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目的通过小分子RNA(siRNA)介导靶基因沉默技术,干扰前列腺癌细胞中果蝇zeste基因增强子的人类同源物的表达,以探讨靶向EZH2基因的RNAi对前列腺癌细胞增殖抑制的效应。方法设计靶向EZH2基因的RNA干扰(RNAi)载体,构建重组表达质粒并且转染人前列腺癌细胞22RV1。利用MTT比色法检测靶向沉默EZH2表达对于22RV1的增殖抑制的影响;利用qRealtime-PCR鉴定转染组与空白组细胞中EZH2基因mRNA的表达量;提取各组细胞的总蛋白,利用Western Blotting检测EZH2蛋白的表达情况。结果在转染了靶向EZH2基因siRNA的细胞株中,可以检测到EZH2表达呈明显的下调趋势;而在空白对照组中其表达很稳定。另外,诱导EZH2基因沉默可以抑制该前列腺肿瘤细胞的增殖和生长。上述结果在各组细胞间呈现明显的统计学差异(P<0.05)。结论siRNA诱导的RNAi可以有效的抑制其靶基因EZH2的表达,同时可以抑制前列腺癌细胞的增殖和生长,是一种潜在的基因治疗新方法。
OBJECTIVE: To investigate the effect of RNAi targeting EZH2 gene on the proliferation inhibition of prostate cancer cells by interfering with the expression of human homologues of zeste gene enhancer in prostate cancer cells by siRNA-mediated target gene silencing . Methods RNA interference (RNAi) vectors targeting EZH2 gene were designed, constructed recombinant expression plasmids and transfected into human prostate cancer cells 22RV1. The effect of targeted silencing of EZH2 on the proliferation inhibition of 22RV1 was detected by MTT assay. The expression of EZH2 mRNA was detected by qRealtime-PCR. The total protein of each group was extracted and analyzed by Western Blotting Detection of EZH2 protein expression. Results EZH2 expression was significantly down-regulated in the cell lines transfected with siRNA targeting EZH2 gene, whereas it was stable in the blank control group. In addition, induction of EZH2 gene silencing can inhibit the proliferation and growth of the prostate tumor cells. The above results showed significant statistical differences among the groups of cells (P <0.05). Conclusion RNAi induced by siRNA can effectively inhibit the expression of its target gene EZH2 and inhibit the proliferation and growth of prostate cancer cells. It is a potential new gene therapy method.