miR-155对HBV蛋白表达的抑制作用及机制  

Inhibitory effect of miR-155 on expression of hepatitis B virus and its mechanism

作　　者：蔡启茵 任广立[1] 张卫云[1] 冯宇鹏[1] 马恒颢[1] 

机构地区：[1]广州军区广州总医院儿科,广州510010

出　　处：《解放军医学杂志》2015年第11期902-905,共4页Medical Journal of Chinese People's Liberation Army

基　　金：广州市科技计划项目(2013J4100116)~~

摘　　要：目的观察mi R-155对SOCS1 m RNA及蛋白水平的影响,探究mi R-155对HBs Ag、HBe Ag表达的抑制作用。方法从Hep G2.2.15细胞中提取基因组,经PCR扩增得到mi R-155前体序列,纯化并连接入pm R-m Cherry载体,构建重组质粒pmi R-155,经去内毒素后转染至Hep G2.2.15细胞。将细胞分为重组组(转染pmi R-155质粒)、空载组(转染pm Rm Cherry质粒)、转染试剂组、空白组。采用荧光实时定量PCR检测各组细胞内mi R-155的表达,RT-PCR检测各组细胞SOCS1 m RNA的表达,Western blotting检测各组细胞SOCS1 蛋白的表达,ELISA检测各组细胞HBs Ag、HBe Ag的表达。结果荧光实时定量PCR结果显示,以空白组细胞内mi R-155的表达量为基准,重组组mi R-155表达量(519.43±52.10)明显高于空载组(24.24±16.70)及转染试剂组(35.04±26.09,P<0.05);RT-PCR结果显示,重组组SOCS1 m RNA表达量(0.63±0.91)显著低于空白组(P<0.05);Western blotting结果显示,重组组SOCS1蛋白表达量显著低于空白组。以空白组的表达量为基准,重组组HBs Ag和HBe Ag蛋白表达的抑制率分别为55.62%±3.77%和47.87%±2.46%(P<0.01)。结论过表达的mi R-155可抑制细胞内SOCS1和HBV蛋白的表达。Objective To investigate the mechanism of mi R-155 regulating SOCS1 in the inhibition of HBs Ag and HBe Ag. Methods The pre-mi R-155 was amplified from total DNA of Hep G2.2.15 by PCR. The target gene fragment was digested and cloned into the pm R-m Cherry plasmid. Pmi R-155 was transfected into Hep G2.2.15 cells（recombinant group） by liposome-mediated method. The empty plasmid, the reagent group and untreated cells were set as control. Firstly the expression of mi R-155 was detected by the real-time quantitative PCR. Secondly, the expression of SOCS1 m RNA was detected by RT-PCR, and then the expression of SOCS1 protein was determined by Western blotting. Finally, the expression of HBs Ag and HBe Ag was determined by ELISA. Results The pmi R-155 eukaryotic over-expression vector was successfully constructed. Mi R-155 level of Hep G2.2.15 cells which was transfected with the recombinant plasmid（519.43±52.10） was obviously higher than those of the empty plasmid（24.24±16.70） and reagent groups（35.04±26.09, P0.05）. RT-PCR showed the expression of SOCS1 m RNA was lower in recombinant group than in untreated group. The expression of SOCS1 protein markedly decreased as shown with Western blotting. Over-expression of mi R-155 could inhibit the expressions of HBs Ag and HBe Ag（55.62±3.77）% and（47.87±2.46）%（P0.01） respectively. Conclusions Over-expression of mi R-155 can down regulate the expression of SOCS1, and inhibit the expressions of HBs Ag and HBe Ag.

关 键 词：乙型肝炎 慢性 微RNAS 乙型肝炎表面抗原 乙型肝炎核心抗原 

分 类 号：R512.6[医药卫生—内科学]