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Extracellular matrix in liver cirrhosis : its abnormality and influence on liver functions.

肝硬化细胞外基质的异常及其对肝功能的影响。

基本信息

批准号：
09670512
负责人：
OGATA Itsuro
金额：
$ 2.11万
依托单位：
University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09670512/
关键词：
extracellular matrix Rho-kinase cirrgotic liver phospholylation Hepatic stellate cell Rho Rho-kinase Rho Kinase

项目摘要

In cirrhotic liver, abundant extracellular matrices and deranged lobular architecture deteriorate the organ-specific function. Hepatic stellate cells (HSCs) transform into myofibroblast-like cells and play an important role in the hepatic fibrogenesis by producing an excess amount of extracellular matrix proteins and contracting the healing wound. We have cloned a serine/threonine protein kinase from a cDNA library prepared from HSC line derived from a CCI_4 induced cirrhotic rat liver. It is identical to the recently-reported Rho-kinase, one of Rho effectors, through which several actomyosin-based cellular processes such as adhesion, cytokinesis, and contraction are regulatcd. This study mainly aimed to clarify the functions of Rho and Rho-kinase in HSCs.Immunoblot analysis confirmed the protein expression and auto-phosphorylation of Rho-kinase by HSCs, as well as their regulation by Rho-stimulator lysophosphatidic acid (LPA). Contraction assay using type I collagen lattice showed that LPA induced contraction of HSCs in a dose-dependent manner, and its effect was reduced by the addition of C3 exoenzyme, an inactivator of Rho. Protein expression of alpha-smooth muscle actin, a cytoskeletal protein in activated HSCs, was not affected by addition of LPA or C3 exoenzyme. When HSCs were added with fibronectin-coated beads, the subplasmalemmal assembly of focal adhesion complex and related proteins including Rho and Rho-kinase were observed arround the beads, and C3 exoenzyme abrogated the assembly.From these data, Rho and Rho-kinase might be involved in the intracellular transduction of the signal from extracellular matrices in HSCs.

在肝硬化中，丰富的细胞外基质和紊乱的小叶结构使器官特异性功能恶化。肝星状细胞（Hepatic stellate cells, hsc）转化为肌成纤维细胞样细胞，通过产生过量的细胞外基质蛋白和收缩愈合伤口，在肝纤维化过程中发挥重要作用。从CCI_4诱导的肝硬化大鼠肝HSC细胞系cDNA文库中克隆出丝氨酸/苏氨酸蛋白激酶。它与最近报道的Rho效应器之一Rho激酶相同，通过Rho效应器调节一些基于肌动球蛋白的细胞过程，如粘附、细胞质分裂和收缩。本研究主要旨在阐明Rho和Rho激酶在造血干细胞中的功能。免疫印迹分析证实了造血干细胞对rho激酶的蛋白表达和自身磷酸化，以及rho刺激剂溶血磷脂酸（LPA）对其的调节。I型胶原晶格收缩实验表明，LPA诱导hsc呈剂量依赖性收缩，加入Rho灭活剂C3外酶可减弱其作用。在活化的造血干细胞中，α -平滑肌肌动蛋白（一种细胞骨架蛋白）的蛋白表达不受LPA或C3外泌酶的影响。当造血干细胞中加入纤维连接蛋白包被的微球时，在微球周围观察到局灶黏附复合物和Rho、Rho激酶等相关蛋白的质下组装，C3外泌酶破坏了这种组装。从这些数据来看，Rho和Rho激酶可能参与了造血干细胞细胞外基质信号的细胞内转导。