HCV核心蛋白对HepG2细胞microRNA表达谱的影响  

The effects of HCV core protein on the expression of microRNAs in HepG2 cells

作　　者：吴燕[1] 曹炬[1] 黄世峰[1] 文阳安[1] 张莉萍[1] 

机构地区：[1]重庆医科大学附属第一医院检验科,重庆400016

出　　处：《基础医学与临床》2012年第6期618-622,共5页Basic and Clinical Medicine

基　　金：国家自然科学基金(30973378;81071621);重庆市自然科学基金(CSTC2009BB5257;2010BB5390);重庆医科大学附属第一医院院内医学科学基金(YXJJ2009-12)

摘　　要：目的分析HCV核心蛋白(HCV-Core)对HepG2细胞microRNA表达谱的影响。方法将构建好的重组真核表达质粒pCDNA3.1(+)/HCV-Core用脂质体转染HepG2细胞,经G418筛选得到稳定转染HCV-Core的HepG2细胞,命名为HepG2-HCV Core细胞系。然后用反转录聚合酶链反应(RT-PCR),细胞免疫荧光和Western blot对HCV核心蛋白在HepG2细胞里mRNA和蛋白水平的表达情况进行验证。利用博奥生物公司的miRNA Array基因芯片,分析稳定表达HCV核心蛋白的HepG2细胞和转染空质粒pCDNA3.1(+)的HepG2细胞二者microRNA表达谱的差异,最后用Taqman探针荧光定量PCR法进行验证。结果感染HCV-Core后,HepG2细胞表达有差异的microR-NA有8种,用Taqman探针荧光定量PCR法进行分析,发现HCV核心蛋白可以上调miR-29、miR-146a、miR-149、miR-192、miR-221、miR-222和miR-193b,下调miR-196a。结论 HCV核心蛋白可以诱导肝细胞microRNA表达谱发生改变,在肝癌发生发展过程中发挥重要作用。Objective To investigate the effects of HCV core protein（HCV-Core）on the expression of microRNAs in HepG2 cells.Methods pCDNA3.1（＋）/HCV-Core eukaryotic expression plasmid was transfected into HepG2 cells.The levels of mRNA and protein expression of HCV core protein in HCV-Core transfected HepG2 cells were validated by RT-PCR,immunohistochemistry and Western blot.The differences of microRNAs expression between pCDNA3.1（＋）/HCV-Core and pCDNA3.1（＋）transfected HepG2 cells were analyzed by miRNA Array GeneChip（Affymetrix,USA）,followed by the verification by real-time tagman-fluoresence quantitative RT-PCR.ResultsmiRNA array analysis demonstrated that there were eight microRNAs altered in HepG2 cells transfected with HCV core protein,and quantitative RT-PCR confirmed that miR-29,miR-146a,miR-149,miR-192,miR-221,miR-222,miR-193b were up-regulated,while miR-196a was down-regulated.Conclusion HCV core protein induced up-regulation or down-regulation of some microRNAs in HepG2 cells,which may contribute to the pathogenesis of hepatocellular carcinoma.

关 键 词：HCV核心蛋白 HEPG2细胞 MICRORNAS 基因芯片 

分 类 号：R512.63[医药卫生—内科学]