【摘要】 目的研究系统性红斑狼疮(SLE)相关基因IFIT1的功能。方法首先将IFIT1从穿梭表达克隆转入真核表达克隆,然后转导进入Jurkat细胞系,用有限稀释法筛选单克隆;再用CD3和CD28刺激单克隆,检测膜分子的表达、细胞凋亡、增生、细胞因子分泌等免疫生物学指标;最后应用RNAi技术,下调IFIT1的表达,检测细胞因子分泌,比较分析IFIT1的功能。结果IFIT1的过表达在T细胞中能诱导白细胞介素(IL)-4和IL-10的分泌增加,下调IFIT1的转录水平后能部分下调IL-4和IL-10表达,而与细胞膜分子的表达、细胞的增生、凋亡无关。结论SLE相关新基因IFIT1在细胞因子的产生中起了重要作用,可能与Th1/Th2的失衡有关。更多还原

【Abstract】 Objective To study the function of IFIT1, a novel gene associated with the systemic lupus erythematosus (SLE). Methods First we generate an expression clone of IFIT1 by performing an LR recombination reaction between the entry clone from Genecopoiea and a GatewayTM destination vector. Then transfect the vector into the Jurkat T cells. Single clone cells expressing EGFP and IFIT1-EGFP were selected by standard limiting dilution method. The stable transfectants of EGFP and IFIT1-EGFP were stimulated with anti-CD3 and anti-CD28. Cell surface antigen expression were dectected by flow cytometry. The apoptosis and expansion of each transfectant were detected according to the manufacture′s instruction. The culture supernatants were assayed for cytokines by sandwich ELISA. Furthermore transfection of psilence vector which could produce anti-ifitl-specifics small RNA duplexes partially blocked the expression of overexpressed ifitl in Jurkat T cell. The same assays were done by stimulation with anti-CD3 and anti-CD28. Results There were no significant difference in the expression level of the various cell surface molecules, apoptosis and expansion among Jurkat, Jurkat/EGFP, and Jurkat/IFIT1-EGFP transfectants. But we find the Jurkat/IFIT1-EGFP can enhance the production of IL-4 and IL-10. After knock down the expression of IFIT1 by RNAi, the levels ofIL-4 and IL-10 production were partially decreased. Conclusion The results indicate the SLE related-geneIFIT1 may play important role in the production of cytokines and interfere the Th1/Th2 balance.更多还原