口服抗生素对NASH大鼠肝组织TLR4信号通路的影响

The changes of hepatic Toll-like receptor 4 signaling by oral administration of ciprofloxacin in rats with nonalcoholic steatohepatitis induced by high-fat diet

目的观察口服抗生素对非酒精性脂肪性肝炎大鼠代谢性内毒素血症激活肝脏4型Toll样受体(TLR4)信号通路的影响。方法将30只SD大鼠随机分为正常对照组、模型组和干预组,后者给予高脂饮食喂养,正常组予普通饲料喂养。干预组大鼠自第9周起给予环丙沙星灌胃。造模12周末,比较各组门静脉血内毒素、谷丙转氨酶、谷草转氨酶、甘油三酯和空腹血糖水平;计算非酒精性脂肪性肝病评分;采用Real time PCR及Westernblot法检测大鼠肝脏TLR4、胰岛素受体底物-1(IRS-1)mRNA及蛋白表达水平;采用ELISA法检测血清和肝匀浆肿瘤坏死因子-α(TNF-α)和白细胞介素-6(IL-6)水平。结果模型组动物门静脉内毒素为0.361±0.018EU/ml,而正常组为0.324±0.013EU/m(l P<0.05);肝脏TLR4 mRNA水平为正常组的7.7倍,IRS-1 mRNA水平下降69%,血清及肝脏TNF-α分别为105.7±30.1pg/ml和608.7±78.8pg/ml,IL-6分别为60.9±12.5pg/ml和756.4±90.8pg/ml,均显著高于正常组(P<0.05);与模型组比,干预组门静脉血内毒素为0.341±0.016EU/m(lP<0.05),NAS积分为4.40±0.26(P<0.05),肝脏TLR4 mRNA和蛋白表达水平分别下降44%和14%,肝脏IRS1 mRNA和蛋白表达水平分别增加2.3倍和1.6倍,TNF-α和IL-6水平分别为531.1±64.6pg/ml和575.1±84.5pg/m(lP<0.05)。结论 NASH大鼠肝脏TLR4信号通路被激活,口服抗生素可减少肠源性内毒素血症,减轻肝脏炎症。

Objective To observe the influence of oral administration of ciprofloxacin .on hepatic TLR4 signaling activated by metabolic endotoxemia in rats with nonalcoholic steatohepatitis （NASH） induced by high-fat diet. Methods Thirty male SD rats were divided randomly into model and intervention group fed with high-fat diet and normal group fed with normal diet. Rats in intervention group were administrated with oral ciprofloxacin from 9th week. At the end of 12th week,the endotoxin level in portal vein,fasting blood glucose and serum lipid were detected. NAS score was evaluated. The protein and mRNA levels of hepatic TLR4 and IRS-1 were detected by real time PCR or Western bloting. Liver and serum pro-inflammatoly cytokines levels were tested by ELISA. Results Serum endotoxin level in model group was significantly higher than that in normal group （0.361±0.018 EU/ml vs. 0.324±0.013 EU/ml,P〈0.01）;NAS score in ciprofloxacin-intervened group declined（4.40 ±0.26 vs. 6.9± 0.3 in model,P〈0.05）;TLR4 mRNA levels and its protein expression in model groups were 8.7 times and 1.4 times higher than that in normal group,and declined by 51% and 14% in intervention group;Compared with the normal group,IRS-I mRNA and its protein expressions in model group declined by 69% and 47%,and increased 2.3 times and 1.6 times in intervention group;Serum and hepatic TNF-α and IL-6 levels in model rats increased significantly （P〈O.05）,which decreased in intervention groups. Conclusion Metabolic endotoxemia activates TLR4 signaling in liver and might play a role in the onset of NASH. Oral ciprofloxacin administration could reduce gutderived endotoxin.