siRNA干扰MDM2基因对人肝癌HepG2细胞增殖和凋亡的影响

摘要

目的 将靶向MDM2的siRNA转染人肝癌HepG2细胞,观察转染后细胞内p21基因的表达变化及其对HepG2细胞增殖和凋亡的影响.方法 将人工合成的针对MDM2基因的siRNA片段通过Lipofectamine<'TM>2000转染人肝癌细胞HepG2.实验分为MDM2 siRNA转染组、阴性对照组、脂质体组、正常对照组.分别用RT-PCR和Western blot法检测各组细胞中MDM2、p21的Mrna和蛋白质的表达量,并用MTT法检测各组细胞的增殖活力,流式细胞仪检测细胞周期和凋亡的变化.结果 与其他3组比较,siRNA转染组细胞中MDM2 Mrna及蛋白表达减少(P<0.01),而p21 Mrna及蛋白表达增高(P<0.01).HepG2转染MDM2 siRNA后,增殖能力减弱,凋亡显著.结论 MDM2基因可能通过影响p21控制肝癌细胞的生长.%Objective To transfect siRNA targeting MDM2 gene into human hepatoma cell line HepG2 and to observe the expression of p21 and its effect on the proliferation and apoptosis of HepG2 cells.Methods The MDM2 siRNA was transfected into HepG2 by means of LipofectamineTM 2000.There were four groups in this study including siRNA group, negative control (NC) group, Lipofectamine (Lip) group and normal cell group.Reverse transcriptase PCR and Western blot were used to measure the MDM2 and p21 expression at mRNA and protein levels, respectively.The cell proliferation was assessed by MTT assay and the changes in cell cycle and apoptosis were evaluated by flow cytometry.Results Compared with normal cell group, NC group and Lip group, the expression of MDM2 mRNA and protein decreased significantly in siRNA group ( P ＜ 0.01 ), the expression of p21 mRNA and protein enhanced significantly ( P ＜ 0.01 ).The proliferation of HepG2 cells was markedly inhibited and the apoptosis of HepG2 cells was increased after transfected by MDM2 siRNA (P ＜ 0.01 ).Conclusion MDM2 gene may regulate the cell proliferation by interacting with p21.