靶向Cyclin D1基因RNA干扰对肝癌细胞增殖和Mdm2等基因表达影响的研究

目的 探讨Cyclin D1表达下调对Mdm2等基因表达及人肝癌细胞增殖的影响.方法 用脂质体将Cyclin D1-siRNA转染人肝癌细胞株Hep3B细胞.实验分为空白对照组、阴性对照siRNA组、Cyclin D1-siRNA组.采用RT-PCR和Western blot检测Cyclin D1、Mdm2、Mdm4、P53和P21表达,流式细胞仪检测细胞周期,MTT检测细胞活力,TUNEL检测细胞凋亡.结果 与空白对照组和阴性对照siRNA组相比,Cyclin D1-siRNA组P53和P21表达上调(P＜0.05),Cyclin D1、Mdm2和Mdm4表达下调(P＜0.01);3组细胞周期G1、S及G2期均无明显差异(P＞0.05);Cyclin D1-siRNA组较其他两组细胞活力明显减弱(P＜0.01),细胞凋亡明显增强(P＜0.01).结论 Cyclin D1表达下调能抑制Mdm2和Mdm4的表达,并能上调P53和P21的表达;Cyclin D1表达下调能抑制肝癌细胞的增殖并促进其凋亡.

Effects of RNA interference of targeting Cyclin D1 gene on hepatoma cell proliferation and Mdm2 gene expression

Objective To investigate the effects of down-regulating Cyclin D1 expression on Mdm2 gene expression and proliferation of human hepatoma cells.Methods Cyclin D1-siRNA was transfected into the hu man hepatoma cell line Hep3B s with liposome.The experiment was divided into the blank control group,negative control siRNA (NC-siRNA) group and Cyclin D1-siRNA group.RT-PCR and Western blot were used to detect the expressions of Cyclin D1,Mdm2,Mdm4,P53 and P21.The cell cycle was measured by flow cytometer;the cellular activity was tested with MTT;the cell apoptosis was examined by TUNEL.Results Compared with the blank control group and NC-siRNA group,the expressions of P53 and P21 in the Cyclin D1-siRNA group were increased (P＜0.05),while the expressions of Cyclin D1,Mdm2 and Mdm4 were decreased (P＜0.01);there were no significant differences in the G1,S and G2 phases among 3 groups(P＞0.05);the cell vitality in the in the Cyclin D1-siRNA group was significantly weakened compared with the other two groups(P＜0.01),while the cell apoptosis was significantly enhanced(P＜0.01).Conclusion The down-regulation of Cyclin D1 gene expression could inhibit the expressions of Mdm2 and Mdm4,and up-regulates the expressions of P53 and P21.Down-regulating Cyclin D1 expression can inhibit the proliferation of liver cancer cells and promotes their apoptosis.