靶向Survivin基因microRNA诱导肝癌细胞凋亡的实验

Experiment on microRNA target survivin induced liver cancer cell apoptosis

目的:探讨靶向Survivin的microRNA转染肝癌细胞阻抑Survivin表达,及其对肝癌细胞凋亡、增殖的影响。方法:设计合成特异性靶向Survivin的microRNA的序列。肝癌细胞株HepG2接种于细胞培养板内,分为5组,空白对照组、脂质体组和低、中、高浓度转染组(分别给予50、100和200 nmol/L microRNA转染);作用后收集各组细胞。四氮唑盐(MTT)法检测不同浓度的microRNA对细胞的生长抑制率。流式细胞术检测各组细胞增殖率和凋亡指数。逆转录-聚合酶链反应(RT-PCR)检测靶向Survivin的mRNA表达水平。结果:通过MTT检测,不同浓度microRNA转染组细胞的抑制率明显高于对照组(P<0.05)。各浓度mi-croRNA转染组细胞凋亡指数明显高于对照组(P<0.05),高浓度转染率最明显(P<0.05)。各浓度microRNA转染组细胞增殖指数明显低于对照组(P<0.05),高浓度转染组最明显(P<0.05)。各浓度microRNA转染组细胞Sur-vivin的mRNA转录水平有不同程度减弱。结论:不同浓度Survivin的microRNA转录能下调Survivin的mRNA表达,诱导肝癌细胞凋亡,抑制细胞增殖。

Objective：To investigate the effect of microRNA target survivin on transfected human liver cancer cell apoptosis, proliferation and survivin expression. Methods ： MicroRNA targeting survivin was synthesized and transfected into HepG2 cells by lipdectin. HepG2 cells were cultured in the culture dish 24 hours before transfection and the cells were divided into control, liposome group and low, medium or high dose microRNA group with a dosage of 50, 100 or 200 umol/L microRNA, respectively, for the transfection in each group. After transfection, the cultured cells were harvested for the following tests. MTT assay was used to detect cell rate of inhibition and flow cytometry was for examining apoptotic index （AI） and proliferative index （PI）, and the expression of survivin mRNA was detected by reverse transcriptase PCR. Results： MTT assay showed that the rate of inhibition in microRNA groups were significantly higher than that of control groups, especially evident in the high dose group（ P 〈 0.05）. The Apoptotic index of the different dose microRNA groups were also significantly higher compared with control groups（ P 〈 0.05 ）, but the expression of survivin mRNA in low, medium and high dose microRNA groups decreased to a certain degree. Conclusion ： Survivin microRNA can effectively decrease the expression of survivin mRNA, induce liver cancer cell apoptosis and inhibit cell proliferation.