磺脲类药物抑制KATP通道增强内质网应激加剧2型糖尿病脑卒中风险性研究

Sulfonylureas are the most widely used oral hypoglycemic agents with a controversially increasing risk of stroke. Our preliminary meta-analysis results showed that sulfonylurea significantly increased the risk of stroke in T2DM patients. The further experiments are being needed to elucidate the role of sulfonylureas in increasing the risk of stroke in T2DM and the relevant molecular mechanism. Besides, our previous experiments confirmed that diabetes exacerbated stroke injury in mice and that sulfonylureas and ATP-sensitive potassium channel(KATP)openers aggravated or attenuated cerebral ischemic brain damage in mice, respectively. Neurovascular unit dysfunction is a key alteration in stroke, and its constituent cells all express KATP endogenously. Current studies have shown that KATP can inhibit endoplasmic reticulum stress and alleviate brain damage caused by high glucose or ischemia. Therefore, we hypothesized that sulfonylureas inhibit KATP channels enhancing endoplasmic reticulum stress, and thus to increase the risk of stroke in T2DM. This project aimed to study the effects of sulfonylureas on the risk of T2DM stroke and the relationship between KATP expression in neurovascular unit and stroke in T2DM through establishing the mice stroke model in T2DM mice and OGD cell model, combining KATP opener with knockdown of key proteins mediated endoplasmic reticulum stress to uncover the underlying molecular mechanism. The findings of this project will provide evidence for the clinical rational administer of sulfonylureas.

磺脲类药物为应用最广泛的口服降糖药，其是否增加脑卒中风险仍有争议。申请者前期Meta分析显示磺脲类药物显著增加T2DM患者脑卒中风险，但其作用及其分子机制仍不清楚。申请者前期研究表明磺脲类药物和ATP敏感性钾通道（KATP）开放剂分别加剧及减轻小鼠缺血性脑损伤。神经血管单元功能异常是脑卒中关键病变，其组成细胞均内源性表达KATP，而KATP能抑制内质网应激而减轻高糖或缺血所致脑损伤。据此，申请者提出研究假说磺脲类药物通过抑制KATP，增强内质网应激而增加T2DM脑卒中风险。本项目拟通过建立T2DM脑卒中小鼠模型和高糖条件下缺血缺氧细胞损伤模型，结合KATP开放剂和沉默内质网应激关键蛋白研究磺脲类药物增加T2DM脑卒中风险及其与神经血管单元KATP表达的关系，并揭示增强内质网应激为其关键分子机制，为磺脲类药物临床合理应用提供依据。

我们的研究成果显示，不同代数磺脲类药物对T2DM脑卒中发生的风险存在差异，其中甲苯磺丁脲、格列本脲可增加pMCAO后小鼠脑梗塞面积，而格列美脲显示中性结果。同时我们通过体内实验证实磺脲类药物甲苯磺丁脲做为KATP抑制剂通过加强血脑屏障的通透性，促进神经血管单元组成细胞凋亡及炎症反应，增加内质网应激相关蛋白表达进而产生脑卒中脑损伤。.我们阐明了T2DM小鼠神经血管单元KATP（Kir6.x、SUR1/2）表达谱。最后我们制备了各个神经血管单元组成细胞高糖条件下细胞缺血缺氧体外细胞模型，经甲苯磺丁脲处理后进行OGD，通过检测内质网应激相关蛋白表达；检测ROS水平；WB检测凋亡相关蛋白在体外水平证实磺脲类药物增加 T2DM 脑卒中风险的主要机制是通过抑制神经血管单元KATP，进而增强内质网应激GSK3β通路，从而介导炎症反应、氧化应激和细胞凋亡。我们的研究为临床应用磺脲类药物治疗T2DM所存在的潜在脑卒中发生的风险提供了实验依据。