Epigallocatechin-3-gallate, a green-tea polyphenol, suppresses Rho signaling in TWNT-4 human hepatic stellate cells

Epigallocatechin-3-gallate (EGCG), a major constituent of the polyphenoids in green tea, has been reported to possess a wide range of biologic activities, including antifibrogenesis. Activated hepatic stellate cells (HSCs) are central to hepatic fibrosis, and Rho (a small GTPase)-signaling pathways have been implicated in the activation and proliferation of HSCs. In this study, we investigated the effect of EGCG on Rho-signaling pathways in activated human HSC-derived TWNT-4 cells. EGCG inhibited stress-fiber formation, an indicator of Rho activation, and changed the distribution of alpha-smooth-muscle actin. These inhibitory effects of EGCG were restored by overexpression of constitutively active Rho. A pull-down assay revealed that activated Rho (GTP-bound state) was strongly inhibited by ECGC and accompanied by suppressed phosphorylation of focal adhesion kinase, which is a regulator of Rho-signaling pathways. 5-Bromo-2'-deoxy-uridine incorporation demonstrated that ECGC 100 mu mol/L suppressed cell growth by 80%, and terminal deoxynucleotidyl transferase viohn-deoxyruidine triphosphate nick end-labeling revealed that EGCG (100 mu mol/L) caused apoptosis in half of the total cells. EGCG also strongly inhibited lysophoaphatidic acid (an activator of Rho) and induced phosphorylation of mitogien-activated protein kinases (Erk1/2, c-jun kinase, and p38). These findings demonstrate that EGCG regulates the structure and growth of HSCs by way of Rho-signaling pathways and suggest that EGCG has therapeutic potential in the setting of liver fibrosis.