The role of Tim-3/Galectin-9 pathway in T-cell function and prognosis of patients with human papilloma virus-associated cervical carcinoma.

The interaction between Tim-3 on T cell and its ligand, Galectin-9, negatively regulates cellular immune responses. However, the role of Tim-3/Galectin-9 pathway in the immune evasion of cervical cancer remains unknown. This study is to investigate the expression, function, and regulation of Tim-3/Galectin-9 signaling pathway in human papilloma virus (HPV) positive cervical cancer. Flow cytometry showed that Tim-3 expression on T cell and Galectin-9 expression on monocytes in HPV positive cervical cancer patients were significantly higher compared to cervical intraepithelial neoplasia and benign uterine fibroids Tim-3 + CD4+ Th1 cells and Tim-3 + CD8+ T cells in HPV positive cervical cancer patients were significantly reduced after surgery. Serum TGF-ß and IL-10 levels were positively correlated with Tim-3 + Treg cells, while IFN-γ and IL-2 were negatively correlated with Tim-3 + Th1 cells. Additionally, Tim-3 + CD4+ T cells were positively correlated with Galectin-9 + monocytes. Survival curve analysis showed that Tim-3 + CD4+ T cells were negatively correlated with patient survival, and closely related to FIGO stage, degree of differentiation, and lymph node metastasis of HPV positive cervical cancer. In vitro experiments showed that by blocking the Tim-3/Galectin-9 pathway, the proliferation of T cells and their ability to express IFN-γ, IL-2, perforin, and granzyme B was significantly restored. In conclusion, high levels of Tim-3 and Galectin-9 in HPV positive cervical cancer patients play roles in the progression of disease by promoting Treg cells to inhibit the cytotoxic function of Th1 and CD8+ T cells. Tim-3/Galectin-9 may serve as a new immunotherapy target for patients with HPV positive cervical cancer.

Activated Galectin-9/Tim3 promotes Treg and suppresses Th1 effector function in chronic lymphocytic leukemia.

Tim-3 is a negative immunoregulator in anti-tumor response, but its mechanism in chronic lymphocytic leukemia (CLL) is not yet clear. The aim of this study was to understand the role of Galectin-9/Tim-3 signaling pathway in the regulation of CD4+ T cell subsets in CLL patients. Flow cytometry results showed that the number of Treg cells obviously increased, and there was a significant Treg/Th17 imbalance in CLL patients. In addition, Tim-3 overexpressed on the surface of Th1 and Treg cells in CLL patients. The levels of Galectin-9 and IL-10 were significantly elevated in patients of CLL, especially in stages of Binet B, and C. However, IFN-γ decreased. Moreover, Galectin-9 in CLL patients was positively correlated with the number of Tim-3+ Treg cells and the level of IL-10. Interestingly, when the Tim-3/Galectin-9 pathway was blocked in vitro, the level of IL-10 in the culture supernatant of CD4+ T was significantly reduced, while the levels of IFN-γ and TNF-α were increased. After co-culture with activated Th1 cells, the apoptosis of CLL cells was significantly increased, and this effect was reversed after treatment with Tim-3+ Tregs. In summary, Galectin-9/Tim-3 are elevated in CLL and associated with disease progression. By the negative regulation of CD4+ T cells, activated Galectin-9/Tim-3 suppresses Th1 effector function and also promotes Treg to be involved in immune escape of CLL. This pathway might become the potential target of immunotherapy in CLL patients.

Tim-3/Galectin-9 signaling pathway is involved in the cytokine changes in mice with alveolar echinococcosis.

BACKGROUND: Tim-3/Galectin-9 is involved in the immune escape of many pathogens. However, the role of Tim-3/Galectin-9 in persistent infection of Echinococcus multilocularis (Em), which is related to immune escape, is still unclear. OBJECTIVE: To investigate the role of Tim-3/Galectin-9 and related cytokines in mice with persistent infection of Em. METHODS: Em infection model was established by injecting the protoscoleces. Serum was collected at days 2, 8, 30, 60, 90, 180 and 270 after infection. Lymphocytes were isolated from liver tissue samples with Ficoll. Tim-3 + CD4 + T percentage was analyzed by flow cytometry. CD4 + T cells were isolated from liver tissues of Em infected mice and cultured in vitro. The mRNA levels of Tim-3, Galectin-9, IFN-γ and IL-4 were detected by qRT-PCR. Cytokine levels in serum and culture supernatant (IFN-γ and IL-4) were analyzed by cytometric bead array. RESULTS: The expression of Tim-3 and Galectin-9 mRNA significantly increased after 30 days of infection, reached peak on day 90, and then decreased slightly on days 180-270. The expression of IFN-γ mRNA, increased on day 2 and 8 after infection, slightly decreased on days 30-60, and obvious decreased on days 90-270, but were still higher than those of the control group. The expression of IL-4 mRNA gradually increased along with the time of infection. In serum of Em infected mice, level of IFN-γ peaked at day 30 and then gradually decreased; whereas IL-4 level peaked at day 90 and then gradually decreased. In vitro experiment found that Tim-3/Galectin-9 directly caused the changes in the levels of IFN-γ and IL-4. CONCLUSIONS: Tim-3/Galectin-9 signaling pathway may be involved in the development of persistent infection of Em by regulating the production of Th1 and Th2 cytokines.

Role of Regulatory B Cells in the Progression of Cervical Cancer.

This study is to investigate the role of regulatory B (Breg) cells in cervical cancer. In total, 70 cases of cervical cancer, 52 cases of cervical intraepithelial neoplasia (CIN), and 40 normal controls were enrolled. The percentage of Breg cells was detected by flow cytometry. Serum levels of IL-10 were measured by ELISA. The correlation between Breg cells and the clinical characterizations of cervical cancer was analyzed. The inhibition effect of Breg cells on CD8+ T cells was tested by blocking IL-10 in vitro. The percentage of CD19+CD5+CD1d+ Breg cells and the level of IL-10 of patients with cervical cancer or CIN were significantly higher than those in the control group (P < 0.05). And the postoperative levels of Breg cells and IL-10 were significantly lower than the preoperative levels (P < 0.05). Breg cells and the IL-10 level were positively correlated in cervical cancer patients (r = 0.516). In addition, the Breg cell percentage was closely related to the FIGO stages, lymph node metastasis, tumor differentiation, HPV infection, and the tumor metastasis of cervical cancer (P < 0.05). The Breg cell percentage was negatively correlated with CD8+ T cells of cervical cancer patients (r = -0.669). The level of IL-10 in the culture supernatant of Bregs treated with CpG was significantly higher than that of non-Bregs (P < 0.05). After coculture with Bregs, the quantity of CD8+ T cells to secrete perforin and Granzyme B was significantly decreased, and this effect was reversed after blocking IL-10 by a specific antibody. Breg cells are elevated in cervical cancer and associated with disease progression and metastasis. Moreover, they can inhibit the cytotoxicity of CD8+ T cells.

Photodegradation of fluazaindolizine in aqueous solution with graphitic carbon nitride nanosheets under simulated sunlight illumination.

The photodegradation of fluazaindolizine (FZDL) under simulated sunlight irradiation was accelerated by the catalysis of graphitic carbon nitride (g-C3N4). Under optimum conditions, such as 5 mg of amount and dispersion, the photodegradation half-life was dramatically enhanced to 2.7 h. More importantly, the pathway of degradation by g-C3N4 was different from both direct photolysis and the catalysis by titanium oxide, with particular negative ions of m/z 221 and 195, corresponding to the cleavage of sulfamide bond and the ring opening of imidazole, respectively. In addition, hydroxyl and superoxide radicals played important roles in photodegradation. The results enriched not only the study of FZDL photodegradation but also the application of g-C3N4. It also suggested the possibility of the water purification by photodegradation for pesticide removal in real life.

Residue behaviours, dissipation kinetics and dietary risk assessment of pyaclostrobin, cyazofamid and its metabolite in grape.

BACKGROUND: Grape is an important fruit consumed either fresh or processed, therefore, fungicide misuse of grape has become an issue of global food safety and human health. Pyraclostrobin, and cyazofamid have been applied to grape frequently. RESULTS: Here a simple QuEChERS (quick, easy, cheap, effective, rugged, and safe) liquid chromatography mass spectrometry technique has been developed and validated for the determination of pyraclostrobin, cyazofamid and its metabolite CCIM in open field grape samples. The recoveries of these three in the range of 0.01 to 5 mg kg-1 (n = 5) ranged from 73.1% to 97.9%. The relative standard deviations (RSDs) were below 12% for all cases. The limits of quantitation of each analyte was 0.005 mg kg-1 , which was lower than maximum residue limits of not only pyraclostrobin but also cyazofamid. Not only dissipation kinetics but also residue determination was obtained in grape for those three pesticides. Furthermore, their half-lives in grapes were 10.7-30.1 days, recommending the pre-harvest intervals for these three of 14 days. The calculated hazard quotient and acute hazard index lower than 100% illustrated the safety of intake of grape for the Chinese population for not only long-term but also short-term dietary risk assessment. CONSLUSIONS: The less than 30 day half-life illustrated that pyraclostrobin and cyazofamid could degrade relatively easily in the environment. The long-term and short-term dietary risk assessment also illustrated the intake safety of these three. Thus, a 14 day pre-harvest interval was safe and recommended. The results of this study contributed to environmental protection, food safety and human health. © 2019 Society of Chemical Industry.

Elevated Expression of Programmed Death-1 and Programmed Death Ligand-1 Negatively Regulates Immune Response against Cervical Cancer Cells.

The present study is to measure the expression of programmed death-1 (PD-1) and programmed death ligand-1 (PD-L1), as well as its clinical significance in cervical cancer patients. Our results showed that different T cell subsets in patients with cervical cancer had high expression of PD-1, and DCs had high expression of PD-L1. High expression of PD-1 on Treg cells in cervical cancer patients facilitated the production of TGF-ß and IL-10 but inhibited the production of IFN-γ. Cervical cancer elevated the expression of PD-1 and PD-L1 in mRNA level. PD-1 expression in peripheral blood of cervical cancer patients was related with tumor differentiation, lymph node metastasis, and invasiveness. PD-1/PD-L1 pathway inhibited lymphocyte proliferation but enhanced the secretion of IL-10 and TGF-ß in vitro. In summary, our findings demonstrate that elevated levels of PD-1/PD-L1, TGF-ß, and IL-10 in peripheral blood of cervical cancer patients may negatively regulate immune response against cervical cancer cells and contribute to the progression of cervical cancer. Therefore, PD-1/PD-L1 pathway may become an immunotherapy target in the future.

CCR7(lo)PD-1(hi) CXCR5(+) CD4(+) T cells are positively correlated with levels of IL-21 in active and transitional cystic echinococcosis patients.

BACKGROUND: In our study, we investigated whether circulating T follicular helper (Tfh) and the related cytokines are involved in human cystic echinococcosis (CE). METHODS: A total of 64 patients with CE and 30 healthy controls were enrolled in this study. Percentages of CCR7(lo)PD-1(hi) cells within CXCR5(+) CD4(+) T cells (circulating Tfh cells) were detected by flow cytometry. Levels of IL-21 and IL-4 in peripheral blood were detected by cytometric bead array. The mRNA expression of IL-21, IL-4, Bcl-6, and Blimp-1 in peripheral blood mononuclear cells (PBMCs) were measured by real-time PCR. Levels of IgG1, IgG2, IgG3, and IgG4 in the patients' sera were measured using enzyme-linked immunosorbent assay. RESULTS: Percentages of circulating Tfh cells were significantly increased in the CE1, CE2, and CE3 groups (p < 0.05). The concentrations of IL-21 and IL-4 in the serum were significantly increased in CE1, CE2, and CE3 groups (p < 0.05). IL-21 was positively correlated with circulating Tfh cells in CE3 group (r = 0.779, p < 0.05). The mRNA levels of IL-21, IL-4, and Bcl-6 were increased in CE1, CE2, and CE3 groups. Levels of IgG1 and IgG4 in patients' sera were increased in CE1, CE2, and CE3 groups. Levels of IgG2 and IgG3 were increased in CE4-5 group. Additionally, after stimulation with hydatid fluid in vitro, the levels of circulating Tfh cells, IL-21 and IL-4 in PBMCs isolated from CE patients were significantly increased (p < 0.05). CONCLUSIONS: The levels of circulating Tfh and related cytokines were significantly increased in CE patients, suggesting that they are involved in human CE.

Rapid and subnanomolar assay of recombinant human erythropoietin by capillary electrophoresis using NanoOrange precolumn labeling and laser-induced fluorescence detection.

Because of less functionally critical carbohydrate sectors that contributed to the stability, efforts have been made to quantify intact recombinant human erythropoietin. A simple, rapid capillary electrophoresis with laser-induced fluorescence method for the assay of recombinant human erythropoietin was developed, with a limit of detection of intact recombinant human erythropoietin at subnanomolar concentration (up to 10 ng/mL or 3 × 10(-10) M), which is among the lowest reported. High sensitivity was accomplished by precolumn derivatization with the noncovalent dye NanoOrange. Capillary electrophoresis separation and reaction conditions were carefully manipulated for avoiding microheterogeneity of glycoforms and inhomogeneity of multiple labeling products. The fluorescence signal was linear over the range of 10 ng/mL-10 µg/mL, corresponding to the detection requirement of recombinant human erythropoietin in biofluids and pharmaceutical samples, as demonstrated by a real sample analysis. Although the salt in reaction mixtures showed a detrimental effect on the fluorescence of the derivatives, this method could tolerate a certain amount of salt, extending its application in biofluid analysis. In addition, zero-order fluorescence emission kinetics was obtained, indicating that the rapid decay of recombinant human erythropoietin was derived from a self-quenching effect.

Th9/IL-9 profile in human echinococcosis: their involvement in immune response during infection by Echinococcus granulosus.

Th9 cells have been reported to contribute to immune responses; however, the role of Th9 cells in Echinococcus granulosus infection is unknown. This study is to determine whether Th9 cells and IL-9 are involved in human Echinococcus granulosus infection. Compared with healthy controls (HC group), the mRNA levels of PU.1, IL-9, and GATA-3 were significantly increased in patients before therapy (CE group), as revealed by qRT-PCR. Flow cytometry analysis showed that the percentages of Th9 and Th2 cells in CE group were significantly higher. The levels of IL-9, IL-4, IL-10, and TGF- ß in CE group were also significantly increased, as detected by CBA assay. The percentages of Th9 and Th2 cells in CE group were positively correlated. After treatments of surgery in combination with albendazole, the PU.1 and GATA-3 mRNA levels were significantly decreased in patients after therapy (PCE group) compared with CE group. The numbers of Th9 and Th2 cells and levels of IL-9, IL-4, IL-10, and TGF- ß were also significantly decreased in PCE group. In conclusion, the ratios of Th9 cells and IL-9 levels were significantly decreased after treatment, suggesting that Th9/IL-9 may be involved in immune response induced by Echinococcus granulosus infection.

The effect of calcium oxalate stones and uric acid stones on male sexual function.

PURPOSE: This study compared the effects of calcium oxalate stones and uric acid stones on male sexual function. METHODS: We enrolled 100 patients with ureteral stones. According to the composition of the stones, they were divided into the calcium oxalate stone group and the uric acid stone group. All patients underwent ureteroscopic holmium laser lithotripsy. General data such as age, body mass index, course of disease, stone diameter, and degree of renal hydronephrosis were compared. Sperm parameters, including sperm density, sperm viability, and sperm deformity rate, as well as International Index of Erectile Function-5 questionnaire (IIEF-5) scores, and Quality of Life (QOL) scores, were measured and compared before and 6 weeks after the surgery. RESULTS: There were no statistically significant differences in general data and sperm parameters between the two groups before the surgery (P > 0.05). However, there were significantly lower IIEF scores but significantly higher QOL scores in the uric acid stone group. In the calcium oxalate stone group, there were no statistically significant differences in sperm parameters, IIEF score, and QOL score before and after the surgery (P > 0.05). In the uric acid stone group, there were no statistically significant differences in sperm parameters before and after surgery (P > 0.05), whereas there were significantly higher IIEF scores but significantly lower QOL scores after the surgery (P < 0.05). The prevalence of erectile dysfunction (ED) in the uric acid stone group was 38.18% (21/55), which was significantly higher compared to 20.00% (9/45) in the calcium oxalate stone group (P < 0.05). The multivariate binary logistic regression analysis showed that the independent risk factor related to ED was uric acid stones (odds ratio: 2.637, 95% confidence interval 1.040-6.689, P = 0.041). No statistically significant differences were found in sperm parameters between patients with and without ED. CONCLUSION: Compared with the calcium oxalate stone group, patients with uric acid stones had a higher prevalence of ED and poorer sexual performance.

Serum TNF-α level and probing depth as a combined indicator for peri-implant disease.

Peri-implant disease (PID) is a general term for inflammatory diseases of soft and hard tissues that occur around implants, including peri-implant mucositis and peri-implantitis. Cytokines are a class of small molecule proteins, which have various functions such as regulating innate immunity, adaptive immunity, and repairing damaged tissues. In order to explore the characteristics and clinical significance of tumor necrosis factor (TNF)-α, interleukin (IL)-6, IL-10, and tumor growth factor (TGF)-ß1 expression levels in serum of patients with peri-implant disease, 31 patients with PID and 31 patients without PID were enrolled. The modified plaque index (mPLI), modified sulcus bleeding index (mSBI), and peri-implant probing depth (PD) were recorded. The levels of serum TNF-α, IL-6, IL-10, and TGF-ß1 were detected by ELISA. TNF-α, mPLI, mSBI, and PD levels were significantly higher in the PID group. TGF-ß1 levels were significantly higher in the control group. There was a significant positive correlation between TNF-α and mPLI, mSBI, and PD. TGF-ß1 was negatively associated with TNF-α, mPLI, mSBI, and PD. Multiple logistic regression analysis showed that TNF-α and PD were risk factors for the severity of PID. The receiver operating curve analysis showed that high TNF-α levels (cut-off value of 140 pg/mL) and greater PD values (cut-off value of 4 mm) were good predictors of PID severity with an area under the curve of 0.922. These results indicated that TNF-α and PD can be used as a biological indicator for diagnosing the occurrence and progression of PID.

Galectin-9 alleviates acute graft-versus-host disease after haplo-hematopoietic stem cell transplantation by regulating regulatory T cell/effector T cell imbalance.

BACKGROUND: Acute graft-versus-host disease (aGVHD) arises from the imbalance of host T cells. Galectin-9 negatively regulates CD4 effector T cell (Th1 and Th17) function by binding to Tim-3. However, the relationship between Galectin-9/Tim-3 and CD4+ T subsets in patients with aGVHD after Haplo-HSCT (haploidentical peripheral blood hematopoietic stem cell transplantation) has not been fully elucidated. Here, we investigated the role of Galectin-9 and CD4+ T subsets in aGVHD after haplo-HSCT. METHODS: Forty-two patients underwent Haplo-HSCT (26 without aGVHD and 16 with aGVHD), and 20 healthy controls were included. The concentrations of Galectin-9, interferon-gamma (IFN-γ), interleukin (IL)-4, transforming growth factor (TGF)-ß, and IL-17 in the serum and culture supernatant were measured using enzyme-linked immunosorbent assay or cytometric bead array. The expression levels of Galectin-9, PI3K, p-PI3K, and p-mTOR protein were detected by western blot analysis. Flow cytometry was used to analyze the proportions of CD4+ T cell subsets. Bioinformatics analysis was performed. RESULTS: In patients with aGVHD, regulatory T (Treg) cells and Galectin-9 decreased, and the Th1, Th17, and Treg cells were significantly imbalanced. Moreover, Treg and Galectin-9 were rapidly reconstituted in the early stage of patients without aGVHD after Haplo-HSCT, but Th17 cells were reconstituted slowly. Furthermore, Tim-3 upregulation on Th17 and Th1 cells was associated with excessive activation of the PI3K/AKT pathway in patients with aGVHD. Specifically, in vitro treatment with Galectin-9 reduced IFN-γ and IL-17 production while augmenting TGF-ß secretion. Bioinformatics analysis suggested the potential involvement of the PI3K/AKT/mTOR pathway in aGVHD. Mechanistically, exogenous Galectin-9 was found to mitigate aGVHD by restoring the Treg/Teffs (effector T cells) balance and suppressing PI3K. CONCLUSION: Galectin-9 may ameliorate aGVHD after haplo-HSCT by modulating Treg/Teffs balance and regulating the PI3K/AKT/mTOR pathway. Targeting Galectin-9 may hold potential value for the treatment of aGVHD.

[Expression and Clinical Significance of Helper T Cells 9 and Its Sytokines Interleukin 9 in Chronic Lymphocytic Leukemia].

OBJECTIVE: To investigate the expression and clinical significance of T helper cell 9 (Th9) and its cytokine interleukin 9(IL-9) in peripheral blood of patients with chronic lymphocytic leukemia(CLL). METHODS: A total of 43 newly diagnosed patients with chronic lymphocytic leukemia in the First Affiliated Hospital of Xinjiang Medical University from June 2021 to June 2022 were selected as the case group. The patients were divided into Binet A group (13 cases), Binet B group (20 cases) and Binet C group (10 cases) by Binet staging system, and 20 healthy volunteers who underwent physical examinationin in our hospital in the same period served as control group. The proportion of Th9 cells in peripheral blood was detected by flow cytometry, the expression level of Th9 specific transcription factors PU.1 and IRF4 was detected by Western blot, and the expression level of serum cytokine IL-9 was detected by ELISA. The proportion of Th9, the expression of PU.1, IRF4 and IL-9 in each group were compared, and the correlation between the proportion of Th9, IL-9 and clinicopathological indexes of CLL patients was analyzed. RESULTS: The proportion of Th9, the expression of PU.1, IRF4 and IL-9 in CLL group were significantly higher than those in control group (P<0.05), the proportion of Th9 and the expression of IL-9 in Binet B and C group were higher than those in Binet A group (P<0.05), but there was no significant difference in the proportion of Th9 cells between Binet B group and C group (P>0.05). The expression of IL-9 in Binet C group was significantly higher than that in Binet B group (P<0.05) . The proportion of Th9 cells and IL-9 were highly expression in patients with ß2 microglobulin abnormality, IGHV unmutation, P53 abnormality and hepatosplenic lymph node enlargement(P<0.05), but not related to age and sex (P>0.05). The results of Spearman correlation analysis showed that the proportion of Th9 in patients with CLL was negatively correlated with the lymphocytic account and lymphocyte proportion(rs=-0.32，rs=-0.34). The proportion of Th9 and IL-9 were positively correlated with Binet stage, Rai stage and CLL-IPI Scoring (rs=0.79，rs=0.54，rs=0.58； rs=0.72，rs=0.63，rs=0.45), but not with WBC, CD4+ T cells and CD8+T cells (P>0.05). The proportion of Th9 was positively correlated with IL-9 (rs=0.53). CONCLUSION: Th9 cells and IL-9 are abnormally highly expressed in CLL, which is related to the poor prognosis of CLL.

Characteristics of Serum Autoantibody Repertoire and Immune Subgroup Variation of Tuberculosis-Associated Obstructive Pulmonary Disease.

Background: Studying the potential etiology and pathogenesis of tuberculosis-associated chronic obstructive pulmonary disease (TOPD) from an autoimmunity perspective may provide insights into peripheral blood autoantibodies and immune cells, as well as their interactions. Methods: This study examined the serum autoantibody repertoire in healthy individuals, patients with chronic obstructive pulmonary disease (COPD), patients with pulmonary tuberculosis (TB), and TOPD patients using the HuProtTM protein chip. Autoantigens in the peripheral blood of TOPD patients were verified using ELISA assay. Various epitopes and immune simulation were predicted using bioinformatic methods. Flow cytometry was employed to detect macrophages(Mφ), T cells, and innate lymphoid cells (ILCs) in the peripheral blood. Results: COPD patients displayed distinct alterations in their IgG and IgM autoantibodies compared to the other groups. GeneOntology (GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses revealed that these autoantibodies were associated with regulating macrophages, T cells, and B cells. ELISA results confirmed the upregulation of expression of proliferating cell nuclear antigen (PCNA), Mitogen-Activated Protein Kinase 3 antigen (MAPK3), and threonine protein kinase 1 antigen (AKT1) proteins in the peripheral blood of TOPD patients. Bioinformatic analysis predicted multiple potential epitopes in Th, CTL, and B cells. Immune simulation results demonstrated that PCNA, MAPK3, and AKT1 can activate innate and adaptive immune responses and induce the expression of different cytokines, such as IFN-g and IL-2. Furthermore, data obtained from flow cytometry assay revealed an upregulation in the face of Th1 cells in the peripheral blood of TOPD patients. Conclusion: Tuberculosis infection can effectively induce autoimmune responses, contributing to increased expression of Th1 cells and associated cytokines, ultimately leading to immune dysregulation. Furthermore, the accumulation of pulmonary inflammatory response facilitates the progression of TOPD and is helpful for the clinical diagnosis and the development of targeted therapeutic drugs for this disease.

The Activation of M1 Macrophages is Associated with the JNK-m6A-p38 Axis in Chronic Obstructive Pulmonary Disease.

Background: Excessive activation of M1 macrophages affects the chronic inflammatory response of the airways and leads to the development of chronic obstructive pulmonary disease (COPD). Therefore, it needs to be closely monitored and investigated. MAPK signaling pathway is involved in the activation of M1 macrophages, and N6-methyladenosine (m6A) is involved in the pathogenesis of COPD. However, it is unknown whether activation of the MAPK signaling pathway is mediated by m6A in M1 macrophages in COPD. Methods: The GEO data were analyzed using bioinformatics techniques to assess the differences between COPD and healthy individuals in the levels of M1 macrophages, their secreted cytokines, and m6A regulators. The MAPK signaling pathway was significantly enriched in the list of differentially regulated genes between COPD and healthy individuals. We further analyzed the correlation between M1 macrophages, m6A, and the MAPK signaling pathway. Next, blood samples from COPD and healthy individuals were collected and analyzed by using flow cytometry, ELISA, and RT-PCR. Western blotting was performed using CSE-induced THP-1 cells. COPD and healthy mice were used for Me-RIP sequencing and flow cytometry experiments. Validation of the results of the above bioinformatics analysis by molecular biology experiments and sequencing techniques. Results: We found that GEO data and blood specimens from COPD patients showed increased M1 macrophages, higher levels of IL-6 and TNF-α, and higher mRNA expression of key mediators of the MAPK signaling pathway (p38, ERK, and JNK). Western blotting showed increased expression of p38, ERK, and JNK in the CSE group. In contrast, the expression of m6A regulators was low. Also, M1 macrophages in COPD mice were hyperactivated and had reduced m6A modifications of p38, ERK, and JNK compared with control. Conclusion: m6A may be involved in M1 macrophage hyperactivation by regulating the MAPK signaling pathway, thereby influencing the development of COPD.

Galectin-9 and myeloid-derived suppressor cell as prognostic indicators for chronic lymphocytic leukemia.

BACKGROUND: Galectin-9 and myeloid-derived suppressor cells (MDSCs) have an important role in tumors, but their clinical values in chronic lymphocytic leukemia (CLL) have not been fully elucidated. This study aimed to analyze the prognosis values of Galectin-9 and MDSCs in CLL. METHODS: The concentrations of Galectin-9, argininase-1, and inducible nitric oxide synthase in serum were detected by enzyme-linked immune sorbent assay. The expression of Tim-3 protein in peripheral blood mononuclear cell was detected by Western blot. Flow cytometry was used to analyze the percentages of Tim-3 on T-cells (CD3+ T, CD4+ T, and CD8+ T cells) and MDSCs. RESULTS: Our results showed that Galectin-9 and MDSCs significantly increased in CLL patients and were closely related to the disease progression. Patient's receiver operating characteristic, progression-free survival, and Cox regression analysis showed that Galectin9 and MDSCs were poor prognostic factors of CLL. CONCLUSION: Galectin-9 and MDSCs were associated with clinical progression and could be important prognostic indicators for CLL.

The level of Tim-3+CD8+ T cells can serve as a potential marker for evaluating the severity of acute graft-versus-host disease after haplo-PBSCT.

Early and accurate diagnosis of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation is crucial for the prognosis of patients. This study identified a potential biomarker for the severity of aGVHD after human leukocyte antigen (HLA)-haploidentical peripheral blood hematopoietic stem cell transplantation (haplo-PBSCT). We included 20 healthy subjects and 57 patients who underwent haplo-PBSCT. Of these patients, 22 developed aGVHD after haplo-PBSCT. The results showed that patients with aGVHD had significantly increased levels of Tim-3+/Perforin+/Granzyme B+CD8+ T cells, but significantly decreased Galectin-9. The differences in Galectin-9 and Tim-3+/Granzyme B+CD8+ T cells between grade I-II aGVHD and III-IV aGVHD were also significant. In vitro, the apoptosis of CD8+ T cells from aGVHD patients was significantly increased after Tim-3/Galectin-9 pathway activation, which decreased Granzyme B secretion. As revealed by univariate analysis, the level of Tim-3+CD8+ T cells was a risk factor for severe aGVHD. ROC analysis demonstrated that high levels of Tim-3+CD8+ T cells had a significant diagnostic value for severe aGVHD, with an area under the curve of 0.854 and cut-off value of 14.155%. In conclusion, the binding of Tim-3 with exogenous Galectin-9 can promote apoptosis of CD8+ T cells and affect the secretion of Granzyme B. Tim-3+CD8+ T cells have the potential to serve as immunological markers for assessing the severity of aGVHD after haplo-PBSCT and identifying patients at a higher risk for severe aGVHD.

Unique Reduced-Intensity Conditioning Haploidentical Peripheral Blood Stem Cell Transplantation Protocol for Patients with Hematologic Malignancy.

Reduced-intensity conditioning (RIC) haploidentical (haplo-) hematopoietic stem cell transplantation (HSCT) requires more hematopoietic progenitor and stem cells (HPSCs) to promote engraftment and immune reconstitution and needs a stronger graft-versus-leukemia effect. Peripheral blood stem cells (PBSCs) offer advantages over bone marrow; however, the use of higher-dose non-T cell-depleted (non-TCD) in vitro PBSCs may increase the occurrence of severe graft-versus-host disease (GVHD). This prospective, single-arm clinical study was performed to investigate using high-dose non-TCD in vitro PBSCs as the graft source, using fludarabine/Ara-C/busulfan (FAB) as the conditioning regimen, using rabbit antithymocyte globulin to remove T cells in vivo, and enhancing GVHD prophylaxis with an IL-2 receptor antagonist in RIC-haplo-HSCT in patients with hematologic malignancies age 50 to 70 years or <50 years with comorbidities (Hematopoietic Cell Transplantation Comorbidity Index score ≥2) classified as intermediate to high risk. The primary endpoint was day 100 acute GVHD (aGVHD). A total of 47 patients were enrolled; the median age was 52 years (range, 30 to 68 years), the median duration of follow-up was 34 months (range, 2 to 99 months), and the medium-infused doses of mononuclear cells, CD34+ cells, and CD3+ cells were 15.93 × 108/kg, 8.68 × 106/kg, and 5.57 × 108/kg, respectively. The cumulative incidence of grade II-IV aGVHD at day 100 was 30.3% (95% confidence interval [CI], 15.9% to 44.8%), and that of grade III-IV aGVHD was 10.2% (95% CI, .6% to 19.8%). The 2-year cumulative incidence of chronic GVHD (cGVHD) was 34.9% (95% CI, 19.0% to 50.8%). The 2-year cumulative incidences of localized and extensive cGVHD were 26.1% (95% CI, 11.80% to 40.40%) and 8.7% (95% CI, 3.26% to 20.65%), respectively. The 2-year cumulative incidence of relapse was 17.3% (95% CI, 5.1% to 29.5%), the 2-year overall survival rate was 71.2% (95% CI, 57.9% to 84.5%), and the 2-year disease-free survival rate was 66.2% (95% CI, 52.1% to 80.3%). The incidence of aGVHD was not high, and the overall efficacy was good. This study demonstrates that this unique RIC-haplo-PBSC transplantation protocol was effective in treating hematologic malignancies. Nonetheless, larger prospective multicenter clinical trials and experimental studies should be performed to further confirm our findings.

[Dynamic change of IL-10 and TGF-beta1 in the liver of Echinococcus multilocularis-infected mice].

OBJECTIVE: To observe the dynamic expression and function of IL-10 and TGF-beta1 in liver of BALB/c mice infected with Echinococcus multilocularis (Em). METHODS: Sixty female BALB/c mice were randomly divided into experiment group and control group. Mice in the experiment group were each injected with 0.2 ml Em protoscolex suspension (containing about 400 protoscoleces) , while those in control group received same volume of normal saline. At 2, 8, 30, 90, 180, and 360 d after infection, 5 mice from each group were sacrificed and liver specimens were collected for pathological examination and immunohistochemical detection for IL-10 and TGF-beta1. RESULTS: In mice of the experiment group, Em cysts in different sizes were found in the abdominal cavity and the liver tissue, which gradually enlarged with the time. HE staining showed infiltration of lymphocytes in liver tissue, pathological change between the cyst wall and hepatic cells. In the control, the liver lobules showed integrity and inflammatory cells were seen occasionally. The level of IL-10 expression in liver tissue of the infected mice increased with the time, and reached a peak [(1639 +/- 1.73) %] at 90 d post-infection and maintained a high level thereafter. The expression of TGF-beta1 also reached the highest level [(23.69 +/- 2.29) %] . Both were significantly higher than the control (P < 0.01), though a low level expression was found in the control at 90d post-injection. CONCLUSION: The expressions of IL-10 and TGF-beta1 both increase in the middle and late stages of the infection. Besides, their inhibited functions do not be helpful for clearing and controlling Echinococcus multilocularis infection in livers.