微小RNA-518b对肝癌细胞株增殖凋亡及侵袭的影响及其机制  

The mechnism and effect of microRNA-518b on cell proliferation, apoptosis and invasion in hepatocellular carcinoma cell

作　　者：张日新[1] 刘磊[2] 肖朝文[1] 李凯[1] 郑直[1] 朱岭[1] Zhang Rixin;Liu Lei;Xiao Chaowen;Li Kai;Zheng Zhi;Zhu Ling(Department of Hepatobiliary & Pancreatic Surgery,the Central Hospital of Wuhan,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430014,China;Department of GastrointestinalSurgery,the Central Hospital of Wuhan,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430014,China)

机构地区：[1]华中科技大学同济医学院附属武汉市中心医院肝胆胰外科,430014 [2]华中科技大学同济医学院附属武汉市中心医院胃肠外科,430014

出　　处：《中华实验外科杂志》2019年第7期1208-1210,共3页Chinese Journal of Experimental Surgery

摘　　要：目的研究微小RNA(miRNA,miR)-518b对肝癌细胞增殖、凋亡及侵袭的影响.方法实时定量反转录聚合酶链反应(RT-qPCR)检测miR-518b在肝癌和正常肝细胞株的表达.HepG2细胞株分成miR-518b过表达组(miR-518b组)、阴性对照组(miR-Con组)及空白对照组(Blank组).噻唑蓝(MTT)测定增殖,流式细胞术测定凋亡,Transwell测定侵袭.分光光度比色法测定半胱氨酰天冬氨酸特异性蛋白酶(Caspase)-3、Caspase-8、Caspawe-9活性,蛋白质印迹法(Westernblot)测定Rap1b蛋白表达,计量资料以均值±标准差(Mean±SD)表示,组间比较采用t检验.结果miR-518b在肝癌细胞株中表达下调.转染后72、96h,miR-518组吸光度(A)490nm低于miR-Con组(0.66±0.07比0.94±0.08,t=3.291,P<0.05)及(0.86±0.07比1.47±0.10,t=3.943,P<0.01),凋亡率高于miR-Con组及Blank组[(21.4±3.2)%比(7.5±1.2)%比(6.8±1.1)%,t=13.287,P<0.05].miR-518b组Caspase-3、Caspase-9相对活力高于miR-Con组和Blank组(2.10±0.20比1.02±0.04比1.04±0.05,t=16.287,P<0.05),(2.20±0.20比1.03±0.03比1.04±0.03,t=16.298,P<0.05).3组Caspase-8相对活力差异无统计学意义(t=8.288,P>0.05).miR-518b组侵袭细胞数少于miR-Con组和Blank组(74±10比151±15比153±12,t=17.276,P<0.05),Rap1b表达量低于miR-Con组和Blank组(0.38±0.03比1.03±0.06比1.05±0.04,江19.642,P<0.05).结论miR-518b在肝癌细胞株中下调表达,过表达miR-518b可抑制增殖和侵袭,诱导凋亡,其机制与Caspase-3、Caspase-9上调及Rap1b下调表达有关.Objective To investigate the effect of microRNA (miRNA, miR)-518b on proliferation, apoptosis and invasion of hepatocellular carcinoma cells. MethodsThe miR-518b expression was measured by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR) in hepatocellular carcinoma and normal hepatocyte lines. HepG2 cell line was divided into miR-518b over-expressed group (miR-518b group), negative control group (miR-Con group) and blank control group (Blank group). Methyl thiazol tetrazolium (MTT) assay, flow cytometry and Transwell assay were used to measure cell proliferation, apoptosis and invasion. The activity of cysteinyl aspartate-specific protease (Caspase)-3, Caspase-8 and Caspase-9 was determined by spectrophotometry, and the expression of Rap1b protein was determined by Western blotting. T-test was used for comparison of measurement data. ResultsThe expression of miR-518b in hepatocellular carcinoma cell lines was lower than that in normal hepatocellular cell line (P< 0.05). The A490 nm of miR-518 group was lower than miR-Con group at 72, 96 h after transfection (0.66±0.07 vs. 0.94±0.08, t=3.291, P<0.05) and (0.86±0.07 vs. 1.47±0.10, t=3.943, P<0.01). The apoptotic rate in the miR-518b group was higher than that in miR-Con group and Blank group [(21.4±3.2)% vs.(7.5±1.2)% vs.(6.8±1.1)%, t=13.287, P<0.05]. The relative activity of Caspase-3 and Caspase-9 in miR-518b group was highest in three groups (2.10±0.20 vs. 1.02±0.04 vs. 1.04±0.05, t=16.287, P<0.05),(2.20±0.20 vs. 1.03±0.03 vs. 1.04±0.03, t=16.298, P<0.05). There was no significant difference in the relative activity of Caspase-8 between miR-518b group, miR-Con group and Blank group (t=8.288, P>0.05). The number of invasive cells in miR-518b group was less than that in miR-Con group and Blank group (74±10 vs. 151±15 vs. 153±12, t=17.276, P<0.05). The expression of Rap1b protein in miR-518b group was lowest in three groups (0.38 ± 0.03 vs. 1.03±0.06 vs. 1.05±0.04, t=19.642, P<0.05). ConclusionThe down-regulation o

关 键 词：中文微小RNA-518b 肝癌 增殖 凋亡 侵袭 

分 类 号：R735.7[医药卫生—肿瘤]