miR-152和miR-200s在炎性因子诱导肝胰岛素抵抗发生中的作用及其机制研究

Insulin resistance and dysfunction of β-cells play a central role in the pathogenesis of type 2 diabetes. Recent studies showed that microRNAs regulate β-cell development and insulin secretion. However, the relation between microRNAs and insulin resistance is poorly understood. Our previous studies indicated that miR-152 and miR-200s were inhibited in NCTC1469 cells exposure to inflammatory factors. Moreover, we found that miR-200s expression was regulated by miR-152. Therefore, we hypothesize that under inflammatory condition, miR-152 impaires insulin signaling pathway and induces the insulin resistance by regulating the expression of miR-200s and their target genes. To test this hypothesis, bio-information, real time PCR, Western blot, adenovirus vector-mediated gene transfer and RNA interference will be used to investigate the role miR-152 and miR-200s in inflammatory factor-induced hepatic insulin resistance, and molecular mechanisms by which miR-152 regulates the expression of miR-200s and their target genes. We will provide insight into pathogenesis of hepatic insulin resistance induced by inflammatory factors and novel ideals for type 2 diabetes prevention.

2型糖尿病的发生与胰岛素抵抗和胰岛β细胞功能障碍密切相关。研究表明，microRNA调节胰岛β细胞发育与胰岛素分泌。但microRNA与胰岛素抵抗关系的研究鲜有报道。我们前期发现miR-152和miR-200在炎性因子诱导产生的肝胰岛素抵抗中显著降低，而miR-200s受miR-152调控。据此提出假说：在炎性因子作用下，miR-152通过调控miR-200s及其下游靶基因影响胰岛素信号通路，导致肝胰岛素抵抗的发生。本课题拟采用生物信息学、real time PCR、Western blot、腺病毒载体转染、RNA干扰等手段，从分子、细胞、组织及动物水平等多层次明确miR-152和miR-200s在炎性因子诱导肝胰岛素抵抗发生中的作用；阐明 miR-152调控miR-200s以及影响胰岛素信号通路的机制。本研究将从microRNA新角度探讨胰岛素抵抗的发生机制，为2型糖尿病防治提供新思路。

肝脏胰岛素抵抗与2型糖尿病密切相关。研究表明microRNA参与调节胰岛素信号通路的转导。本课题明确了miR-152 和miR-200s 在炎性因子诱导肝胰岛素抵抗发生中的作用；阐明了miR-152和miR-200s 影响胰岛素信号通路的机制。圆满完成原定的研究计划，实现了预期目标，并取得了突出进展，发表SCI论文17篇，在国际学术会议做报告6次，全国学术会议做报告21次，培养5名博士生。.1. 发现miR-152和miR-200s在炎性因子诱导肝胰岛素抵抗发生中起重要作用：1）在db/db小鼠、高脂喂养小鼠和IL-6诱导的小鼠胰岛素抵抗模型肝脏中miR-152和miR-200s表达降低；PI3K/AKT/GSK信号通路活性降低；糖原合成减少；2）在NCTC1469细胞中过表达miR-152和miR-200s增强PI3K/AKT/GSK信号通路活性，促进糖原合成；低表达miR-152和miR-200s降低PI3K/AKT/GSK信号通路活性，减少糖原合成；3）过表达miR-152和miR-200s能够逆转IL-6对胰岛素信号通路的抑制作用。.2. 阐明miR-152和miR-200s通过调控下游靶基因FOG2和PTEN参与炎性因子诱导肝脏胰岛素抵抗发生的作用机制：1）miR-152和miR-200s下游靶基因为FOG2和PTEN；luciferase assay证明PTEN3’-UTR上有miR-152和miR-200s的结合位点；2）miR-152能够负调节PTEN蛋白水平；miR-200s能够负调节FOG2和PTEN蛋白水平；3）低表达FOG2和PTEN能够逆转IL-6对胰岛素信号通路的抑制作用。.3. 揭示miR-200b和miR-200c在肝脏甘油三酯(TG)蓄积中的作用机制：在Hep1-6和NCTC1469细胞中低表达miR-200b和miR-200c激活TG代谢信号通路，细胞内TG合成增加；上调miR-200b和miR-200c的表达可抑制TG代谢信号通路，细胞内TG含量减少； JUN是miR-200b和miR-200c的靶基因； JUN可以与SREBP1启动子区域结合，进而促进其转录，最终促进脂质合成并引起脂质蓄积增加； .本研究从microRNA这个新视点为揭示胰岛素抵抗的发生机制奠定了基础，为2型糖尿病及其并发症的防治提供了新的思路。