轻断食对2型糖尿病模型大鼠胰岛素抵抗及胰岛β细胞功能的作用及其机制研究

Study on effect and mechanism of light fasting on insulin resistance and pancreaticβ-cell function in rats with type 2 diabetes mellitus model

目的探讨轻断食对2型糖尿病大鼠胰岛素抵抗以及胰岛β细胞AMPK-mTOR-ULK1自噬信号通路的影响及其可能的机制。方法46只雄性Wistar大鼠中随机选取6只作为正常对照组(给予普通饲料饲养4 w),其余40只大鼠建立2型糖尿病模型(给予高脂饲料饲养,每5天腹腔注射1次STZ 25 mg/kg体重,共4 w)。40只2型糖尿病大鼠模型建立成功后,根据干预的不同分为模型组、模型+轻断食组、模型+运动干预组、模型+药物干预组、模型+运动+轻断食组。所有干预组大鼠同时每5天腹腔注射1次小剂量STZ 25 mg/kg体重,正常对照组同时每5天腹腔注射1次生理盐水0.5 mL/100 g体重。干预结束行简易糖耐量试验(OGTT),检测各组大鼠空腹血糖、空腹血清胰岛素水平,计算出胰岛素抵抗指数(HOMA-IR)、胰岛β细胞功能指数(HOMA-β),评价干预效果。Western-blot法检测干预前、后大鼠胰腺组织中AMPK-mTOR-ULK1信号通路及自噬相关蛋白腺苷酸活化蛋白激酶(AMPK)、磷酸化-AMPK(p-AMPK)、哺乳动物类雷帕霉素靶蛋白(mTOR)、磷酸化-mTOR(p-mTOR)、UNC-51样激酶1(ULK1)、微管相关蛋白轻链3(LC3)、卷曲螺旋状Myosin样Bcl-2相互作用蛋白-1(Beclin-1)的表达水平。结果与模型组大鼠比较,模型+轻断食组、模型+运动+轻断食组大鼠的空腹血糖、空腹胰岛素及服糖后2 h血糖水平、HOMA-IR水平显著降低(P均<0.05);大鼠胰腺组织Beclin-1、p-AMPK、ULK1表达均显著降低,p-mTOR表达明显升高(P<0.05)。与模型组大鼠比较,模型+运动+轻断食组的HOMA-β水平显著升高(P<0.05),大鼠胰腺组织中的LC3表达显著降低(P<0.05)。结论轻断食干预对2型糖尿病大鼠具有改善胰岛素抵抗、提升胰岛β细胞功能的作用,并影响细胞自噬AMPK-mTOR-ULK1信号通路。

Objective To explore the effects of light fasting on insulin resistance and pancreaticβ-cells AMPK-mTOR-ULK1 autophagy signaling pathway and its possible mechanisms in type 2 diabetic rats.Methods 6 out of 46 male Wistar rats were randomly selected as the normal control group(given normal chow for 4 w),and the remaining 40 rats were established as the type 2 diabetes model(given high-fat chow and intraperitoneal injections of STZ 25 mg/kg once every 5 days for 4 w).40 type 2 diabetes rats were divided into model group,model+light fasting group,model+exercise intervention group,model+drug intervention group,and model+exercise+light fasting group,according to the different interventions.All rats in the intervention groups were simultaneously injected intraperitoneally with a small dose of STZ 25 mg/kg body weight once every 5 days,and the normal control group was simultaneously injected intraperitoneally with saline 0.5 mL/100 g body weight once every 5 days.At the end of the intervention,a simple glucose tolerance test(OGTT)was performed to detect fasting blood glucose and fasting serum insulin levels in each group,and the insulin resistance index(HOMA-IR)and isletβ-cell function index(HOMA-β)were calculated to evaluate the effect of the intervention.Western-blot assay was performed to detect the AMPK-mTOR-ULK1 signaling pathway and autophagy-related proteins adenylate-activated protein kinase(AMPK),phosphorylated-AMPK(p-AMPK),mammalian target of rapamycin-like protein(mTOR),phosphorylated-mTOR(p-mTOR),UNC-51-like kinase 1(ULK1),microtubule-associated protein light chain 3(LC3)and coiled spiral Myosin like Bcl-2 interacting protein-1(Beclin-1)in the pancreas before and after intervention.Results Compared with model group rats,the fasting blood glucose,fasting insulin,blood glucose level 2 hours after taking glucose,HOMA-IR level of the model+light fasting group,model+exercise+light fasting group rats were significantly reduced(P<0.05).The expression of Beclin-1,p-AMPK and ULK1 in rat pancreatic tissue was significantly reduced,while the expression of p-mTOR was significantly increased(P<0.05).Compared with the model group rats,the level of HOMA-βin the model+exercise+light fasting group was significantly higher(P<0.05),and the expression of LC3 in the pancreatic tissue of rats was significantly lower(P<0.05).Conclusion Light fasting intervention has the effect of improving insulin resistance and elevating the function of pancreaticβ-cells in type 2 diabetes model rats,and affects the cellular autophagy AMPK-mTOR-ULK1 signaling pathway.