Proinflammatory plasticity towards Th17 paradigm of regulatory T cells consistent with elevated prevalence of TGFBR2 variants in elderly patients with primary immune thrombocytopenia.

BACKGROUND: Primary immune thrombocytopenia (ITP) is characterized for the skewed Th differentiation towards Th1 and Th17 cells as well as the impaired number and function of regulatory T cells (Tregs). Tregs are capable of co-expressing effector Th markers in different inflammatory milieu, which probably indicates Treg dysfunction and incompetence to counter over-activated immune responses. METHODS: Ninety-two primary ITP patients from March 2013 to December 2018 were included, and proinflammatory plasticity in different Treg compartments, age groups, and TGFBR2 variant carrier status were investigated. RESULTS: Patients were categorized into elderly (n = 44) and younger (n = 48) groups according to an age of 50 years at disease onset. The overall remission rate was 82.6% after first-line regimens, including 47.8% complete remission. TGFBR2 variants were found in 7 (7.6%) patients with three V216I and four T340M heterozygote carriers. ITP patients demonstrated elevated co-expression of IL-17 and decreased co-expression of both IFN-γ and IL-13 than health control (all p < 0.01). The elderly group demonstrated elevated prevalence of TGFBR2 variants (p = 0.037) and elevated co-expression of IL-17 (p = 0.017) in Tregs, while female predominance was found in the younger group (p = 0.037). In the elderly group, TGFBR2 variant carriers demonstrated further elevated co-expression of IL-17 (p = 0.023) and decreased co-expression of both IFN-γ (p = 0.039) and IL-13 (p = 0.046) in the aTreg compartment. CONCLUSIONS: Our findings revealed additional aberrations of Treg proinflammatory plasticity in elderly primary ITP patients, and highlighted the potential role of Treg dysfunction and senescence in the pathogenesis and management among these patients.

Coalescence-Induced Droplet Jumping.

When two or more droplets coalesce on a superhydrophobic surface, the merged droplet can jump spontaneously from the surface without requiring any external energy. This phenomenon is defined as coalescence-induced droplet jumping and has received significant attention due to its potential applications in a variety of self-cleaning, anti-icing, antifrosting, and condensation heat-transfer enhancement uses. This article reviews the research and applications of coalescence-induced droplet jumping behavior in recent years, including the influence of droplet parameters on coalescence-induced droplet jumping, such as the droplet size, number, and initial velocity, to name a few. The main structure types and influence mechanism of the superhydrophobic substrates for coalescence-induced droplet jumping are described, and the potential application areas of coalescence-induced droplet jumping are summarized and forecasted.

The abnormal function of CD39+ regulatory T cells could be corrected by high-dose dexamethasone in patients with primary immune thrombocytopenia.

Primary immune thrombocytopenia is an autoimmune disease, characterized with decreased platelet and increased risk of bleeding. Recent studies have shown the reduction and dysfunction of regulatory T (Treg) cells in ITP patients. CD39 is highly expressed on the surface of Treg cells. It degrades ATP to AMP and CD73 dephosphorylates AMP into adenosine. Then adenosine binds with adenosine receptor and suppresses immune response by activating Treg cells and inhibiting the release of inflammatory cytokines from effector T (Teff) cells. Adenosine receptor has several subtypes and adenosine A2A receptor (A2AR) plays a crucial role especially within lymphocytes. The CD39+ Treg cells and the expression of A2AR showed abnormality in some autoimmune disease. But knowledge of CD39+ Treg cells and A2AR which are crucial in the adenosine immunosuppressive pathway is still limited in ITP. Thirty-one adult patients with newly diagnosed ITP were enrolled in this study. CD39 and A2AR expression was measured by flow cytometry and RT-PCR. The function of CD39 was reflected by the change of ATP concentration detected by CellTiter-Glo Luminescent Cell Viability Assay. CD39 expression within CD4+CD25+ Treg cells in ITP patients was decreased compared to normal controls. After high-dose dexamethasone therapy, response (R) group showed increased CD39 expression within Treg cells while non-response (NR) group did not show any difference in contrast to those before treatment. The expression of A2AR in CD4+CD25- Teff and CD4+CD25+ Treg cells was both lower in ITP patients than that of normal controls. After therapy, CD4+CD25- Teff cells had higher A2AR expression while CD4+CD25+ Treg cells did not show any difference in comparison to that before treatment. The enzymatic activity of CD39 was damaged in ITP patients and improved after high-dose dexamethasone therapy. In ITP, there was not only numerical decrease but also impaired enzymatic activity in CD39+ Treg cells. After high-dose dexamethasone treatment, these two defects could be reversed. Our results also suggested that ITP patients had reduced A2AR expression in both CD4+CD25+ Treg cells and CD4+CD25- Teff cells. CD4+CD25- Teff cells had increased A2AR expression after treatment.

The pretreatment platelet count is an independent predictor of tumor progression in patients undergoing transcatheter arterial chemoembolization with hepatitis B virus-related hepatocellular carcinoma.

AIM: To explore the prognostic value of the pretreatment platelet (PLT) count in patients undergoing transcatheter arterial chemoembolization (TACE) with hepatocellular carcinoma (HCC). MATERIALS & METHODS: We prospectively analyzed 317 hepatitis B virus-related HCC patients undergoing TACE. Time to progression (TTP) was selected to evaluate the clinical significance of PLT level in HCC patients. RESULTS: PLT was the only parameter showing statistical significance of all the clinical characteristics between two distinct tumor response groups. After ruling out cirrhosis as a potential major confounding factor, the conclusion was further established. Higher pretreatment PLT level, portal vessel invasion and higher stratification of α-fetoprotein level were independently associated with longer TTP. The prognostic score model combining the three risk factors revealed that higher risk scores might mean shorter TTP. CONCLUSION: The pretreatment PLT level is a potentially useful biomarker to predict the prognostic outcomes in HCC patients undergoing TACE and deserves to be further explored in subsequent works.

Distinct alterations of CD68+CD163+ M2-like macrophages and myeloid-derived suppressor cells in newly diagnosed primary immune thrombocytopenia with or without CR after high-dose dexamethasone treatment.

BACKGROUND: Although impaired myeloid-derived suppressor cells (MDSCs) recently have been studied in immune thrombocytopenia (ITP), another myeloid-derived cell population signified as M2 macrophages has not been investigated properly in ITP patients. In the present study, we intended to determine the features of circulating M2-like macrophages, to examine its relationship with MDSCs, and to explore their prognostic values in ITP. METHODS: Peripheral blood mononuclear cells from healthy controls and primary ITP patients were isolated to test the circulating M2-like macrophages and MDSCs. The circulating M2-like macrophage population defined as CD68+CD163+ and circulating MDSC population as CD11b+CD33+HLA-DR- were determined by flow cytometry. Plasma inflammatory cytokines were measured by multiplex ELISA. RESULTS: The percentages of MDSCs were found to be expanded in newly diagnosed patients of ITP, especially among those of the complete response (CR) group (p < 0.0001). Positive linear correlation was verified between percentages of M2-like macrophages and MDSCs. The same correlation was also determined in the CR group. After treatment, the percentages of M2-like macrophages and MDSCs were both increased significantly in CR group, while those patients among the PR + NR group manifested a significant numeric decrease of MDSCs but only a moderate decrease in M2-like macrophages. MIP-1α/CCL3 was negatively correlated with M2-like macrophages while MCP-1 possessed a positive correlation with M2-like macrophages, eotaxin-1/CCL11 was negatively correlated with MDSCs and interleukin-1ß (IL-1ß) was found to be negatively correlated with both M2-like macrophages and MDSCs. CONCLUSIONS: The present findings indicated critical roles of both circulating M2-like macrophages and MDSCs in ITP. The positive correlation between them might be related to inflammatory factors-mediated bidirectional interactions or partially due to their similar background patterns during differentiation. MIP-1α/CCL3, MCP-1, eotaxin-1/CCL11 and IL-1ß might play a critical role in the expansion of both M2 macrophages and MDSCs population in ITP patients, which deserves further investigation.

MST4 kinase regulates immune thrombocytopenia by phosphorylating STAT1-mediated M1 polarization of macrophages.

Primary immune thrombocytopenia (ITP) is an autoimmune hemorrhagic disorder in which macrophages play a critical role. Mammalian sterile-20-like kinase 4 (MST4), a member of the germinal-center kinase STE20 family, has been demonstrated to be a regulator of inflammation. Whether MST4 participates in the macrophage-dependent inflammation of ITP remains elusive. The expression and function of MST4 in macrophages of ITP patients and THP-1 cells, and of a macrophage-specific Mst4-/- (Mst4ΔM/ΔM) ITP mouse model were determined. Macrophage phagocytic assays, RNA sequencing (RNA-seq) analysis, immunofluorescence analysis, coimmunoprecipitation (co-IP), mass spectrometry (MS), bioinformatics analysis, and phosphoproteomics analysis were performed to reveal the underlying mechanisms. The expression levels of the MST4 gene were elevated in the expanded M1-like macrophages of ITP patients, and this elevated expression of MST4 was restored to basal levels in patients with remission after high-dose dexamethasone treatment. The expression of the MST4 gene was significantly elevated in THP-1-derived M1 macrophages. Silencing of MST4 decreased the expression of M1 macrophage markers and cytokines, and impaired phagocytosis, which could be increased by overexpression of MST4. In a passive ITP mouse model, macrophage-specific depletion of Mst4 reduced the numbers of M1 macrophages in the spleen and peritoneal lavage fluid, attenuated the expression of M1 cytokines, and promoted the predominance of FcγRIIb in splenic macrophages, which resulted in amelioration of thrombocytopenia. Downregulation of MST4 directly inhibited STAT1 phosphorylation, which is essential for M1 polarization of macrophages. Our study elucidates a critical role for MST4 kinase in the pathology of ITP and identifies MST4 kinase as a potential therapeutic target for refractory ITP.

A prognostic model incorporating inflammatory cells and cytokines for newly diagnosed multiple myeloma patients.

Peripheral blood cell counts and cytokines can be used as predictors of multiple myeloma (MM) patients' outcomes. 313 newly diagnosed MM patients treated with novel agents were divided into training and validation cohorts. We selected the common peripheral blood cell counts, including the lymphocyte/monocyte ratio (LMR), neutrophil/lymphocyte ratio (NLR), and platelet/lymphocyte ratio (PLR), systemic inflammation response index (SIRI), and serum cytokines which contained tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), interleukin-2 receptor (IL-2R), interleukin-8 (IL-8), interleukin-6 (IL-6), and interleukin-10 (IL-10) as related variables. The least absolute shrinkage and selection operator (LASSO) regression was conducted to sort the predictor variables in the training cohort, and then the developed nomogram was assessed in the training and validation cohort. Our study showed that SIRI, PLR, and IL-8 were independent prognostic factors for the survival of MM patients. Patients with lower SIRI (≤ 0.87) had superior survival than patients with higher SIRI (> 0.87). Further, according to the LASSO regression, a nomogram embracing LMR (> 3.78), SIRI (> 0.87), PLR (≤ 106.44), and IL-8 was established. The nomogram demonstrated a better correlation with the outcomes of MM patients in the training cohort than International Staging System (ISS) and Revised-International Staging System (R-ISS). The same results were verified in the validation cohort. The nomogram incorporating inflammatory cells and cytokines could be a helpful tool to stratify MM patients in the era of novel agents.

TCR CDR3 Sequencing as a Clue to Elucidate the Landscape of Dysimmunity in Patients with Primary Immune Thrombocytopenia.

Background: Primary immune thrombocytopenia (ITP) is an autoimmune disorder. The existence of autoreactive T cells has long been proposed in ITP. Yet the identification of autoreactive T cells has not been achieved, which is an important step to elucidate the pathogenesis of ITP. Methods: ITP patients' peripheral blood was collected prior to the treatment and one month after initiating dexamethasone treatment per related therapeutic guideline. Serum cytokines were profiled to examine T cell subtypes imbalance using a protein chip. TCR Vß analysis in CD8+T cells of ITP patients, and TCR CDR3 DNA sequencing of CD4+T and CD8+T cells were performed to determine the autoreactive T cells' clones. Results: Cytokine profiling revealed imbalanced distribution of T cells subtypes, which was confirmed by CD4+T and CD8+T cells' oligoclonal expansion of TCR Vß analysis and TCR CDR3 DNA sequencing. VDJ segments were found to be more frequently presented in ITP patients, when compared with health controls. There was an individualized CD4+T cell or CD8+T cell CDR3 sequence in each ITP patient. Conclusions: The present study revealed that T cell clones expanded in ITP patients' peripheral blood, and each clone had an individualized TCR CDR3 sequence. The expanded T cell clones preferred to use some specific VDJ segment. Further studies are warranted to get access to individualized treatment such as Car-T in patients with ITP.

A novel predictive model for distinguishing mediastinal lymphomas from thymic epithelial tumours.

OBJECTIVES: We recently reported a high rate of nontherapeutic thymectomy. Mediastinal lymphomas (MLs) are the malignancies most likely to be confused with thymic epithelial tumours (TETs). This study aimed to establish a predictive model by evaluating clinical variables and positron emission tomography to distinguish those diseases. METHODS: From 2018 to 2021, consecutive patients who were pathologically diagnosed with TETs or MLs were retrospectively reviewed. Univariable and multivariable analyses were used to identify association factors. The Akaike information criterion was used to select variables. A nomogram was developed and validated to differentiate MLs from TETs. RESULTS: A total of 198 patients were included. Compared with TETs, patients with MLs were more likely to be younger with higher metabolic tumour volume (154.1 vs 74.6 cm3), total lesion glycolysis (1388.8 vs 315.2 g/ml cm3), SUVmean (9.2 vs 4.8), SUVpeak (12.9 vs 6.3) and SUVmax (14.8 vs 7.5). A nomogram was established based on the stepwise regression results and the final model containing age and SUVmax had minimal Akaike information criterion value of 72.28. Receiver operating characteristic analyses indicated that the area under the curve of predictive nomogram in differentiating MLs from TETs was 0.842 (95% CI: 0.754-0.907). The internal bootstrap resampling and calibration plots demonstrated good consistence between the prediction and the observation. CONCLUSIONS: Combination of age and SUVmax appears to be a useful tool to differentiate MLs from TETs. The novel predictive model prevents more patients from receiving nontherapeutic thymectomy.

Cytokine Consistency Between Bone Marrow and Peripheral Blood in Patients With Philadelphia-Negative Myeloproliferative Neoplasms.

Background: Inflammation might play a critical role in the pathogenesis and progression of Philadelphia-negative myeloproliferative neoplasms (Ph-MPNs) with elevated inflammatory cytokines in peripheral blood (PB). However, the inflammatory status inside the bone marrow (BM), which is the place of malignancy origin and important microenvironment of neoplasm evolution, has not yet been elucidated. Methods: Inflammatory cytokine profiles in PB and BM of 24 Ph-MPNs patients were measured by a multiplex quantitative inflammation array. Cytokines that correlated between PB and BM were selected and then validated by ELISA in a separate cohort of 52 MPN patients. Furthermore, a panel of cytokines was identified and examined for potential application as non-invasive markers for the diagnosis and prediction of fibrosis progress of MPN subtypes. Results: The levels of G-CSF, I-309, IL-1ß, IL-1ra, IL-12p40, IL-15, IL-16, M-CSF, MIG, PDGF-BB, and TIMP-1 in BM supernatants were significantly higher than those in PB (all p < 0.05). Linear correlations between BM and PB levels were found in 13 cytokines, including BLC, Eotaxin-2, I-309, sICAM-1, IL-15, M-CSF, MIP-1α, MIP-1δ, RANTES, TIMP-1, TIMP-2, sTNFRI, and sTNFRII (all R > 0.4 and p < 0.05). Levels of BLC, Eotaxin-2, M-CSF, and TIMP-1 in PB were significantly different from those in health controls (all p < 0.05). In PB, levels of TIMP-1 and Eotaxin-2 in essential thrombocythemia (ET) group were significantly lower than those in groups of prefibrotic primary myelofibrosis (pre-PMF) [TIMP-1: 685.2 (322.2-1,229) ng/ml vs. 1,369 (1,175-1,497) ng/ml, p = 0.0221; Eotaxin-2: 531.4 (317.9-756.6) pg/ml vs. 942.4 (699.3-1,474) pg/ml, p = 0.0393] and primary myelofibrosis (PMF) [TIMP-1: 685.2 (322.2-1229) ng/ml vs. 1,365 (1,115-1,681) ng/ml, p = 0.0043; Eotaxin-2: 531.4 (317.9-756.6) pg/ml vs. 1,010 (818-1,556) pg/ml, p = 0.0030]. The level of TIMP-1 in myelofibrosis (MF) >1 group was significantly higher than that in MF ≤ 1 group. Conclusion: Abnormal inflammatory status is present in MPN, especially in its BM microenvironment. Consistency between PB and BM levels was found in multiple inflammatory cytokines. Circulating cytokine levels of BLC, M-CSF, Eotaxin-2, and TIMP-1 reflected inflammation inside BM niche, suggesting potential diagnostic value for MPN subtypes and prognostic value for fibrosis progression.

Interleukin (IL)-1 family cytokines could differentiate primary immune thrombocytopenia from systemic lupus erythematosus-associated thrombocytopenia.

BACKGROUND: Primary immune thrombocytopenia (ITP) is an autoimmune-mediated disorder characterized by a decreased platelet count. Systemic lupus erythematosus (SLE) is also an autoimmune disease in which thrombocytopenia is a common hematologic manifestation. Interleukin (IL)-1 family cytokines are major proinflammatory and immunoregulatory mediators. This study aimed to investigate the role of IL-1 cytokines in patients with ITP and SLE and the potential pathophysiologic mechanism to differentiate SLE-associated thrombocytopenia (SLE-TP) from ITP. METHODS: Multiplex cytokine assay and real-time polymerase chain reaction (RT-PCR) were used to measure IL-1 cytokines in 17 newly diagnosed ITP patients, 17 SLE-TP patients, 19 SLE patients without thrombocytopenia (SLE-NTP), and 10 healthy controls. RESULTS: The serum levels of IL-1ß, IL-18, IL-36α, IL-36ß, IL-36γ, and IL-33 were decreased significantly in ITP patients compared with SLE-TP patients, SLE-NTP patients, and healthy controls (P<0.05). While there was no significant difference in the serum level of IL-37 between ITP and SLE-TP patients, there was a positive correlation between the platelet count and IL-37 level in ITP patients. Our data suggested that serum IL-1ß, IL-18, IL-36α, IL-36ß, IL-36γ, IL-33, and IL-37 could be considered biomarkers in the diagnosis of ITP. CONCLUSIONS: Serum IL-1ß, IL-18, IL-36α, IL-36ß, IL-36γ, and IL-33 could be considered biomarkers to differentiate SLE-TP from ITP patients.

Intestinal microbiota dysbiosis play a role in pathogenesis of patients with primary immune thrombocytopenia.

BACKGROUND: The intestinal microbiota is essential for the maintenance of the physiology of immune homeostasis. Dysbiosis has been described in some autoimmune diseases, however its role is still elusive in primary immune thrombocytopenia (ITP), which is one kind of autoimmune diseases. This study aimed to characterize the phylogenetic diversity of the fecal microbiota and its relationship with the platelet activation status in patients with ITP. METHODS: The platelet activation status was assessed by 2 platelet markers, PAC-1 (antibody that recognizes the activated GPIIb/IIIa complex) and CD62p (Platelet surface P-selectin) by flow cytometry. Total DNA was extracted from fecal samples of ITP patients and healthy controls (HC). Sequencing the V4 hypervariable region of bacterial 16S rRNA genes was used to identify the changes in phylogenetic diversity and composition of the intestinal flora. The obtained sequencing reads were assigned to operational taxonomic units (OTUs, 97% sequence identity) and taxonomically classified to assess composition and diversity. RESULTS: The percentage of PAC-1+ platelets in ITP patients was higher than that in control group (p < 0.001), The percentage of CD62p+ and PAC-1+CD62p+ platelets in ITP patients both higher than those in control group (p < 0.001). At the phylum level, eight different phyla were identified in ITP individuals, with a majority of Bacteroidetes (45.96%) and Firmicutes (38.59%), followed by Proteobacteria (11.43%), Fusobacteria(1.29%), and Actinobacteria (1.22%). While in the Healthy volunteers, ten phyla were detected, with a predominance of Firmicutes (50.92%) and Bacteroidetes (34.26%), came before Proteobacteria (13.60%), and Actinobacteria (0.90%). The gut microbiota was skewed in ITP, with an increased proportion of Proteobacteria, Bacteroidetes and Bacteroidetes/Firmicutes ratio, a decreased proportion of Firmicutes compared with HC. Disease specific alterations in diversity was also identified, especially the potential markers (Anaerorhabdus, sutterella, Peptostreptococcaceae, Clostridium_XI and carnobacteriaceae, p < 0.05) for ITP. CONCLUSIONS: The results suggested that the distinct microbiota dysbiosis in ITP characterized by alterations in biodiversity and composition, which could provide insights for diet therapy and fecal microbiota transplantation treatment to cure ITP. There might be somehow compensatory enhancement of platelet activation in ITP patients. And there is associate between platelet activation and intestinal microbiota in patients with ITP.

Circulating CX3CR1+CD163+ M2 monocytes markedly elevated and correlated with cardiac markers in patients with acute myocardial infarction.

BACKGROUND: Vulnerable plaques have been generally recognized to play a role in the pathogenesis of acute myocardial infarction (AMI), however, the role of circulating CX3CR1+CD163+ M2 monocytes has not been studied properly. We aim to evaluate the features of CX3CR1+CD163+ M2 monocytes and its relationship with cardiac specific markers in AMI patients. METHODS: The circulating M2 monocytes were identified in AMI patients (n=35) and healthy controls (HCs, n=10) by flow cytometry using two staining methods: CD68+CD163+ (cytoplasmic staining) and CX3CR1+CD163+ (surface staining). CX3CR1+ monocytes were purified by magnetic cell sorting. The expression level of peroxisome proliferator-activated receptor γ (PPARγ) and arginase-1 (Arg-1) were measured by real-time quantitative PCR and Western Blot in CX3CR1+ monocytes. RESULTS: Circulating M2 monocytes extremely expanded in AMI patients compared with HCs (P<0.01). Positive linear correlation was confirmed between CD68+CD163+ and CX3CR1+CD163+ cell populations in AMI patients (r=0.39, P=0.02). The percentage of circulating CX3CR1+CD163+ M2 monocytes positively correlated with cardiac specific markers (cTNT, CK-MB) and acute phase markers (glucose, hs-CRP) (cTNT, r=0.63, P<0.01, CK-MB, r=0.54, P<0.01, glucose, r=0.62, P<0.01, hs-CRP, r=0.58, P<0.01). CX3CR1+ monocytes in AMI patients expressed higher levels of PPARγ and Arg-1 than those in HCs (P<0.01). CONCLUSIONS: Circulating M2 monocytes increased in AMI patients and positively correlated with the elevation of both cardiac specific and acute phase markers. CX3CR1+CD163+ M2 monocytes might have application value for the early diagnosis of AMI.

The paradoxical prognostic role of 1q21 Gain/Amplification in multiple myeloma: every coin has two sides.

The prognostic value of 1q21 Gain/Amplification (1q21 Gain/Amp) in multiple myeloma (MM) has always been controversial. A total of 419 newly diagnosed MM patients were included in this retrospective study. The positive rate of 1q21 Gain/Amp was 48.6%. MM patients with 1q21 Gain/Amp were characterized as being in more advanced clinical stages and were more likely to be accompanied by del(13q14), t(4;14) or complex karyotypes (CKs) as well as with more severe anemia and worse renal function. In these patients, the percentage of complete remission (CR) or very good partial response (VGPR) was higher, however, in the early treatment period, the probability of progressive disease (PD) was also higher. No significant difference on progression free survival (PFS) and overall survival (OS) was showed between the group of 1q21Amp and 1q21Gain. The prognostic impact of 1q21 Gain/Amp on PFS of MM patients was heterogeneous and was in accordance with the accompanying parameters.

The prediction value of Treg cell subtype alterations for glucocorticoid treatment in newly diagnosed primary immune thrombocytopenia patients.

BACKGROUND: Primary immune thrombocytopenia (ITP) is an autoimmune heterogeneous disorder of which Treg cells are numerically or functionally deficient. It is known that human FoxP3+CD4+ T cells were composed of 3 phenotypically and functionally distinct subpopulations (resting Treg, rTreg; activated Treg, aTreg; and non-suppressive Treg, n-sTreg). The current study was aimed to determine whether these Treg subtypes are altered in ITP patients and the related potential clinical applications. METHOD: Normal control volunteers and newly diagnosed ITP patients were enrolled in the study. The percentage of Treg cells' subtypes in peripheral blood was examined by flow cytometry before and after the glucocorticoid treatment. The IL-10 production by Treg subtypes was also examined. RESULTS: Treg cell subtypes of aTreg increased, rTreg decreased, and n-s Treg increased in newly diagnosed ITP patients' peripheral blood. The IL-10 production by respective Treg subtype didn't change after the treatment, and aTreg cells had the highest IL-10 yield. Patients who gained remission during follow-up had a higher aTreg cells' percentage than those who did not at the disease diagnosis. CONCLUSION: Tregs cell subtypes percentage was altered when ITP occurred. The increased aTreg cells at disease diagnosis might predict a better glucocorticoid treatment efficacy.

Transcription Factor IRF4 Dysfunction Affects the Immunosuppressive Function of Treg Cells in Patients with Primary Immune Thrombocytopenia.

BACKGROUND: Th17/Treg balance skews towards Th17 in ITP patient. IRF4 has been highlighted for its close relationship to the immunosuppressive function of Treg cells and the IL-17 synthesis in CD4+ T cells. This study was aimed at examining the effects of IRF4 to the Th17/Treg cells in patients with ITP. METHODS: Treg and Teff cells were isolated from PBMCs of newly diagnosed ITP patients. The percentages of CD4+CD25hiFoxp3+Treg cells and the CD3+CD4+IL-17+Th17 cells were detected by flow cytometry. After being cultured, the supernatants of Tregs were collected for IL-10 concentration test. The IRF4 levels of Tregs were measured. Teffs were cultured alone or with Tregs for 24 hours. Then the supernatants were collected for IL-17 concentration test. The binding intensity of IRF4 to the gene IL-10 in Treg cells was detected by ChIP-qPCR. Metabolic assays for Teffs and Tregs were performed with Agilent Seahorse XF96 Analyzer. RESULTS: The secretion of IL-10 by Tregs was decreased in ITP patients. The intensity of IRF4 binding to IL-10 DNA of Tregs in patients was higher than that of normal controls and Teffs in ITP patients. The expressions of IRF4 of Tregs in ITP patients were remarkably lower than that of healthy controls. The percentage of Th17 cells in healthy controls was significantly increased after IRF4 mRNA silencing. Abnormal metabolism of Treg and Teff cells was found in ITP patients. CONCLUSION: The skewed ratio of Th17/Treg cells and dysfunction of Treg cells in newly diagnosed ITP patients was at least partly caused by IRF4 dysfunction. The underlying mechanism might be the impact of IRF4 on the metabolism of Treg and Teff cells.

[A clinical comparative study on treatment of severe newly diagnosed immune thrombocytopenia by recombinant human thrombopoietin combined with glucocorticoid].

OBJECTIVE: To evaluate the efficacy and safety of recombinant human thrombopoietin (rhTPO) combined with glucocorticoid in treatment of severe newly diagnosed primary immune thrombocytopenia (ITP). METHODS: From June 2009 to December 2012, 24 male patients and 38 female patients with the diagnosis of severe primary ITP in our hospital were randomized into trial group (31 cases) or control group (31 cases), the median age was 50 years (range: 21-84 years). Trial group was treated with rhTPO combined with glucocorticoid, and control group was treated with glucocorticoid only. RESULTS: At the day 3, 7 and 14 from the beginning of treatment, the average platelet count (APC) in trial group[(35.5±24.9)×109/L, (135.2±94.9)×109/L and (192.0±109.1)×109/L]were significantly higher than that in control group[(24.5±15.6)×109/L, (78.2±121.9)×109/L and (95.8±60.5)×109/L, P=0.022, 0.009 and 0.001, respectively]. There was no significant difference in APC between the two groups at day 28 and 90 after treatment[(147.8±59.1)×109/L vs (105.1±56.9)×109/L, P=0.243; (137.4±52.3)×109/L vs (104.3±59.8)×109/L, P=0.568, respectively]. At the day 7, 14 and 28, the complete response rates in trial group were 61.3%, 87.1% and 80.6%, which were also significantly higher than that in control group (16.1%, 29.0% and 48.3%, P=0.000, 0.000 and 0.004, respectively). The median time to response in trial group was 3 days while in the control group was 5 days; the median duration of complete response in trial group was 76 days while in the control group was 54 days. In trial group, there were 4 cases treated with platelet transfusion, while in control group there were 11 cases, respectively. CONCLUSION: For patients with severe primary ITP, rhTPO combined with glucocorticoid could rapidly increase the platelet count, significantly improve the complete response rate and prolonged the effect with a low incidence of tolerable adverse events compared to single use of glucocorticoid. rhTPO combined with glucocorticoid could be a new therapeutic choice to those patients.