中国免疫学杂志2011，Vol.27Issue(8)：681-685,695,6.DOI:10.3969/j.issn.1000-484X.2011.08.002
siRNA沉默抗原递呈细胞表面CD86的表达对T淋巴细胞激活的影响
Effect of silencing CD86 expression on antigen presenting cells by siRNA on activation of T lymphocytes
摘要
Abstract
Objective:To investigate the effect of silencing CD 86 expression on antigen presenting cells (APC )by siRNA on proliferation and interleukine(IL)-10,IFN-γ production of Tlymphocytes,and provide new methods for transplant rejection and autoimmune disease.Methods.Three siRNA sequences used for targeted silencing of human CD 86 were designed and chemically synthesized,the transferred into the Raji cells by LipofectamineTM 2000;24h,48h and 72h after transfection,expression of CD 86 on Raji cells was assessed by flow cytometry (FCM),and the mRNA expression was analyzed by fluorescent quantitation PCR;Inhibition the expression of CD 86 on Raji cells by siRNA,blocking CD 86 molecule by anti-CD86 monoclonalantibody and combination of the two methods that result in blocking CD 86-CD 28 signal pathway,the proliferation and IL-10,IFN-γ production of Tlymphocytes were evaluated by MTT assay and ELISA.Results:The results of FCM showed:natural CD 86 expression on Raji cells was 98.0％;24 h after siRNA transfection,only siRNA-2 showed apparent suppression effect,CD 86 expression was 61.7％,inhibition ration was 37.0％;48 h after transfection,CD 86 expression of control and siRNA-1 groups was un changed,that of siRNA-2 and siRNA-3 groups was 39.6％and 72.6％ respectively,inhibition ratio was 59.6％ and 25.9％ respectively;72h after transfection,CD86 expression of each group was:control,90.4％;siRNA-1,83.1％;siRNA-2,15.2％;siRNA-3,46.2％;inhibition ratio was 7.6％,15.2％,84.5％ and 52.8％ respectively.The results of fluorescent quantitation PCR showed:72 h after transfection,relative expression of CD 86 mRNA of siRNA-2 AND siRNA-3 groups had significant deviation with normal group,inhibition ratio was 78.7％ and 45.9％ respectively. Suppression CD 86 expression on Raji cells by siRNA-2 can effectively inhibit the activation,proliferation and IL-10 ,IFN-γ production of Tlymphocytes as well as anti-human CD 86 monoclonal antibody,and they had synergistic effect on Tlymphocyte suppression.Conclusion:Silencing CD 86 expression on APC by siRNA can block CD 86/ CD 28 costimulatory pathway,inhibit the activation,proliferation and IL-10,IFN-γ production of Tlymphocytes,impair the response of Tlymphocytes and induce immune tolerance.关键词
CD86/siRNA/免疫耐受/共刺激分子Key words
CD 86/small interfering RNA/Immune torlerance/Cosimulatory molecules分类
医药卫生引用本文复制引用
黄莉,孙杰,郭静雅,高增燕,邱玉华..siRNA沉默抗原递呈细胞表面CD86的表达对T淋巴细胞激活的影响[J].中国免疫学杂志,2011,27(8):681-685,695,6.基金项目
本文受国家科技部"重大新药创制"科技重大专项(2009ZX09103-705)资助 （2009ZX09103-705）
