theodoropoulou M, Kouraklis A,
clinical study and evaluation of 381 cases over a Karakantza M, Theodorou GL, Zoumbos NC, et
period of 28 years. Annals of Internal Medicine, al. Adult chronic idiopathic thrombocytopenic
1960; 53: 861 . purpura (ITP) is the manifestation of a type-1
polarized immune response. Blood, 2004; 103: 2645–
23. Wadenvik H, Jacobsson S, Kutti J, Syrjala M. 2647.
In vitro and in vivo behavior of 111In-labelled
platelets: an experimental study of healthy male 35. Abboud RM, Laver J, Xu F, Weksler B, Bussel
volunteers. Eur J Haematol 1987; 38:415–425 JB. Serum levels of GM-CSF are elevated in patients
with thrombocytopenia. British Journal of
24. Olsson B, Ridell B, Jernås M, Wadenvik H. Haematology 1996; 92:486–488.
Increased number of B-cells in the red pulp of the
spleen in ITP. Ann Hematol. 2012 Feb;91(2):271-7. 36. Chanock S. The etiology of childhood immune
thrombocytopenic purpura: how complex is it?
25. Vianelli N, Galli M, de Vivo A, Intermesoli T, Journal of Pediatric Hematology/Oncology, 2003;
Giannini B, Mazzucconi MG, et al. Efficacy and 25(Suppl. 1), S7–10.
safety of splenectomy in immune thrombocytopenic
purpura: long-term results of 402 cases. 37. Olsson S, Cagnoni F, Dignetti P, Melioli G,
Haematologica. 2005; 90(1):72-7. Canonica GW. Low concentrations of cytokines
produced by allergen-stimulated peripheral blood
26. Kojouri K, Vesely SK, Terrell DR, George JN. mononuclear cells have potent effects on nasal polyp-
Splenectomy for adult patients with idiopathic derived fibroblasts. Clin Exp Immunol. 2003;
thrombocytopenic purpura: a systematic review to 132(2):254-60.
assess long-term platelet count responses, prediction
of response, and surgical complications. Blood. 2004; 38. Harrington WJ, Minnich V, Hollingsworth JW,
104:2623–2634 Moore CV. Demonstration of a thrombocytopenic
factor in the blood of patients with thrombocytopenic
purpura. J Lab Clin Med. 1951; 38: 1–10.

59
Adly

39. Wang T, Zhao H, Ren H, Guo J, Xu M, Yang R, et 52. Buckner JH. Mechanisms of impaired regulation by
al. Type 1 and type 2 T-cell profiles in idiopathic CD4(+)CD25(+)FOXP3(+) regulatory T cells in
thrombocytopenic purpura. Haematologica. 2005; human autoimmune diseases. Nat Rev Immunol.
90(7):914-23. 2010; 10:849–859.
40. Panitsas FP, Theodoropoulou M, Kouraklis A, 53. Yu J, Heck S, Patel V, Levan J, Yu Y, Bussel
Karakantza M, Theodorou GL, Zoumbos NC, et JB, et al. Defective circulating CD25 regulatory T
al. Adult chronic idiopathic thrombocytopenic cells in patients with chronic immune
purpura (ITP) is the manifestation of a type-1 thrombocytopenic purpura. Blood. 2008; 112:1325–
polarized immune response. Blood. 2004; 1328.
103(7):2645-7 . 54. Zhang XL, Peng J, Sun JZ, Liu JJ, Guo CS,
41. Sakakura M, Wada H, Tawara I, Nobori T, Wang ZG, et al. De novo induction of platelet-
Sugiyama T, Sagawa N, et al. Reduced specific CD4(+)CD25(+) regulatory T cells from
Cd4+Cd25+ T cells in patients with idiopathic CD4(+)CD25(−) cells in patients with idiopathic
thrombocytopenic purpura. Thromb Res. thrombocytopenic purpura. Blood. 2009; 113:2568–
2007;120(2):187-93. 2577.
42. Hu Y, Ma DX, Shan NN, Zhu YY, Liu XG, Zhang 55. McMillan R. Autoantibodies and autoantigens in
L, et al. Increased number of Tc17 and correlation chronic immune thrombocytopenic purpura. Semin
with Th17 cells in patients with immune Hematol. 2000;37:239 –248.
thrombocytopenia. PLoS One. 2011; 6(10):e26522 . 56. He R, Reid DM, Jones CE, Shulman NR.
43. Zhu X, Ma D, Zhang J, Peng J, Qu X, Ji C, et al. Spectrum of Ig classes, specificities, and titers of
Elevated interleukin-21 correlated to Th17 and Th1 serum antiglycoproteins in chronic idiopathic
cells in patients with immune thrombocytopenia. J thrombocytopenic purpura. Blood. 1994;83:1024 –
Clin Immunol. 2010;30(2):253-9 . 1032.
44. Hu Y, Li H, Zhang L, Shan B, Xu X, Li H, et al. 57. Kravitz MS, Shoenfeld Y. Thrombocytopenic
Elevated profiles of Th22 cells and correlations with conditions—autoimmunity and hypercoagulability:
Th17 cells in patients with immune Commonalities and differences in ITP, TTP, HIT,
thrombocytopenia. Hum Immunol. 2012; 73(6):629- and APS. Am J Hematol. 2005; 80:232–242.
35 . 58. Kiefel V, Freitag E, Kroll H, Santoso S,
45. Semple JW. Immune pathophysiology of Mueller-Eckhardt C. Platelet autoantibodies (IgG,
autoimmune thrombocytopenic purpura. Blood Rev IgM, IgA) against glycoproteins IIb/IIIa and Ib/IX in
2002;16:9–12. patients with thrombocytopenia. Ann Hematol.
46. Gruel Y, Boizard B, Daffos F, Forestier F, 1996;72:280 –285.
Caen J, Wautier JL. Determination of platelet 59. Fabris F, Scandellari R, Ruzzon E, Randi ML,
antigens and glycoproteins in the human fetus. Blood. Luzzatto G, Girolami A. Platelet-associated
1986;68:488–92. autoantibodies as detected by a solid-phase modified
47. Filion MC, Proulx C, Bradley AJ, Devine DV, antigen capture ELISA test (MACE) are a useful
Sékaly RP, Décary F, et al. Presence in peripheral prognostic factor in idiopathic thrombocytopenic
blood of healthy individuals of autoreactive T cells to purpura. Blood. 2004;103:4562– 4564.
a membrane antigen present on bone marrow-derived 60. Fillatreau S, Sweenie CH, McGeachy MJ, Gray
cells. Blood. 1996 Sep 15;88(6):2144-50. D, Anderton SM. B cells regulate autoimmunity by
48. Lo D, Freedman J, Hesse S, Brinster RL, provision of IL-10. Nat Immunol. 2002; 3:944–950
Sherman L. Peripheral tolerance in transgenic mice: 61. Moulin V, Andris F, Thielemans K, Maliszewski
tolerance to class II MHC and non- MHC transgene C, Urbain J, Moser M. B lymphocytes regulate
antigens. Immunol Rev 1991;122:87–102. dendritic cell (DC) function in vivo: increased
49. Jonuleit H, Schmitt E. The regulatory T cell interleukin 12 production by DCs from B cell-
family: distinct subsets and their interrelations. J deficient mice results in T helper cell type 1
Immunol 2003;171:6323–7. deviation. J Exp Med. 2000; 192:475–482
50. Zhou B, Zhao H, Yang RC, Han ZC. Multi- 62. Blair PA, Noreña LY, Flores-Borja F, Rawlings
dysfunctional pathophysiology in ITP. Crit Rev DJ, Isenberg DA, Ehrenstein MR, et al.
Oncol Hematol. 2005; 54(2):107-16. CD19(+)CD24(hi)CD38(hi) B cells exhibit
regulatory capacity in healthy individuals but are
51. Sakaguchi S, Miyara M, Costantino CM, Hafler functionally impaired in systemic Lupus
DA. FOXP3+ regulatory T cells in the human Erythematosus patients. Immunity. 2010; 32:129–
immune system. Nat. Rev. Immunol. 2010; 10:490– 140.
500.

60
 Pathophysiology of ITP

63. Carter NA, Vasconcellos R, Rosser EC, 68. Roth R, Nakamura T, Mamula MJ. B7
Tulone C, Muñoz-Suano A, Kamanaka M, et al. costimulation and autoantigen specificity enable B
Mice lacking endogenous IL-10-producing regulatory cells to activate autoreactive T cells. J Immunol
B cells develop exacerbated disease and present with 1996;157:2924–31.
an increased frequency of Th1/Th17 but a decrease in 69. Shevach EM. Organ-specific autoimmunity. In: Paul
regulatory T cells. J Immunol. 2011; 186:5569–5579 WE, editor. Fundamental immunology. 4th ed.
64. Amu S, Saunders SP, Kronenberg M, Mangan Philadelphia, PA: Lippincott-Raven Publishers; 1999.
NE, Atzberger A, Fallon PG. Regulatory B cells p. 1089–125.
prevent and reverse allergic airway inflammation via 70. Cines DB, McKenzie SE, Siegel DL. Mechanisms
FoxP3-positive T regulatory cells in a murine model. of action of therapeutics in idiopathic
J Allergy Clin Immunol. 2010; 125:1114–1124. thrombocytopenic purpura. J Pediatr Hematol Oncol.
65. Chang M, Nakagawa PA, Williams SA, Schwartz 2003;25:S52–6.
MR, Imfeld KL, Buzby JS, et al. Immune 71. Louwes H, Zeinali Lathori OA, Vellenga E, de
thrombocytopenic purpura (ITP) plasma and purified Wolf JT. Platelet kinetic studies in patients with
ITP monoclonal autoantibodies inhibit idiopathic thrombocytopenic purpura. Am J Med.
megakaryocytopoiesis in vitro. Blood. 1999;106:430–4.
2003;102:887– 895.
72. Takahashi R, Sekine N, Nakatake T. Influence of
66. Jessica HR, James BB, Douglas BC, Don LS. monoclonal antiplatelet glycoprotein antibodies on in
Genetic analysis of autoantibodies in idiopathic vitro human megakaryocyte colony formation and
thrombocytopenic purpura reveals evidence of clonal proplatelet formation. Blood. 1999;93:1951–8.
expansion and somatic mutation. Blood.
2002;100:1388–98. 73. Cremer M, Schulze H, Linthorst G, et al.
Serum levels of thrombopoietin, IL-11, and IL-6 in
67. Takai T. Roles of Fc receptors in autoimmunity. Nat pediatric thrombocytopenias. Ann Hematol.
Rev Immunol 2002;2:580–92. 1999;78:401–7.

61