Raf kinase inhibitor protein inhibits cell proliferation but promotes cell migration in rat hepatic stellate cells
Raf kinase inhibitor protein inhibits cell proliferation but promotes cell migration in rat hepatic stellate cells
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DOI：
10.1111/j.1478-3231.2009.01981.x
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发表时间：
2009-04-01
影响因子：
6.7
通讯作者：
Jiang, Huiqing
中科院分区：
文献类型：
--
作者：
Ma, Junji;Li, Fangfang;Jiang, Huiqing
肝星状细胞（HSC）在肝纤维化和肝硬化的发病机制中起重要作用。Raf激酶抑制蛋白（Raf kinase inhibitor protein，RKIP）是细胞外信号调节激酶（extracellular signal-regulated kinases，ERK）/丝裂原活化蛋白激酶（mitogen-activated protein kinase，MAPK）信号通路的抑制剂，具有抑制肿瘤转移的作用。有趣的是，RKIP促进Madin-Darby犬肾上皮细胞的细胞迁移。然而，RKIP对HSC行为的影响尚不清楚。本研究以新鲜分离的HSC和HSC-T6细胞为研究对象，探讨RKIP在HSC增殖、凋亡和迁移中的作用。通过Western印迹分析静止和活化的HSC中RKIP的量、RKIP、Raf和ERK的磷酸化（pRKIP、pRaf和pERK）。用表达RKIP的质粒转染HSC-T6细胞或用RKIP抑制剂locostatin处理HSC-T6细胞。用MTT法、TUNEL法和Transwell细胞迁移实验检测活化后HSC的增殖、凋亡和迁移情况，发现活化后HSC中RKIP蛋白表达下调，pRKIP、pRaf和pERK蛋白表达上调。RKIP过表达显著减轻了RKIP、Raf和ERK的磷酸化。这反过来又抑制了HSC增殖。Locostatin不仅抑制RKIP蛋白的表达，而且在一定程度上逆转了RKIP抑制的RKIP、Raf和ERK的磷酸化。RKIP增强HSC迁移和增强伤口闭合。Raf激酶抑制剂蛋白抑制ERK/MAPK信号通路，抑制HSC增殖。RKIP促进HSC迁移和伤口闭合。
Hepatic stellate cells (HSCs) play an important role in the pathogenesis of liver fibrosis and cirrhosis. Raf kinase inhibitor protein (RKIP), an inhibitor of extracellular signal-regulated kinases (ERK)/mitogen-activated protein kinase (MAPK) signalling pathway, has been proved to suppress tumor metastasis. Interestingly, RKIP promotes cell migration in Madin-Darby canine kidney epithelial cells. However, the effects of RKIP on HSC behaviours are unknown. The purpose of the present study is to investigate the role of RKIP in HSC proliferation, apoptosis and migration.Two types of cells, freshly isolated HSC and HSC-T6 cell line, were used in this study. The amount of RKIP, the phosphorylation of RKIP, Raf and ERK (pRKIP, pRaf and pERK) were analysed in quiescent and activated HSCs by Western blots. HSC-T6 cells were transfected with RKIP-expressing plasmid or treated with locostatin, a RKIP inhibitor. HSC proliferation, apoptosis and migration were evaluated with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labelling (TUNEL) staining and Transwell cell migration assay respectively.In activated HSCs, RKIP protein expression was downregulated whereas pRKIP, pRaf and pERK were upregulated. RKIP overexpression significantly mitigated the phosphorylation of RKIP, Raf and ERK. This in turn inhibited HSC proliferation. Locostatin not only inhibited RKIP protein expression but also, to some extent, reversed the RKIP-inhibited phosphorylation of RKIP, Raf and ERK. RKIP augmented HSC migration and enhanced wound closure. Locostatin reversed the effects of RKIP.Raf kinase inhibitor protein inhibits ERK/MAPK signalling and this inhibition impedes HSC proliferation. RKIP promotes HSC migration and wound closure.