新型酒精性肝病小鼠模型的建立  

Construction of a new alcoholic liver disease mouse model

作　　者：黄东东[1] 沃璐璐[1] 阮昕[1] 许雅芊 龚一鸣[1] 杨琳希 李雪川[1] 康月苧 贺明[1] 

机构地区：[1]上海交通大学基础医学院病理生理学教研室细胞分化与凋亡教育部重点实验室,上海200025

出　　处：《上海交通大学学报（医学版）》2017年第7期906-913,共8页Journal of Shanghai Jiao tong University:Medical Science

基　　金：国家自然科学基金(81470841);上海市浦江人才计划(16PJ1405400)~~

摘　　要：目的·建立一种可靠的、模拟酒精性肝病(ALD)患者饮酒模式的新型ALD小鼠模型(ALDNM)。方法·在Lieber-De Carli模型基础上,结合Gao-Binge模型,使用自主设计的饲喂瓶和饲料配方,通过慢性喂养加急性灌胃法建立ALDNM模型:给予小鼠5 d液体对照饲料适应后,随机分为对照组和乙醇组,每组10只;乙醇组小鼠自第6日饲喂乙醇占热量比为30%的饲料10 d,第16日上午2组小鼠各自以31.5%乙醇或等热量的糊精溶液灌胃,9 h后收集标本。比较Lieber-De Carli模型和ALDNM模型小鼠的一般情况、肝脏病理学改变和血清学指标;利用油红O染色和三酰甘油(TAG)检测,明确ALDNM小鼠肝脏脂质含量;利用realtime PCR检测ALDNM模型肝组织中白介素-6(IL-6)、肿瘤坏死因子α(TNF-α)、脂肪酸合成酶(Fas)、超长链脂肪酸延伸酶6(Elovl6)、硬脂酰辅酶A去饱和酶(Scd1)等基因mRNA水平,Western blotting检测肝组织蛋白中磷酸化信号转导与转录激活因子(p-STAT3)的变化。结果·Lieber-De Carli模型小鼠一般状态较差,血清谷丙转氨酶(GPT)、谷草转氨酶(GOT)无明显变化。ALDNM模型中,乙醇组小鼠肝细胞极度肿胀呈圆形,体积增大,细胞质内充满大量脂肪空泡;油红O染色结果显示,乙醇组小鼠肝细胞内呈现深浅不等的油红O着色;乙醇组小鼠肝重指数、肝脏TAG含量、血清GPT和GOT较对照组显著升高,血清HDL明显降低;而且,ALDNM乙醇组小鼠肝脏中炎症通路和脂质合成代谢通路相关基因表达水平显著升高。结论·成功建立了ALDNM模型;该模型操作简单,构建时间短,费用低,能较好地模拟ALD患者饮酒模式和发病过程,且具有小鼠进食量平稳、重复性好、肝脏损伤明显等优点。Objective · To establish a reliable alcoholic liver disease mouse model （ALDNM） that mimics the drinking pattern of alcoholic liver disease （ALD） patients. Methods · Using the self-designed feeding tubes and liquid diet, ALDNM model was developed through chronic feeding combined with acute gavage of ethanol based on Lieber-DeCarli model and Gao-Binge model. C57BL/6 mice were administered with control liquid diet for adaptation for first 5 d, and then divided into pair-fed group and ethanol-fed group （10 mice each group）. Ethanol-fed mice were fed with the liquid diet in which ethanol accounts for 30% of total energy, while the pair-fed mice were fed with the control diet for 10 d. At the 16th day, ethanol-fed mice and pair-fed mice were respectively gavaged a single dose of 31.5% ethanol or isocaloric maltose dextrin, and euthanized 9 h later. Sera and livers were collected. The general physiological condition, hepatic tissue pathological changes and serum indexes between Lieber-DeCarli models and ALDNM models were compared. The liver lipids of ALDNM mice were determined by Oil red O （ORO） staining and hepatic triacylglyceride （TAG） test. Meanwhile, the mRNA levels of interleukin-6 （IL-6）, tumor necrosis factor α （TNF-α）, fatty acid synthase （Fas）, long chain fatty acid elongase 6 （Elovl6） and stearyl-CoA desaturase （Scd1） were detected by real-time PCR in ALDNM models. Western blotting was used to detect the changes of phosphorylated signal transduction and transcriptional activator （p-STAT3） in the livers. Results · Lieber-DeCarli model mice were generally in poor condition, and there was no significant change in serum glutamic-pyruvic transaminase （GPT） and glutamic-oxaloacetic transaminase （GOT） compared to pair-fed group. However, in ALDNM models, H-E staining showed that the hepatocytes of ethanol-fed mice were extremely swollen with round volume, increased cytoplasm and filled with large amounts of fat vacuoles. ORO staining analyses showed obvious

关 键 词：酒精性肝病 小鼠 动物模型 炎症信号通路 脂质合成信号通路 

分 类 号：R-332[医药卫生] R575.5