肿瘤坏死因子-α和结肠紧密连接蛋白影响急性肝衰竭小鼠肠黏膜屏障功能的分子机制  

Molecular mechanism of tumor necrosis factor alpha and colonic tight junction protein affecting intestinal mucosal barrier function in mice with acute liver failure.

作　　者：冶成贵 朵红梅 魏国月 童芝文 辛晓恩 YE Cheng-gui;DUO Hong-mei;WEI Guo-yue(Department of General Surgery,Fourth People's Hospital of Qinghai Province,Xining Qinghai 810000,China.;Department of Respiratory Medicine,Fourth People's Hospital of Qinghai Province,Xining Qinghai 810000,China.)

机构地区：[1]青海省第四人民医院普外科,青海西宁810000 [2]青海省第四人民医院呼吸内科,青海西宁810000 [3]青海省第四人民医院消化内科,青海西宁810000

出　　处：《临床和实验医学杂志》2021年第18期1924-1928,共5页Journal of Clinical and Experimental Medicine

基　　金：青海省卫生健康委科研指导性课题(编号:2019-wjzdx-86)。

摘　　要：目的探讨肿瘤坏死因子-α(TNF-α)和结肠紧密连接蛋白影响急性肝衰竭(ALF)小鼠肠黏膜屏障功能的分子机制。方法18只SPF级6~8周龄BALB/c雄性小鼠根据实验目的分为对照组(腹膜内注射0.9%氯化钠溶液),模型组(腹膜内注射D-半乳糖胺800 mg/kg体重),抗体治疗组(在ALF诱导前接受抗体治疗,接受抗TNF-αIgG,每只小鼠100μg),每组6只。通过蛋白印迹分析claudin-1、occludin和ZO-1的蛋白表达。通过定量实时聚合酶链反应(qRT-PCR)分析小鼠结肠紧密连接蛋白claudin-1、occludin和ZO-1的mRNA表达。通过酶联免疫吸附试验检测小鼠血清中TNF-α、IL-6、IL-1β、IL-8炎症因子的水平。通过全自动生化分析仪检测小鼠肝损伤指标丙氨酸氨基转移酶(ALT),总胆汁酸(TBA)的浓度。通过组织分析对小鼠肠黏膜完整性和组织损伤程度进行评分。通过细菌易位测定和异黄酮荧光素评估小鼠肠黏膜通透性。结果模型组较对照组claudin-1、occludin和ZO-1的蛋白表达降低(P<0.05),抗体治疗组较模型组表达升高(P<0.05)。模型组较对照组claudin-1、occludin和ZO-1的mRNA表达降低(P<0.05),抗体治疗组较模型组表达升高(P<0.05)。模型组较对照组TNF-α、IL-6、IL-1β、IL-8的血清浓度水平升高(P<0.05),抗体治疗组较模型组血清水平降低(P<0.05)。模型组较对照组ALT和TBA的浓度升高(P<0.05),抗体治疗组较模型组ALT和TBA的浓度降低(P<0.05)。模型组较对照组肠黏膜完整性评分和肠组织损伤评分升高(P<0.05),抗体治疗组较模型组肠黏膜完整性评分和肠组织损伤评分降低(P<0.05)。模型组较对照组的细菌菌落形成和FITC-D升高(P<0.05),抗体治疗组较模型组细菌菌落形成和FITC-D降低(P<0.05)。结论在ALF小鼠中,TNF-α通过抑制ZO-1、claudin-1和occludin表达而增加了肠通透性,破坏肠黏膜屏障功能。Objective To explore the molecular mechanism of tumor necrosis factor-α(TNF-α)and colonic tight junction protein affecting the intestinal mucosal barrier function of acute liver failure(ALF)mice.Methods Eighteen SPF-grade BALB/c male mice aged 6-8 weeks were divided into control groups(intraperitoneal injection of 0.9%sodium chloride solution),model group(intraperitoneal injection of D-galactosamine,GalN;800 mg/kg body weight),antibody treatment group(receiving antibody treatment before ALF induction,receiving anti-TNF-αIgG,100μg per mouse),6 mice in each group according to experimental requirements.The protein expression of claudin-1,occludin and ZO-1 was analyzed by western blot.The mRNA expression of claudin-1,occludin and ZO-1 in mouse colon was analyzed by quantitative real-time polymerase chain reaction(qRT-PCR).The levels of TNF-α,IL-6,IL-1βand IL-8 inflammatory factors in the serum of mice were detected by enzyme-linked immunosorbent assay.The concentration of alanine aminotransferase(ALT)and total bile acid(TBA),which were indicators of liver injury in mice,were detected by an automatic biochemical analyzer.The integrity of the mouse intestinal mucosa and the degree of tissue damage were scored by tissue analysis.The intestinal mucosal permeability of mice was evaluated by bacterial translocation assay and isoflavone fluorescein.Results Compared with the control group,the protein expression of claudin-1,occludin and ZO-1 was lower than that in the model group(P<0.05),and the expression in the antibody treatment group was higher than that in the model group(P<0.05).Compared with the control group,the expression of claudin-1,occludin and ZO-1 in the model group was lower(P<0.05),and the expression of the antibody treatment group was higher than that in the model group(P<0.05).The serum levels of TNF-α,IL-6,IL-1βand IL-8 in the model group were higher than those in the control group(P<0.05),and the serum levels of the antibody treatment group were lower than those in the model group(P<0.05).The conc

关 键 词：小鼠 急性肝衰竭 肿瘤坏死因子-Α 肠紧密连接蛋白 肠黏膜屏障 肠通透性 

分 类 号：R575.3[医药卫生—消化系统]