上调microRNA-101沉默EZH2基因表达对人膀胱癌T24细胞系增殖和凋亡的影响

Effects on Cell Proliferation and Apoptosis of Human Bladder Cancer Cells by Up-regulating the Expression of MiRNA-101 Silence Gene EZH2

目的探讨microRNA-101抑制靶基因EZH2对膀胱癌细胞增殖和凋亡的影响.方法构建靶向EZH2基因的microRNA-101质粒并转染至人膀胱癌细胞中,采用RT-PCR法检测EZH2mRNA的表达情况,利用四甲基偶氮唑盐(MTT)检测细胞增殖抑制,通过Annexin V-FITC/PI流式细胞术实验观察转染后细胞的凋亡情况.结果 microRNA-101阳性对照组EZH2 mRNA较阴性对照组和空白对照组表达下降(P<0.05);在转染5 d后,其生长抑制率明显高于对照组(P<0.01);microRNA-101组较阴性对照组及空白对照组G0/G1期细胞数明显增多,S期细胞数明显减少,凋亡率增加(P<0.01).结论上调microRNA-101可抑制EZH2基因的表达,并有效的抑制人膀胱癌细胞的增殖,促进其凋亡,为深入研究膀胱癌的基因治疗提供理论依据.

Objective To investigate the effects of inhibiting enhancer of zeste homolog 2（EZH2） on cell proliferation and apoptosis by up-regulating the expression of miRNA-101 in bladder cancer cells. Methods The microRNA-101-expressing plasmid targeting EZH2 gene was constructed and transfered into T24 cells. RT-PCR was used to detect the expression of EZH2 mRNA. The proliferation of T24 cells was detected in vivo by MTT,and cell apoptosis was observed by Annexin V-FITC/PI flow cytometric analysis. Results The siRNA-expressing plasmid targeting EZH2 gene successfully inhibited the expression of EZH2 in T24 cells. Compared with control groups, the expression of mRNA in the positive group was significantly inhibited（P 0.05）. After plasmid transfection of 5 days,the cell proliferative activity was significantly lower in the miRNA-101 mimics group than that in the negative control group and blank control group（P〈0.01）. In the miRNA-101 mimics group, cells increased significantly at Go/G1 phase [（80.12±7.8） %,P〈0.01], while decreased at S phase [（11.50±1.2） %,P〈0.01]. And the expression of EZH2 mRNA was down-regulated（P〈0.01）. Conclusion The microRNA-101 silencing EZH2 can significantly inhibit cell proliferation of T24 cells and promote its apoptosis. It provides a theoretical basis for further study of bladder cancer gene therapy.