大黄酸激活SIRT1信号通路对胰岛α细胞的作用及机制研究

Dysfunction of beta-cell and alpha-cell plays a very important role in the development of diabetes. Correcting alpha-cell dysfunction becomes, therefore, an attractive and rational therapeutic approach, particularly in the light of new treatments that may directly act on these pathogenetic mechanisms of type 2 diabetes. SIRT1, one kind of longevity gene, is a key regulator in cell cycle, response to DNA damage, metabolism, apoptosis and autophagy. Our preliminarily data indicates that over expression of SIRT1 decreased the glucagon secretion in alpha TC1-6 cell, but the exact mechanism remains unclear. Rhein, a natural compound from Rheum palmatum L, has been found to protect diabetes and related metabolic disorders. However, the effect of rhein on alpha-cell function has not been explored. In this project, we plan to use the beta-cell deleting mice models and alpha TC1-6 cell, to invest the effect of rhein on glucagon secretion, alpha cell proliferation, inflammation, oxidative stress, and insulin signaling in vivo and vitro. Further, we will explore whether the effect of rhein on alpha-cell function via activating SIRT1 pathway. This finding will contribute to uncover the new molecular mechanism of rhein in diabetes and provide scientific evidence of Chinese traditional medicine for prevention and treatment of diabetes by targeting alpha-cell function.

血糖稳态失衡的本质在于胰岛α和β细胞双重功能障碍，以α细胞为靶点的药物成为糖尿病领域的研究新热点。本课题组长期致力于胰岛α细胞信号通路的基础研究工作。我们在前期研究中发现去乙酰化酶SIRT1，作为一种机体基因转录、能量代谢和细胞衰老的关键调控分子，与α细胞胰高血糖素的分泌密切相关。同时发现中药单体大黄酸能有效抑制胰高血糖素分泌，其作用是独立于β细胞的。据此提出科学假说：大黄酸激活胰岛α细胞SIRT1信号通路，调控下游FOXO1、PPARγ、NF-κB、PTP1B等多个效应分子，通过下调胰高血糖素合成转录因子、抑制α细胞增殖、减轻氧化应激及炎症，促进胰岛素信号转导等途径，改善α细胞功能，维持血糖稳态平衡。本课题将拟用基因转染、离体葡萄糖灌流、体外研究及干扰等技术证实假说，旨在阐明大黄酸改善胰岛α细胞功能的分子机制，为中药针对α细胞治疗糖尿病提供新的理论基础及科学依据。

近年来α细胞功能对血糖稳态及胰岛功能的影响已成为糖尿病领域的研究热点，以α细胞为靶点可能为糖尿病的防治提供新的手段和途径。本项目选取高脂饲养联合链脲佐菌素诱导的糖尿病模型，采用免疫组化、荧光染色、RT-PCR、ELISA等方法证实大黄酸降低糖尿病小鼠空腹血糖、改善糖耐量异常的作用主要是通过抑制胰岛α细胞胰高血糖素的分泌。为了进一步证明大黄酸对胰岛α细胞的直接作用，我们采用高糖孵育α细胞株TC1-6细胞模型，通过膜片钳单通道电流记录技术、流式细胞、Real-Time PCR、ELISA、Western blot及体外干预等方法发现大黄酸能直接抑制胰高血糖素分泌，其机制与降低α细胞膜上ATP敏感钾通道（KATP）电流、减少细胞内钙离子浓度有关。在此基础上，我们进一步探讨了大黄酸降低KATP电流的分子机制，结果表明可能与α细胞的SIRT1表达上调有关。以上研究为大黄酸在糖尿病领域的进一步应用提供药理学依据，也为拓宽中药抗糖尿病提供新的策略。