Thymocyte selection-associated high mobility box protein regulates T lymphocytes exhaustion in patients with myelodysplastic syndromes by inhibiting PI3K/AKT/mTOR pathway.

Myelodysplastic syndromes (MDS) patients often experience CD8+ T lymphocytes exhaustion, which plays a crucial role in the development of MDS. However, the specific role of thymocyte selection-associated high mobility box protein (TOX) in the CD8+ T lymphocytes exhaustion in MDS patients remains unclear. In this study, we investigated the role of TOX in CD8+ T lymphocytes exhaustion in patients with MDS. The expression of TOX, inhibitory receptors (IRs), and functional molecules in peripheral blood T lymphocytes of MDS patients and normal controls were detected using flow cytometry. Lentiviral transduction was used to create stable TOX-knockdown CD8+ T lymphocytes, and small interfering RNA (si-RNA) was used to knock down TOX in Jurkat cells. The expression of TOX was found to be significantly higher in CD8+ T lymphocytes of MDS patients compared to normal controls. This was associated with upregulated IRs and reduced expression of functional molecules such as Granzyme and Perforin. Myelodysplastic syndromes patients with higher TOX expression had poor clinical indicators and shorter survival. Knockdown of TOX using sh-RNA partially reverses the exhausted phenotype and enhances the lethality of CD8+ T lymphocytes. Moreover, the knockdown of TOX using si-RNA in Jurkat cells improved cell proliferation activity, down-regulated IRs and activated PI3K/AKT/mTOR signaling pathway. TOX promotes the exhaustion of CD8+ T lymphocytes by inhibiting PI3K/AKT/mTOR pathway, and targeted inhibition of TOX could partially restore the effector functions and activity of CD8+ T lymphocytes.

A nomogram model for predicting the efficacy of cyclosporine in patients with pure red cell aplasia.

Pure red cell aplasia (PRCA) is a rare bone marrow disorder characterized by a severe reduction or absence of erythroid precursor cells, without affecting granulocytes and megakaryocytes. Immunosuppressive therapies, particularly cyclosporine, have demonstrated efficacy as a primary treatment. This study aims to develop a predictive model for assessing the efficacy of cyclosporine in acquired PRCA (aPRCA). This retrospective study encompasses newly treated aPRCA patients at the General Hospital of Tianjin Medical University. Diagnosis criteria include severe anemia, and absolute reticulocyte count below 10 × 109/L, with normal white blood cell and platelet counts, and a severe reduction in bone marrow erythroblasts. Cyclosporine therapy was administered, with dose adjustments based on blood concentration. Response to cyclosporine was evaluated according to established criteria. Statistical analysis involved logistic multi-factor regression, generating a predictive model. The study included 112 aPRCA patients with a median age of 63.5 years. Patients presented with severe anemia (median Hb, 56 g/L) and reduced reticulocyte levels. Eighty-six patients had no bone marrow nucleated erythroblasts. Primary PRCA accounted for 62 cases (55.4%), and secondary PRCA accounted for 50 cases (44.6%). Univariate analysis revealed that ferritin, platelet to lymphocyte ratio (PLR), and CD4/CD8 ratio influenced treatment response. Multivariate analysis further supported the predictive value of these factors. A prediction model was constructed using ferritin, PLR, and CD4/CD8 ratio, demonstrating high sensitivity and specificity. The ferritin, PLR, and CD4/CD8-based nomogram showed good predictive ability for aPRCA response to cyclosporine. This model has potential clinical value for individualized diagnosis and treatment of aPRCA patients.

Aponermin or placebo in combination with thalidomide and dexamethasone in the treatment of relapsed or refractory multiple myeloma (CPT-MM301): a randomised, double-blinded, placebo-controlled, phase 3 trial.

BACKGROUND: Aponermin, a circularly permuted tumor necrosis factor-related apoptosis-inducing ligand, is a potential death receptor 4/5-targeted antitumour candidate. Previous phase 1/2 studies have demonstrated the efficacy of aponermin in patients with relapsed or refractory multiple myeloma (RRMM). To confirm the superiority of aponermin plus thalidomide and dexamethasone (aponermin group) over placebo plus thalidomide and dexamethasone (placebo group) in RRMM, a randomized, double-blinded, placebo controlled phase 3 trial was performed. METHODS: Four hundred seventeen patients with RRMM who had previously received at least two regimens were randomly assigned (2:1) to receive aponermin, thalidomide, and dexamethasone or placebo, thalidomide, and dexamethasone. The primary endpoint was progression-free survival (PFS). Key secondary endpoints included overall survival (OS) and overall response rate (ORR). RESULTS: A total of 415 patients received at least one dose of trial treatment (276 vs. 139). The median PFS was 5.5 months in the aponermin group and 3.1 months in the placebo group (hazard ratio, 0.62; 95% confidence interval [CI], 0.49-0.78; P < 0.001). The median OS was 22.4 months for the aponermin group and 16.4 months for the placebo group (hazard ratio, 0.70; 95% CI, 0.55-0.89; P = 0.003). Significantly higher rates of ORR (30.4% vs. 13.7%, P < 0.001) and very good partial response or better (14.1% vs. 2.2%, P < 0.0001) were achieved in the aponermin group than in the placebo group. Treatment with aponermin caused hepatotoxicity in some patients, as indicated by the elevated alanine transaminase, aspartate transaminase, or lactate dehydrogenase levels (52.2% vs. 24.5%, 51.1% vs. 19.4% and 44.9% vs. 21.6%, respectively), mostly grade 1/2, transient and reversible. The main grade 3/4 adverse events included neutropenia, pneumonia and hyperglycemia. The incidence of serious adverse events was similar between the two groups (40.6% vs. 37.4%). There was no evidence that aponermin leads to hematological toxicity, nephrotoxicity, cardiotoxicity, or secondary tumors. CONCLUSIONS: Aponermin plus thalidomide and dexamethasone significantly improved PFS, OS and ORR with manageable side effects in RRMM patients who had received at least two prior therapies. These results support the use of aponermin, thalidomide, and dexamethasone as a treatment option for RRMM patients. TRIAL REGISTRATION: The trial was registered at http://www.chictr.org.cn as ChiCTR-IPR-15006024, 17/11/2014.

High TOX expression on CD8+ T cells in pure red cell aplasia.

Thymocyte selection-associated high-mobility group box protein (TOX) is an important molecule regulating the development and exhaustion of T lymphocytes. Our aim is to investigate the role of TOX in the immune pathogenesis of pure red cell aplasia (PRCA). TOX expression of CD8+ lymphocytes from the peripheral blood of patients with PRCA was detected by flow cytometry. Additionally, the expression of immune checkpoint molecules PD1 and LAG3 and cytotoxic molecules perforin and granzyme B of CD8+ lymphocytes was measured. The quantity of CD4+CD25+CD127low T cells was analyzed. TOX expression on CD8+ T lymphocytes in PRCA patients was significantly increased (40.73 [Formula: see text] 16.03 vs. 28.38 [Formula: see text] 12.20). The expression levels of PD1 and LAG3 on CD8+ T lymphocytes in PCRA patients were significantly higher than those in the control group (34.18 [Formula: see text] 13.26 vs. 21.76 [Formula: see text] 9.22 and 14.17 [Formula: see text] 13.74 vs. 7.24 [Formula: see text] 5.44, respectively). The levels of perforin and granzyme in CD8+ T lymphocytes of PRCA patients were 48.60 [Formula: see text] 19.02 and 46.66 [Formula: see text] 25.49, respectively, which were significantly higher than those of the control group (31.46 [Formula: see text] 7.82 and 16.17 [Formula: see text] 4.84, respectively). The number of CD4+CD25+CD127low Treg cells in PRCA patients was significantly decreased (4.30 [Formula: see text] 1.27 vs. 1.75 [Formula: see text] 1.22). In PRCA patients, CD8+ T cells were activated and exhibited overexpression of TOX, PD1, LAG3, perforin, and granzyme B, while regulatory T cells decreased. These findings suggest that T cell abnormality plays a critical role in the pathogenesis of PRCA.

Cofilin-1 participates in the hyperfunction of myeloid dendritic cells in patients with severe aplastic anaemia.

Cofilin-1 interacts with actin to regulate cell movement. The importance of cofilin-1 in immunity has been established, and its involvement in a number of autoimmune diseases has been confirmed. However, its role in severe aplastic anaemia (SAA) remains elusive. Thus, the aim of the current study was to investigate the role of cofilin-1 in patients with SAA. Flow cytometry, Western blotting and real-time quantitative reverse transcription-polymerase chain reaction were performed to detect the mRNA and protein expression of cofilin-1 in myeloid dendritic cells (mDCs) from patients with SAA. The expression of cofilin-1 was then suppressed via siRNA, and its effects on mDCs and downstream effector T-cell function were evaluated. Cofilin-1 expression was higher in mDCs from patients with SAA and correlated with routine blood and immune indexes. Moreover, cofilin-1 knockdown in mDCs from patients with SAA reduced their phagocytic capacity, migration capacity, and CD86 expression through F-actin remodelling, downregulating the stimulatory capacity of mDCs on CD4+ and CD8+ T lymphocytes. Collectively, these findings indicate that cofilin-1 participates in the hyperfunction of mDCs in patients with SAA and that the downregulation of cofilin-1 in mDCs from patients with SAA could be a novel treatment approach for SAA.

HDAC11 deficiency disrupts oncogene-induced hematopoiesis in myeloproliferative neoplasms.

Protein acetylation is an important contributor to cancer initiation. Histone deacetylase 6 (HDAC6) controls JAK2 translation and protein stability and has been implicated in JAK2-driven diseases best exemplified by myeloproliferative neoplasms (MPNs). By using novel classes of highly selective HDAC inhibitors and genetically deficient mouse models, we discovered that HDAC11 rather than HDAC6 is necessary for the proliferation and survival of oncogenic JAK2-driven MPN cells and patient samples. Notably, HDAC11 is variably expressed in primitive stem cells and is expressed largely upon lineage commitment. Although Hdac11is dispensable for normal homeostatic hematopoietic stem and progenitor cell differentiation based on chimeric bone marrow reconstitution, Hdac11 deficiency significantly reduced the abnormal megakaryocyte population, improved splenic architecture, reduced fibrosis, and increased survival in the MPLW515L-MPN mouse model during primary and secondary transplantation. Therefore, inhibitors of HDAC11 are an attractive therapy for treating patients with MPN. Although JAK2 inhibitor therapy provides substantial clinical benefit in MPN patients, the identification of alternative therapeutic targets is needed to reverse MPN pathogenesis and control malignant hematopoiesis. This study establishes HDAC11 as a unique type of target molecule that has therapeutic potential in MPN.

MicroRNA-21-5p Regulates CD3+T Lymphocytes Through VCL and LTF in Patients with Immune Thrombocytopenia.

BACKGROUND: Immune thrombocytopenia (ITP) is an autoimmune hemorrhagic disease with distinct clinical manifestations such as extensive skin petechiae, mucosal bleeding, and even visceral hemorrhage. In this study, CD3+T lymphocytes from ITP patients were screened for differentially expressed genes. The expression of miR-21 and miR-155 in T lymphocytes of ITP patients were investigated. The downstream target genes of miR-21 and miR-155 were also searched for the correlation between differentially expressed genes of ITP. METHODS: Differential gene screening was performed using the GSE43177 data set in the GEO database, and the expression of miR-21 and miR-155 in T lymphocytes of ITP patients was verified by qPCR. The interaction network of core downstream target genes and ITP differentially expressed genes of miR-21 and miR-155 were constructed with the STRING database, and the associated factors were verified by qPCR. RESULTS: In ITP patients, the expression of CD8+T lymphocytes increased, the expression of CD4+T lymphocytes decreased, and the ratio of CD4+/CD8+T cells decreased. Fourteen genes were differentially expressed in CD3+T lymphocytes, all of which were upregulated, and the expression of S100A8 was increased in ITP patients. The expression of miR-21-5p and miR-155-5p increased in CD3+T lymphocytes of initial ITP patients. The core down-stream target gene VCL of miR-21 was associated with the differentially expressed genes such as LTF, LCN2, and DEFA4 in the interaction network. VCL expression was decreased and LTF expression was increased in ITP patients. CONCLUSIONS: S100A8 plays an important role in the regulation of CD3+T lymphocytes in ITP patients. MiR-21-5p regulates the differentially expressed gene LTF by inhibiting the core downstream target gene VCL and participates in the immune mechanism of T lymphocytes in ITP patients. MiR-155-5p is also involved in the immunoregulatory mechanism of T lymphocytes in ITP patients.

Abnormal expression of histone acetylases in CD8+ T cells of patients with severe aplastic anemia.

INTRODUCTION: We aimed to investigate the balance between the mRNA levels of histone acetyltransferases (HATs) and histone deacetylases (HDACs) in CD8+ T cells of patients with severe aplastic anemia (SAA). METHODS: Twenty untreated SAA patients, 18 remission SAA patients (R-SAA), and 22 normal controls were evaluated. The mRNA expression levels of HATs, HDACs, and IFNG in CD8+ T cells were measured by real-time quantitative reverse transcription polymerase chain reaction. RESULTS: Histone acetylase EP300 and CREBBP mRNA levels were significantly elevated in CD8+ T cells of SAA patients compared with the normal controls (both p < 0.05). No significant differences were observed in HDAC1 and HDAC7 mRNA between SAA patients and the normal controls. There was an obvious positive correlation between IFNG and EP300 (r = 0.5126, p < 0.01), and CREBBP (r = 0.4663, p < 0.05), respectively, in SAA and R-SAA patients. In addition, EP300 and CREBBP mRNA levels were clearly correlated with clinical parameters of peripheral blood and bone marrow in those patients. CONCLUSION: Our findings suggest that EP300 and CREBBP are increased in CD8+ T cells of SAA patients and are correlated with disease severity. The imbalances in HATs and HDACs may play a role in activating CD8+ T cells to promote the immune pathogenesis of SAA.

Identification of potential pathogenic genes for severe aplastic anemia by whole-exome sequencing.

BACKGROUND: Severe aplastic anemia (SAA) is a syndrome of severe bone marrow failure due to hyperfunction of CD8+ T cells. While, the genetic background of SAA is still unknown. In this study, we tried to explore the possible genetic variants in CD8+ T cells of SAA patients. METHODS: We performed whole-exome sequencing (WES) in CD8+ T cells of 4 SAA patients and 7 normal controls. The mutations that existed in SAA but not in NCs were identified as candidate genes. Then, we compared them with genes in the enriched KEGG pathway of differently expressed genes (DEGs) from previous RNA-seq. After analyzing the types of mutations, we identified possible pathogenic genes and validated them by RT-PCR. Finally, we compared them with the autoimmune disease-related genes in DisGeNET database to select the most possible pathogenic genes. RESULTS: We found 95 candidate mutant genes in which, 4 possible pathogenic genes were identified: PRSS1, KCNJ18, PRSS2, and DGKK. RT-PCR results showed that compared with NCs, PRSS1 and KCNJ18 mRNA expression was significantly increased in SAA patients (p < 0.05), PRSS2 was also increased in SAA patients but without statistical difference, and DGKK gene could not be detected by RT-PCR in SAA patients. In addition, PRSS1 was associated with autoimmune diseases from the DisGeNET database. CONCLUSION: The mutations of PRSS1, KCNJ18, PRSS2, and DGKK, especially PRSS1 in CD8+T cells, may be involved in the immune pathogenesis of SAA.

Bone marrow free immune checkpoints as a potential biomarker for differential diagnosis of acquired bone marrow failures.

OBJECTIVE: Clinically, to make a definite diagnosis of aplastic anemia (AA), idiopathic cytopenia of undetermined significance (ICUS) or myelodysplastic syndrome (MDS), they should be distinguished from each other. AA and ICUS have some incidence to transform into MDS. Immunosuppressive therapy (IST) is effective in AA and partial ICUS patients, while other ICUSs are more likely to progress to MDS without response to IST. To date, we neither found a technical method that could easily identify AA from hypoproliferative MDS, nor a simple parameter that could indicate ICUS with a response to IST. Here, we detected the concentration of free immune checkpoints in bone marrow supernatant of AA, ICUS, and MDS patients, analyzed the differences of immune status among these three diseases, to try to find a way to predict the response to IST in ICUSs. METHODS: Seventy-four novel patients were enrolled with newly diagnosed acquired bone marrow failure (including 29 AA patients, 11 ICUS patients, and 34 MDS patients), bone marrow supernatants were collected. Luminex liquid suspension array technology was used to measure the concentrations of 17 immune checkpoints to analyze the differences of immune status among these three diseases. RESULTS: The levels of 17 free immune checkpoints were elevated in MDS and showed a strong correlation with each other, followed by ICUS, and with the weakest in AA. By drawing the ROC curve, we found eight immune checkpoints, including sCD40, sCD86/B7-2, sCTLA-4, sGITR, sHVEM, sPD-1, sTIM-3, and sTLR-2, could easily distinguish AA from hypoproliferative MDS. ICUSs with lower concentrations of these eight free immune checkpoints predicted a better IST response. CONCLUSION: In conclusion, we found that there were notable differences in the immune status of AA, ICUS, and MDS. The concentrations of sCD40, sCD86/B7-2, sCTLA-4, sGITR, sHVEM, sPD-1, sTIM-3, and sTLR-2 could be used to identify AA and hypoproliferative MDS patients, as well as to distinguish ICUS patients who could benefit from IST.

Clinical observation of low-dose combination chemotherapy in refractory/recurrent paroxysmal nocturnal hemoglobinuria patients: A single-center retrospective analysis.

BACKGROUND: We performed a retrospective analysis to investigate the clinical characteristics and therapeutic strategies of 20 refractory/recurrent PNH patients, including the clinical efficacy of chemotherapy treatment, safety, and survival. METHODS: The clinical data of 20 classic PNH patients who were refractory/recurrent or had glucocorticoid dependence in our hospital were analyzed, including clinical manifestations, laboratory examinations, treatment efficacy, and survival. RESULTS: Seventeen patients had a marked improvement in anemia after chemotherapy, 14 patients acquired blood transfusion independence, and the Hb of 3 patients increased to normal levels. Although 6 patients still needed blood transfusion, the transfusion interval was significantly prolonged. The percentages of LDH, TBIL, and RET, which are indicators of hemolysis, were significantly lower than those before chemotherapy. The dosage of adrenal glucocorticoids was reduced by more than half compared with that before chemotherapy. CONCLUSIONS: Chemotherapy can reduce PNH clones, promote normal hematopoiesis, and control hemolytic attack. It is a promising and widely used therapeutic method.

Clinical study on empirical and diagnostic-driven (pre-emptive) therapy of voriconazole in severe aplastic anaemia patients with invasive fungal disease after intensive immunosuppressive therapy.

The aim of this study is to compare the curative effect of empirical with diagnostic-driven (pre-emptive) therapy of voriconazole in severe aplastic anaemia patients (SAAs) with invasive fungal disease (IFD) after intensive immunosuppressive therapy (IST). Patients undergoing voriconazole antifungal therapy were randomized to empirical therapy group and diagnostic-driven therapy group. Empirical therapy group accounted for 48.5% of all cases, and diagnostic-driven therapy group accounted for 51.5%. The morbidity of IFD (probable and proven cases) was slightly increased in diagnostic-driven therapy group compared with empirical therapy group (P > 0.05). The total effective rate was 62.1%. The effective rate in empirical therapy group was 78.1%, which was significantly increased compared with diagnostic-driven therapy group (47.1%) (P < 0.05). This value was especially significant in possible IFD cases (P < 0.05). The efficacy of possible IFD cases in empirical therapy group was the best (84%) followed by the probable and proven cases in empirical therapy group (57.1%). In diagnostic-driven therapy group, the efficacy of possible IFD cases was 50%, and the efficacy of probable and proven cases was only 37.5%. The difference was statistically significant (P < 0.05). Absolute neutrophil count (ANC) is the key anti-infection factor. The efficacy of patients whose ANC ˂ 0.1 × 109/L was 39.28%, which was significantly reduced compared with that of patients whose ANC ≥ 0.1 × 109/L (78.95%) (P < 0.05). This finding was especially obvious in diagnostic-driven therapy group. As empirical therapy is superior to diagnostic-driven therapy, we recommend that empirical therapy should be started for high-risk patients, and efforts should be made to definitively diagnose the disease.

Abnormal Macrophage Polarization in Patients with Myelodysplastic Syndrome.

BACKGROUND: This study is aimed at assessing the subsets of bone marrow macrophages in patients with myelodysplastic syndrome (MDS) and exploring the role of macrophages in the pathogenesis of MDS. METHODS: Thirty-eight newly diagnosed MDS patients were enrolled in the Department of Hematology of General Hospital of Tianjin Medical University from June 2015 to June 2016. Bone marrow monocytes and macrophage subsets (M1/M2) were detected in patients with MDS and normal controls by flow cytometry. M1 macrophages were cultured in vitro, and the expression of IL-1ß and TNF-α mRNA was measured using real-time polymerase chain reaction. RESULTS: Compared with the normal control group, the proportion of bone marrow monocytes was higher (2.11 ± 0.93% vs. 3.66 ± 3.38%), and the mean fluorescence intensity of surface molecule CD14 was lower in the higher-risk (HR) MDS group (639.05 ± 359.78 vs. 458.26 ± 306.72, p < 0.05). The ratio of M2 macrophages to monocytes was higher in patients with HR-MDS (1.82 ± 2.47% vs. 3.93 ± 3.81%, p < 0.05). The ratio of M1 to M2 macrophages was lower in the HR-MDS group (3.50 ± 3.22 vs. 1.80 ± 0.88, p < 0.05). The expression of IL-1ß and TNF-α mRNA in M1 macrophages was significantly lower in the MDS group (p < 0.05). CONCLUSIONS: Patients with MDS had abnormal macrophage polarization, which may be involved in the alteration of bone marrow microenvironments.

Myeloid dendritic cells in severe aplastic anemia patients exhibit stronger phagocytosis.

BACKGROUND: A deeper understanding of the pathogenesis of severe aplastic anemia (SAA) is urgently warranted to achieve better therapeutic effects. The objective of this study was to investigate the phagocytosis of myeloid dendritic cell (mDC) in SAA patients. METHODS: Myeloid dendritic cells were induced in vitro from bone marrow mononuclear cells from 26 SAA patients and 12 normal controls (HCs). The phagocytosis of mDCs was detected by flow cytometry using FITC-Dextran (40KD), and its correlation with the immune status and severity of the disease was analyzed. RESULTS: The phagocytosis of mDC from untreated SAA patients was significantly stronger than that from complete remission group and HC group (p < 0.05). There was no statistical difference between the latter two groups (p > 0.05). The phagocytosis of mDC from SAA patients correlated positively with the concentration of interleukin (IL)-2 (r = 0.389, p < 0.05), and IL-4 (r = 0.556, p < 0.05), negatively with CD4+ /CD8+ ratio (r = -0.421, p < 0.05). It also had negative correlations with the level of hemoglobin (r = -0.393, p < 0.05), white blood cell (r = -0.436, p < 0.05), platelet (r = -0.431, p < 0.05), and reticulocyte (r = -0.447, p < 0.05). The phagocytosis of mDC does not correlate with the response to IST. CONCLUSIONS: The increased phagocytosis of mDC in untreated SAA patients may contribute to abnormal activation of T helper (Th) and subsequent cytotoxic T lymphocyte (CTL) activation in these patients. It may be involved in the immune pathogenesis of SAA.

Immunological characteristics and effect of cyclosporin in patients with immune thrombocytopenia.

OBJECTIVE: Immune thrombocytopenia (ITP) is well-known as an antibody-mediated autoimmune disease, and it is easy to get response but often turns to relapse or refractory. Cyclosporin is a traditional immunosuppressant and had a good effect on ITP patients. In this paper, we retrospectively analyze the immunological characteristics and therapeutic effect of cyclosporin in 220 patients with ITP. METHODS: All newly diagnosed ITP patients in the Department of Hematology, Tianjin Medical University General Hospital from June 2018 to December 2020 were enrolled and divided into four groups according to the expression of autoantibodies and the occurrence of prodromal infection. The basic data and immune indexes of ITP patients in each group were collected. The clinical immunological characteristics of patients in each group and the therapeutic effect of cyclosporin in each group were analyzed. RESULTS: Multi-autoantibody ITP patients were more likely to have low serum albumin and high gamma globulin, and the ratio of albumin to globulin decreased. In addition, the level of IgA and IgG increased and the level of complement C3 and C4 decreased more frequently than those in other groups. The number of CD3+T lymphocytes, especially CD3+CD4+T lymphocytes, decreased in ANA+ITP patients. The number of CD16+CD56+NK cells, pDC/DC ratio, and pDC/mDC ratio were higher than those in other groups. The expression of IL-6 and the proportion of CD19+B lymphocytes increased in two groups of ITP patients with abnormal autoantibodies. The patients of pro-infected group were more likely to suffer from coagulation disorder. After treatment with cyclosporin, the response rate increased and the 3-month relapse rate decreased in all ITP patients, and the therapeutic effect of patients with high megakaryocyte number was significantly higher than that of patients with low megakaryocyte number. The impact factors that influence the effect of glucocorticoid and(or) IVIG were the number of CD3+CD8+T lymphocytes, CD4/CD8 cell ratio, and the number of CD19+B lymphocytes. The independent impact factor of cyclosporin therapeutic response rate was the number of CD3+T lymphocytes. CONCLUSIONS: ITP is a heterogeneous disease, recurrence may occur during or rapidly after treatment. Cyclosporine included treatment can improve the effective rate of ITP and reduce the relapse rate within 3 months. The number of CD3+T lymphocytes was the only impact factor that influence the therapeutic effect of cyclosporin in ITP patients.

CD8+ T cells exhaustion induced by myeloid-derived suppressor cells in myelodysplastic syndromes patients might be through TIM3/Gal-9 pathway.

CD8+ T cells play a central role in antitumour immunity, which often exhibit 'exhaustion' in the setting of malignancy and chronic viral infection due to T cell immunoglobulin and mucin domain 3 (TIM3) and myeloid-derived suppressor cells (MDSCs). Our team previously found that overactive MDSCs and exhausted TIM3+ CD8+ T cells were observed in myelodysplastic syndromes (MDS) patients. However, it is not obvious whether MDSCs suppress CD8+ T cells through TIM3/Gal-9 pathway. Here, Gal-9, as the ligand of TIM3, was overexpressed in MDSCs. The levels of Gal-9 in bone marrow supernatants, serum and culture supernatants of MDSCs from MDS patients were elevated. CD8+ T cells from MDS or normal controls produced less perforin and granzyme B and exhibited increased early apoptosis after co-culture with MDSCs from MDS. Meanwhile, the cytokines produced by CD8+ T cells could be partially restored by TIM3/Gal-9 pathway inhibitors. Furthermore, CD8+ T cells produced less perforin and granzyme B after co-culture with excess exogenous Gal-9, and the function of CD8+ T cells was similarly restored by TIM3/Gal-9 pathway inhibitors. Expression of Notch1, EOMES (associated with perforin and granzyme B secretion), p-mTOR and p-AKT (associated with cell proliferation) was decreased in CD8+ T cells from MDS after co-culture with excess exogenous Gal-9. These suggested that MDSCs might be the donor of Gal-9, and TIM3/Gal-9 pathway might be involved in CD8+ T cells exhaustion in MDS, and that TIM3/Gal-9 pathway inhibitor might be the promising candidate for target therapy of MDS in the future.

Roles of immune responses in the pathogenesis of immunorelated pancytopenia.

Some patients with pancytopenia do not conform to any diagnostic criteria of known haematological or non-haematological diseases; however, they respond well to corticosteroid, high-dose intravenous immunoglobulin and rituximab treatment. This abnormality is termed immunorelated pancytopenia (IRP). Later studies indicated that IRP might be a kind of autoimmune disease in which T helper (Th) type 2 cell function is enhanced, resulting in the hyperfunction of B lymphocytes, which then produce excess autoantibodies that attack the bone marrow (BM) and cause cytopenia. Hypofunction of regulatory T (Treg) cells and enhanced Th17 cell function, an elevated percentage of plasmacytoid dendritic cells (pDCs) and a decreased percentage of natural killer (NK) cells help to promote the process. Moreover, increased expression of a synergistic stimulator of B lymphocytes, CD70 and the reactive overexpression of the BCR inhibitory coreceptor CD22 also support this claim. Candidate autoantigens targeted by autoantibodies on haematopoietic cell membranes have also been reported in IRP. This review is focused on studies that demonstrate the role of immune responses in the pathogenesis of IRP. Current diagnostic criteria and treatments for IRP are also referenced to provide a thorough understanding. Distinguishing IRP from idiopathic cytopenias of undetermined significance (ICUS) and other haematological disorders, for example myelodysplastic syndrome (MDS), aplastic anaemia (AA), paroxysmal nocturnal hemoglobinuria (PNH) and Evans syndrome, may help patients with pancytopenia benefit from proper treatment. Further studies are required to achieve new insight into the pathophysiology of IRP with regard to the immune system, which will be instrumental for the development of novel therapies for inhibiting disease initiation and/or progression.

Dose-Reduced Decitabine Might Bring Benefits for Elderly AML Patients with Complex Karyotype: a Case Report.

BACKGROUND: Elderly patients with acute leukemia have high mutation frequency and are often accompanied by various underlying diseases, so treatment is difficult and mortality is high. METHODS: We report an elderly acute myeloid leukemia patient with a complex karyotype who received a reduced dose of decitabine in induction chemotherapy due to a combination of severe heart disease. RESULTS: After a course of induction chemotherapy, the patient achieved complete cytogenetic remission. Cardiac function did not deteriorate during or after chemotherapy. CONCLUSIONS: Dose-reduced decitabine alone can be effective and safe in the induction chemotherapy for elderly AML patients with complex karyotype.

Effects of Shikonin on the Functions of Myeloid Dendritic Cells in a Mouse Model of Severe Aplastic Anemia.

This study is aimed at investigating the effects of shikonin, a pyruvate kinase M2 (PKM2) inhibitor, on the functions of myeloid dendritic cells (mDCs) in a mouse model of severe aplastic anemia (AA) generated by total body irradiation and lymphocyte infusion. Flow cytometry and qPCR were used to determine the proportions of PKM2+ mDCs and other immune indicators in the AA mice. Glucose consumption level, pyruvate generation level, and ATP content were used to determine the level of glycolytic metabolism in the mDCs. The survival rates of AA mice were evaluated after the administration of shikonin or the immunosuppressive agent cyclosporin A. The AA mice displayed pancytopenia, decreased CD4+/CD8+ cell ratio, increased perforin and granzyme levels in CD8+ cells, increased costimulatory CD80 and CD86 expressions, and inadequate regulatory T cell number. In vivo animal experiments showed that the shikonin-mediated inhibition of the PKM2 expression in mice was associated with high survival rates. In addition, the administration of cyclosporin A or shikonin decreased the expression of cytotoxic molecules and costimulatory CD80 and CD86 on CD8+ cells. Taken together, the results of this study indicated that shikonin could inhibit the activation and proliferation of mDCs as well as the activation of downstream cytotoxic T cells by reducing the PKM2 level in mDCs.

Infection risk in autoimmune hematological disorders with low-dose rituximab treatment.

BACKGROUND: Rituximab has been widely used in many autoimmune diseases. AIM: To evaluate the infection risk of rituximab in autoimmune hematological disorders. METHODS: Retrospectively studied and compared the clinical data of 89 patients in our hospital who used low-dose rituximab (group R) or pulse cyclophosphamide (group C) for their refractory/relapsed autoimmune hematological diseases from January 2011 to January 2017. The kinds of their diseases included autoimmune hemolytic disease (AIHA), Evans syndrome, and idiopathic thrombocytopenic purpura (ITP). All patients chose either rituximab treatment or cyclophosphamide treatment on their own considerations. FINDINGS: The median follow-up time was six months in group R and four months in group C. After treatments, the patients in group R showed higher white blood cell (WBC) count and neutrophil count than group C (P = .020, P = .037). CD20-positive B cells in group R remained at a very low level after rituximab treatment and need about 15 months to return to normal level, which was longer than group C (six months). The incidence of infection in these two groups has no significant difference, which was 34.7% (17/30) in group R and 32.5% (13/28) in group C (P = .976). Tuberculosis infections after rituximab treatment were found in three patients for the first time. CONCLUSION: The G-CSF, nadir WBC count, and IgA level were protective factors of infection during rituximab treatment. Low-dose rituximab therapy in autoimmune hematological diseases does not increase infection risk compared with cyclophosphamide.