化学修饰的siRNA对乳腺癌细胞VEGF基因表达抑制及凋亡的诱导作用

目的：体外水平探讨化学修饰的小干扰RNA（siRNA）介导的RNAi治疗乳腺癌的可行性。方法：选用阳离子脂质体Lipofectamine2000^TM作为转染试剂，将化学修饰的针对人VEGF基因的siRNA转染人类乳腺癌细胞株MCF-7。半定量RT—PCR和蛋白印迹试验分别检测转染前后细胞中VEGF、Bcl2和baxmRNA和蛋白水平表达的变化，Hoechst33258荧光染色分析细胞凋亡。结果：与未转染的正常细胞组相比，靶向VEGF基因的siRNA转染MCF-7后，细胞中VEGF及Bcl-2mRNA和蛋白表达水平显著降低，P〈0．01。bax基因的表达未发生明显变化，Bcl-2和bax比值下降。Hoechst33258荧光染色显示转染siRNA72h后细胞内可见凋亡小体。结论：化学修饰的siRNA介导的RNAi下调靶基因VEGF的表达，并对MCF-7细胞有明显的凋亡诱导作用。

Chemically modified siRNA inhibited VEGF expression and induced apoptosis in human breast carcinoma cell line MCF-7

OBJECTIVE: To investigate the feasibility of gene therapy using chemically modified siRNA for breast cancer in vitro.METHODS: VEGF siRNA was transfected into MCF-7 cells to induce RNAi by using cationic liposome Lipofectamine 2000TM as transfecting agent.After the transfection,the expression leves of mRNA and protein VEGF,Bcl-2 and bax were evaluated by RT-PCR and Western blot,respectively.The cells were observed with an invert microscope in shape and with a fluorescence microscope in apoptosis dyed by Hoechst 33258.RESULTS: Compared with the control groups,experiment data showed that siRNA directed against VEGF gene down-regulated the expression of VEGF in the level of mRNA and protein (P<0.01),and Bcl-2 expression changed obviously.The ratio of Bcl-2 to bax decreased.Seventy-two hours after siRNA being transfected,apoptosis bodies were observed in stained cells by Hoechst 33258 under a fluorescent microscope.CONCLUSION: RNAi mediated by chemically modified siRNA efficiently inhibits VEGF gene expression and induces apoptosis in MCF-7.