Role In Pathogenesis Of Immune Thrombocytopenia Research Articles

Genetic variants in canonical Wnt signaling pathway associated with pediatric immune thrombocytopenia

AbstractThrough the use of genetic sequencing, molecular variants driving autoimmunity are increasingly identified in patients with chronic and refractory immune cytopenias. With the goal of discovering genetic variants that predispose to pediatric immune thrombocytopenia (ITP) or increase risk for chronic disease, we conducted a genome-wide association study in a large multi-institutional cohort of pediatric patients with ITP. A total of 591 patients were genotyped using an Illumina Global Screening Array BeadChip. Six variants met genome-wide significance in comparison between children with ITP and a cohort of healthy children. One variant in NAV2 was inversely associated with ITP (adjusted odds ratio [aOR], 0.52; P = 3.2 × 10−11). Two other variants in close proximity to NKD1 were also inversely associated with ITP (aOR, 0.43; P = 8.86 × 10−15; aOR, 0.48; P = 1.84 × 10−16). These genes have been linked to the canonical Wnt signaling pathway. No variants met genome-wide significance in comparison of those with ITP that self-resolved in <1 year versus those who developed chronic ITP. This study identifies genetic variants that may contribute to ITP risk and raises a novel pathway with a potential role in ITP pathogenesis.

Recent progress in immune thrombocytopenia pathophysiology and treatment

Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by increased platelet destruction and impaired platelet production from bone marrow megakaryocytes. The details of different platelet destruction mechanisms resulting from differences in autoantibodies and autoantibody-independent direct platelet destruction remain unclear although antiplatelet autoantibodies directed against platelet glycoproteins, such as GPIIb/IIIa and GPIb, play a central role in ITP pathogenesis. ITP is diagnosed by excluding other causes of thrombocytopenia due to the lack of standard tests or biomarkers for its confirmation. Plasma thrombopoietin level measurement and reticulated platelet ratio are useful in distinguishing the cause of thrombocytopenia and will be included in the new "Diagnostic reference guide of adult immune thrombocytopenia." Currently, the treatment of refractory chronic ITP is mainly based on thrombopoietin receptor agonists, but ITP drugs with novel mechanisms of action are actively developed. New therapeutic agents are expected to be selected based on an accurate diagnosis and tailored to the pathophysiology of each case in ITP treatment.

Impact of TNFAIP3 Genetic Polymorphisms on Primary Immune Thrombocytopenia in Egyptian Adults: Case-control Study

BACKGROUND: Immune Thrombocytopenia (ITP) is a common acquired hematological disease. Genetic polymorphisms play an important role in ITP pathogenesis and prognosis. TNF-α-induced protein 3 (TNFAIP3) is a negative regulator of NF-kB in many signaling pathways. Several variants of TNFAIP3 have been associated with various inflammatory autoimmune disorders. AIM: Our study aimed to study the association of TNFAIP3 single nucleotide polymorphisms (SNPs); rs2230926 &amp; rs5029939 with ITP susceptibility, as well ITP prognosis by follow up the cases for 18 months. METHODS: One hundred and ten ITP patients as well 110 matched unrelated normal controls were enrolled in our study. The polymorphisms were assessed by real-time polymerase chain reaction (real time PCR). RESULTS: There were a significant difference between cases and control groups regarding rs2230926 T&gt;G and rs5029939 C&gt;G frequencies with p &lt; 0.05. Linkage disequilibrium (LD) analysis of the two variants revealed that there was a significant LD (p &lt; 0.001). Non-cutaneous bleeding manifestations were observed mainly in the mutant genotypes of rs2230926 and rs5029939. The ITP patients with mutant genotypes of rs5029939 showed more need to use 2nd line immunosuppressive therapy as well the mutant genotypes of rs2230926 showed more steroid dependence and less complete recovery. CONCLUSION: Our data concluded the presence of LD between rs5029939 and rs2230926. The mutant genotypes of both variants were associated with increase the susceptibility to ITP and accompanied by worse clinical manifestations and poor response to the treatment in the adult Egyptian patients.

Predictive Value of High ICAM-1 Level for Poor Treatment Response to Low-Dose Decitabine in Adult Corticosteroid Resistant ITP Patients.

Primary immune thrombocytopenia (ITP) is an autoimmune hemorrhagic disease. Endothelial cell activation/injury has been found in some autoimmune diseases including SLE, systemic sclerosis, and rheumatoid arthritis, but its role in ITP pathogenesis remains unclear. This study attempted to elucidate the correlation between endothelial dysfunction and disease severity of ITP and find related markers to predict response to low-dose decitabine treatment. Compared with healthy volunteers, higher plasma levels of soluble intercellular adhesion molecule-1 (ICAM-1), vascular endothelial growth factor (VEGF), and Angiopoietin-2 were found in adult corticosteroid resistant ITP patients. Notably, ICAM-1 levels were negatively correlated with the platelet count, and positively associated with the bleeding score. Recently, we have reported the efficacy and safety of low-dose decitabine in adult patients with ITP who failed for the first line therapies. Here, we evaluated the correlation of plasma ICAM-1 level with the efficacy of low-dose decitabine therapy for corticosteroid resistant ITP. A total of 29 adult corticosteroid resistant ITP patients who received consecutive treatments of low-dose decitabine were enrolled in this study. Fourteen patients showed response (nine showed complete response and five showed partial response). The levels of ICAM-1 before and after treatment were significantly higher in the non-responsive ITP patients than in the responsive patients. As shown in the multivariable logistic regression model, the odds of developing no-response to low-dose decitabine increased by 36.8% for per 5 ng/ml increase in plasma ICAM-1 level [odds ratio (OR) 1.368, 95% confidence interval (CI): 1.060 to 1.764]. In summary, this was the first study to elucidate the relationship between endothelial dysfunction and corticosteroid resistant ITP and identify the potential predictive value of ICAM-1 level for response to low-dose decitabine.

A Landscape of Transcriptome Sequencing in Childhood Immune Thrombocytopenia(ITP)

Background: Immune thrombocytopenia(ITP) is a complex autoimmune disease characterized by an isolated thrombocytopenia. It has long been known that B cells produce autoantibodies that are directed against various platelet membrane receptors, eventually resulting in platelet destruction by the reticuloendothelial system. T cell has recently been shown to play a critical role in ITP pathogenesis. In addition to platelet destruction, platelet production can be decreased by an immune-mediated megakaryocyte maturation abnormality and abnormal apoptosis. Despite all this, the pathogenesis of childhood ITP has been proven complex, better knowledge of the childhood ITP pathogenesis continues to be a major objective for the near future. And to our knowledge, there is no specialized gene expression study by means of RNA sequencing (RNA-seq) to characterize the whole transcriptome change of childhood ITP. Thus, we performed RNA-Seq of two childhood ITP patients to identify differentially expressed genes and key biological processes involved in childhood ITP.Methods: Mononuclear cells were separated from the bone marrow samples of two childhood ITP patients. Then we used bioinformatics methods to identify statistically differentially expressed genes and performed functional enrichment analysis to characterize the important functions or signal pathways. To investigate the change of whole transcriptome between ITP and control group and identify potential new functions associated with ITP pathogenesis, we performed Gene Set Enrichment Analysis (GSEA) through multiple functional databases including GO, KEGG and Reactome to determine statistically significant functional gene sets.Results: We obtained more than 16 million paired-end reads in two ITP patients. Each sample could well map to reference genome with average 85.31% rate of unique mapping reads. By comparing the differences of transcriptome between ITP and control group, we identified 2197 significantly differentially expressed genes, including 1396 up-regulated genes and 801 down-regulated genes (Figure A). Among these genes, we found 12 genes were not only known disease genes of ITP but also significantly differentially expressed between ITP and control group, such as FOXP3, HES1 and ITGA2B. These differentially expressed genes were involved in GO functions associated with immune, blood coagulation and platelet development, for example, platelet aggregation, platelet degranulation and immune system development (Figure B). Moreover, these genes were also significantly involved in KEGG pathways, such as oxidative phosphorylation and cell cycle pathway.To analyze the function impact of B cells and T cells in ITP patients, we performed GSEA for gene expression profile. The results showed that genes were significantly enriched with B cell and T cell-mediated immunity and the core enrichment genes were clearly up-regulation in ITP patients. In addition, the differentially expressed genes between ITP and control group were enriched in functions related with cytokines (Figure C). By comparing the differential expression genes with known genes encoding cytokines, 9 cytokine genes were significantly differentially expressed between ITP patients and controls, such as CXCL12, BMP3 and IL-24. Further more, we found that the gene expression profile was positively enriched with genes involved in megakaryocyte development and platelet production (Figure D). 5 megakaryopoiesis-associated genes showed significantly abnormal expression, including GATA1, MYB, CXCL12, ANGPT2 and COL1A1. Last but not least, several potential new pathways were also discovered, such as ECM receptor interaction, cell cycle, oxidative phosphorylation, focal adhesion and leukocyte transendothelial migration.Conclusions: Our results characterized the whole transcriptome change of childhood ITP patients and elucidated the genes and pathways consistently aberrant in ITP. These abnormal expressed genes, cytokines and pathways may play important roles in the pathogenesis of childhood ITP patients, and may serve as potential targets of diagnosis or treatment. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Role of circulating myeloid-derived suppressor cells in pathogenesis of immune thrombocytopenia in children and adolescents

Context Regulatory T cells have an immunosuppressive function on T-cell activation. They are involved in pathophysiology and treatment of immune thrombocytopenia (ITP). Circulating myeloid-derived suppressor cells (MDSCs) are involved in immune dysregulation in ITP. Aim To determine the mean level of MDSCs in acute, persistent, and chronic ITP and its effect on treatment modalities and prognosis. Patients and methods A total of 41 patients with ITP were recruited from the pediatric hematology clinic of Ain Shams University. They were classified into acute, persistent, and chronic and were compared with 20 age-matched and sex-matched healthy controls. All patients were subjected to history taking with emphasis on age of presentation, disease duration, and treatment modalities, as well as thorough clinical examination. Mean values of C-reactive protein, alanine aminotransferase, and serum creatinine were collected from the patients’ files. All study participants underwent complete blood count (Coulter) and MDSCs by flow cytometry. Secondary thrombocytopenia was excluded. Results Acute ITP was detected in 29%, 24% had persistent, and 46% had chronic ITP. Their age ranged from 1 to 16 years at the study entry, and 51.2% were male. Active disease was found in 58.5%, whereas 41.4% in remission. No treatment was offered to 53%, whereas 24% of patients were on steroids. MDSCs decreased significantly in patients with ITP compared with the control group (P&lt;0.001), whereas there was no significant difference among the patient groups regarding MDSCs level (P=0.325) or with different treatment modalities. Conclusion Reduced numbers of MDSCs play a role in pathogenesis of ITP. Yet, MDSCs did not differ according to disease duration or treatment modalities.

Gut Microbiome and Metabolome Were Altered and Strongly Associated With Platelet Count in Adult Patients With Primary Immune Thrombocytopenia.

Gut microbiota has been implicated in the pathogenesis of many autoimmune diseases. This is still an area of active research given that the role of gut microbiota on the primary immune thrombocytopenia (ITP) remains unclear. In this study, fecal samples of 30 untreated adult primary ITP patients and 29 healthy controls (HCs) were used to investigate the gut microbial community and metabolite profiles. Our results show that fecal bacteria such as Blautia, Streptococcus, and Lactobacillus are enriched, whereas bacteria such as Bacteroides are depleted in ITP patients. Notably, fecal metabolites such as fatty acyls and glycerophospholipids are enriched and strongly correlate with discrepant gut microbiota. Furthermore, combinations of Weissella and Streptococcus anginosus, or Cer (t18:0/16:0), Cer (d18:1/17:0), and 13-hydroxyoctadecanoic acid could provide good diagnostic markers for ITP. Moreover, a strong negative correlation was found between platelet count and altered gut microbiota such as S. anginosus and gut metabolites such as Cer (t18:0/16:0) in ITP. In conclusion, dysbiosis of both gut microbiota and metabolome develops in ITP patients compared to HCs. Several ITP-altered gut bacteria and metabolites can be diagnostic biomarkers for ITP, and are highly correlated with platelet count, suggesting that they may also play a role in ITP pathogenesis.

Investigation of Dynamic Thiol/Disulfide Homeostasis in Children With Acute Immune Thrombocytopenia.

Oxidative stress may play a role in the pathogenesis of immune thrombocytopenia (ITP), but the role of dynamic thiol/disulfide homeostasis has not been studied. The objective of this study was to assess whether there is a change in thiol/disulfide homeostasis in children with acute ITP. A total of 40 children with acute ITP and 50 healthy age-matched and sex-matched controls were included in this study. Serum total thiol and native thiol levels have been measured with a novel automatic spectrophotometric method. The amount of dynamic disulfide bonds and related ratios were calculated from these values. The average total thiol and native thiol levels of the patient group were found to be significantly lower than those levels of controls (P<0.01). However, intravenous immunoglobulin (IVIG) treatment with 1 g/kg/d prevented these reductions. disulfide level was slightly, but not significantly, depressed in ITP patients, but it recovered following IVIG treatment. We detected no marked changes in disulfide/total thiol, disulfide/native thiol, and native thiol/total thiol ratios between groups. These results are the first to demonstrate that thiol/disulfide homeostasis plays a role in ITP pathogenesis, and IVIG treatment can prevent the reduced thiol levels in children.

Involvement of Interferon-γ + 874A/T Polymorphism in the Pathogenesis of and Therapeutic Response to Immune Thrombocytopenia.

Immune thrombocytopenia (ITP) is an autoimmune disease characterized by symptoms of thrombocytopenia and bleeding due to production of autoantibodies against platelets. Recently, the occurrence of polymorphisms has been identified as one of the main causes of disease onset. To conduct this study, we recruited 140 patients and control individuals with no history of platelet loss. After collection of specimens, the prevalence of interferon-γ polymorphism was evaluated using the allele-specific oligonucleotide-polymerase chain reaction (ASO-PCR) technique and confirmed by sequencing techniques. The results showed that the frequency of the AA genotype was higher in the control group, compared with patients with ITP; however, in the acute and chronic groups, the frequency of the AT genotype was higher than that of the AA genotype. We also discovered that there was no significant correlation between platelet counts before and after treatment, nor in its related parameters with interferon (IFN)-γ polymorphism. rs2430561 does not seem to have any role in ITP pathogenesis and treatment response.

T helper 1/T helper 2-associated chemokine and chemokine receptor expression in immune thrombocytopenia

Background Immune thrombocytopenia (ITP) is an acquired immune disorder. Chemokines have complicated role in different autoimmune disorders including ITP. C-C motif chemokine ligand 2 (CCL2) chemokine represents a T helper (Th)2 polarizing chemokine, whereas C-X-C motif chemokine receptor 3 (CXCR3) and C-C motif chemokine receptor 2 (CCR2) represent chemokine members with Th1 polarization effect on the immune system. ITP is associated with an imbalance in Th1/Th2 ratio and favors Th1 polarization. This study aimed to explore the role of CCL2 and its receptor CCR2 in addition to CXCR3 receptor gene expression in the pathogenesis and severity of ITP.Participants and methods Expression of CCL2, CCR2, and CXCR3 was assayed using real-time quantitative polymerase chain reaction in peripheral blood mononuclear cells of 21 normal healthy participants and 68 patients with ITP: 24 acute cases, 25 chronic responder cases, and 19 chronic non-responder cases.Results Acute ITP group showed a 1.85 median fold change in CCL2 gene expression from the control group. CCR2 and CXCR3 showed a higher median fold change from the control group in acute ITP (7.36 and 5.42, respectively) and in chronic non-responder patients (3.38 and 4.32, respectively), whereas the chronic responder patients showed the least changes (1.63 and 2.35, respectively). There was no significant difference in chemokine or chemokine receptors gene expression between different ITP groups (P>0.05). Statistically significant positive correlations were detected between CCL2 and CCR2 (r=0.453, P<0.001) and between CXCR3 and CCR2 (r=0.583, P<0.001) among patients with ITP.Conclusion CCR2 and CXCR3 but not CCL2 may have a role in ITP pathogenesis. Further studies investigating the role of the complicated chemokine network may help better understanding of ITP pathogenesis.

Alterative Expression Pattern of T Cell Immunosuppressive Receptors in Peripheral Blood of Patients with ITP

Immune Thrombocytopenia (ITP) is an autoimmune disorder characterized by production of auto-antibodies against platelet antigens. Dysregulation of T cells is a major role involved in this disease, while anti-platelet antibodies induce platelets destruction due to an imbalanced immune response. Increasing data showed that abnormal expression of T cell immunosuppressive receptors such as Programmed death 1 (PD-1) and Cytotoxic T lymphocyte antigen-4 (CTLA-4) may relate to autoimmune disease pathogenesis. In this study, we analyzed the expression levels of T cell immunosuppressive receptors including PD-1, PD-L1, CTLA-4, T cell immunoglobulin mucin 3 (Tim-3), lymphocyte activation gene-3 (LAG-3) and B and T lymphocyte attenuator (BTLA) in RNA from peripheral blood mononuclear cells (PBMCs) of ITP patients (18 cases, male: 12, female: 6, median age: 38.5 years, range 24-68 years), PBMCs from 20 healthy individuals were served as control group, by real-time PCR. The results showed that significant lower expression levels of PD-1 (median: 0.0015) and PD-L1 (median: 0.0572) were showed in ITP group in comparison with healthy group (PD-1, median: 0.0117, p < 0.0001; PD-L1, median: 0.5428, p < 0.0001), while the expression levels of Tim-3 (median: 0.1352), LAG-3 (median: 0.1403) and BTLA (median: 0.1403) were significant higher than that from healthy group (Tim-3, median: 0.0751, p < 0.007; Lag-3, median: 0.0155, p < 0.0001; BTLA, median: 0.0315, p < 0.0001). There was not significant different on the CTLA-4 expression level between ITP group (median: 0.0818) and healthy group (median: 0.1667) (p = 0.219). In conclusion, we firstly described the alterative expression pattern of T cell immunosuppressive receptors in ITP, which may have a role in ITP pathogenesis. DisclosuresLi:The National Natural Science Foundation of China (81370605): Research Funding.

Soluble Programmed Death 1 (PD-1) Is Decreased in Patients With Immune Thrombocytopenia (ITP): Potential Involvement of PD-1 Pathway in ITP Immunopathogenesis.

Immune thrombocytopenia (ITP) is an autoimmune disease characterized by dysregulation of T cells. Programmed death (PD) 1 and programmed death 1 ligand 1 (PD-L1) are cosignaling molecules, and the major role of the PD-1 pathway is the inhibition of self-reactive T cells and to protect against autoimmune diseases. We measured levels of serum soluble PD 1 (sPD-1) and serum soluble PD-L1 (sPD-L1) in 67 patients with ITP (24 newly diagnosed ITP [ndITP], 43 chronic ITP [cITP]) and 21 healthy controls (HCs). We determined decreased serum sPD-1 levels both in patients with ndITP and in patients with cITP when compared to HC. Moreover, there was a positive correlation between sPD-1 levels and platelet counts. The sPD-L1 levels were decreased in patients with ndITP when compared to patients with cITP. This is the first study investigating PD-1 signaling pathway in ITP. Decreased sPD-1 levels may have a role in ITP pathogenesis as without the inhibitory regulation of PD-1, sustained activation of T cells may cause inflammatory responses which is the case in ITP.

Preferential splenic CD8+ T-cell activation in rituximab-nonresponder patients with immune thrombocytopenia

AbstractThe pathogenic role of B cells in immune thrombocytopenia (ITP) has justified the therapeutic use of anti-CD20 antibodies such as rituximab (RTX). However, 60% of ITP patients do not respond to RTX. To decipher the mechanisms implicated in the failure of RTX, and because the spleen plays a well-recognized role in ITP pathogenesis, 12 spleens from ITP patients who had been nonresponders to RTX therapy were compared with 11 spleens from RTX-untreated ITP patients and 9 controls. We here demonstrate that in RTX-nonresponder ITP patients, preferential Th1 and Tc1 T lymphocyte polarizations occur, associated with an increase in splenic effector memory CD8+ T-cell frequency. Moreover, in the RTX- nonresponder patient group, the CD8+ T-cell repertoire displays a restricted pattern. In the blood, the phenotype of CD8+ T cells before and after RTX treatment is not modified in responders or nonresponders. Altogether, these results demonstrate for the first time an activation of splenic CD8+ T cells in ITP patients who did not respond to RTX and suggest their involvement in platelet destruction in these patients.

Expression of TIM-1 and TIM-3 in spleen mononuclear cells and their role in Th1 polarization in primary immune thrombocytopenia patients

To explore the expression and clinical significance of T cell immunoglobulin mucin (TIM)-1, TIM-3 and T cell-specific transcription factors T-bet and GATA-3 in spleen mononuclear cells in patients with primary immune thrombocytopenia (ITP). The spleen samples were obtained from 17 active ITP patients and 10 controls with spleen traumatic rupture. By using real-time quantitative polymerase chain reaction, the mRNA expressions of TIM-3, TIM1, T-bet and GATA-3 were studied in all subjects. TIM-3 mRNA levels of active ITP patients were significantly decreased to (29 ± 16)% of that of control, TIM-1 mRNA levels of active ITP patients increased to (3.20 ± 2.18) folds of that of control, but the difference was not significant. The ratio of TIM-1/ TIM-3 was elevated in active ITP patients. T-bet mRNA levels were up-regulated in ITP patients by (2.82 ± 1.57) folds (P<0.05) and the expression of GATA3 was decreased by 14% folds (P<0.05) compared to controls. The ratio of T-bet/GATA3 were significantly elevated in ITP patients. The imbalance between TIM-3 and TIM-1 expression might play an important role in pathogenesis of ITP.