Spleen In Immune Thrombocytopenia Research Articles

When immune thrombocytopenia gets on your nerves: β2-adrenergic receptor agonists as a potential treatment?

When immune thrombocytopenia gets on your nerves: β2-adrenergic receptor agonists as a potential treatment?

Secondary spleen in immune thrombocytopenia: Not so accessory after all….

Immune thrombocytopenia (ITP) is an autoimmune bleeding disorder characterized by an isolated thrombocytopenia. The pathophysiology is complex but involves platelet-autoantibodies and/or cytotoxic T cells, with the spleen playing an important regulatory role. Accessory spleen (AcS) may possibly contribute to ITP relapse following splenectomy; however, the microenvironment of AcS has not been directly compared to the main spleen. Pizzi et al. conducted a histological study of adult ITP patients where they compared eight matched AcS to main spleens, and they observed a similar immunological composition in both groups. This supports the possibility of AcS-mediated ITP relapse post splenectomy. Commentary on: Pizzi et al. Accessory spleens recapitulate the immune microenvironment of the main spleen in immune thrombocytopenia. Br J Haematol 2023;202:147-152.

Accessory spleens recapitulate the immune microenvironment of the main spleen in immune thrombocytopenia.

Accessory spleens (AcS) may play a relevant role in immune thrombocytopenia (ITP) and possibly contribute to ITP relapse following splenectomy. Little is known about the immune microenvironment of AcS in ITP. To address this issue, we compared the histological features of eight matched AcS and main spleen (MS) samples, obtained from adult patients with primary ITP. AcS and MS had overlapping immune architectural features and lymphoid composition, suggesting that similar immunologic events occur in AcS and MS of ITP. These findings may have implications for the potential mechanisms of AcS-mediated ITP relapse. Further studies are needed to confirm these results and to dissect spleen immunity in ITP.

Histology of the spleen in immune thrombocytopenia: clinical-pathological characterization and prognostic implications.

Immune thrombocytopenia (ITP) is an acquired disorder, characterized by immune-mediated platelet destruction. The spleen plays a key pathogenic role in ITP and splenectomy is a valuable second-line therapy for this disease. Little is known on ITP spleen histology and response to splenectomy is unpredictable. This study aims to characterize ITP spleen histology and assess possible predictors of splenectomy outcome. A series of 23 ITP spleens were retrospectively assessed for the following histological parameters: density of lymphoid follicles (LFs), marginal zones (MZs), T helper and cytotoxic T cells; presence of reactive germinal centers (GCs); width of perivascular T cell sheaths; and red pulp features. Clinical and histological data were matched with postsplenectomy platelet counts to assess their prognostic relevance. Three histological patterns were documented: a hyperplastic white pulp pattern, a non-activated white pulp pattern (lacking GCs), and a white pulp-depleted pattern. Poor surgical responses were associated with presplenectomy high-dose steroid administration, autoimmune comorbidities and low T follicular helper cell density. The combination of such parameters stratified patients into different splenectomy response groups. The removal of accessory spleens was also associated with better outcome. ITP spleens are histologically heterogeneous and clinical-pathological parameters may help predict the splenectomy outcome.

LncRNA GAS5 inhibits Th17 differentiation and alleviates immune thrombocytopenia via promoting the ubiquitination of STAT3

BackgroundThe increased differentiation of T helper 17 cells (Th17) accelerates the development of immune thrombocytopenia (ITP), which is a common autoimmune disease with limited therapeutic methods. Recent studies have revealed that long non-coding RNAs (lncRNAs) play a critical role in autoimmune diseases, thus this study aims to investigate the effect of lncRNA GAS5 on the differentiation of Th17 cells in ITP. MethodsThe expression of GAS5 in peripheral blood mononuclear cells (PBMCs) of ITP patients and spleen tissues of ITP mice was measured by qRT-PCR. The percentage of Th17 cells in CD4+ cells was measured by flow cytometry. The combination between GAS5 and STAT3 was confirmed by RNA pull-down assay and RNA Binding Protein Immunoprecipitation (RIP). The ubiquitination of STAT3 was detected by ubiquitination assay and the interaction between STAT3 and TRAF6 was measured by Co-Immunoprecipitation (Co-IP). Finally, the effect of GAS5 on Th17 differentiation was investigated in vitro and in vivo using lentivirus (lenti)-GAS5. ResultsGAS5 expression was downregulated both in PBMCs of ITP patients and spleen tissues of ITP mice. Overexpression of GAS5 suppressed Th17 differentiation while had no effect on Treg differentiation in naïve CD4+ cells. RNA pull-down and RNA immunoprecipitation assays confirmed the interaction between GAS5 and STAT3. Further studies showed GAS5 accelerated the degradation of STAT3 via promoting TRAF6-mediated ubiquitination. Overexpressing GAS5 suppressed Th17 differentiation in vitro and alleviated ITP in vivo via reducing STAT3. ConclusionLncRNA GAS5 inhibited Th17 differentiation through promoting the TRAF6-mediated ubiquitination of STAT3, thus relieving ITP.

The spleen in immune thrombocytopenia

The spleen in immune thrombocytopenia

High‐dose dexamethasone or all‐trans‐retinoic acid restores the balance of macrophages towards M2 in immune thrombocytopenia

Background Immune thrombocytopenia (ITP) is an autoimmune disorder. Deficiency of immune tolerance in antigen-presenting cells and cross-communication between antigen-presenting cells and T cells are involved in the pathogenesis of ITP. Macrophages can polarize into proinflammatory M1 or anti-inflammatory M2 phenotypes in response to different environmental stimuli, and have diverse immunologic functions. Objectives To investigate the M1/M2 imbalance in ITP and whether high-dose dexamethasone (HD-DXM) or all-trans-retinoic acid (ATRA) could restore this imbalance. Methods The numbers of M1 and M2 macrophages in the spleens of ITP patients and patients with traumatic spleen rupture were analyzed by immunofluorescence. Monocyte-derived macrophages were cultured and induced with cytokines and drugs. The expression of M1 and M2 markers and functions of M1 and M2 macrophages before and after modulation by HD-DXM or ATRA were evaluated with flow cytometry and ELISA. Results There was preferred M1 polarization in ITP spleens as compared with healthy controls. Monocyte-derived macrophages from ITP patients had increased expression of M1 markers and impaired immunosuppressive functions. Either HD-DXM or ATRA corrected this imbalance by decreasing the expression of M1 markers and increasing the expression of M2 markers. Moreover, HD-DXM-modulated orATRA-modulated macrophages suppressed both CD4+ and CD8+ T-cell proliferation and expanded CD4+ CD49+ LAG3+ type1 T-regulatory cells. HD-DXM or ATRA modulated macrophages to shift the T-cell cytokine profile towards Th2. Treating patients with HD-DXM or ATRA revealed that macrophages induced from responders showed a predominant M2-like phenotype and immunosuppressive function. Conclusions Aberrant macrophage polarization is involved in the pathogenesis of ITP. Either HD-DXM or ATRA is able to correct this imbalance.

The spleen dictates platelet destruction, anti-platelet antibody production, and lymphocyte distribution patterns in a murine model of immune thrombocytopenia.

For many years, splenectomy has been used to treat immune thrombocytopenia (ITP), and this procedure benefits approximately two-thirds of the treated patients. Although splenectomy may raise platelet counts, antibody-coated platelets and cytotoxic T lymphocytes appear to persist or can change over time. To better understand how the spleen may affect anti-platelet immune responses, we used a murine model of ITP demonstrating both antibody-mediated and T lymphocyte-mediated thrombocytopenia. Mice with severe combined immunodeficiency (SCID) were either splenectomized or not and transfused with splenocytes from CD61 (GPIIIa) knockout mice immunized against CD61(+) platelets. Platelet counts and anti-platelet antibody levels were performed weekly. After 4weeks, the mice were sacrificed, and lymphoid organs were harvested and examined by flow cytometry to quantify CD4(+)CD25(+)FoxP3(+) Tregs and conventional cross-presenting XCR1(+) and tolerizing SIRPα+ dendritic cells. The results indicate that compared with control non-splenectomized mice, thrombocytopenia was improved and anti-platelet antibody production was significantly diminished in all splenectomized mice that received immune splenocytes. Splenectomized SCID mice also had a marked reduction in Tregs in the thymus together with an increased proportion of both thymic dendritic cell subsets that correlated with increased platelet counts. Of interest, although splenectomy diminished anti-platelet antibody production and raised platelet counts, marrow megakaryocyte densities were still significantly reduced in mice that received immune splenocytes. These results suggest that the spleen in murine ITP not only is the primary site responsible for platelet destruction, but it also controls, to a significant extent, the antibody response against platelets and the migration patterns of lymphocyte subsets.

The role of Epstein-Barr virus (EBV) and cytomegalovirus (CMV) in immune thrombocytopenia

BackgroundAlthough Epstein-Barr virus (EBV) and cytomegalovirus (CMV) are known to trigger immune thrombocytopenia (ITP), the manifestations of EBV and CMV in ITP spleen tissues continue to be poorly understood.MethodsOur research retrospectively reviewed a total of 42 ITP patients and 20 healthy control cases from the West China Hospital of Sichuan University from 2008 to 2012. Patients' characteristics, preoperative platelet counts, and the expression of EBV and CMV in spleen tissue were analyzed.ResultsNo statistically significant differences were identified for age and gender between the ITP group and the healthy control group. The comparison of these two categories (the ITP group and the control group) showed substantial variations in the expression of EBV and CMV: nine (21.4%) and eight (19%) ITP patients had a positive expression of EBV and CMV, compared with none and one patient in the control group, respectively. In the EBV-ITP group, patients with a positive EBV expression revealed appreciably decreased preoperative platelet count compared with patients with a negative EBV expression. No other statistically significant difference was found in the CMV group.ConclusionWe have demonstrated the presence of both EBV and CMV infections in the spleen tissue of ITP patients. EBV infections were implicated in the reduced platelet counts in ITP.

OP0198 Proliferative Lymphoid Nodules and not Germinal Centers are the Sites of Autoantigen Stimulation in Human Spleen

ObjectivesThis study focused on understanding the cellular mechanisms involved in the breakdown of tolerance and resulting in the production of autoantibodies in human spleen. Therefore, spleens of patients with immune...

Characterization of Th1- and Th2-associated Chemokine Receptor Expression in Spleens of Patients with Immune Thrombocytopenia

In view of the numerous clinical observations and laboratory studies that suggest a critical role for the spleen in immune thrombocytopenia (ITP) pathophysiology, we aimed to characterize Th1-associated chemokine receptors CXCR3 and CCR5 and Th2-associated chemokine receptor CCR3 in spleens of ITP patients and assess the significance of their differential expression in the clinical setting. The histopathology of spleens was observed using hematoxylin-eosin staining (HE), and the positive rate of CXCR3, CCR5 and CCR3 expression in spleens of 24 ITP patients and 12 patients with traumatic splenic rupture as normal controls was detected by immunohistochemistry using the SP method. CXCR3, CCR5 and CCR3 protein expression was analyzed by Western blot and mRNA levels were investigated by real-time polymerase chain reaction (RT-PCR). Reactive hyperplasia could be seen in follicles of the white pulp, the germinal central zone was enlarged and the marginal zone was thickened, the central arteries were thickened and fibrotic, and the density of the capillary vessel was increased in ITP patients. ITP group displayed a higher rate of expression of Th1-associated chemokine receptors CXCR3 and CCR5 (83.3% vs. 75%, 100% vs. 83.3%) but lower rate of expression of Th2-associated chemokine receptor CCR3 (50% vs. 66.7%) compared with the controls (P < 0.05, respectively). Western blot analysis revealed that CXCR3 and CCR5 protein expression was significantly increased in ITP patients while CCR3 was significantly reduced (P < 0.05, respectively). Meanwhile, ITP patients displayed increased mRNA levels of CXCR3 and CCR5 but decreased gene expression of CCR3 (P < 0.05, respectively). The data suggested that the abnormal expression of Th1/Th2 chemokine receptors may participate in splenic immune disorder in patients with ITP. Using corresponding inhibitors may inhibit Th1-dominant expression and mitigate the progress of the disease.

B cell depletion in immune thrombocytopenia reveals splenic long-lived plasma cells

Primary immune thrombocytopenia (ITP) is a disorder caused by autoantibody-mediated platelet destruction and decreased platelet production. Rituximab, a B cell-depleting agent, has become the first-line treatment for ITP; however, patients with refractory disease usually require splenectomy. We identified antibody-secreting cells as the major splenic B cell population that is resistant to rituximab. The phenotype, antibody specificity, and gene expression profile of these cells were characterized and compared to those of antibody-secreting cells from untreated ITP spleens and from healthy tissues. Antiplatelet-specific plasma cells (PC) were detected in the spleens of patients with ITP up to 6 months after rituximab treatment, and the PC population displayed a long-lived program similar to the one of bone marrow PC, thus explaining for most of these patients the absence of response to rituximab and the response to splenectomy. When analyzed by multiplex PCR at the single-cell level, normal splenic PC showed a markedly different gene expression profile, with an intermediate signature, including genes characteristic of both long-lived PC and proliferating plasmablasts. Surprisingly, long-lived PC were not detected in untreated ITP spleens. These results suggest that the milieu generated by B cell depletion promotes the differentiation and settlement of long-lived PC in the spleen.

Splenic proliferative lymphoid nodules distinct from germinal centers are sites of autoantigen stimulation in immune thrombocytopenia

AbstractTo understand more specific abnormalities of humoral autoimmunity, we studied 31 spleens from immune thrombocytopenia (ITP) patients and 36 control spleens. Detailed analysis identified at least 2 different splenic structures accommodating proliferating B cells, classic germinal centers (GCs), and proliferative lymphoid nodules (PLNs). PLNs were characterized by proliferating Ki67+ B cells close to follicular dendritic cells (FDCs) and lacked polarization into dark and light zones. As opposed to cells in GCs, proliferating B cells in PLN lacked expression of Bcl6. In both PLNs and GCs of ITP spleens, the density of T cells was significantly reduced. Both T follicular helper cells (TFH) and regulatory T cells were reduced within PLNs of ITP spleens suggesting a defect of tolerance related to a loss of T-cell control. Within PLNs of ITP, but not controls, abundant platelet glycoprotein (GP) IIb/IIIa autoantigens was found in IgM containing immune complexes tightly bound to FDCs and closely approximated to proliferating B cells. GPIV was found less often, but not in the same PLNs as GPIIb/IIIa. Autoantigens were not found in the GCs of ITP or controls indicating that PLNs are the sites of autoantigen stimulation in ITP potentially related to a lack of control by T cells and/or the present autoantigen.

Immunologic effects of rituximab on the human spleen in immune thrombocytopenia

AbstractImmune thrombocytopenia (ITP) is an autoimmune disease with a complex pathogenesis. As in many B cell–related autoimmune diseases, rituximab (RTX) has been shown to increase platelet counts in some ITP patients. From an immunologic standpoint, the mode of action of RTX and the reasons underlying its limited efficacy have yet to be elucidated. Because splenectomy is a cornerstone treatment of ITP, the immune effect of RTX on this major secondary lymphoid organ was investigated in 18 spleens removed from ITP patients who were treated or not with RTX. Spleens from ITP individuals had follicular hyperplasia consistent with secondary follicles. RTX therapy resulted in complete B-cell depletion in the blood and a significant reduction in splenic B cells, but these patients did not achieve remission. Moreover, whereas the percentage of circulating regulatory T cells (Tregs) was similar to that in controls, splenic Tregs were reduced in ITP patients. Interestingly, the ratio of proinflammatory Th1 cells to suppressive Tregs was increased in the spleens of patients who failed RTX therapy. These results indicate that although B cells are involved in ITP pathogenesis, RTX-induced total B-cell depletion is not correlated with its therapeutic effects, which suggests additional immune-mediated mechanisms of action of this drug.

Increased number of B-cells in the red pulp of the spleen in ITP

Platelets are targeted by autoantibodies and destroyed in the reticuloendothelial system in the spleen, liver and bone marrow in patients with immune thrombocytopenia (ITP). Other mechanisms such as destruction by cytotoxic T-cells and defective production of platelets in the bone marrow also exist. Splenectomy normalizes the platelet count in 70% of ITP patients, however, precious little is known about the spleen in this disease. Our aim was therefore to investigate the splenic morphology and especially the number and localization of splenic leukocytes in patients with ITP and controls and to evaluate factors predicting outcome of splenectomy. Spleen sections from 29 ITP patients and 11 individuals splenectomized due to trauma were analyzed by immunohistochemistry. All except one of the ITP patients had a normalized platelet count 12months after splenectomy and the platelet count was inversely correlated with age. ITP patients had an increased number of B-cells in the red pulp. The number of white pulp B-cells and number of T-cells in both compartments was unchanged. In conclusion, B-cells are increased in the red pulp of the spleen and together with cytotoxic T-cells, helper T-cells and macrophages line the sinusoids enabling the immunological attack on platelets in ITP.

Increased Number of B-Cells In the Red Pulp of the Spleen In ITP

AbstractAbstract 1447 Background.Platelets are targeted by autoantibodies and destroyed in the reticuloendothelial system, e.g. the spleen, in patients with chronic immune thrombocytopenia (ITP). However, other pathophysiologic mechanisms such as platelet destruction by cytotoxic T-cells and defective bone marrow production of platelets also contribute to the thrombocytopenia in ITP. Splenectomy normalizes the platelet count in around 70% of chronic ITP patients, however, precious little is known about the spleen micromorphology in this disease. We examined splenic pathology in patients with ITP, focusing on the intrasplenic distribution of lymphocyte subsets. Patients and Methods.The paraffin-embedded block containing representative material of the splenic tissue from 29 splenectomized chronic ITP patients (15 females and 14 males, mean age 45±21 (SD) years) and 11 controls, splenectomized because of traumatic splenic rupture (2 females and 9 males, mean age 48±19 (SD) years), was used to produce serial sections with a thickness of 5 μm. These sections were stained with conventional haematoxylin-eosin, and immunostained to identify CD3, CD4, CD8, CD20 and CD68 using the Dako Envision System in a Techmate Horizon Autostainer (Glostrup, Denmark). Using micromorphometry the lymphocyte subsets were enumerated in the red and white pulp of these spleens. Results.All ITP patients were preoperatively optimized, using corticosteroids and/or IVIg, and the immediate pre-splenectomy platelet count ranged 5–357 × 109/l.Significantly more T-cells than B-cells were seen in the red pulp, both in ITP and control spleens (p<0.01). Conversely, more B-cells than T-cells were observed in the white pulp. There was no difference in the number of T-cells in the red and white pulp, between ITP patients and controls. However, the number of B-cells was increased in the red pulp of patients with ITP compared with the controls. Furthermore, numerous B-cells and CD8+ (cytotoxic) T-cells were located in the splenic cords, lining the sinusoids in the red pulp, a microenvironment where they readily can interact with the other blood cells, e.g. platelets. Conclusion.Chronic ITP is associated with an increased number of CD20+ B-cells in the red pulp of the spleen. The red pulp of the spleen also provides a microenvironment where close interaction between platelets, CD3+, CD4+, and CD8+ T-cells and B-cells can take place. Trafficking of regulatory and effector cells into this microenvironment might be a potential therapeutic target. Disclosures:No relevant conflicts of interest to declare.

A weapon of mass destruction: the spleen in immune thrombocytopenia

A weapon of mass destruction: the spleen in immune thrombocytopenia