黄芪-葛根药对通过PI3K/Akt/FoxO1通路调控糖异生作用治疗糖尿病大鼠作用机制

Mechanism of Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) Herb Pair Regulating Gluconeogenesis Through PI3K/Akt/FoxO1 Pathway in Diabetic Rats

目的 观察黄芪-葛根(芪葛)药对对2型糖尿病(T2DM)大鼠肝组织磷酯酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/叉头框转录因子1(FoxO1)通路及磷酸烯醇式丙酮酸羧基酶(PEPCK)、限速酶葡萄糖6-磷酸酶(G6Pase)蛋白表达的影响以探讨其治疗T2DM作用机制。方法 采用高脂饲料联合一次性小剂量尾静脉注射链脲佐菌素(STZ)方法建立T2DM大鼠模型;成模后除正常组10只外,设芪葛低、中、高剂量组,阳性药组及模型组5组,将成模大鼠随机平均分入。各组大鼠连续药物干预4周后,麻醉,取材后对大鼠血清FBG、FINS、肝功、血脂及糖原水平进行检测;苏木素-伊红法染色观察肝脏组织病理形态学改变;蛋白免疫印迹法检测肝脏组织PI3K/Akt/FoxO1信号通路中相关蛋白及G6Pase、PEPCK蛋白水平。结果 相比于模型组,芪葛高、中剂量组FBG及FINS水平均降低,糖原水平显著升高(P<0.05,P<0.01);各给药组甘油三酯(TG)、谷草转氨酶(AST)、谷丙转氨酶(ALT)及低密度脂蛋白(LDL-C)均显著降低(P<0.01,P<0.05)且与阳性药组差异无统计学意义(P>0.05);芪葛高、低剂量组TC水平降低(P<0.05)且与阳性药组差异无统计学意义(P>0.05)。HE染色结果发现,与模型组相比,各给药组大鼠肝脏组织体积及色泽均有明显改善,虽仍有不同程度的肝组织结构紊乱、肝细胞脂肪变性、肝细胞肿胀、胞浆内可存在大小、数量均不等的脂滴空泡,但显著优于模型组大鼠肝脏组织状态。与模型组相比,芪葛高剂量组FoxO1、G6Pase及PEPCK蛋白表达降低,Akt、PI3K、p-Akt、p-PI3K表达升高(P<0.05)。结论 芪葛药对可能是通过调控PI3K/Akt/FoxO1信号通路,降低G6Pase、PEPCK的表达,抑制糖异生作用,调节糖脂代谢,修复肝损伤,从而发挥治疗T2DM作用的。

Objective To explore the mechanism of Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) herb pair in the treatment of type 2 diabetes mellitus(T2 DM) by observing the effects of Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) herb pair on PI3 K/Akt/FoxO1 pathway and the expressions of phosphoenolpyruvate carboxykinase(PEPCK) and glucose 6-phosphatase(G6 Pase) protein in liver tissue of T2 DM rats.Methods T2 DM rat model was established by high-fat diet combined with one-time tail vein injection of streptozotocin(STZ).The rats with successful model replication were randomly divided into model group,positive drug group,Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) herb pair high,medium and low dose groups,and normal group.Rats in each group received continuous administration for 4 weeks,fasted 12 hours after the last administration.The rats can drink water and then they were anesthetized.After taking materials,the levels of fasting blood glucose(FBG),fasting insulin(FINS),triglyceride(TG),total cholesterol(TC),high density lipoprotein cholesterol(HDL-C),low density lipoprotein cholesterol(LDL-C) and glycogen were detected.HE staining was used to observe the pathomorphological changes of liver.The expression levels of PI3 K/Akt/FoxO1 signal protein,PEPCK and G6 Pase protein in liver were detected by Western blot.Results Compared with those of the normal group,the levels of FBG and FINS in the model group were significantly higher(P0.05).The levels of TC in Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) herb pair high and low dose groups were decreased(P0.05).The results of HE staining showed that compared with those of the model group,the volume and color of liver tissue in each administration group were significantly improved.Although there were still different degrees of liver tissue structural disorder,hepatocyte steatosis,hepatocyte swelling,and different sizes and quantities of lipid droplets vacuoles in the cytoplasm,it was significantly better than the state of liver tissue in the model group.Compared with those of the model group,the expressions of Akt,p-Akt,PI3 K and p-PI3 K protein were increased and the expressions of FoxO1,PEPCK and G6 Pase protein decreased in Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) herb pair high dose group(P<0.05).Conclusion Huangqi(Astragali Radix)-Gegen(Puerariae Lobatae Radix) herb pair may inhibit the expressions of PEPCK and G6 Pase and hinder liver gluconeogenesis by regulating PI3 K/Akt/FoxO1 signal pathway,so as to protect the liver of T2 DM rats and regulate T2 DM.