Objective : Investigate the effect of zebularine( Zeb ) on methylation and expression of RASSF1A gene in human ovarian cancer cell line A2780 and to discuss the regulating effect of demethylation agent on its expression. Methods : A2780 cells were cultured in RPMI 1640 and were treated with different concentrations ( 50, 100 ，200 μmol·L-1 ) of DNA methyltransferase inhibitor Zeb. Methylation- specific PCP( MSP ) was used to detect promoter methylation of RASSF1A gene. RT-PCR and Western blotting were used to detect expression of mRNA and protein of RASSF1A gene before and after treatment with Zeb respectively. Results : Promoter hypermethylation of RASSF1A gene was detected in ovarian cancer cell A2780 ， and no expression was detected before treatment. After treated with Zeb ，the promoter region of RASSF1A gene exhibit demethylation , and its expression occured at mRNA and protein level. Conclusion: Zeb can induce demethvlation of RASSF1A gene in human ovarian cancer cellA2780 and regulate its reexpression.%目的:通过观察应用去甲基化药物zebularine(Zeb)后人卵巢癌细胞系A2780中抑癌基因RASSF1A甲基化状态及表达水平的改变,探讨 Zeb对卵巢癌细胞RASSF1A基因表达的调控.方法:应用不同浓度(50、100、200 μmol·L-1) 的Zeb处理体外培养的卵巢癌细胞A2780后,甲基化特异性PCR(MSP)法检测用药前后细胞中RASSF1A基因的甲基化状态,RT-PCR法及 Western-blotting法检测用药前后细胞中RASSF1A 基因mRNA及蛋白表达的变化.结果:RASSF1A基因在卵巢癌细胞A2780中启动子区呈异常甲基化状态,在mRNA及蛋白水平表达阴性.经过Zeb处理后,RASSF1A基因启动子区呈去甲基化状态,其mRNA及蛋白重新表达.结论:去甲基化药物Zeb能使卵巢癌细胞RASSF1A基因呈去甲基化状态,从而调控RASSF1A基因重新表达.
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