转基因Fah-/-肝损伤小鼠模型的构建  

Construction of Transgenic Fah^(-/-)Liver Injury Mouse Model

作　　者：代路路 王弋[1] 龙淑娴 严家荣[1] 刘月姝[1] 姚欣伶 楼彩霞[1] 邝少松[1] DAI Lulu;WANG Yi;LONG Shuxian;YAN Jiarong;LIU Yueshu;YAO Xinling;LOU Caixia;KUANG Shaosong(Guangdong Medical Laboratory Animal Center,Foshan 528248,China)

机构地区：[1]广东省医学实验动物中心,佛山528248

出　　处：《实验动物科学》2024年第1期18-24,共7页Laboratory Animal Science

基　　金：广东省科技计划项目(2019A030317015)。

摘　　要：目的 建立转基因Fah-/-肝损伤小鼠模型,探讨其肝损伤的机制。方法 选取SPF级C57BL/6J野生型小鼠作为WT组。取转基因Fah+/-肝损伤杂合子小鼠培育后,根据基因鉴定结果选取6~8周龄的Fah-/-纯合子小鼠24只,分为正常给予NTBC饮水组、停NTBC饮水1周组和停NTBC饮水2周组,称量体质量,采血分离血清,进行肝功能生化指标检测,处死动物,取肝组织进行病理学检查、免疫组化检测Fah蛋白酶的表达及采用Western blot方法检测Fah蛋白表达。结果 经基因鉴定,成功繁育Fah-/-纯合子小鼠24只,后续用于肝损伤机制研究结果表明,停NTBC 1周组、停NTBC 2周组肝功能生化指标ALT、AST、TBIL与正常给予NTBC组比较显著升高,而ALB显著降低,差异有统计学意义。肝组织病理HE染色结果表明,WT组及正常给予NTBC组肝组织结构正常,而停NTBC组出现肝细胞肥大、坏死及炎性细胞浸润等不同程度的病变,停NTBC时间越长,肝损伤的程度越严重;肝组织免疫组化结果表明,WT组Fah蛋白酶表达为强阳性,正常给予NTBC组Fah蛋白酶表达阴性,停NTBC 1周组和停NTBC 2周组肝细胞Fah蛋白酶表达弱阳性。Fah蛋白表达结果与免疫组化结果基本符合。结论 成功建立Fah-/-小鼠模型并对其肝损伤机制研究,为后续研究提供实验数据参考。Objective To establish a transgenic Fah-/-liver injury mouse model and explore the mechanism of liver injury.Method SPF C57BL/6J wild-type mice were selected as WT group.According to the result of gene identification,24 Fah-/-homozygous mice aged 6-8 weeks were selected and divided into the NTBC drinking water group,the NTBC drinking water stop group for one week,and the NTBC drinking water stop group for two weeks.The weight was weighed,blood was collected,serum was isolated,liver function biochemical indexes were tested,and the animals were sacrificed.The expression of Fah protease and Fah protein were detected by immunohistochemistry and Western blot.Result After gene identification,24 Fah-/-homozygous mice were successfully bred.The result of subsequent study on the mechanism of liver injury showed that ALT,AST and TBIL of liver function in the one-week and two-week NTBC cessation groups were significantly increased compared with those in the normal NTBC group,while ALB was significantly decreased,with statistical significance.Pathological HE staining result of liver tissue showed that the structure of liver tissue in WT group and normal NTBC group was normal, while different degrees of lesions such as hepatocyte hypertrophy, necrosis and inflammatory cell infiltration were observed in NTBC group. The longer the time of NTBC cessation, the more serious the degree of liver injury. The result of liver immunohistochemistry showed that the expression of Fah protease in WT group was strongly positive, the expression of Fah protease in normal NTBC group was negative, and the expression of Fah protease in hepatocytes of the one-week and two-week NTBC stop groups was weakly positive. The expression of Fah protein by Western blot was consistent with that by immunohistochemistry. Conclusion The Fah- / - mouse model was successfully established and the mechanism of liver injury was studied, so as to provide experimental data reference for subsequent studies.

关 键 词：Fah-/-小鼠 NTBC 肝损伤 延胡索酸乙酰乙酸水解酶(Fah) 

分 类 号：Q95-3[生物学—动物学]