Studying The Effect Of Epigenetic Modification On The Differentiation Of Islet Cells And The Expression Of Associated Genes

Objective:To study the effect of epigenetic modification on the differentiation of islet cells and the expression of associated genes (Pdx-1,Pax4,MafA,Nkx6.1,etc). Method:Mouse embryonic stem cells(1×107), NIH3T3 cells(1×107) and NIT-1 cells(1×107) were cultured and collected. The promoter methylation status of islet differentiation-associated genes (Pdx-1,Pax4,MafA and Nkx6.1),Oct4 and MLH1 genes among different cell types were measured by using methylated DNA immunoprecipitation-real time quantitative PCR methods (MeDIP-qPCR). The promoter histone modification status of these genes among different cell types were measured by using chromatin immunoprecipitation-real time quantitative PCR methods. The expression of these genes in the three cell lines were measured by realtime quantitative PCR. Analyze the relationship between the differences of epigenetic modification(DNA methylation, H3 acetylation, H3K4m3 and H3K9m3) and gene expressions. Results:(1) In NIH3T3 cell, the transcription-initiation-site of Pdx-1, MafA and Nkx6.1 are high extent of methylaion status. Compared with mES cell and NIT-1 cell, NIH3T3 cell has a sigificantly higher level of DNA methylation modification (P<0.05). There are the Pearson correlation between expressions of Pdx-1,MafA,Nkx6 genes and methylation modification.(2) Compared with mES cell and with NIH3T3 cell, NIT-1 cell has a sigificantly higher level of H3K4m3 modification in the transcription-initiation-site of Pdx-1,Pax4,MafA and Nkx6.1 genes (p<0.05), with sigificant level of gene expression. (3) Compared with mES cell and with NIT-1 cell, NIH3T3 cell has a sigificantly higher level of H3K9m3 modification in the transcription-initiation-site of Pdx-1,Pax4,MafA and Nkx6.1 genes (p<0.05), with no gene expression. Conclusion:Histone modification (H3K4m3 and H3K9m3) and DNA methylation can regulate one another, and synergtically regulate the expression of islet differentiation-associated genes. These epigenetic modifications pose significant importance in the differentiation process from embryonic stem cell into islet cell.