2型糖尿病相关非酒精性脂肪性肝病老龄小鼠模型的建立

Establishment of type 2 diabetes with non-alcoholic fatty liver disease in aged mice

目的:探讨2型糖尿病(T2DM)合并非酒精性脂肪性肝病(NAFLD)老龄小鼠模型的建立方法。方法:18周龄C57BL/6J雄性小鼠随机分为正常对照组与模型组,每组8只。模型组小鼠全程给予高脂饲料(HFD,60%Fat)喂养6周,对照组采用普通饲料喂养。第3周开始,模型组小鼠连续5 d腹腔注射50 mg/kg链脲佐菌素(STZ),对照组给予等体积柠檬酸/柠檬酸钠溶液腹腔注射。注射结束后3 d和1周分别尾尖采血测禁食4 h后血糖,>11.0 mmol/L即为糖尿病造模成功。监测小鼠体重与一般情况,继续HFD饲喂至第6周造模结束。腹腔麻醉、眼球采血,分离心脏、肝脏及肾脏,进行血浆生化与组织病理检测。结果:6周HFD喂养联合STZ造模后,模型组小鼠肝脏体积增大,H&E染色显示肝细胞内出现脂肪空泡与气球样改变,小叶结构破坏,并伴有炎症细胞浸润增加;心肌与肾皮质中炎症细胞浸润、组织结构紊乱。同时,血浆中谷丙转氨酶、谷草转氨酶及胱抑素C水平升高,提示肝肾功能损伤发生。血浆葡萄糖、总胆固醇、三酰甘油、低密度脂蛋白及高密度脂蛋白胆固醇水平均显著增加。肝脏油红O染色显示脂滴大量蓄积,肝内总胆固醇与三酰甘油含量增加。结论:老龄小鼠中通过HFD喂养联合低剂量STZ多次注射能够诱发糖尿病与肝脂肪变性,从而快速有效地建立T2DM合并NAFLD的老龄小鼠模型。

Objective:To investigate the establishment of type 2 diabetes mellitus(T2DM)with non-alcoholic fatty liver disease(NAFLD)in aged mice.Methods:A total of 18-week-old C57BL/6J male mice was randomly divided into a control group and model group(n=eight mice per group).Mice in the model group were fed with high fat diet(HFD,60%Fat)for six weeks,while mice in the control group fed with normal chow diet.At the start of the third week,mice in the model group were intraperitoneally injected with 50 mg/kg streptozotocin(STZ)for five consecutive days,while mice in the control group were injected with an equal volume of citric acid/sodium citrate solution.Blood samples were collected from the tail tip three days and one week after the last injection and the blood glucose was measured after fasting for four hours,which>11.0 mmol/L was considered as modeling success.The body weight and general condition of mice were monitored until the end of modeling at week six.After anesthesia,the blood,heart,liver,and kidney were collected,and plasma biochemical and histopathological examinations were conducted.Results:After HFD feeding combined with multiple injections of low-dose STZ for six weeks,the liver mass of mice in the model group were increased.H&E staining showed abundant lipid droplet vacuoles and balloon-like changes in hepatocytes,destruction of lobule structure,and infiltration of inflammatory cells.The myocardium and renal cortex also suffer from inflammation and structure disorders.Plasma levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),and cystatin C(CysC)elevated remarkably,indicating the occurrence of hepatic and renal injury.Plasma glucose,total cholesterol(TC),triglyceride(TG),LDL-C,and HDL-C levels were also significantly increased.Oil red O staining revealed deposition of lipid droplets in the liver,combined with increased contents of hepatic TC and TG.Conclusion:HFD combined with multiple injections of low-dose STZ could induce diabetes mellitus and hepatic steatosis in aged mice,so as to rapidly and effectively establish the aged mice model of T2DM complicated with NAFLD.