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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。
Objectives To establish a novel ALD mouse model (ALDNM) that models alcohol consumption in patients with alcoholic liver disease (ALD). Methods · Based on the Lieber-De Carli model, combined with the Gao-Binge model, the self-designed feeding bottle and feed formula were used to establish ALDNM model by chronic feeding plus acute gavage: mice were given 5 d liquid control diet , And were randomly divided into control group and ethanol group, with 10 mice in each group. The mice in ethanol group were fed 10% ethanol on the 6th day for 10 days, and the mice in the ethanol group were 31.5% Or other heat dextrin solution gavage, 9 h after the collection of specimens. The general situation, liver pathological changes and serological indexes of Lieber-De Carli model and ALDNM model mice were compared. The lipids of liver of ALDNM mice were determined by oil red O staining and triglyceride (TAG) assay. The levels of IL-6, TNF-α, Fas, Elovl6, stearoyl-CoA desaturase (Scd1) and other gene mRNA levels, Western blotting detection of liver tissue protein phosphorylation signal transducers and activators of transcription (p-STAT3) changes. Results · The mice in Lieber-De Carli model had poor general condition, and there was no significant change in serum GPT and GOT. In the model of ALDNM, the hepatocytes in mice in ethanol group were extremely swollen and round in shape, and their volume increased and the cytoplasm was filled with a large number of fat vacuoles. The results of oil red O staining showed that the red, The hepatic index, hepatic TAG content, serum GPT and GOT in ethanol group were significantly higher than those in control group, while HDL in serum was significantly decreased. Moreover, the expressions of inflammatory pathways and lipid metabolism pathway genes in liver of ALDNM ethanol group Significantly increased levels. Conclusions · The ALDNM model has been established successfully. The model has the advantages of simple operation, short construction time and low cost. It can simulate the pattern of alcohol consumption and pathogenesis in ALD patients well and has the advantages of stable food intake, good repeatability and obvious liver damage .