日本血吸虫热休克蛋白60通过诱导CD4~+CD25~+Treg细胞表达IL-10和TGF-β增强其免疫抑制功能
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摘要
目的探讨日本血吸虫热休克蛋白60(Sj HSP60)增强小鼠CD4+CD25+调节性T细胞(Treg)免疫抑制功能的可能机制。方法采用体外研究探索Sj HSP60对Treg细胞功能的影响,通过体外细胞增殖实验检测Sj HSP60促进Treg细胞免疫抑制功能增强的途径,利用流式细胞术检测Treg细胞表达IL-10和TGF-β的水平以及Treg细胞Foxp3和CTLA-4表达水平。结果体外实验证实,Sj HSP60可增强Treg细胞的免疫抑制功能(t=6.466,P=0.006);体外细胞增殖实验表明,Sj HSP60刺激后的Treg细胞可通过分泌可溶性细胞因子IL-10(t=7.363,P=0.002)和TGF-β(t=11.262,P=0)发挥免疫抑制功能;流式细胞术检测显示,Sj HSP60刺激后的Treg细胞IL-10(t=12.367,P=0)、TGF-β(t=8.431,P=0.001)、Foxp3(比例:t=5.225,P=0.006;平均荧光强度:t=8.319,P=0.001)和CTLA-4(比例:t=3.518,P=0.024;平均荧光强度:t=4.164,P=0.014)表达水平显著增加。结论 Sj HSP60可通过促进Treg细胞表达IL-10和TGF-β增强其免疫抑制功能,并可能与Sj HSP60诱导Treg细胞表达Foxp3和CTLA-4有关。
Objective To explore the possible mechanisms by which Schistosoma japonicum heat shock protein 60(SjHSP60)enhances CD4+CD25+regulatory T cell(Treg)immunosuppressive function. Methods An in vitro method was used to investigate the effect of Sj HSP60 on Treg immunosuppressive activity. Co-cultures in transwells and in vitro suppression assay were performed to investigate how Sj HSP60 enhanced the immunosuppressive function of Tregs. Intracellular cytokine staining combined with flow cytometry was used to detect Treg-expressing IL-10 and TGF-β,and flow cytometry was also used to analyze the expressions of Foxp3 and CTLA-4 in Tregs. Results Sj HSP60 enhanced the immunosuppressive function of Tregs. Soluble cytokines IL-10 and TGF-β mediated inhibitory activity of Sj HSP60-triggered Tregs. Sj HSP60 induced significant increases in both IL-10 and TGF-β expressions of Tregs. Further investigation showed significant increased Foxp3 and CTLA-4 in Sj HSP60-trggered Tregs. Conclusion Sj HSP60 enhances Treg immunosuppressive function by promoting the expressions of IL-10 and TGF-β,possibly due to SjHSP60-mediated induction of Foxp3 and CTLA-4 in Tregs.
Objective To explore the possible mechanisms by which Schistosoma japonicum heat shock protein 60(SjHSP60)enhances CD4+CD25+regulatory T cell(Treg)immunosuppressive function. Methods An in vitro method was used to investigate the effect of Sj HSP60 on Treg immunosuppressive activity. Co-cultures in transwells and in vitro suppression assay were performed to investigate how Sj HSP60 enhanced the immunosuppressive function of Tregs. Intracellular cytokine staining combined with flow cytometry was used to detect Treg-expressing IL-10 and TGF-β,and flow cytometry was also used to analyze the expressions of Foxp3 and CTLA-4 in Tregs. Results Sj HSP60 enhanced the immunosuppressive function of Tregs. Soluble cytokines IL-10 and TGF-β mediated inhibitory activity of Sj HSP60-triggered Tregs. Sj HSP60 induced significant increases in both IL-10 and TGF-β expressions of Tregs. Further investigation showed significant increased Foxp3 and CTLA-4 in Sj HSP60-trggered Tregs. Conclusion Sj HSP60 enhances Treg immunosuppressive function by promoting the expressions of IL-10 and TGF-β,possibly due to SjHSP60-mediated induction of Foxp3 and CTLA-4 in Tregs.
引文
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[2]Sakaguchi S,Yamaguchi T,Nomura T,et al.Regulatory T cells and immune tolerance[J].Cell,2008,133(5):775-787.
[3]Taylor MD,van der Werf N,Maizels RM.T cells in helminth infection:the regulators and the regulated[J].Trends Immunol,2012,33(4):181-189.
[4]Finlay CM,Walsh KP,Mills KH.Induction of regulatory cells by helminth parasites:exploitation for the treatment of inflammatory diseases[J].Immunol Rev,2014,259(1):206-230.
[5]Maizels RM,Yazdanbakhsh M.T-cell regulation in helminth parasite infections:implications for inflammatory diseases[J].Chem Immunol Allergy,2008,94:112-123.
[6]Bashi T,Bizzaro G,Ben-Ami Shor D,et al.The mechanisms behind helminth’s immunomodulation in autoimmunity[J].Autoimmun Rev,2015,14(2):98-104.
[7]Dunne DW,Cooke A.A worm’s eye view of the immune system:consequences for evolution of human autoimmune disease[J].Nat Rev Immunol,205,5(5):420-426.
[8]Vignali DA,Collison LW,Workman CJ.How regulatory T cells work[J].Nat Rev Immunol,2008,8(7):523-532.
[9]金鑫,陈晓军,朱继峰,等.日本血吸虫热休克蛋白60 k Da的免疫原性及保护性研究[J].中国血吸虫病防治杂志,2016,28(1):45-50.
[10]Wang X,Zhou S,Chi Y,et al.CD4+CD25+Treg induction by an HSP60-derived peptide SJMHE1 from Schistosoma japonicum is TLR2 dependent[J].Eur J Immunol,2009,39(11):3052-3065.
[11]Hossain DM,Panda AK,Manna A,et al.Fox P3 acts as a cotranscription factor with STAT3 in tumor-induced regulatory T cells[J].Immunity,2013,39(6):1057-1069.
[12]Hryniewicz A,Boasso A,Edghill-Smith Y,et al.CTLA-4 blockade decreases TGF-beta,IDO,and viral RNA expression in tissues of SIVmac251-infected macaques[J].Blood,2006,108(12):3834-3842.
[13]Chen W,Jin W,Wahl SM.Engagement of cytotoxic T lymphocyteassociated antigen 4(CTLA-4)induces transforming growth factor beta(TGF-beta)production by murine CD4+T cells[J].J Exp Med,1998,188(10):1849-1857.
[14]Mc Sorley HJ,Maizels RM.Helminth infections and host immune regulation[J].Clin Microbiol Rev,2012,25(4):585-608.
[15]Zhou S,Jin X,Chen X,et al.Heat shock protein 60 in eggs specifically induces Tregs and reduces liver immunopathology in mice with schistosomiasis japonica[J].PLo S One,2015,10(9):e0139133.
[16]Di Ianni M,Falzetti F,Carotti A,et al.Tregs prevent GVHD and promote immune reconstitution in HLA-haploidentical transplantation[J].Blood,2011,117(14):3921-3928.
[17]Masteller EL,Warner MR,et al.Expansion of functional endogenous antigen-specific CD4+CD25+regulatory T cells from nonobese diabetic mice[J].J Immunol,2005,175(5):3053-3059.
[18]Marek-Trzonkowska N,Mysliwiec M,Dobyszuk A,et al.Therapy of type 1 diabetes with CD4+CD25highCD127-regulatory T cells prolongs survival of pancreatic islets-results of one year follow-up[J].Clin Immunol,2014,153(1):23-30.
[19]Desreumaux P,Foussat A,Allez M,et al.Safety and efficacy of antigen-specific regulatory T-cell therapy for patients with refractory Crohn’s disease[J].Gastroenterology,2012,143(5):1207-1217.
[20]Trzonkowski P,Bacchetta R,Battaglia M,et al.Hurdles in therapy with regulatory T cells[J].Sci Transl Med,2015,7(304):304-318.
[21]Sakaguchi S,Miyara M,Costantino CM,et al.FOXP3+regulatory T cells in the human immune system[J].Nat Rev Immunol,2010,10(7):490-500.
[22]Buckner JH.Mechanisms of impaired regulation by CD4+CD25+FOXP3+regulatory T cells in human autoimmune diseases[J].Nat Rev Immunol,2010,10(12):849-859.
[23]Shevach EM,Mc Hugh RS,Piccirillo CA,et al.Control of T-cell activation by CD4+CD25+suppressor T cells[J].Immunol Rev,2001,182:58-67.
[24]Jonuleit H,Schmitt E,Stassen M,et al.Identification and functional characterization of human CD4+CD25+T cells with regulatory properties isolated from peripheral blood[J].J Exp Med,2001,193(11):1285-1294.