Yang Zesong, Liu Fang, Qu Hua, Wang Hang, Xiao Xiaoqiu, Deng Huacong
Department of Endocrinology, The First Affiliated Hospital of Chongqing Medical University, 400016 Chongqing, PR China.
Laboratory of Lipid and Glucose Metabolism, The First Affiliated Hospital of Chongqing Medical University, 400016 Chongqing, PR China.
Mol Cell Endocrinol. 2015 Oct 15;414:111-9. doi: 10.1016/j.mce.2015.07.023. Epub 2015 Jul 26.
Diabetes mellitus is a leading cause of death and disability worldwide, which presents a serious public health crisis in China nowadays. It has been well recognized that excessive β-cell apoptosis is the key pathogenesis of diabetes, of which the mammalian target of rapamycin (mTOR) serves as the critical signaling pathway. Emerging evidence indicates that vitamin D deficiency acts as a potential risk factor for diabetes. The present study aims to test the hypothesis that 1 alpha, 25-dihydroxyvitamin D(3) [1, 25(OH)2D3] can inhibit β-cell apoptosis via the suppression of mTOR signaling pathway. β-cells (INS-1) were cultured in the context of normal glucose or high glucose media with or without 1, 25(OH)2D3 treatment. β-cell apoptosis was evaluated by inverted fluorescence microscope, flow cytometry and electron microscope, respectively. Quantitative RT-PCR and Western blotting were performed to assess the possible perturbations in mTOR signaling pathway. High glucose significantly increased β-cell apoptosis. Of importance, RT-PCR and Western blotting demonstrated that high glucose inhibited DNA-damage-inducible transcript 4 (DDIT4) and TSC1/TSC2, up-regulated Rheb/mTOR/p70S6K and enhanced expression of the apoptosis regulating proteins, such as phospho-Bcl-2, cytochrome C and cleaved caspase. Interestingly, 1, 25(OH)2D3 treatment reversed high glucose induced pathological changes in mTOR signaling pathway, restored expression of DDIT4 and TSC1/TSC2, blocked aberrant up-regulation of Rheb/mTOR/p70S6K and the apoptosis regulating proteins, and effectively inhibited β-cell apoptosis. Therefore, 1, 25(OH)2D3 treatment can effectively protects β cell against high glucose-induced apoptosis mainly via the suppression of mTOR signaling pathway, which may be considered as a potential therapy for patients with diabetes.
糖尿病是全球死亡和残疾的主要原因，目前在中国呈现出严重的公共卫生危机。人们已经充分认识到，β细胞过度凋亡是糖尿病的关键发病机制，其中哺乳动物雷帕霉素靶蛋白（mTOR）是关键的信号通路。新出现的证据表明，维生素D缺乏是糖尿病的潜在危险因素。本研究旨在验证1α,25-二羟基维生素D3[1,25(OH)2D3]可通过抑制mTOR信号通路来抑制β细胞凋亡这一假说。β细胞（INS-1）在正常葡萄糖或高葡萄糖培养基中培养，并进行或不进行1,25(OH)2D3处理。分别通过倒置荧光显微镜、流式细胞术和电子显微镜评估β细胞凋亡情况。进行定量逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹法（Western blotting）以评估mTOR信号通路中可能存在的扰动。高葡萄糖显著增加了β细胞凋亡。重要的是，RT-PCR和Western blotting结果表明，高葡萄糖抑制了DNA损伤诱导转录本4（DDIT4）和结节性硬化复合物1/2（TSC1/TSC2），上调了小G蛋白Rheb/mTOR/核糖体蛋白S6激酶（p70S6K），并增强了凋亡调节蛋白的表达，如磷酸化Bcl-2、细胞色素C和裂解的半胱天冬酶。有趣的是，1,25(OH)2D3处理逆转了高葡萄糖诱导的mTOR信号通路病理变化，恢复了DDIT4和TSC1/TSC2的表达，阻断了Rheb/mTOR/p70S6K和凋亡调节蛋白的异常上调，并有效抑制了β细胞凋亡。因此，1,25(OH)2D3处理主要通过抑制mTOR信号通路可有效保护β细胞免受高葡萄糖诱导的凋亡，这可能被视为糖尿病患者的一种潜在治疗方法。
