Activation of hepatocyte growth factor promotes apoptosis of hepatic stellate cells via the Rho pathway

Objective To investigate the role of activated hepatocyte growth factor (HGF) in apoptosis of hepatic stellate cells (HSCs) and in modulating the Rho signaling pathway.Methods HSCs were divided into the following groups:blank control,consisting of HSCs without treatment; two treatment controls,consisting of HSCs exposed to exogenous HGF at 50 ng/ml and HSCs exposed to exogenous HGF activator (HGFA) at 70 ng/ml; three experimental groups,consisting of HSCs exposed to both exogenous HGF and HGFA,HSCs pre-incubated with the HGF inhibitor c-met at 500 ng/ml for 6 hours and then exposed to exogenous HGF and HGFA,and HSCs preincubated with the Rho pathway inhibitor Y-27632 at 10 ng/ml and then exposed to exogenous HGF and HGFA.Activation status of the cultured HSCs was determined by change in expression of alpha-smooth muscle actin (SMA).The optimal intervention concentration ofY-27632 was determined by MTT assay.The apoptotic status of HSCs was determined by flow cytometry.Expression of the HGF-alpha chain was detected by immunofluorescence.The expression of RhoA was evaluated by PCR (for mRNA) and by immunohistochemical staining and Western blot analysis (for protein).Results Exposure to 10 μmol/L Y-27632 led to obvious growth inhibition ofHGF + HGFA-induced HSCs,compared with the other concentrations tested (P ＜ 0.05).HGF + HGFA induced the expression of the HGF-alpha chain in a time-dependent manner (P ＜ 0.01); however,the increases in expression of HGF-alpha chain induced by HGF alone and HGFA alone were not significantly different from the level in the blank controls (P＞ 0.05).Exposure to HGF alone and HGFA alone led to a time-dependent increase in apoptosis (24 h,48 h,72 h) but exposure to HGF + HGFA led to the highest levels of apoptosis (P ＜ 0.05).Exposure to HGF + HGFA led to a time-dependent decrease in RhoA mRNA and protein expression (P ＜ 0.01).Conclusion Activation of hepatocyte growth factor promotes apoptosis of hepatic stellate cells by suppressing RhoA expression and down-regulating the Rho signaling pathway. Key words: Hepatocyte growth factor;  Hepatocyte growth factor activator;  Hepatic stellate cells; RhoA