知识与财富的链接
目的:探讨转录因子维A酸相关孤独受体γt(RORγt)对T细胞亚群产生IFN-γ的调控作用。方法:健康成人外周血单个核细胞用CD3单克隆抗体和IL-2激活和扩增T细胞,活化T细胞用脂质体法转染人工合成的RORγt基因特异性短干扰RNA(siRNA)序列3个片段(RORγt-siRNA-982,-1026,-1197),RT-PCR半定量法检测活化T细胞转染siRNA后RORγt基因表达,流式细胞仪检测不同T细胞亚群产生IFN-γ细胞的比例。结果:活化T细胞转染RORγt-siRNA后,对RORγt基因表达的检测显示,RORγt-siRNA-982序列无抑制作用,而RORγt-siRNA-1026和-1197序列均有明显抑制作用。活化T细胞转染RORγt-siRNA-1026后,产生IFN-γ细胞百分数在CD4+ T细胞(32.78±3.41)中,明显低于未转染组(44.54±1.75)(P<0.01)。结论:转录因子RORγt对T细胞,特别是对CD4+ T细胞产生IFN-γ有正调控作用。
Objective：To investigate the regulation of transcription factor RORγt on the IFN-γ production in different T cell subpopulations.Methods：Peripheral blood mononuclear cells were isolated from peripheral blood of healthy donors by density gradient centrifugation,and were stimulated with CD3 monoclonal antibody and cultured in IL-2 containing medium for 12 days.The activated T cells were transfected with chemical synthesized small interfering RNA（siRNA）sequences for RORγt gene using liposomes.The expression of RORγt gene was detected in RORγt-siRNA transfected T cells by RT-PCR assay.The percentages of IFN-γ producing cells in total T cells,CD4^＋ and CD8^＋ T cells were measured in the transfected T cells by flowcytometry.Results：By using semi-quantity RT-PCR assay,the RORγt gene expression was inhibited in activated T cells that transfected with RORγt-siRNA-1026 and-1197,but not inhibited in that with RORγt-siRNA-982.Among the RORγt-siRNA-1026 transfected activated T cells,the percentage of IFN-γ producing cells in CD4^＋ T cells（32.78±3.41）significantly decreased,compared to untransfected group（44.54±1.75）（P〈0.01）.Conclusions：Transcription factor RORγt is involved in the positive regulation for IFN-γ production in CD4^＋ T cells.
