RNA干扰抑制MGMT表达对人脑胶质瘤化疗的影响

Effect of Inhibiting MGMT Gene Expression by RNA Interference on the Chemosensitivity of Giloma Cell Line

目的应用小干扰RNA(siRNA)质粒载体抑制人脑胶质瘤T98细胞系中O6-甲基鸟嘌呤-DNA甲基转移酶(MGMT)基本的表达,了解其对胶质瘤细胞化疗敏感性的影响。方法通过LipofectimineTM2000将针对MG-MT的siRNA质粒载体转入T98细胞中。采用实时定量PCR法测定MGMT mRNA的表达,Western blot测定MGMT蛋白的表达。MTT法测定转染前后T98细胞系对卡氮芥(BCNU)的敏感性变化。结果成功构建针对MGMT基因的siRNA质粒载体;质粒载体转染T98细胞后,对其MGMT mRNA的抑制率达87%,MGMT蛋白表达量明显减少,转染后的T98细胞对BCNU的敏感性增加,提高了约6倍。结论针对MGMT基因的siRNA质粒载体可以靶向抑制MGMT基因在T98细胞系中的表达,增加T98细胞对BCNU的敏感性。

Objective To investigate the effect of combinative samll interference RNA（siRNA） vectors targeting O6-methylguanine DNA methyltransferase（MGMT） gene on chemosensitivity of the glioma cell line.Methods T98 cells with MGMT siRNA vectors were transfected by LipofectamineTM 2000.Real-time quantitative polymerase chain reaction was used to detect the expression of MGMT mRNA.Expression of MGMT protein was detected by Western blot.And MTT was used to assesse the sensitivity of T98 cell to alkylating agent.Results The combinative siRNA vectors were successfully constructed.MGMT mRNA and protein expression level was dramatically suppressed in the transfected T98 cells,which was 87%.The sensitivity of T98 cells to BCNU had increased after transfected with MGMT siRNA vectors,which was 6 times.Conclusion RNA interference mediated by the combinative siRNA vectors can significantly down-regulate the expression of MGMT in glioma cell line T98 and increases its chemosensitivity to BCNU.