左归降糖清肝方对db/db小鼠2型糖尿病合并非酒精性脂肪肝病脂质代谢的影响

Effect of Zuogui Jiangtang Qinggan Formula on lipid metabolism in db/db mouse model of type 2 diabetes mellitus complicated with non⁃alcoholic fatty liver disease

基于INSR/AMPK/SREBP⁃2信号通路,观察左归降糖清肝方对db/db小鼠2型糖尿病(type 2 diabetes mellitus,T2DM)合并非酒精性脂肪肝病(non⁃alcoholic fatty liver disease,NAFLD)脂质代谢的影响,探讨其治疗T2DM合并NAFLD的作用机制。随机将24只db/db小鼠分为4组:模型组,阳性药物组,左归降糖清肝方低、高剂量组。采用6只C57小鼠作为空白组。左归降糖清肝方高、低剂量组分别予左归降糖清肝方15、7.5 g·kg^(-1)灌胃,阳性药物组予二甲双胍0.067 g·kg^(-1)灌胃,空白组和模型组予等体积蒸馏水灌胃,连续6周。灌胃结束后,检测小鼠空腹血糖(fasting blood glucose,FBG),进行口服葡萄糖耐量实验以及血脂、肝功能水平,肝组织病理学分析脂质蓄积情况。Western blot检测胰岛素受体(insulin receptor,INSR)、AMP依赖的蛋白激酶[adenosine 5′⁃monophosphate(AMP)⁃activated protein kinase,AMPK]、p⁃AMPK、甾醇调节元件结合蛋白2(sterol⁃regulatory element⁃binding protein 2,SREBP⁃2)蛋白表达。结果显示,与模型组相比,经左归降糖清肝方治疗的小鼠体质量、肝重系数、空腹血糖、血清总胆固醇(total cholesterol,TC)、甘油三酯(triglyceride,TG)和低密度脂蛋白(low⁃density lipopro⁃tein,LDL)以及肝功能水平降低。肝组织病理学显示,与模型组相比,经左归降糖清肝方治疗的小鼠肝组织脂滴空泡明显减少、紫红色糖原颗粒明显增加。Western blot检测结果表明,与空白组比较,模型组INSR、p⁃AMPK/AMPK蛋白表达水平均显著下调,SREBP⁃2蛋白表达水平均显著上调。与模型组比较,左归降糖清肝方高剂量组INSR、p⁃AMPK/AMPK蛋白表达水平均显著上调,SREBP⁃2蛋白表达水平显著下调;左归降糖清肝方低剂量组INSR、p⁃AMPK/AMPK蛋白表达水平有上调趋势,SREBP⁃2蛋白表达水平有下调趋势,但差异无统计学意义。上述研究表明,左归降糖清肝方可能通过激活INSR/AMPK/SREBP⁃2信号通路,改善db/db小鼠胰岛素抵抗和脂质代谢。

This study aims to explore the effect of Zuogui Jiangtang Qinggan Formula(ZGJTQGF)on the lipid metabolism in the db/db mouse model of type 2 diabetes mellitus(T2DM)complicated with non⁃alcoholic fatty liver disease(NAFLD)via the insulin receptor(INSR)/adenosine 5′⁃monophosphate(AMP)⁃activated protein kinase(AMPK)/sterol⁃regulatory element⁃binding protein 2(SREBP⁃2)signaling pathway.Twenty⁃four db/db mice were randomized into positive drug(metformin,0.067 g·kg^(-1))and low⁃(7.5 g·kg^(-1))and high⁃dose(15 g·kg^(-1))ZGJTQGF groups.Six C57 mice were used as the blank group and administrated with an equal volume of distilled water.The mice in other groups except the blank group were administrated with corresponding drugs by gavage for 6 consecutive weeks.At the end of drug administration,fasting blood glucose(FBG)and blood lipid levels were measured,and oral glucose tolerance test was performed.Compared with the blank group,the mice treated with ZGJTQGF showed decreased body mass and liver weight coefficient,lowered levels of FBG,total cholesterol(TC),triglyceride(TG),and low⁃density lipoprotein(LDL),and weakened liver function.The pathological changes and lipid accumulation in the liver tissue were examined.Western blot was employed to measure the protein levels of INSR,AMPK,p⁃AMPK,and SREBP⁃2.Compared with the blank group,the model group showed down⁃regulated protein levels of INSR and p⁃AMPK/AMPK and up⁃regulated protein level of SREBP⁃2.Compared with the model group,high⁃dose ZGJTQGF up⁃regulated the protein levels of INSR and p⁃AMPK/AMPK and down⁃regulated the protein level of SREBP⁃2.Low⁃dose ZGJTQGF slightly up⁃regulated the protein levels of INSR and p⁃AMPK/AMPK and down⁃regulated the protein level of SREBP⁃2,without significant differences.The results suggested that ZGJTQGF may alleviate insulin resistance and improve lipid metabolism in db/db mice by activating the INSR/AMPK/SREBP⁃2 signaling pathway.