西方饮食联合小剂量四氯化碳构建非酒精性脂肪性肝炎小鼠模型研究

Establishment of non-alcoholic steatohepatitis mouse model induced by Western diet combined with low-dose carbon tetrachloride

目的·采用西方饮食(Western diet,WD)联合小剂量四氯化碳(CCl4)构建非酒精性脂肪性肝炎(non-alcoholic steatohepatitis,NASH)小鼠模型,并探究发生典型NASH病理改变的时间节点。方法·雄性8周龄C57BL/6小鼠采用WD饲料喂养联合每周1次腹腔注射CCl4(0.2 μL/g)的方法构建NASH模型,在造模后不同时间点检测小鼠空腹血糖、三酰甘油、谷丙转氨酶、谷草转氨酶水平,并于第24周行葡萄糖耐量试验;肝脏取材后计算肝指数,并通过油红O染色、天狼猩红染色、苏木精-伊红染色及TUNEL法等技术评估肝脏病理改变。结果·模型组与对照组相比,小鼠的空腹血糖、葡萄糖耐量试验结果间均无明显差异,但是在造模第8、12、24周2组肝指数间存在显著差异(均P<0.05)。第24周时,模型组三酰甘油、谷丙转氨酶、谷草转氨酶水平均显著高于对照组(均P<0.05)。油红O染色、天狼猩红染色、苏木精-伊红染色及TUNEL法结果显示,造模第8周,小鼠肝脏组织出现大量小脂滴堆积,肝细胞以凋亡为主;第16周,肝脏组织中出现大脂滴堆积、肝细胞气球样变性和点状坏死灶,肝细胞凋亡仍然较多;第24周,肝组织出现3期纤维化,点状坏死灶数目增多而脂肪变性减弱,肝细胞以增殖为主。结论·采用WD联合小剂量CCl4方法成功建立小鼠NASH模型,该模型能够在较短时间内模拟出NASH的病理特征。

Objective·To construct a non-alcoholic steatohepatitis(NASH)mouse model by the combination of Western diet(WD)and low-dose carbon tetrachloride(CCl4),and explore the time nodes of typical NASH pathological changes.Methods·Male 8-week C57BL/6 mice were fed WD and intraperitoneally injected with CCl4 at a dose of 2μL/g of body weight per week to construct NASH models.At different time points,the fasting blood glucose,and the levels of triacylglyceride,glutamic-pyruvic transaminase and glutamic-oxaloacetic transaminase were tested;glucose tolerance was tested at the 24th week.Besides,the liver index was calculated and oil red O staining,Sirius red staining,hematoxylin-eosin staining and TUNEL test were conducted to evaluate liver pathological changes after liver sampling.Results·Between the control group and model group,there was no significant difference in fasting blood glucose and glucose tolerance test result,while the significant differences of liver index were observed at the 8th,12th and 24th week(P<0.05).And at the 24th week,the levels of triacylglyceride,glutamic-pyruvic transaminase and glutamic-oxaloacetic transaminase were higher in the model group than those in the control group(P<0.05).According to the results of oil red O staining,Sirius red staining,hematoxylin-eosin staining and TUNEL test,in the model group,a large amount of small lipid droplets accumulation in the liver tissues was detected and hepatocytes were mainly in apoptotic state at the 8th week;large lipid droplets,hepatocellular ballooning and spot-like necrosis were observed,and hepatocyte apoptosis persisted at the 16th week;stage 3 fibrosis of liver was observed,and the number of spot-like necrosis increased but lipid droplets decreased,while hepatocytes were mainly in a proliferative state at the 24th week.Conclusion·The mouse model of NASH can be established successfully by WD combined with low-dose CCl4,which can simulate the pathologic features of NASH in a short time.