丹参素对肝星状细胞RhoA/RockⅠ信号通路的影响

The effect of Tanshinol on RhoA/RockⅠsignaling parthway in hepatic stellate cells

目的:观察丹参素对肝星状细胞(Hepatic stellate cells,HSC)RhoA/RockⅠ信号通路的影响。方法:体外分离、培养大鼠肝HSC,以RockⅠ特异性抑制剂Y-27632 30μmol/L、0.062 5～0.25 mmol/L丹参素分别与纤维连接蛋白(Fibronectin,FN)刺激的HSC共同孵育。四甲基偶氮唑盐法测定HSC增殖,酶联免疫吸附试验测定Ⅰ型胶原分泌,Western blot测定RhoA,p-Mlc(Thr18/Ser19)p,-FAK(Tyr397)蛋白表达。结果:FN作用24h能刺激HSC增殖,Ⅰ型胶原分泌,且上调了RhoAp、-Mlc(Thr18/Ser19)蛋白,p-FAK(Tyr397)蛋白的表达;丹参素和RockⅠ特异性抑制剂Y-27632均能明显抑制上述效应。结论:FN刺激活化了HSC的RhoA/RockⅠ信号通路、整合素信号通路;丹参素抑制HSC增殖,胶原分泌的作用与阻断RhoA/RockⅠ信号通路有关;RhoA/RockⅠ信号通路阻断后对整合素信号通路关键酶FAK下调效应为丹参素经RhoA/RockⅠ信号通路抗纤维化的分子机制之一。

Objective：To investigate the role of Tanshinol on RhoA/RockⅠsignaling parthway in Hepatic Stellate Cells.Methods：The mice of HSCs were isolated.After stimulation of FN,HCSs were cultured in vitro with Tanshinol（0.062 5～0.25 mmol/L）or Y-27632（30 μmol/L）,a specific RockⅠinibitor.The cell proliferation was assessed by MTT assay;the secretion of collagen Ⅰwere examined by methods of Enzyme Linked Immunosorbent Assay;the protein levels of RhoA,p-Mlc（Thr18/Ser19）,and p-FAK（Tyr397） were analyzed by Western Blot.Results：FN promoted the proliferation of HSC significantly and provoked the secretion of collagenⅠmarkedly,meanwhile the protein levels of RhoA,p-Mlc（Thr18/Ser19）,and p-FAK（Tyr397） also increased.The interventions of Y-27632 and Tanshinol could inhibits the above effects in HSC distinctly.Conclusion：FN activates the RhoA/RockⅠsignal parthway and the intergin signal parthway in HSC;Interventing the RhoA/RockⅠsignal parthway might be involved with the effect of Tanshinol that inhibits the HSC proliferation and decreases the secretion of collagenⅠ.The down regulation of FAK by RockⅠis one of the anti-fibrosis mechanisms of Tanshinol.