摘要
目的 通过合成及转染微小RNA （miRNA）-3178模拟物（mimic）,观察上调miR-3178表达对肝癌血管内皮细胞（HCC TECs）增殖、凋亡及细胞周期的影响.方法 实时定量聚合酶链反应（Real-time PCR）验证肝窦内皮细胞（HSECs）和肝癌血管内皮细胞中miR-3178的差异表达.miR-3178 mimic转染肝癌血管内皮细胞上调miR-3178表达,实验分为3组：空白对照组（CON组）、miR-3178上调组（Mimic组）、阴性对照组（NC组）,Real-time PCR验证转染前后肝癌血管内皮细胞miR-3178表达.荧光显微镜下观察转染效率.流式细胞仪检测细胞周期及细胞凋亡,噻唑蓝（MTT）法检测细胞增殖.结果 Real-time PCR结果显示HCC TECs中miR-3178表达明显低于HSECs（P ＜0.01）.转染后HCC TECs中miR-3178表达明显高于转染前（P＜0.01）,细胞转染效率达到90％以上.MTT结果显示Mimic组在转染后24、36、48、72 h增殖率均明显低于其他组（P＜0.05）,在72 h增殖率达到最低.细胞凋亡结果显示,Mimic组[（18.19±2.25）％]凋亡率明显高于CON组[（6.43±1.23）％]和NC组[（6.41±1.56）％],差异有统计学意义（P＜0.01）.细胞周期结果显示,Mimic组[（69.12±4.87）％]G0/G1期细胞明显多于CON组[（55.12±4.13）％]和NC组[（54.08±4.72）％],差异有统计学意义（P＜0.05）.Mimic组[（10.98士1.76）％]S期细胞明显少于CON组[（24.23±2.13）％]和NC组[（24.78±2.24）％],差异有统计学意义（P＜0.01）.结论 HCC TECs中miR-3178明显低表达,上调miR-3178表达能明显抑制HCC TECs细胞增殖,促进细胞凋亡及G0/G1期细胞阻滞.
Objective To research the effect of growth, apoptosis and cell cycle of hepatocellular carcinoma （HCC） tumor endothelial cells （TECs） caused by up-regulate the expression of microRNA-3178 （miR-3178） through transfection of miR-3178 mimic.Methods Real-time polymerase chain reaction （Real-time PCR） was used to identify differential expression of miR-3178 in normal hepatic sinusoidal endothelial cells （HSECs） and HCC TECs.Furthermore, up-regulation of miR-3178 expression was achieved using miR-3178 mimic transfected into HCC TECs, HCC TECs were divided into 3 groups： control （CON） group, miRNA-3178 up-regulation （Mimic） group, negative control （NC） group.Real-time PCR was used to detect expression of miR-3178 in HCC TECs before and after transfection.Transfection efficiency was observed by using an inverted fluorescence microscope.Proliferation of HCC TECs were detected by methyl thiazol tetrazolium （MTT） assay.Flow cytometry was used to detect the apoptosis and cell cycle of HCC TECs.Results The results of Real-time PCR showed that miR-3178 was significantly down-regulated in HCC TECs compared to HSECs （P 〈 0.01）, exspression of miR-3178 was significantly increased after trsansfecion （P 〈 0.01）.The transfection efficiency in HCC TECs was higher than 90%.MTT assay showed that the profilation rate of HCC TECs of mimic group at 24, 36, 48, 72 h were decreased significantly than other groups （P 〈 0.05） and achieved the lowest at 72 hours.The results of apoptosis showed that mimic group （18.19 ± 2.25）% showed significant increased apoptosis compared to CON group （6.43 ±1.23）% and NC group （6.41 ± 1.56）% （P〈0.01）.The results of cell cycle showed that mimic group （69.12 ±4.87）% showed significant increased percentage of G0/G1 phase cells compared to CON group （55.12 ±4.13）% and NC group （54.08 ±4.72）% （P〈0.05）,Mimic group （10.98 ± 1.76）% showed decreased percentage of S phase cells compared to CON group （24.23 ±2.13） % and NC group （24.78 ±2.24）% （P 〈0.01）.Conclusion MiR-3178 was low expression in HCC TECs and specifically inhibit proliferation and promote apoptosis and G1-phase arrest of HCC TECs in vitro.Thus, miR-3178 might be a valid target for the treatment of HCC.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2015年第11期2695-2698,共4页
Chinese Journal of Experimental Surgery
基金
武汉市科技攻关计划资助项目（201260523178-2）
关键词
肝癌血管内皮细胞
微小RNA-3178
增殖
凋亡
细胞周期
Tumor vascular endothelial cells of hepatocellular carcinoma
Proliferation
Apoptosis
Cell cycle
