RNA干扰抑制人肝癌细胞N-ras基因表达的研究

Suppression of N-ras expression by interference RNA in BEL7402 hepatocellular carcinoma cells

目的研究RNA干扰(RNAi)对BEL7402细胞的N-ras基因表达的干扰效应。方法建立装载靶向N-ras基因小干扰性RNA(siRNA)的表达载体;用脂质体法将此表达载体转染BEL7402细胞株,以转染空载细胞作为对照;采用RT-PCR和Western blot检测N-ras基因的表达;流式细胞仪检测细胞凋亡情况。结果转染siRNA的表达载体可以抑制内源N-ras基因在转录和转译上的表达。N-ras基因的表达抑制与BEL7402细胞的凋亡相关联。结论转染siR-NA的表达载体可明显抑制肝癌细胞株BEL7402N-ras基因的表达,并伴有细胞的凋亡。其工作的开展将为RNAi治疗、肝癌N-ras基因功能研究、新药开发提供研究基础。

Objective To study the inhibitory effect of RNA interference （RNAi） on N-ras expression in hepatocellular carcinoma cell line, BEL7402. Methods Expression vector of N-ras gene targeting small interference RNA （siRNA） was constructed （psilencer-l-N-ras） and transfected into BEL7402. cells by lipofectamine, and the unloaded vector was used as control （mock）. The expression of N-ras mRNA and protein was identified by RT - PCR and Western blot. Apoptosis of the transfected cells was examined by flow cytometry. Results After BEL7402 cells were transfected with psilencer2-N-ras,the expression of-N-ras mRNA and protein was suppressed. Apoptosis was identified in the transfected BEL7402 cells. Conclusion The expression of N-ras at transcriptional and translational levels in BEL7402 cells transfected with siRNA is markedly inhibited, which may be associated with the induction of apoptosis.