PKC介导的自噬在肝纤维化中的作用

The role of PKC-mediated autophagy in liver fibrosis

目的探讨在PKC信号通路介导的自噬在肝纤维化小鼠肝组织中的变化及意义。方法本研究将C57BL/6小鼠随机分为两组:模型组和假手术对照组,其中模型组采用胆管结扎方法诱导小鼠肝纤维化,手术4周后取材。HE染色、Masson染色显示小鼠肝组织纤维化程度;免疫组化检测肝组织内LC3蛋白的表达;Western Blot法检测肝组织中α-SMA、PKCα、p-PKCα和LC3蛋白的表达,最后将人正常肝细胞L02细胞转染PKCαsiRNA后检测LC3蛋白的表达变化并分析二者之间的关系。结果 HE和Masson染色结果显示胆管结扎诱导的肝纤维化模型组有明显的纤维增生。Western Blot结果显示模型组肝组织中α-SMA、PKCα和p-PKCα蛋白表达明显高于对照组(P<0.01),而Western Blot和免疫组化结果均显示LC3蛋白的表达则明显降低(P<0.05)。最后,L02细胞在转染PKCαsiRNA后显示LC3蛋白表达上调(P<0.01)。结论 PKCα信号通路的激活可以抑制自噬的发生,二者的相互作用可能与肝纤维化的发生和发展密切相关。

Objective To explore the change and importance of PKC-mediated autophagy in mice liver fibrosis tissue.Methods The C57BL/6mice were randomly divided into two groups：the bile duct ligation model and sham-operated groups.Mice in the model group were treated with bile duct ligation for induced liver fibrosis.After 4weeks of operation,the degree of hepatic fibrosis was evaluated by HE and Masson staining,firstly.Subsequently,the expression of LC3 in liver tissues was detected by immunohistochemistry assay,and expressions ofα-SMA,PKCα,p-PKCαand LC3 in liver tissues were also determined by Western Blot.Finally,the expression of LC3 was detected in L02 cells transfected with PKCαsiRNA and the relationship between PKCαand LC3 was assessed.Results More fibrotic liver tissues were found in model group according to HE and Masson staining.As a result,the expression ofα-SMA,PKCαand pPKCαwere significantly increased（P〈0.01）,while that of LC3 was decreased（P〈0.01）in liver tissues from model group as compared with those from the sham-operated group by Western Blot and Immunohistochemistry assay.After transfection of PKCαsiRNA in L02 cells,the level of LC3 was upregulated（P〈0.01）.Conclusion The inhibition of autophage by PKCαpathway may contribute to the occurrence and development of liver fibrosis.