A characterized saponin extract of Panax japonicus suppresses hepatocyte EMT and HSC activation in vitro and CCl4-provoked liver fibrosis in mice: Roles of its modulatory effects on the Akt/GSK3ß/Nrf2 cascade.

BACKGROUND AND PURPOSE: Liver fibrosis constitutes a pathologic condition resulting in a series of advanced liver diseases. Oleanane-type saponins are distinctive active constituents in the medicinal plant Panax japonicus C. A. Mey (P. japonicus). Herein, we assessed protective effects of a characterized saponin extract of rhizomes of P. japonicus (SEPJ) on hepatocyte EMT and HSC activation in vitro and liver fibrosis in mice. We also investigated molecular mechanisms underlying the hepatoprotective activity of SEPJ. METHODS: EMT of AML-12 hepatocytes was evaluated by observing morphology of cells and quantifying EMT marker proteins. Activation of LX-2 HSCs was assessed via scratch assay, transwell assay, and EdU-incorporation assay, and by quantifying activation marker proteins. Liver fibrosis in mice was evaluated by HE, SR, and Masson staining, and by measuring related serum indicators. Immunoblotting and RT-PCR were performed to study mechanisms underlying the action of SEPJ. RESULTS: SEPJ inhibited TGF-ß-induced EMT in AML-12 hepatocytes and activation of LX-2 HSCs. SEPJ elevated Akt phosphorylation at Ser473 and GSK3ß phosphorylation at Ser9 in these cells, giving rise to a descent of the catalytic activity of GSK3ß. These events increased levels of both total and nuclear Nrf2 protein and upregulated expressions of Nrf2-responsive antioxidative genes. In addition, enhanced phosphorylation of Akt and GSK3ß acted upstream of SEPJ-mediated activation of Nrf2. Knockdown of Nrf2 or inhibition of Akt diminished the protective activity of SEPJ against TGF-ß in both AML-12 and LX-2 cells. Our further in vivo experiments revealed that SEPJ imposed a considerable alleviation on CCl4-provoked mouse liver fibrosis. Moreover, hepatic Akt/GSK3ß/Nrf2 cascade were potentiated by SEPJ. Taken together, our results unveiled that SEPJ exerted protective effects against fibrogenic cytokine TGF-ß in vitro and ameliorated liver fibrosis in mice. Mechanistically, SEPJ regulated the Akt/GSK3ß/Nrf2 signaling which subsequently enhanced intracellular antioxidative capacity. CONCLUSIONS: SEPJ inhibits hepatocyte EMT and HSC activation in vitro and alleviates liver fibrosis in mice. Modulation of the Akt/GSK3ß/Nrf2 cascade attributes to its hepatoprotective effects. Our findings support a possible application of SEPJ in the control of liver fibrosis.