蒲黄总黄酮对长期游离脂肪酸诱导胰岛β细胞GPR40信号通路影响的研究

Impaired insulin secretion of pancreatic β-cells caused by chronically elevated free fatty acid (FFA) is strongly linked decreased activity of G protein coupled receptor-40 (GPR40) signaling which is activated by FFA. According to traditional Chinese medical pathogenesis of high-fatty-diet-induced diabetes and chronic illness leading to blood stasis，Pollen Typhae, a Chinese herb that performs to promote blood circulation and to remove blood stasis, has been widely used to treat type 2 diabetes in clinical pratice, but the mechanisms keep to be fully elucidated. Our study group found that Pollen Typhae total flavone (PTF),the main active ingredient of Pollen Typhae, ameliorated impaired insulin secretion of pancreatic β-cells induced by FFA in preliminary experiment, which was similar to that of GW9508, a GPR40 agonist, implicating that PTF had closed relationship with GPR40 signaling. Based on this results, the present study aims to observe the effects of PTF on insulin secretion in chronic FFA-induced pancreatic β-cells in vitro, and to explore the potential molecular mechanisms according to the effects of PTF on signal molecules in GPR40 signaling including GPR40, PLC, PKC by methods such as Western blot, Real-time PCR, which will provide scientific evidence for the clinical application of pollen Typhae and developing new forms of PTF in preventing and treating type 2 diabetes, and suggest lead compound to develop new drug to mediate insulin secretion.

长期高水平游离脂肪酸（FFA）损伤胰岛β细胞胰岛素分泌与FFA特异性G蛋白偶联受体-40（GPR40）信号通路活性下降密切相关。基于高脂饮食致消，久病入络中医2型糖尿病病因病机理论，活血化瘀中药蒲黄已广泛用于治疗2型糖尿病，但机制尚未完全清楚。课题组在预试验中发现蒲黄主要活性成分蒲黄总黄酮(PTF)能改善长期FFA诱导损伤胰岛β细胞胰岛素分泌，效应与GPR40激动剂GW9508相似，提示PTF与GPR40信号通路密切相关。本项目旨在前期研究基础上，体外研究探讨PTF对长期FFA诱导损伤胰岛β细胞胰岛素分泌的影响，并基于GPR40信号通路，应用Western blot，Real-time PCR等技术观察PTF对GPR40、PLC、PKC等信号分子的影响，从而揭示PTF潜在的分子机制，为临床应用蒲黄和开发PTF新剂型防治2型糖尿病提供科学依据，也为开发新的胰岛功能调节剂提供潜在的先导物。

2型糖尿病往往伴有高游离脂肪酸（FFA）血症，长期高水平FFA可损伤胰岛β细胞胰岛素分泌功能，导致其葡萄糖刺激胰岛素分泌（GSIS）减少，这与G蛋白偶联受体-40（GPR40）信号通路功能障碍密切相关。活血化瘀中药蒲黄已广泛用于治疗2型糖尿病。项目研究显示，蒲黄提取物蒲黄总黄酮（PTF）能够预防FFA诱导损伤胰岛β细胞GSIS，提高FFA诱导细胞GPR40蛋白和基因表达，增加细胞内三磷酸肌醇（IP3）、二酰基甘油（DAG）水平和一定程度促进蛋白激酶C（PKC）蛋白表达，并提高磷脂酶C（PLC）和PKC活性。此外，PTF还能提高FFA诱导胰岛β细胞ATP水平。结果提示，PTF极可能是通过GPR40信号通路预防FFA诱导损伤胰岛β细胞胰岛素分泌功能。项目研究揭示了蒲黄防治2型糖尿病的部分机制，为临床应用蒲黄和开发PTF新剂型防治2型糖尿病提供科学依据，也为开发新的胰岛功能调节剂提供潜在的先导物。