靶向抑制滑膜细胞TNF-α表达的siRNA筛选及鉴定  

Screening and identification of siRNA-mediated TNF-α expression in synovial cells of Human

作　　者：沈孝龙[1] 廖琦[1] 郝亮[1] 

机构地区：[1]南昌大学第二附属医院骨科,南昌330006

出　　处：《重庆医学》2014年第31期4230-4231,4234,共3页Chongqing medicine

基　　金：江西省自然(青年)科学基金项目(2010GZY0250)

摘　　要：目的设计并筛选能高效抑制创伤性关节炎滑膜细胞肿瘤坏死因子α(TNF-α)表达的最佳干扰序列,为进一步研究TNF-α基因对创伤性关节炎的临床应用奠定基础。方法根据人源TNF-αmRNA的序列,设计合成3组不同的且能干扰TNF-α基因表达的小型干扰RNA(siRNA)序列,将其转染人创伤性关节炎滑膜细胞,与阴性对照组(NC组)比较。培养48h后,采用实时灾光定量PCR(RT-PCR)检测滑膜细胞TNF-αmRNA表达水平,采用ELISA法检测细胞上清液TNF-α表达水平。结果与NC组相比,3组siRNA中有2组siRNA可有效使人滑膜细胞中TNF-αmRNA表达减少(P<0.01);同时细胞上清液中TNF-α分泌量下降,与NC组比较差异有统计学意义(P<0.01)。结论成功设计合成了特异且高效阻断TNF-α表达的siRNA。Objective To design and identify small interfering RNA （siRNA） targeting tumor necrosis factor‐α （TNF‐α） ex‐pression ,siRNAs were electroporated into synovial cells of Human to screen those which can effectively suppress TNF‐α expres‐sion .Methods Three TNF‐α specific double stranded siRNA were designed targeting different regions of TNF‐α mRNA（compared with negative control group） .RT‐PCR and Elisa were applied to detect TNF‐α ,mRNA expression and the secretion of TNF‐α in cell supernatant ,respectively .Results Two of the three customized TNF‐α siRNA could inhibit the expression of TNF‐α mRNA in synovial cells of Human （P〈 0 .01） .At the same time ,the secretion of TNF‐α decreased in cell supernatant ,the difference was sig‐nificant statistically compared with the control group（P〈 0 .01） .Conclusion TNF‐α siRNA can be successfully designed and syn‐thesized ,which can specifically and effectively suppress TNF‐α mRNA expression .

关 键 词：肿瘤坏死因子Α RNA 小分子干扰 滑膜细胞 创伤性关节炎 

分 类 号：R346[医药卫生—基础医学]