靶向cyclinE小干扰RNA对肝癌HepG2细胞增殖和凋亡的影响

Effects of small RNA interference to target cyclinE gene on apoptosis and proliferation of HepG2 hepatocellular carcinoma cells

目的:以cyclinE基因编码区为靶位,构建表达小干扰RNA(siRNA)的质粒载体,观察转染后对HepG2细胞的影响.方法:针对cyclinE基因序列构建表达siRNA的真核表达载体pSilencer3.1-H1hygro,利用脂质体Metafectene转染CyclinE基因高表达的肝癌细胞株HepG2.流式细胞仪检测细胞周期及凋亡率,MTT法检测细胞增殖活性,RT-PCR和Western blot法观察转染后细胞cyclinE基因表达.结果:成功构建了表达siRNA的真核质粒载体,转染后cyclinE基因mRNA及蛋白表达水平分别下降了79%和65%.结论:靶向cyclinE基因的siRNA可有效沉默HepG2细胞高表达的cyclinE基因,从而抑制肝癌细胞的增殖并促进凋亡.

AIM： To design pSilencer3.1-Hlhygro plasmid expressing small interfering RNA （siRNA） that targets the coding region of cyclinE, and to evaluate the inhibitory effect of this siRNA on HepG2 cells after transfection. METHODS： pSilencer3.1-Hlhygro vector of the siRNA based on the coding region of cyclinE was constructed. The plasmid and liposome metafectene were cotransfected into HepG2 cells, which overexpressed cyclinE. Cell proliferation and apoptosis were assessed by MTT assay and flow cytometry, respectively, and expression of cyclinE was detected by RT-PCR and Western blotting. RESULTS： The plasmid expressing siRNA targeted on the coding region of cyclinE was successfully constructed. Expression of cyclinE mRNA and protein was suppressed by up to 79% and 65%, respectively. CONCLUSION： siRNA targeted on cyclinE can silence cyclinE overexpression, and subsequently suppress proliferation and promote apoptosis of HepG2 cells.