TLR4在LPS诱导的大鼠肝内胆管组织损伤中的作用及机制

发布时间：2019-05-11 11:22

【摘要】：目的探讨TOLL样受体4(TLR4)在脂多糖(LPS)诱导的大鼠肝内胆管组织损伤中的作用及机制。方法将48只SD大鼠随机均分为四组,LPS+siRNA转染组和LPS+阴性病毒组经尾静脉分别注入siRNA-TLR4纯化腺病毒、阴性对照病毒,对照组和LPS组均经尾静脉注射等量生理盐水。给药后24 h,LPS组、LPS+siRNA转染组和LPS+阴性病毒组均经胆总管内缓慢注入LPS,对照组注入等量生理盐水。制模后48、72 h各组分别取6只大鼠,采集肝门部包含胆管的肝组织标本。采用HE染色观察肝门部肝内胆管组织炎性细胞浸润情况及胆管形态等病理变化。采用免疫组化法检测胆管上皮细胞TLR4、细胞角蛋白19(CK19)、波形蛋白(Vimentin)蛋白表达,采用RT-PCR法检测胆管组织TLR4、CK19、Vimentin mRNA表达。结果 LPS组造模48、72 h时胆管周围炎性细胞浸润且胆管间质水肿,伴周围纤维结缔组织轻度增生;LPS+阴性病毒组表现与LPS组相似;LPS+siRNA转染组造模48 h时胆管周围仅有少量炎性细胞,造模72 h时胆管周围炎性细胞稍增多,但较LPS组显著减轻。与对照组比较,其他三组造模48、72 h时胆管上皮细胞CK19蛋白及mRNA表达均降低,TLR4、Vimentin蛋白及mRNA表达均升高,组间比较P均0.01;LPS+siRNA转染组造模48、72 h时胆管上皮细胞CK19蛋白及mRNA表达均高于LPS组、LPS+阴性病毒组,TLR4、Vimentin蛋白及mRNA表达均低于LPS组、LPS+阴性病毒组,组间比较P均0.01。结论 LPS诱发肝内胆管组织发生炎症反应及纤维化损伤过程中产生大量TLR4,TLR4可导致肝胆管上皮细胞发生上皮间质转化,从而加重纤维化损伤程度。
[Abstract]:Aim to investigate the role and mechanism of TOLL-like receptor 4 (TLR4) in lipopolysaccharide (LPS)-induced intrahepatic bile duct injury in rats. Methods Forty-eight SD rats were randomly divided into four groups:, LPS siRNA transfection group and LPS negative virus group. SiRNA-TLR4 purified adenovirus was injected into the tail vein, and the negative control virus was injected into the tail vein respectively. The control group and the LPS group were injected with the same amount of normal saline through the tail vein. At 24 h after administration, LPS siRNA-transfected group and LPS-negative virus group were injected with the same amount of normal saline into the common bile duct by slow injection into the common bile duct of LPS, control group. At 48 h and 72 h after model, 6 rats in each group were taken and the liver tissue specimens containing bile duct in the hilum were collected. HE staining was used to observe the infiltration of inflammatory cells in the intrahepatic bile duct and the morphological changes of the bile duct. The expression of cytokeratin 19 (CK19) and vimentin (Vimentin) in bile duct epithelial cells (TLR4,) was detected by immunohistochemical method, and the expression of TLR4,CK19,Vimentin mRNA in bile duct tissue was detected by RT-PCR method. Results at 48 h and 72 h after LPS, inflammatory cells infiltrated around the bile duct and edema of the bile duct stroma, with slight proliferation of the surrounding fibrous connective tissue, and the expression of LPS-negative virus group was similar to that of the LPS group. There were only a few inflammatory cells around the bile duct in the LPS siRNA transfection group at 48 h, and a little more inflammatory cells in the bile duct around the bile duct at 72 h after the model was made, but decreased significantly than that in the LPS group. Compared with the control group, the expression of CK19 protein and mRNA in bile duct epithelial cells decreased and the expression of TLR4,Vimentin protein and mRNA increased in the other three groups (P < 0.01). The expression of CK19 protein and mRNA in bile duct epithelial cells of LPS siRNA transfection group was higher than that of LPS group, LPS negative virus group, TLR4,Vimentin protein and mRNA expression of LPS negative virus group were lower than that of LPS group, LPS negative virus group at 72 h after transfection, and the expression of TLR4,Vimentin protein and mRNA was significantly higher than that of LPS negative virus group (P < 0.01). Conclusion the inflammatory reaction of intrahepatic bile duct tissue induced by LPS and the production of a large number of TLR4,TLR4 in the process of fibrosis injury can lead to epithelial stroma transformation of hepatobiliary epithelial cells, thus aggravating the degree of fibrosis injury.
【作者单位】：河南省省立医院;遵义医学院附属医院;
【基金】：国家自然科学基金资助项目(81260085)
【分类号】：R575.62

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