抑癌基因Smad4对HepG2生长的影响及其机制探讨  

The growth effects of tumor suppressor Smad4 gene on human hepatocellular carcinoma cell line HepG2

作　　者：刘萍[1] 田德安[1] 夏丽敏[1] 晏维[1] 徐倩[1] 徐湖波[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院消化内科,湖北武汉430030

出　　处：《胃肠病学和肝病学杂志》2008年第12期975-978,共4页Chinese Journal of Gastroenterology and Hepatology

摘　　要：目的研究抑癌基因Smad4对人肝癌细胞株HepG2生长的影响,并探讨其作用机制。方法采用脂质体瞬时转染法转染PFTX-5 Smad4质粒至人肝癌细胞株HepG2(实验组),以转染空质粒PFTX-5的HepG2细胞为对照组,野生型HepG2细胞为空白组,利用免疫印迹(Western blot)和逆转录聚合酶链反应(RT-PCR)检测Smad4及血管内皮生长因子(VEGF)表达的变化;用MTT法检测细胞增殖及用FITC-Annexin V、碘化吡啶(propidiumiodide,PI)双染后流式细胞仪(FCM)检测细胞周期和凋亡情况,Westernblot检测细胞周期素D1(Cyclin D1)表达差异。结果PFTX-5 Smad4瞬时转染到肝癌细胞后,实验组与对照组和空白组相比,Smad4 mRNA和蛋白表达明显上升(P<0.05),而VEGF mRNA和蛋白的表达显著降低(P<0.05)。实验组细胞CyclinD1表达明显下降(P<0.05),细胞周期时相分布出现合成前期(G0/G1期)阻滞。实验组细胞凋亡率明显升高(P<0.01),增殖受到明显抑制。结论Smad4高表达可能通过降低肝癌细胞中VEGF的表达,抑制cyclinD1转录合成,并诱导强烈的G1期阻滞和细胞凋亡,对细胞的增殖有明显的抑制作用,为进一步研究肝癌生长增殖机制提供了实验基础。Objective To study the growth effects of transfected tumor suppressor Smad4 gene on human hepatocellular carcinoma cell line HepG2. Methods Plasmid of PFTX-5 Smad4 was transfected into human hepatocellular carcinoma cell line HepG2. The HepG2 cells transfected with this plasmid （experimental group） and those treated with PFTX-5 vector （control group） and without any treatment （blank group） were compared. The expression of Smad4 and VEGF were detected by RT-PCR and Western blot. Cell proliferation was detected by MTT and then assessed the cell cycle distribution and apoptosis by flow cytometry with fluoreseein isothiocyanate-conjugated Annexin V （FITC-Annexin V ） and propidium iodide （PI） staining. The expression of CyclinD1 was detected by Western blot. Results After being transfected with PFTX-5 Smad4, with the decrease in VEGF gene expression （P 〈 0.05 ） , the growth rate of HepG2 was also inhibited compared with those of the control and blank groups. Cell cycle blocked at G0/G1 stage and hepatic cell apoptosis was found （P 〈 0.01 ）. CyclinD1 expressed lower （P 〈 0.05）. Conclusion Growth of HepG2 can be controled by increasing Smad4 expression. These provide potent theoretical ground for studying the mechanism of cell proliferation in hepatoma.

关 键 词：癌 肝细胞 HEPG2细胞 SMAD4基因 

分 类 号：R735.7[医药卫生—肿瘤]