牙髓细胞成牙本质向分化过程中DSPP基因的表观遗传学修饰

Epigenetic Modifications of DSPP Gene During Odontoblastic Differentiation of Dental Pulp Cells

目的:通过检测牙髓细胞(DPC)体外成牙本质向分化过程中牙本质涎磷蛋白(DSPP)基因的表观遗传学修饰模式的变化。探讨表观遗传学机制在DPC成牙本质向分化中的作用。方法:在体外诱导DPC成牙本质向分化,采用Real-time PCR检测DSPP基因表达的改变,采用重亚硫酸盐测序法检测DSPP基因的甲基化水平改变,染色质免疫共沉淀检测DSPP启动子区组蛋白修饰(H3K27me3和H3K4me3)修饰的变化。结果:在DPC诱导分化后,DSPP的表达逐渐增高,伴随着基因启动区甲基化水平的降低,同时与转录激活相关的组蛋白修饰H3K4me3升高,而与转录抑制相关的组蛋白修饰H3K27me3降低。结论:DPC在成牙本质分化过程中DSPP表达的上调与DSPP启动子区域出现开放基因表达的表观遗传学修饰模式有关。这提示,表观遗传学调控参与到DPC成牙本质向分化的过程。

Objective： To study the epigenetic mechanisms of DNA methylation and histone modification on the regulation of DSPP gene during odontoblastic differentiation of dental pulp ceils （DPCs）. Methods： The human DPCs were isolated and treated with odontoblastic induction medium. The expression DSPP in DPCs during odontoblastic differentiation was evaluated by Real-time PCR. The DNA methylation of DSPP gene was analyzed with bisulfite sequencing PCR. The DSPP gene-associated HSK4me3 and H3K27me3 were quantified by using chromatin immunoprecipitation. Results： Odontoblastic differentiation of human DPCs was confirmed by mineralization nod- ules formation. The expression of DSPP was significantly up-regulated during odontoblastic differentiation of DPCs with a decrease in DSPP gene associated H3K27me and CpG sites methyla-tion, and a increase in DSPP gene associated H3K4me3 on the day 14 after odontoblastic induction. Conclusion： The DNA methylation and histone modification are possible involved in the regulation of DSPP during odontoblastic differentiation, which suggests epigenetic mechanisms have the effects on odontoblastic differentiation of DPCs.