Licorice Compounds Glycyrrhizin and 18β-Glycyrrhetinic Acid Are Potent Modulators of Bile Acid-induced Cytotoxicity in Rat Hepatocytes

Eric Gumpricht,Rolf Dahl,Michael W Devereaux,Ronald J Sokol

doi:10.1074/jbc.m411673200

Eric Gumpricht, Rolf Dahl + Show 2 more

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https://doi.org/10.1074/jbc.m411673200

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Abstract

The accumulation of hydrophobic bile acids results in cholestatic liver injury by increasing oxidative stress, mitochondrial dysfunction, and activation of cell signaling pathways. Licorice root and its constituents have been utilized as antihepatotoxic agents. The purpose of this study was to evaluate the potential modulation by a primary component of licorice root, glycyrrhizin (GL), and its metabolite, 18beta-glycyrrhetinic acid (GA), in a hepatocyte model of cholestatic liver injury. Preincubation of fresh rat hepatocyte suspensions with GL or GA reduced glycochenodeoxycholic acid (GCDC)-dependent reactive oxygen species generation, with GA more potent than GL. Interestingly, GL and GA had opposing effects toward GCDC-induced cytotoxicity; GA prevented both necrosis and apoptosis, whereas GL enhanced apoptosis. GCDC promoted activation of caspase 10, caspase 3, and PARP; all were inhibited by GA but not GL. Induction of apoptosis by GCDC was also associated with activation of JNK, which was prevented by GA. Activation of caspase 9 and dissipation of mitochondrial membrane potential were prevented by GA but not GL. In liver mitochondrial studies, GL and GA were both potent inhibitors of the mitochondrial permeability transition, reactive oxygen species generation, and cytochrome c release at submicromolar concentrations. Results from this study suggest that GL exhibits pro-apoptotic properties, whereas GA is a potent inhibitor of bile acid-induced apoptosis and necrosis in a manner consistent with its antioxidative effect.

Highlights

Cholestatic liver disorders are characterized by impaired bile flow resulting in the retention of bile constituents and hepatocellular damage
GL and glycyrretinic acid (GA) Reduce glycochenodeoxycholic acid (GCDC)-stimulated reactive oxygen species (ROS) Generation—To determine the effect of licorice compounds on bile acid-induced ROS generation, freshly isolated rat hepatocyte suspensions were incubated for 4 h with 100 ␮M GCDC, and DCF fluorescence was measured
Concentration-effect relationships of the two compounds after 4 h of incubation with GCDC (Fig. 1c) showed that both GA and GL function at low concentrations to reduce bile acid-induced oxidative stress, that maximal effect was achieved at low concentrations, and that GA was superior to GL in this effect

Summary

Introduction

Cholestatic liver disorders are characterized by impaired bile flow resulting in the retention of bile constituents and hepatocellular damage. Mechanistic studies have revealed that several factors, including physicochemical properties [6] and death receptor activation [7], account for the pro-apoptotic effects of bile acids. Extensive evidence supports the involvement of mitochondrial pathways in bile acid-induced hepatocyte toxicity, including induction of the mitochondrial permeability transition (MPT) [9]. Oxidative stress generated by mitochondria plays a role in bile acid-induced cellular toxicity, as demonstrated in liver mitochondria [10, 11] and rat hepatocytes [12, 13] as well as in in vivo studies with whole animals exposed to bile acids [14]. The purpose of this study was to examine the effects of GA and GL on cell pathways of bile acid-induced cytotoxicity in both freshly isolated rat hepatocyte suspensions and purified liver mitochondrial fractions

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