miRNAs-DNMT1调节网络在皮肤角质形成细胞衰老中的作用及机制研究

Studies have shown that miRNA could regulate aging. At the same time, we found out in our preliminary experiments that the expression of miR-217、miR-377 and miR-342 increased, while DNA methyltransferase 1 (DNMT1) markedly decreased in the senescent keratinocytes (KC). According to the prediction database, all of the above three miRNAs can combine with DNMT1 mRNA at adjacent positions. And the expression of the above three miRNAs decreased when DNA methylation inhibitor was added. In our present study, we will firstly understand the role of DNMT1 in keratinocyte senescence. Then, we will investigate the mechanism of how the above three miRNAs individually or jointly effect the DNMT1 expression and KC senescence by using luciferase reporter gene assay. Moreover, we will explore the regulation of DNMT1 on the expression level and promoter methylation level of the miRNAs by using the bisulfite sequencing method. Finally, we will screen some senescence related genes that are both regulated by miRNAs/DNMT1 and modified by DNA methylation, and investigate their functions in keratinocyte senescence. Our study aims to explore the mechanism of miRNAs-DNMT1 epigenetics network on the KC senescence and provide a new direction in the research field and therapeutic field of skin aging.

已有研究表明衰老的发生受到了miRNA的调节。我们的预实验发现miR-217、miR-377及miR-342在衰老的角质形成细胞（KC）中表达上升而DNA甲基转移酶1（DNMT1）明显下降；数据库预测发现上述3个miRNA均可结合DNMT1的mRNA且位置相近，DNA甲基化抑制剂处理后，KC中这3个miRNA的表达均下降。本研究中，我们将先明确DNMT1在KC衰老中的作用，再利用荧光素酶报告基因等技术探讨上述3个miRNA单独或联合作用对DNMT1表达和KC衰老的影响和机制；然后利用重亚硫酸氢盐测序等技术了解DNMT1对miRNAs表达量和启动子甲基化水平的调节；最后筛选出受miRNAs/DNMT1通路调节并受DNA甲基化修饰的衰老相关基因并探讨其对KC衰老的影响。本研究旨在探讨miRNAs-DNMT1表观遗传学修饰网络对皮肤KC衰老的影响及机制，为皮肤衰老的研究和治疗领域提供新的方向。

已有研究表明衰老的发生受到了 miRNA 的调节。我们实验发现 miR-377、miR-217在衰老的人皮肤成纤维细胞（ HSFs）中表达上升而 DNA 甲基转移酶 1（ DNMT1）明显下降；数据库预测发现miR-377、miR-217均可结合 DNMT1 的3’UTR区。我们利用荧光素酶报告基因等技术探讨上述两个microRNAs作用对 DNMT1表达和 HSFs衰老的影响和机制；最后利用重亚硫酸氢盐测序筛选出受 miRNAs/DNMT1 通路调节并受 DNA甲基化修饰的衰老相关基因并探讨其对 HSFs衰老的影响。本研究探讨了microRNA-DNMT1信号通路调控人皮肤成纤维细胞细胞衰老的这一新机制，找到了人皮肤成纤维细胞中靶向调控DNMT1从而影响其衰老的两个上游microRNA分子(miR-377和miR-217)。该研究深入探究了皮肤成纤维细胞细胞衰老的表观遗传学机制，也为人类抗衰老研究提供了潜在的治疗靶点。