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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 To observe the epigenetic modification of insulin-like growth factor 2 (IGF2) during the replication-induced senescence of cultured human embryo lung fibroblasts and the persistence phase of hydrogen peroxide-induced premature aging. Methods The mRNA expression of IGF2 was detected by real-time PCR. The methylation of promoter region was detected by methylation-specific PCR. Histone modification was detected by chromatin immunoprecipitation combined with quantitative PCR, including histone H3, H4 acetylation, H3 (Lys4) And H4 (Lys20) methylation modification. Results During the process of cell-induced senescence, the mRNA expression of IGF2 increased in the middle-aged cell group and the replicative senescent cell group, and increased significantly in the premature aging group. In the cell senescence process, only the senescent cell group in the promoter region -658 ~ -456bp with a certain degree of methylation; histone modification of replicative senescent cells in the promoter region -856 ~ -634bp with H4 acetylation, H3 methylation modified mainly; +9 ~ +145 bp modification, replication The senescent cell group was modified by methylation of Histone H3 and H4, and the methylation of H3 was the main one in the persistent premature aging group. Conclusions During the process of cell senescence, the IGF2 promoter region regulates the mRNA expression of IGF2 promoter, and there are differences in the regulation mechanism between replicative senescence and premature senility.