5-氮杂胞苷对宫颈癌细胞FHIT基因去甲基化及抑制增殖作用

目的:探讨5-氮杂胞苷对Hela宫颈癌细胞脆性组氨酸三联基因(Fragile histidine triad gene,FHIT)启动子去甲基化作用及对其增殖的影响。方法:采用低浓度和高浓度5-氮杂胞苷作用Hela宫颈癌细胞,分别采用BSP和RT-PCR检测处理前后FHIT基因启动子甲基化状态和mRNA表达,应用MTT法检测细胞增殖改变。结果:Hela宫颈癌细胞FHIT基因启动子表现高度甲基化状态(CG位点甲基化率为80%～100%),mRNA表达完全受到抑制,细胞呈指数增殖状态;低浓度5-氮杂胞苷作用后的甲基化率显著降低(10%～40%),细胞增殖速度降低;高浓度组FHIT基因启动子甲基化状态被完全逆转,mRNA表达明显高于低浓度组,细胞增殖速度显著低于正常组和低浓度组。结论:5-氮杂胞苷可逆转Hela宫颈癌细胞FHIT基因甲基化状态,使基因恢复表达而抑制细胞增殖,去甲基化和抑制增殖作用随5-氮杂胞苷剂量增加而增加。

Effect of 5-azacitidine on demethylation of FHIT gene and its inhibiting effect on proliferation of cervical cancer cells

Objective:To explore the effect of 5-azacitidine on demethylation of fragile histidine triad(FHIT) gene promoter and its inhibiting effect on proliferation of cervical cancer cells.Methods:Low concentration 5-azacitidine and high concentration 5-azacitidine were used to treat cervical cancer HeLa cells,bisulfate-sequencing PCR(BSP)and RT-PCR were used to detect the methylation statuses and mRNA expressions of FHIT gene promoter before and after treatment,MTT method was used to detect the change of cell proliferation.Results:FHIT gene promoter in cervical cancer HeLa cells showed high methylation status(the rate of methylation in CG situs was 80%-100%),mRNA expression was inhibited completely,the cervical cancer HeLa cells showed exponential proliferation pattern;after treatment with low concentration 5-azacitidine,the rate of methylation decreased significantly(10%-40%),the speed of cell proliferation decreased;after treatment with high concentration 5-azacitidine,the methylation status of FHIT gene promoter were reversed completely,mRNA expression level was significantly higher than that in low concentration 5-azacitidine group,the speed of cell proliferation was significantly lower than those in normal concentration 5-azacitidine group and low concentration 5-azacitidine group.Conclusion:5-azacitidine can reverse the methylation status of FHIT gene promoter in cervical cancer HeLa cells,recover the gene expression,and inhibit cell proliferation,the demethylation and inhibiting effect on proliferation will strengthen with the increase of 5-azacitidine dose.