短链脂肪酸通过激活GPR43上调AMPK信号通路抑制小鼠胰岛β细胞凋亡

Short-chain fatty acid activation of GPR43 upregulates AMPK signaling pathway and inhibits apoptosis of mouse islet β cells

目的探究短链脂肪酸对小鼠胰岛β细胞的调节作用及相关机制。方法将C57BL/6J小鼠随机分为正常组、模型组、乙酸钠治疗组、丙酸钠治疗组,每组10只。其中模型组、乙酸钠治疗组、丙酸钠治疗组腹腔注射链脲佐菌素(STZ)诱导1型糖尿病。乙酸钠治疗组、丙酸钠治疗组分别每日灌胃乙酸钠和丙酸钠,正常组、模型组灌胃同等剂量生理盐水。监测各组小鼠体重、血糖变化、口服糖耐量和血清中胰岛素水平。实时荧光定量聚合酶链反应(RT-qPCR)检测小鼠胰岛GPR43、bcl-2、caspase3表达水平,Western blotting检测小鼠胰岛GPR43、AMPK、p-AMPK、bcl-2、caspase3表达水平。体外培养小鼠胰岛β细胞系min6,通过脂质体转染法将si-GPR43和si-NC转染至min6细胞,给予乙酸钠和丙酸钠共孵育,分组为si-NC组、si-NC+乙酸钠和丙酸钠组、si-GPR43组、si-GPR43+乙酸钠和丙酸钠组。RT-qPCR和Western blotting检测各组细胞bcl-2、caspase3表达水平,Annexin V-FITC流式细胞术检测各组细胞凋亡水平。结果与正常组相比,模型组小鼠体重、口服糖耐量和胰岛素水平降低、血糖升高(P<0.05);胰岛中caspase3表达水平升高,GPR43、bcl-2表达水平降低,p-AMPK蛋白表达水平降低(P<0.05)。与模型组相比,乙酸钠治疗组和丙酸钠治疗组小鼠体重、口服糖耐量和胰岛素水平升高、血糖降低(P<0.05);胰岛中caspase3表达水平降低,GPR43、bcl-2表达水平升高,p-AMPK蛋白表达水平升高(P<0.05)。与si-NC组相比,si-GPR43组p-AMPK蛋白表达水平降低(P<0.05)。与si-NC组相比,si-NC+乙酸钠和丙酸钠组caspase3表达水平降低,bcl-2表达水平升高,细胞凋亡率降低(P<0.05)。与si-NC组、si-NC+乙酸钠和丙酸钠组相比,si-GPR43组、si-GPR43+乙酸钠和丙酸钠组caspase3表达水平升高,bcl-2表达水平降低,细胞凋亡率升高(P<0.05)。结论短链脂肪酸可通过激活GPR43上调AMPK信号通路,从而抑制小鼠胰岛β细胞凋亡,缓解1型糖尿病。

Objective To investigate the regulation of short chain fatty acids on mouse isletβcells and its related mechanism.Methods C57BL/6J mice were randomly divided into 4 groups:normal group,model group,sodium acetate treatment group and sodium propionate treatment group,with 10 mice in each group.Model group,sodium acetate treatment group,sodium propionate treatment group intraperitoneal injection of streptozotocin(STZ)induced type 1 diabetes.Sodium acetate treatment group and sodium propionate treatment group were gastric injected with sodium acetate and sodium propionate,respectively,while normal group and model group were gastric injected with the same dose of normal saline.Body weight,blood glucose,oral glucose tolerance and serum insulin level were monitored.Real-time fluorescence quantitative polymerase chain reaction(RT-qPCR)was used to detect the expression levels of GPR43,bcl-2 and caspase3 in the islets of mice,and Western blotting was used to detect the expression levels of GPR43,AMPK,p-AMPK,bcl-2 and caspase3 in the islets of mice.Mouse isletβcell line min6 was cultured in vitro,si-GPR43 and si-NC were transfected into min6 by liposome transfection method,and co-incubated with sodium acetate and sodium propionate.The cells were divided into si-NC group,si-NC+sodium acetate and sodium propionate group,si-GPR43 group,si-GPR43+sodium acetate and sodium propionate group.The expression levels of bcl-2 and caspase3 were detected by RT-qPCR and Western blotting,and apoptosis levels were detected by Annexin V-FITC flow cytometry.Results Compared with normal group,body weight,oral glucose tolerance and insulin level of model group were decreased,and blood glucose was increased(P<0.05).The expression level of caspase3 increased,the expression level of GPR43 and bcl-2 decreased,and the expression level of p-AMPK protein decreased in islets(P<0.05).Compared with model group,body weight,oral glucose tolerance and insulin level were increased and blood glucose was decreased in sodium acetate and sodium propionate treatment groups(P<0.05).The expression level of caspase3 was decreased,the expression level of GPR43 and bcl-2 was increased,and the expression level of p-AMPK protein was increased(P<0.05).Compared with si-NC group,p-AMPK protein expression level in si-GPR43 group was decreased(P<0.05).Compared with si-NC group,the expression level of caspase3 was decreased,the expression level of bcl-2 was increased,and the apoptosis rate was decreased in si-NC+sodium acetate and sodium propionate groups(P<0.05).Compared with si-NC group,si-NC+sodium acetate and sodium propionate group,the expression level of caspase3 in si-GPR43 group,si-GPR43+sodium acetate and sodium propionate group was increased,the expression level of Bcl-2 was decreased,and the apoptosis rate was increased(P<0.05).Conclusion Short-chain fatty acids could inhibit the apoptosis of mouse isletβcells and alleviate type 1 diabetes by activating GPR43 and upregulating AMPK signaling pathway.