THE HEPATOTOXICITY OF RHEIN INVOLVES IMPAIRMENT OF MITOCHONDRIAL FUNCTIONS

Metabolism of rhein (4,5-dihydroxyanthraquinone-2-carboxylic acid) in primary cultures of rat hepatocytes caused production of oxygen-derive d free radicals by redox cycling; this was shown as an increased rate of superoxide-dismutasesensitive NAD(P)H oxidation and NAD(P)H-cytochr ome c reduction. Furthermore, rhein caused a depletion of intracellula r reduced glutathione and an immediate, almost 10-fold increase in int racellular free Ca2+. Exposure to rhein also induced the following: a decrease in the mitochondrial membrane potential, as analyzed by uptak e of rhodamine 123 (Rh 123); initiation of lipid peroxidation, measure d as accumulation of malondialdehyde and 4-hydroxyalkenals; and cell d eath (LD(50) = 20 mu M). Pretreatment of cell cultures with dithiothre itol (DTT), nifedipin or N',N'-diphenyl-p-phenylenediamine (DPPD) incr eased the intracellular free Ca2+ concentration 5-fold but inhibited r hein-induced cytotoxicity. Moreover, addition of these protecting subs tances maintained the level of ATP and glutathione (GSH) and prevented accumulation of lipid peroxidation products. Depletion of intracellul ar glutathione by pretreatment with buthionine sulfoximine (BSO), or i nhibition of glutathione reductase with 1,3-bis-2-chloroethyl-1-nitros ourea (BCNU) decreased cell viability (LD(50) = 2.5 mu M). On the othe r hand, increasing GSH by pretreatment with L-2-oxothiazolidine-4-carb oxylic acid (OTC)did not provide complete protection. In summary, rhei n undergoes redox cycling that gives rise to oxygen metabolites that a ffect the mitochondrial membranes (recorded as a decreased membrane po tential) and alter the plasma membrane (i.e. induced the formation of surface blebs). Mitochondrial malfunction also causes changes in Ca2homeostasis and depletion of ATP, which eventually lead to cell death. (C) 1997 Elsevier Science Ireland Ltd.