抑制STAT5A基因的表达对人肝癌HepG2细胞增殖的影响及其机制研究  

The influence of inhibiting STAT5A expression on proliferation of human hepatocellular carcinoma cell line HepG2 and its mechanism

作　　者：杨华秀[1] 曾永秋[2] 曹洋[3] 黄燕[2] 李洁[2] 

机构地区：[1]泸州医学院附属中医院肝病科,四川泸州646000 [2]泸州医学院医学细胞生物学与遗传学教研室,四川泸州646000 [3]泸州医学院生理学教研室,四川泸州646000

出　　处：《泸州医学院学报》2012年第6期559-563,共5页Journal of Luzhou Medical College

基　　金：四川省教育厅重点项目(07ZC028)

摘　　要：目的:研究抑制STAT5A基因的表达对人肝癌HepG2细胞增殖的影响,并对其影响肝癌细胞增殖的机制进行探讨,为STAT5A基因在肝细胞癌中的功能研究提供实验依据。方法:构建针对STAT5A基因的shRNA真核表达载体,脂质体法转染人肝癌细胞HepG2,采用半定量RT-PCR的方法检测细胞中STAT5A mRNA的变化;蛋白质印迹技术检测转染前后细胞中STAT5A蛋白、细胞周期蛋白Cyclin D1蛋白表达的变化;采用MTT法检测细胞增殖的情况。结果:酶切及测序结果显示shRNA表达载体均构建成功,转染HepG2细胞48h后,与对照组相比,RT-PCR结果显示实验组细胞中STAT5A mRNA的抑制率为72.03%(P<0.05),蛋白质印迹法显示STAT5A蛋白的抑制率为66.27%(P<0.05),细胞周期蛋白Cyclin D1蛋白表达下降47.16%(P<0.05),同时MTT法显示细胞生长速度减慢、增殖抑制明显。结论:抑制STAT5A基因的表达可有效抑制HepG2细胞的增殖,其机制可能是通过下调Cyclin D1的表达实现的。Objective： To study the effect of suppression of expression of STAT5A gene on proliferation of human hepatocellular carcinoma cell line HepG2.Methods： The RNA interference eukaryotic expression vector specific for STAT5A gene was constructed and transfected into HepG2 cells by the liposome method.The expression of STAT5A mRNA was detected by semi-quantitative RT-PCR,and the changes of STAT5A and Cyclin D1 protein were detected by Western blotting.MTT method was used to detect cell proliferation.Results： The RNA interference eukaryotic expression vectors specific for STAT5A gene was confirmed to be successfully constructed by restriction enzymes and DNA sequencing.Compared with control group,the inhibitory ratios of STAT5A mRNA in experimental group was 72.03%,and the STAT5A protein and Cyclin D1 protein was 66.27% and 47.16% respectively 48h after transfection（P0.05）.The result of MTT showed that the cell growth velocity of the experiment group stepped down and the proliferation inhibition rate was inhibited significantly（P〈0.05）.Conclusion： Inhibiting the expression of STAT5A gene can inhibit the proliferation of HepG2 cell efficiently,and its mechanism may be downregulating the level of Cyclin D1 protein.

关 键 词：STAT5A基因 RNA干扰 HEPG2细胞 细胞增殖 CYCLIN D1 

分 类 号：R735.7[医药卫生—肿瘤]