MBD2 promotes Th2 differentiation in ovalbumin-induced CD4+T cells

Introduction:Allergen-specific CD4+T cells play a central role in autoimmune disorders,allergies and asthma,with Th2-type immunity being the typical functional response of CD4+T cells.This study aimed to investigate the role of MBD2 in regulating Th2 cell differentiation.Methods:Splenic mononuclear cells were extracted from C57BL/6 mice,and CD4+T cells were isolated using magnetic beads and confirmed through flow cytometry.Lentivirus was employed to construct MBD2-silenced CD4+T cells.In vitro experiments were performed to treat splenogenic mononuclear cells and CD4+T cells with Ovalbumin(OVA),and Th2 cell ratios and IL-4 levels were assessed using flow cytometry and ELISA.Results:The purity of the isolated CD4+T cells was 95.73%,confirming successful isolation of primary CD4+T cells.Compared to the control group,the Th2 cell ratio exhibited an increase in the Th2-induced group.Treatment with 5-Aza(concentrations,1-100μM)promoted Th2 cell differentiation and increased IL-4 levels.Notably,when combined with Th2 induction and 10μM 5-Aza treatment,silencing MBD2 further amplified Th2 cell ratios and elevated IL-4 levels in cell supernatants.Furthermore,OVA(concentration,200μg/mL)induced the differentiation of CD4+T cells into Th2 cells and increased IL-4 secretion.Interestingly,silencing MBD2 significantly increased the Th2 cell ratio and IL-4 levels in OVA-treated CD4+T cells.Conclusion:In summary,OVA promoted CD4+T cell differentiation into Th2 cells and enhanced IL-4 levels.MBD2 was identified as a mediator of Th2 cell differentiation in splenic-derived CD4+T cells,influenced by OVA or 5-Aza treatment.

Integration of scRNA-Seq and Bulk RNA-Seq to analyze the heterogeneity ofcolorectal cancer immune cells and establish a molecular risk model

Background:Colorectal cancer(CRC)is a highly heterogeneous malignant tumor that significantly impacts clinical diagnosis and treatment.Single-cell RNA sequencing is an innovative method for exploring tumor heterogeneity and understanding its role at cellular and genetic levels.Method:The colorectal cancer Single-cell RNA sequencing data were analysed on the immune.RNA-seq data in bulk form was utilized to assess the major genes of the immune cell subsets linked to CRC.We conducted an analysis of the abundance of immune cells in the microenvironment of CRC,and also performed weighted gene co-expression network analysis.Gene set enrichment analysis helped perform two analytical procedures of subtype groups.Furthermore,Least absolute shrinkage and selection operator regression was employed to analyse and screen for a gene signature.Finally,quantitative PCR Was performed to detect the expression levels of signature genes in CRC.Results:The Single-cell RNA sequencing(GSE146771)dataset was integrated to obtain 9 cell clusters.The Single-sample gene set enrichment analysis showed that the related gene expression of T-cell subsets of different functional statuses could vary greatly between patients with GSE146771.Immune cell analysis of TCGA-CRC indicated an improved overall survival rate for patients with elevated Th2 cell abundance.Five-gene signature(Risk Score=-0.205×CDC25C-0.231×GSTCD-0.010×KPNA2-0.002×KIF15-0.171×ORC1)was developed by weighted correlation network analysis,and lasso Cox regression.Then,the risk prediction efficacy of the signature was validated in four GSE datasets.Furthermore,the expression of five genes was reduced in CRC tissue by quantitative PCR.Conclusion:Five-gene signature based on CRC heterogeneity was developed as a prognosis predictor,which can serve as a potential treatment target.

Rapamycin ameliorates experimental autoimmune uveoretinitis by inhibiting Th1/Th2/Th17 cells and upregulating CD4+CD25+ Foxp3 regulatory T cells

· AIM: To determine the effects of rapamycin on experimental autoimmune uveoretinitis(EAU) and investigate of role of rapamycin on T cell subsets in the disease.·METHODS: EAU was induced in rats using peptides1169 to 1191 of the interphotoreceptor binding protein(IRBP). Rapamycin(0.2 mg/kg/d) was administrated by intraperitoneal injection for a consecutive 7d after immunization. Th1/Th2/Th17 cytokines, TGF-β1, and IL-6produced by lymphocyteswere measured by ELISA, while Th17 cells and CD4 +CD25 + regulatory T cells(Tregs)from rat spleen were detected by flow cytometry.·RESULTS: Intraperitoneal treatment immediately after immunization dramatically ameliorated the clinical course of EAU. Clinical responses were associated with reduced retinal inflammatory cell infiltration and tissue destruction. Rapamycin induced suppression of Th1/Th2/Th17 cytokines, including IFN-γ, IL-2, IL-17, IL-4, and IL-10 release from T lymphocytes of EAU rats, in vitro.Rapamycin also significantly increased TGF-β1production but had no effect on IL-6 productionof T lymphocytes from EAU rats in vitro. Furthermore,rapamycin decreased the ratio of Th17 cells/CD4 +T cells and upregulated Tregs in EAU, as detected by flow cytometry.·CONCLUSION: Rapamycin effectively interferes with T cell mediated autoimmune uveitis by inhibiting antigen-specific T cell functions and enhancing Tregs in EAU.Rapamycin is a promising new alternative as an adjunct corticosteroid-sparing agent for treating uveitis.

Impact of Bisphenol A (BPA) and Free Fatty Acids (FFA) on Th2 Cytokine Secretion from INS-1 Cells

Bisphenol A (BPA) is used in huge amounts for many plastic products and is a hormone (estrogen) disrupting agent. BPA as well as FFAs may be deleterious for the immune system. The aim was to identify Th2 cytokines and some of their signal transduction mechanisms in INS-1 cells, an insulin secreting cell line. Screening using a proteome profile indicated an increase of IL-1, IL-2, IL-4, IL-6, IL-10, IL-13 and IL-17 by BPA. Also FFAs (in combination with LPS) were positive. In detailed quantitative measurements, these results were confirmedly indicating a complex array of pro-and anti-inflammatory potential. The interaction of BPA with 17β-estradiol was non-additive with respect to IL-4 and IL-6 release and additive with respect to FFA interaction indicating same and different mechanisms of action, respecttively. As signal transduction PI3K (Wortmannin-sensitive) and STAT-3/6 (Tofacitinib-sensitive) are involved in various effects, INS-1 cells release several cytokines due to BPA and FFA attack which may be involved in disturbance of glucose homoeostasis and type 1 diabetes.

GATA-3 promotes Th2 responses through three different mechanisms： induction of Th2 cytokine production, selective growth of Th2 cells and inhibition of Thl cell-specific factors

Naive CD4 T cells can differentiate into at least two different types ofT helpers, Thl and Th2 cells. Th2 cells, capable of producing IL-4, IL-5 and IL-13, are involved in humoral immunity against extracellular pathogens and in the induction of asthma and other allergic diseases. In this review, we summarize recent reports regarding the transcription factors involved in Th2 differentiation and cell expansion, including StatS, Gfi- 1 and GATA-3. Stats activation is necessary and sufficient for IL-2-mediated function in Th2 differentiation. Enhanced Stats signaling induces Th2 differentiation independent of IL-4 signaling; although it does not up-regulate GATA-3 expression, it does require the presence of GATA-3 for its action. Gfi-1, induced by IL-4, promotes the expansion of GATA-3-expressing cells. Analysis of conditional Gata3 knockout mice confirmed the critical role of GATA-3 in Th2 cell differentiation （both IL-4 dependent and IL-4 independent） and in Th2 cell proliferation and also showed the importance of basal GATA-3 expression in inhibiting Thl differentiation.

All-transRetinoic Acid Regulates Th1/Th2 Balance in CD4+T cells When GATA-3 is Deficient

The essential effect of vitamin A on immune function occurs through various mechanisms including direct effect on ThloTh2 balance modulation. However, it is unclear whether or not vitamin A can regulate Thl-Th2 balance under a strong Thl-polarizing condition. Therefore, the purpose of our study was to examine the effect of vitamin A metabolite allotrans retinoic acid （ATRA） on ThloTh2 differentiation in CD4~ T cells under GATA-3 deficiency, which can induce Thl-polarizing condition. In the present study, GATA-3 deficiency T cells were induced by siRNA and checked by real-time quantitative PCR and western blot. GATA-3 deficiency CD4＋ T cells and normal CD4＋ T were treated for 48 h with or without ATRA.

Activated rat hepatic stellate cells influence Th1/Th2 profile in vitro

AIM: To investigate the effects of activated rat hepatic stellate cells(HSCs) on rat Th1/Th2 profile in vitro.METHODS: Growth and survival of activated HSCs and CD4+ T lymphocytes cultured alone or together was assessed after 24 or 48 h. CD4+ T lymphocytes were then cultured with or without activated HSCs for 24 or 48 h and the proportion of Th1 [interferon(IFN)-γ+] and Th2 [interleukin(IL)-4+] cells was assessed by flow cytometry. Th1 and Th2 cell apoptosis was assessed after 24 h of co-culture using a caspase-3 staining procedure. Differentiation rates of Th1 and Th2 cells from CD4+ T lymphocytes that were positive for CD25 but did not express IFN-γ or IL-4 were also assessed after 48 h of co-culture with activated HSCs. Galectin-9 expression in HSCs was determined by immunofluorescence and Western blotting. ELISA was performed to assess galectin-9 secretion from activated HSCs.RESULTS: Co-culture of CD4+ T lymphocytes with activated rat HSCs for 48 h significantly reduced the proportion of Th1 cells compared to culture-alone conditions(-1.73% ± 0.71%; P < 0.05), whereas the proportion of Th2 cells was not altered; the Th1/Th2 ratio was significantly decreased(-0.44 ± 0.13; P < 0.05). In addition, the level of IFN-γ in Th1 cells wasdecreased(-65.71 ± 9.67; P < 0.01), whereas the level of IL-4 in Th2 cells was increased(82.79 ± 25.12; P < 0.05) by co-culturing, as measured by mean fluorescence intensity by flow cytometry. Apoptosis rates in Th1(12.27% ± 0.99%; P < 0.01) and Th2(1.71% ± 0.185%; P < 0.01) cells were increased 24 h after co-culturing with activated HSCs; the Th1 cell apoptosis rate was significantly higher than in Th2 cells(P < 0.01). Galectin-9 protein expression was significantly decreased in HSCs only 24 h after coculturing(P < 0.05) but not after 48 h. Co-culture for 48 h significantly increased the differentiation of Th1 and Th2 cells; however, the increase in the proportion of Th2 cells was significantly higher than that of Th1 cells(1.85% ± 0.48%; P < 0.05).CONCLUSION: Activated rat HSCs lower the Th1/Th2 profile, inhibiting the Th1 response and enhancing the Th2 response, and this may be a novel pathway for liver fibrogenesis.

Mesenchymal stromal cells as potential immunomodulatory players in severe acute respiratory distress syndrome induced by SARSCoV- 2 infection

Severe acute respiratory syndrome coronavirus-2 and the related coronavirus disease-19(COVID-19)is a worldwide emerging situation,which was initially reported in December 2019 in Wuhan,China.Currently,more than 7258842 new cases,and more than 411879 deaths have been reported globally.This new highly transmitted coronavirus is responsible for the development of severe acute respiratory distress syndrome.Due to this disorder,a great number of patients are hospitalized in the intensive care unit followed by connection to extracorporeal membrane oxygenation for breath supporting and survival.Severe acute respiratory distress syndrome is mostly accompanied by the secretion of proinflammatory cytokines,including interleukin(IL)-2,IL-6,IL-7,granulocyte colony-stimulating factor(GSCF),interferon-inducible protein 10(IP10),monocyte chemotactic protein-1(MCP1),macrophage inflammatory protein 1A(MIP1A),and tumor necrosis factor alpha(TNF-α),an event which is known as“cytokine storm”.Further disease pathology involves a generalized modulation of immune responses,leading to fatal multiorgan failure.Currently,no specific treatment or vaccination against severe acute respiratory syndrome coronavirus-2(SARS-CoV-2)has been developed.Mesenchymal stromal cells(MSCs),which are known for their immunosuppressive actions,could be applied as an alternative co-therapy in critically-ill COVID-19 patients.Specifically,MSCs can regulate the immune responses through the conversion of Th1 to Th2,activation of M2 macrophages,and modulation of dendritic cells maturation.These key immunoregulatory properties of MSCs may be exerted either by produced soluble factors or by cell-cell contact interactions.To date,several clinical trials have been registered to assess the safety,efficacy,and therapeutic potential of MSCs in COVID-19.Moreover,MSC treatment may be effective for the reversion of ground-glass opacity of damaged lungs and reduce the tissue fibrosis.Taking into account the multifunctional properties of MSCs,the proposed stem-cell-based therapy may be proven significantly effective in critically-ill COVID-19 patients.The current therapeutic strategy may improve the patient’s overall condition and in parallel may decrease the mortality rate of the current disease.

Effects of the Overall Alkaloid of a Traditional Chinese Medicine “Tongbiling” on the Cytokine Expression in Th1 and Th2 Cells of Rheumatoid Arthritis and Their Signaling Pathway

To investigate the effects of overall alkali of a traditional Chinese medicine “Tongbiling” (brucine and strychnine alkaloids in main) on the cytokines expression in Th1 and Th2 cells in the synovial fluid of patients with rheumatism arthritis and their signal pathway, the mononuclear cells in the synovial fluid (SFMC) of patients were isolated by Ficoll-Hypaque gradient centrifugation, and the CD3 + CD69 + and CD3 + HLA-DR antigen were analyzed by flow cytometry in comparison with those of the peripheral blood. The rest of cells were cultured after resuspension with RPMI 1640 culture medium. Phorbol 12,13-dibutyrate (PDB) and ionomycin were added successively into the culture with various concentration of overall alkali Tongbiling (TBL). After 4 h of cultivation, the expression of IFN-γ and IL-4 in CD3 + cells were analyzed by flow cytometry. The influence of overall alkali TBL (100?mg/L) on the intracellular calcium was investigated after Fluo-3/AM labeling and stimulation with PDB and ionomycin at 1, 2, 4 and 10?min, and the influence of TBL on the expression of CD3 + CD69 + cells were determined with stimulation of PDB for 24?h in the whole blood lymphocytes culture. It was found that the percentage of T cells bearing CD69 was significantly up-regulated (77%), while that of T cells bearing HLA-DR was 44% in the synovial mononucleated cells. After PDB and ionomycin stimulation, the expression of IFN-γ in CD3 + cells were up-regulated, but there was no change on the expression of IL-4 in CD3 + cells, indicating that ratio of Th1/Th2 was significantly increased and Th cells differentiate to Th1 cells in mainly. Four concentrations of overall alkaloid of TBL (200?mg/L, 100?mg/L, 50?mg/L, 25?mg/L) could down-regulated the expression of IFN-γ in CD3 + cells and the Th1/Th2 ratio obviously, but all the concentrations of the overall alkaloids had no effect on the expression of IL-4 in CD3 + cells. 100?mg/L concentration of the overall alkaloid did not down-regulate the intracellular calcium level. Each concentration of the overall alkaloid could down-regulated the expression CD69 obviously on the PDB-activated mouse T cells. It concluded from the above observations that the overall alkaloid of TBL could relieve the inflammatory and immune damages by suppressing the expression of Th1 type cytokines and Th1 cell differentiation, regulating the imbalance of Th1/Th2 cells and inhibiting the early activation of the T lymphocytes bearing CD69. There was no remarkable influence on the intracellular calcium signaling transduction pathway. The inhibitory effected on T cells to express IFN-γ might be due to the suppression of PKC-MAPK signaling pathway. From the standpoint of traditional Chinese medicine, this might be due to the regulation of “Yin” and “Yang” imbalance of joints to modify the pathological status in rheumatoid arthritis. This study provided an experimental basis for the application of overall alkaloids of TBL in the treatment of rheumatoid arthritis.

Influence of the invasion of peripheral blood mononuclear cells by hepatitis B virus on immune response of the patients with chronic hepatitis B

Objective:To exploretheinfluenceof HBVinvasionintoperipheralbloodmononuclearcells(PBMC)on theimmuneresponseof patientswithchronichepatitisB.Method s:Thecytokinelevelsintheculturesupernatantof PBMCfrom56patientswithchronichepatitisB weredeterminedby ELISA,andPCRwasemployedto amplifythe HBVDNA.Results:Thelevelsof IFN-γinpatientswithhepatitisB waslowerthanthosetof thecontrol,butthe differencewasnotstatisticallysignificant,whilethelevelsof IL-4weresignificantlyhigherthanthoseof thecontrol(P<0.01).Theserumlevelsof HBVDNAwerenegativelycorrelatedwiththatof IFN-γin culturesupernatantsof PBMC.Thirty-fivepatientspositiveof HBVDNA inthePBMCswereidentifiedfrom56patientswithhepatitisB,andtheirIFN-γlevelprovedto be significantlydifferent.Conclusions:Th2cell-mediatedimmuneresponseis predo-minantin chronichepatitisB whichis associatedwiththechronicityof HBVinfection.HBVinvasionintothe PBMCsmayaffectTh1andTh2cell-mediatedimmuneresponseof thepatientswithchronichepatitisB.

CD_4^(+) EFFECTOR CELLS DEFAULT TO THE TH2 PATHWAY IN INTERFERONγ-DEFICIENT MICE INFECTED WITH LEISHMANIA MAJOR

Mice with homologous disruption of the interferon γ(IFN-γ) gene on the C57BL/6 background were infected witly Leishmania major and the immune response assessed. In contrast to wild-type or heterozygous knockout mice,deficient animals were unable to restrict growth of the parasite and suffered lethal infection over 6￣8 wk.Although wild-type and heterozygous littermates developed CD4+ cells that contained transcripts for IFN-γ and lymphotoxin,tipical of T helper type 1(Th1) cells, the knockout mice developed CD+4 cells that contained transcripts for interleukin 4(IL-4),IL-5,and IL13,typical of Th2 cells. ELISPOT assays confirmed the reciprocal patterns of IFN-γ or IL-4 production by T cells in similar frequencies in the respective groups of mice,and antibody analysis confirmed the presence of Th2-mediated isotype switching in the knockout mice. These data suggest that CD4+ T cells that normally respond to antigens by differentiation to Th1 cells default to the Th2 pathway in the absence of endogenous IFN-γ.

ECT2通过调控PI3K/AKT信号通路对胰腺癌细胞恶性行为、糖酵解及TH细胞分化的影响

目的探讨上皮细胞转化序列2(ECT2)通过调控PI3K/AKT信号通路对胰腺癌细胞恶性行为、糖酵解及TH细胞分化的影响。方法RT⁃qRCR检测人脐静脉内皮细胞系HUVEC及胰腺癌细胞中ECT2 mRNA表达,将胰腺癌Panc⁃1细胞株分为胰腺癌组(Panc⁃1细胞株正常培养)、NC组(Panc⁃1细胞株转染ECT2阴性对照)、ECT2 siRNA组(Panc⁃1细胞株转染ECT2 siRNA)、抑制剂组(Panc⁃1细胞株转染加入PI3K/AKT抑制剂LY294002),ECT2 siRNA+抑制剂组(Panc⁃1细胞株转染ECT2 siRNA加入PI3K/AKT抑制剂LY294002),采用RT⁃qRCR各组细胞中ECT2 mRNA表达;Transwell法检测各组Panc⁃1细胞侵袭能力;划痕实验检测迁移;流式细胞仪检测各组细胞IFN⁃γ及IL⁃4表达;免疫印迹分别检测糖酵解代表蛋白及PI3K/AKT表达。结果胰腺癌组、NC组、ECT2 siRNA组、抑制剂组及ECT2 siRNA+抑制剂组的ECT2 mRNA比较分别为1.00±0.00、0.95±0.03、0.41±0.08、0.65±0.05及0.20±0.04,组间比较,差异有统计学意义(F=310.700,P<0.05);胰腺癌组、NC组、ECT2 siRNA组、抑制剂组及ECT2 siRNA+抑制剂组Panc⁃1细胞侵袭数目分别为(256.30±28.36)个、(241.18±24.05)个、(155.48±17.56)个、(178.90±18.44)个及(95.15±12.10)个,组间比较,差异有统计学意义(F=59.310,P<0.01);胰腺癌组、NC组、ECT2 siRNA组、抑制剂组及ECT2 siRNA+抑制剂组Panc⁃1细胞迁移距离分别为(14.02±1.36)mm、(13.42±1.29)mm、(9.25±0.85)mm、(8.50±0.45)mm及(4.25±0.53)mm,组间比较差异有统计学意义(F=101.200,P<0.05);ECT2 siRNA组细胞IFN⁃γ升高及IL⁃4占比降低,与NC组比较,差异有统计学意义(P<0.05),与抑制剂组相比,ECT2 siRNA+抑制剂组细胞IFN⁃γ升高及IL⁃4占比降低,组间比较差异有统计学意义(P<0.05);ECT2 siRNA组细胞HIF⁃1α、GLUT1、HK⁃Ⅱ和PFK蛋白均降低,与抑制剂组无意义(P>0.05),与抑制剂组相比,ECT2 siRNA+抑制剂组细胞HIF⁃1α、GLUT1、HK⁃Ⅱ和PFK蛋白降低,组间比较差异有统计学意义(P<0.05);ECT2 siRNA组细胞HIF⁃1α、GLUT1、HK⁃Ⅱ和PFK蛋白均降低,与NC组差异有统计学意义(P<0.05),与抑制剂组相比,ECT2 siRNA+抑制剂组细胞HIF⁃1α、GLUT1、HK⁃Ⅱ和PFK蛋白降低,组间比较差异有统计学意义(P<0.05)。结论抑制ECT2可通过降低胰腺癌细胞糖酵解,提高Th细胞中IFN⁃γ表达而抑制恶性侵袭及迁移,其机制可能与抑制PI3K/AKT信号通路活性相关。

Acute and persisting Th2-like immune response after fractionated colorectal γ-irradiation

AIM： To investigate if an immune imbalance may account for the development and progression of chronic radiation enteritis. We analyzed the Th1/Th2 immune response profile early and 6 mo after fractionated colorectal irradiation. METHODS： A rat model of fractionated colorectal γ-irradiation （4-Gy fractions, 3 fractions per week） was designed to investigate the effects of cumulative dose on inflammatory mediators （cytokines and chemo- kines） and immune response （Th1/Th2 profile and immunosuppressive mediator IL-10） during acute （early） response and 6 mo after the end of fractionated irradiation （chronic response）. Analyses were performed 1 d after the cumulative doses of 16 Gy and 36 Gy and 1 d, 3 d, and 26 wk after the cumulative dose of 52 Gy. RESULTS： Without causing histological damage, fractionated radiation induced elevated expression of IL-1β, TNFα, MCP-1, and iNOS in distal colonic mucosa during the early post-irradiation phase. At that time, a Th2 profile was confirmed by expression of both the Th2- specific transcription factor GATA-3 and the chemokine receptor CCR4 and by suppression of the Thl cytokine IFNγ/IP-10 throughout the irradiation protocol. After 6 mo, despite the 2-fold reduction of iNOS and MCP-1 levels, the Th2 profile persisted, as shown by a 50% reduction in the expression of the Thl transcription factor T-bet, the chemokine receptor CCXCR3, and the IFNγ/ STAT1 pathway. At the same time-point, the immunosuppressive IL-10/STAT3 pathway, known to regulate the Th1/Th2 balance, was expressed, in irradiated rats, at approximately half its level as compared to controls.This suppression was associated with an overexpression of SOCS3, which inhibits the feedback of the Thl polarization and regulates IL-10 production. CONCLUSION： Colorectal irradiation induces Th2 polarization, defective IL-10/STAT3 pathway activation and SOCS3 overexpression. These changes, in turn, maintain a immunological imbalance that persists in the long term.

Activation of natural killer T cells contributes to Th1 bias in the murine liver after 14 d of ethinylestradiol exposure

BACKGROUND As the main component of oral contraceptives(OCs),ethinylestradiol(EE)has been widely applied as a model drug to induce murine intrahepatic cholestasis.The clinical counterpart of EE-induced cholestasis includes women who are taking OCs,sex hormone replacement therapy,and susceptible pregnant women.Taking intrahepatic cholestasis of pregnancy(ICP)as an example,ICP consumes the medical system due to its high-risk fetal burden and the impotency of ursodeoxycholic acid in reducing adverse perinatal outcomes.AIM To explore the mechanisms and therapeutic strategies of EE-induced cholestasis based on the liver immune microenvironment.METHODS Male C57BL/6J mice or invariant natural killer T(iNKT)cell deficiency(Jα18-/-mice)were administered with EE(10 mg/kg,subcutaneous)for 14 d.RESULTS Both Th1 and Th2 cytokines produced by NKT cells increased in the liver skewing toward a Th1 bias.The expression of the chemokine/chemokine receptor Cxcr6/Cxcl16,toll-like receptors,Ras/Rad,and PI3K/Bad signaling was upregulated after EE administration.EE also influenced bile acid synthase Cyp7a1,Cyp8b1,and tight junctions ZO-1 and Occludin,which might be associated with EEinduced cholestasis.iNKT cell deficiency(Jα18-/-mice)robustly alleviated cholestatic liver damage and lowered the expression of the abovementioned signaling pathways.CONCLUSION Hepatic NKT cells play a pathogenic role in EE-induced intrahepatic cholestasis.Our research improves the understanding of intrahepatic cholestasis by revealing the hepatic immune microenvironment and also provides a potential clinical treatment by regulating iNKT cells.

Exposure to ephedrine attenuates Th1/Th2 imbalance underlying OVA-induced asthma through airway epithelial cell-derived exo-somal lnc-TRPM2-AS

Although various anti-inflammatory medications,such as ephedrine,are employed to manage cough-variant asthma,their underlying mechanisms are yet to be fully understood.Recent studies suggest that exosomes derived from airway epithelial cells(AECs)contain components like messenger RNAs(mRNAs),micro-RNAs(miRNAs),and long noncoding RNA(lncRNA),which play roles in the occurrence and progression of airway inflammation.This study investigates the influence of AEC-derived exosomes on the efficacy of ephedrine in treating cough-variant asthma.We established a mouse model of asthma and measured airway resist-ance and serum inflammatory cell levels.Real-time polymerase chain reaction(RT-qPCR),Western blotting,and enzyme-linked im-munosorbent assay(ELISA)analyses were used to assess gene and protein expression levels.Exosomes were isolated and character-ized.RNA immunoprecipitation(RIP)and RNA pull-down assays were conducted to examine the interaction between hnRNPA2B1 and lnc-TRPM2-AS1.In the ovalbumin(OVA)-challenged mouse model,ephedrine treatment reduced inflammatory responses,air-way resistance,and Th1/Th2 cell imbalance.Exosomes from OVA-treated AECs showed elevated levels of lnc-TRPM2-AS1,which were diminished following ephedrine treatment.The exosomal lnc-TRPM2-AS1 mediated the Th1/Th2 imbalance in CD4^(+)T cells,with its packaging into exosomes being facilitated by hnRNPA2B1.This study unveils a novel mechanism by which ephedrine ameli-orates OVA-induced CD4^(+)T cell imbalance by suppressing AEC-derived exosomal lnc-TRPM2-AS1.These findings could provide a theoretical framework for using ephedrine in asthma treatment.

Correlation of serum nutrient levels with immune cell differentiation and inflammatory response activation in children with pneumonia

Objective:To investigate the correlation of serum nutrient levels with immune cell differentiation and inflammatory response activation in children with pneumonia.Methods:200 children with pneumonia who were treated in our hospital between April 2015 and August 2017 were selected as the pneumonia group, and 100 healthy children who were vaccinated in this hospital during the same period were selected as the normal control group. The differences in serum levels of nutrients, Th1/Th2 cytokines, Th17/Treg cytokines and inflammatory mediators were compared between the two groups. Pearson test was used to assess the relationship between serum nutrient levels and disease severity.Results: Serum Vit A, Fe and Zn levels of pneumonia group were lower than those of control group. The differences in serum Vit D, Ca and Mg levels were not statistically significant between the two groups of children. Serum Th1 cytokines IL-2 and TNF-β contents of pneumonia group were lower than those of control group whereas Th2 cytokines IL-4 and IL-5 contents were higher than those of control group;Th17 cytokines IL-17, IL-21 and IL-22 contents were higher than those of control group whereas Treg cytokines IL-10 and TGF-β contents were lower than those of control group;inflammatory mediators CRP, PCT and MCP-1 contents were significantly higher than those of control group. Pearson test showed that serum nutrients Vit A, Fe and Zn levels in children with pneumonia were directly correlated with the degree of immune cell differentiation and inflammatory response.Conclusion: Serum Vit A, Fe, Zn and other nutrient levels abnormally decrease in children with pneumonia, and the specific level are directly correlated with the severity of the disease.

Silencing of <i>ERK2</i>with Small Interference RNA Regulates the Expression of CXCL1 in Th17 Cell

Patients with steroid-resistant asthma had their monocyte-derived TH17 cells collected. The expression levels of ERK2 in the TH17 were silenced and inhibited using ERK2 specific small interfering RNA (siRNA). By screening of CXCL1 and IL-17A in the TH17 culture supernatant, the expression levels of ERK2 and CXCL1 were determined. Using targeted siRNA to inhibit ERK2, the expression of ERK2 in the TH17 was reduced. Furthermore, inhibiting ERK2 hindered CXCL1 expression and decreased CXCL1 and IL-17A production. These findings suggest that ERK2 is involved in the synthesis of CXCL1 and IL-17A, two proteins that play a key role in the pathogenesis of hormone-resistant asthma.

全血培养检测Th_1/Th_2细胞

目的 建立全血培养刺激法检测外周血单核细胞 ( PBMC) Th1/Th2 的方法。方法 无菌抽取 2 0名健康检体者静脉血 ,分别按叶氏和该法 (先加入刺激剂全血培养 5 h后分离淋巴细胞 ,其余步骤按叶氏报道的免疫组化方法操作 )检测 Th1/Th2 细胞。结果 该方法与叶氏法相比 ,Th1/Th2 阳性细胞稍高。结论 该方法结果可靠 ,简便易行 ,适合临床实验室。

Th－LAK细胞与rIL－2联合治疗恶性骨肿瘤

应用胎儿胸腺淋巴细胞诱导制备Ｔｈ－ＬＡＫ细胞，与ｒＩＬ－２联合治疗中晚期恶性骨肿瘤７例。治疗前后动态检测外周血淋巴细胞、Ｔ淋巴细胞亚群、ＮＫ细胞活性。结果，患者的一般症状和免疫状态明显改善，化疗药物的毒副作用降低，病情缓解，无明显毒副反应。

Analysis of Th1/Th2 Response Pattern for Erythrodermic Psoriasis

As one of the most serious types of psoriasis, pathogenesis of erythrodermic psoriasis（EP） is unclear so far. In this study, we aimed to detect the levels of Th1/Th2 cytokine-associated transcription factors and T-lymphocyte clone in peripheral blood mononuclear cells（PBMCs） derived from EP patients, and gene expression level of T-bet/GATA-3 in skin lesion. The potential role of Th1/Th2 reaction pattern played in the pathogenesis of EP was also discussed. Serum levels of IFN-γ, IL-2, IL-4 and IL-10 were quantified by ELISA among 16 EP patients, 20 psoriasis vulgaris（PV） patients and 15 healthy controls. The expression levels of T-bet/GATA-3 in the skin lesion and PBMCs were examined by real-time qPCR. The ratio of Th1/Th2 was measured by flow cytometry. The levels of IFN-γ, IL-2, IL-4 and IL-10 were higher in EP patients than in the healthy controls. The levels of IL-4 and IL-10 were 69.44±11.45 and 12.62±4.57 pg/mL, respectively, in EP patients, significantly higher than those in PV patients and healthy controls（P〈0.05）. Flow cytometry revealed the levels of both Th1 and Th2 in PBMCs from EP patients were higher than those in healthy controls, and the Th1/Th2 ratio was dramatically lower than in PV patients（P〈0.01）. The ratios of IFN-γ/IL-4 and T-bet/GATA-3 in EP patients were both less than 1.0, suggesting a reversal when compared with the other two groups. Our study indicated that the EP patients exerted a Th1/Th2 bidirectional response pattern, and the balance of Th cell subsets inclines to Th2, which might be one of the important mechanisms of EP pathogenesis.