Effect of RhoA on transforming growth factor β1-induced rat hepatic stellate cell migration
Effect of RhoA on transforming growth factor β1-induced rat hepatic stellate cell migration
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DOI：
10.1111/j.1478-3231.2012.02809.x
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发表时间：
2012-08-01
影响因子：
6.7
通讯作者：
Jiang, Wei
中科院分区：
文献类型：
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作者：
Li, Lei;Wang, Ji-Yao;Jiang, Wei
背景：尽管肝星状细胞(HSCs)的迁移在肝纤维化反应中是必不可少的，但调节其迁移的细胞内和细胞外信号知之甚少。目的探讨Rho鸟苷三磷酸酶(Rho GTPase)信号转导途径在转化生长因子β1(TGFβ1)诱导的HSC迁移中的作用。方法采用原代大鼠肝星状细胞和HSC-T6大鼠肝星状细胞系。用Transwell Boyden小室实验评估细胞迁移，而用激光共聚焦显微镜观察细胞骨架的变化。Western blotting检测Rho GTP酶(RhoA、rac1和CDC42)在HSC中的表达，谷胱甘肽S转移酶(GST)下拉实验检测Rho GTP酶的活性。最后，分析了RhoA在稳定表达成分活性(CA，Q63L)或显性阴性(Dn，T19N)RhoA突变体的HSC-T6细胞中对转化生长因子β1诱导的细胞迁移的特异性影响。结果RhoA激活后，转化生长因子β1诱导大鼠肝星状细胞骨架重塑和迁移。RhoA的激活水平决定了HSCs的运动性。结论这些发现加深了我们对调节HSC迁移的细胞内和细胞外信号的理解。此外，RhoA可能是治疗肝纤维化的候选靶点。
Background Although the migration of hepatic stellate cells (HSCs) is essential to the hepatic fibrotic response, the intracellular and extracellular signals that regulate their migration are poorly understood. Aims To investigate the role of Rho guanosine triphosphatase (Rho GTPase) signalling, specifically via RhoA, in transforming growth factor beta 1 (TGF beta 1)-induced HSC migration. Methods Both primary rat HSCs and the HSC-T6 rat hepatic stellate cell line were used in this study. Cell migration was evaluated using the Transwell Boyden Chamber assay, whereas cytoskeletal changes were observed using laser confocal microscopy. Western blotting was used to detect the expression of Rho GTPases (RhoA, Rac1 and Cdc42) in HSCs, and their activation was determined using glutathione S-transferase (GST) pull-down assays. Finally, the specific effects of RhoA on TGF beta 1-induced cell migration were analysed in HSC-T6 cells stably transfected with constitutively active (CA, Q63L) or dominant-negative (DN, T19N) RhoA mutants. Results Transforming growth factor beta 1 induced cytoskeletal remodelling and migration of rat HSCs following RhoA activation. The level of RhoA activation determined the motility of the HSCs. Conclusions These findings broaden our understanding of the intracellular and extracellular signals that regulate HSC migration. Furthermore, RhoA may be a candidate therapeutic target for hepatic fibrosis.