沉默线粒体核糖体蛋白L35基因对人食管癌TE-1细胞生长的影响

Effect of silencing mitochondrial ribosomal protein L35 gene on growth of human esophageal cancer TE-1 cells

目的:探讨慢病毒介导沉默线粒体核糖体蛋白L35 (MRPL35)基因对人食管癌TE-1细胞生长的影响,并阐明其机制。方法:选择3种人食管癌TE-1、ECA109和KYSE150细胞,采用实时定量PCR法检测3种细胞中MRPL35mRNA相对表达水平。食管癌TE-1细胞分为shMRPL35组和shCtrl组,分别加入沉默靶基因带有嘌呤霉素抗性的si-RNA慢病毒和带有嘌呤霉素抗性的阴性对照si-RNA慢病毒进行感染,建立稳定沉默MRPL35基因的食管癌细胞系,实时定量PCR及Western blotting法检测各组细胞中MRPL35基因沉默效率,采用CCK-8法检测各组细胞生长曲线,AnnexinⅤ-PE/7AAD双染色后流式细胞术检测细胞凋亡率。结果:3种食管癌细胞均表达MRPL35基因,3种细胞中MRPL35mRNA表达水平比较差异无统计学意义(P>0.05)。实时定量PCR法和Western blotting法检测,shMRPL35组TE-1细胞中MRPL35mRNA和蛋白表达水平明显低于shCtrl组(P<0.05)。与shCtrl组比较,shMRPL35组TE-1细胞生长速度明显降低(P<0.05),凋亡率明显升高(P<0.01)。结论:沉默MRPL35基因可抑制食管癌细胞TE-1增殖,并通过凋亡途径发挥作用。

Objective:To investigate the effect of lentivirus-mediated silencing of mitochondrial ribosomal protein L35(MRPL35)gene on the growth of human esophageal cancer TE-1 cells,and to clarify its mechanism.Methods:Three kinds of human esophageal cancer cells,TE-1,ECA109 and KYSE150,were selected.The relative expression levels of MRPL35 mRNA in three kinds of cells by real-time quantitative PCR.The esophageal cancer TE-1 cells were divided into shMRPL35 group and shCtrl group,and the cells were infected with si-RNA lentivirus and si-RNA lentivirus;the esophageal cancer cell line stably silenting the MRPL35 gene was established.Real-time quantitative PCR and Western blotting methods were used to detect the efficiency of MRPL35 gene silencing.The cell growth curves in various groups were detected by CCK-8 method,and the apoptotic rates were detected by flow cytometry after AnnexinⅤ-PE/7AAD doub le staining.Results:Three kinds of esophageal cancer cells expressed MRPL35 gene,and the expression levels were not statistically significant between them(P>0.05).The results of real-time quantitative PCR and Western blotting methods showed that the mRNA and protein levels of MRPL35 in the TE-1 cells in shMRPL35 group were significantly lower than those in shCtrl group(P<0.05).Compared with shCtrl group,the cell growth speed in shMRPL35 group was decreased(P<0.05),and the apoptotic rate was significantly increased(P<0.01).Conclusion:Silencing MRPL35 gene can inhibit the proliferation of esophageal cancer TE-1cells and plays a role through the apoptotic pathway.