人胚肺成纤维细胞衰老过程中胰岛素样生长因子2表观遗传活化作用

Epigenetic activation of IGF2 during cellular senescence

目的观察体外培养人胚肺成纤维细胞复制性衰老过程中及过氧化氢诱导早衰持续阶段胰岛素样生长因子2(IGF2)表观遗传学修饰。方法荧光定量PCR检测IGF2的mRNA表达,甲基化特异PCR检测其启动子区甲基化改变,染色质免疫沉淀结合定量PCR检测组蛋白修饰,包括组蛋白H3、H4乙酰化,H3(Lys4)及H4(Lys20)甲基化修饰。结果细胞复制性衰老过程中,中年细胞组及复制性衰老细胞组IGF2的mRNA表达升高,早衰持续组升高显著;细胞衰老过程中,仅复制性衰老细胞组在启动子区-658～-456bp具有一定的甲基化水平;复制性衰老细胞组的组蛋白修饰在启动子区-856～-634bp以H4乙酰化,H3甲基化修饰为主;+9～+145bp的修饰,复制性衰老细胞组受组蛋白H3及H4甲基化联合修饰,早衰持续组以H3甲基化修饰为主。结论细胞衰老过程中,IGF2启动子区组蛋白修饰联合调控其mRNA表达,复制性衰老与早衰的调控机制存在差异。

Objective Epigenetic regulations of insulin-like growth factor 2（IGF2） were observed during cellular replicative senescence and premature senescence induced by hydrogen peroxide of human embryonic lung fibroblasts （HEFs）.Methods The mRNA level of IGF2 was detected by Q-PCR.The methylation status in the promoter region was observed by methylation-specific PCR.The histone modifications was detected by chromatin immunoprecipitation-Q-PCR assay,including acetylation for H3,H4 and methylation for H3（Lys4）and H4（Lys20）.Results In the process of cellular senescence,the mRNA level of IGF2 increased in both mid-aged and replicative senescent cells,but increased obviously in premature senescent cells compared with that of young cells.In the promotor region from-658 bp to-456 bp,the methylation level for IGF2 was detected only in replicative senescent cells.About the main histone modifications,IGF2 in the region （-856 bp--634 bp） was H4 acetylation and H3K4 methylation in replicative senescence,while in the region （＋9 bp-＋145 bp） was H3K4 and H4K20 methylation in replicative senescence and H3K4 methylation in premature senescence.Conclusion The histone modifications take part in regulating the mRNA expression for IGF2 during cellular senescence and different mechanisms exist between two types of senescence.