A Ginsenoside Metabolite, 20-O-β-D-Glucopyranosyl-20(S)-protopanaxadiol, Triggers Apoptosis in Activated Rat Hepatic Stellate Cells via Caspase-3 Activation

Abstract

We investigated the apoptotic effects of the protopanaxadiol ginsenosides, Rb₁ and Rb₂, and their intestinal bacterial metabolite, 20-O-β-D-glucopyranosyl-20(S)-protopanaxadiol (M1), and of the protopanaxatriol ginsenoside, Rg₁, and its intestinal bacterial metabolite, 20(S)-protopanaxatriol, in activated rat hepatic stellate cells (HSCs) transformed by Simian virus 40 (T-HSC/Cl-6). As HSCs play a central role in liver fibrosis, agents that selectively induce apoptosis of HSCs could be used to treat this disease. Apoptosis was measured using cell viability tests, DNA fragmentation analysis, and immunoblot analysis of poly(ADP-ribose) polymerase cleavage. M1 (40 μM for 24 h) significantly induced apoptosis in activated rat HSCs. M1 induced apoptosis in a dose-dependent manner as shown by DNA fragmentation, an increased population of cells in the sub-G1 phase, and reduced mitochondrial transmembrane potential. M1 induced caspase-3 activity in a dose- and time-dependent manner. A specific inhibitor of caspase-3 prevented induction of apoptosis by M1 as shown by DNA fragmentation analysis. It is concluded that M1 induces apoptosis in T-HSC/Cl-6 cells via caspase-3 activation.

References

• 1 Singh V K, Agarwal S S, Gupta B M. Immunomodulatory activity of Panax ginseng extract.  Planta Med. 1984;  50 462-5
• 2 Matsuda H, Samukawa K, Kubo M. Anti-inflammatory activity of ginsenoside Ro.  Planta Med. 1990;  56 19-23
• 3 Panwar M, Kumar M, Samarth R, Kumar A. Evaluation of chemopreventive action and antimutagenic effect of the standardized Panax ginseng extract, EFLA400, in Swiss albino mice.  Phytother Res. 2005;  19 65-71
• 4 Choi C H, Kang G, Min Y D. Reversal of P-glycoprotein-mediated multidrug resistance by protopanaxatriol ginsenosides from Korean red ginseng.  Planta Med.. 2003;  69 235-40
• 5 Hasegawa H. Proof of the mysterious efficacy of ginseng: basic and clinical trials: metabolic activation of ginsenoside: deglycosylation by intestinal bacteria and esterification with fatty acid.  J Pharmacol Sci. 2004;  95 153-7
• 6 Wakabayashi C, Murakami K, Hasegawa H, Murata J, Saiki I. An intestinal bacterial metabolite of ginseng protopanaxadiol saponins has the ability to induce apoptosis in tumor cells.  Biochem Biophys Res Commun. 1998;  246 725-30
• 7 Oh S H, Lee B H. A ginseng saponin metabolite-induced apoptosis in HepG2 cells involves a mitochondria-mediated pathway and its downstream caspase-8 activation and Bid cleavage.  Toxicol Appl Pharmacol. 2004;  194 221-9
• 8 Lee J Y, Shin J W, Chun K S, Park K K, Chung W Y, Bang Y J. et al . Antitumor promotional effects of a novel intestinal bacterial metabolite (IH-901) derived from the protopanaxadiol-type ginsenosides in mouse skin.  Carcinogenesis. 2005;  26 359-67
• 9 Lee H U, Bae E A, Han M J, Kim N J, Kim D H. Hepatoprotective effect of ginsenoside Rb1 and compound K on tert-butyl hydroperoxide-induced liver injury.  Liver Int. 2005;  25 1069-73
• 10 Jung S H, Woo M S, Kim S Y, Kim W K, Hyun J W, Kim E J. et al . Ginseng saponin metabolite suppresses phorbol ester-induced matrix metalloproteinase-9 expression through inhibition of activator protein-1 and mitogen-activated protein kinase signaling pathways in human astroglioma cells.  Int J Cancer. 2006;  118 490-7
• 11 Lee S H, Seo G S, Ko G, Kim J B, Sohn D H. Anti-inflammatory activity of 20(S)-protopanaxadiol: enhanced heme oxygenase 1 expression in RAW 264.7 cells.  Planta Med. 2005;  71 1167-70
• 12 Friedman S L. Liver fibrosis - from bench to bedside.  J Hepatol. 2003;  38 S38-53
• 13 Park W H, Lee S K, Kim C H. A Korean herbal medicine, Panax notoginseng, prevents liver fibrosis and hepatic microvascular dysfunction in rats.  Life Sci. 2005;  76 1675-90
• 14 Allen R T, Hunter W J, Agrawal D K. Morphological and biochemical characterization and analysis of apoptosis.  Pharmacol Toxicol Methods. 1997;  37 215-28
• 15 Kim J Y, Kim K M, Nan J X, Zhao Y Z, Park P H, Lee S J. et al . Induction of apoptosis by tanshinone I via cytochrome c release in activated hepatic stellate cells.  Pharmacol Toxicol. 2003;  92 195-200
• 16 Park P H, Nan J X, Park E J, Kang H C, Kim J Y, Ko G. et al . Effect of tetrandrine on experimental hepatic fibrosis induced by bile duct ligation and scission in rats.  Pharmacol Toxicol. 2000;  87 261-8
• 17 Jakopec S, Dubravcic K, Polanc S, Kosmrlj J, Osmak M. Diazene JK-279 induces apoptosis-like cell death in human cervical carcinoma cells.  Toxicol In Vitro. 2006;  20 217-26
• 18 Park E J, Zhao Y Z, Kim Y H, Lee B H, Sohn D H. Honokiol induces apoptosis via cytochrome c release and caspase activation in activated rat hepatic stellate cells in vitro .  Planta Med. 2005;  71 82-4