Received for publication July 15, 2008.
Revised August 5, 2008.
Accepted for publication August 6, 2008.

L-Methionine-dl-sulfoxide metabolism and toxicity in freshly isolated mouse hepatocytes: Gender differences and inhibition with aminooxyacetic acid

Abstract

L-Methionine-dl-sulfoxide (MetO) is an L-methionine (Met) metabolite, but its role in Met metabolism and toxicity is not clear. In this study, MetO uptake, metabolism to Met, cytotoxicity, and GSH and GSSG status were characterized in freshly isolated mouse hepatocytes incubated at 37°C with 0-30 mM MetO for 0-5 h. In male hepatocytes, dose-dependent cytotoxicity concomitant with GSH depletion without GSSG formation occurred after exposure to 20 or 30 mM MetO, but not 10 mM MetO. Interestingly, female hepatocytes exposed to 30 mM MetO showed no cytotoxicity and exhibited increased intracellular GSH levels compared to control hepatocytes. Male hepatocytes had approximately 2-fold higher levels of intracellular Met-d-O or Met-l-O after MetO (30 mM) exposure for 0-1.5 h compared to female hepatocytes. In hepatocytes of both genders, Met-l-O was detected at nearly 5-fold higher levels than Met-d-O, and no significant increase in cellular Met levels was detected. Addition of aminooxyacetic acid (AOAA), an inhibitor of transamination reactions, to MetO-exposed male hepatocytes resulted in higher cellular Met-d-O and Met-l-O levels and decreased the cytotoxicity of MetO. Interestingly, exposure of control male hepatocytes to AOAA selectively increased cellular Met-d-O levels to levels similar to those observed after exposure to MetO (30mM). Analysis of MetO transamination activity by glutamine transaminase K in mouse liver cytosol revealed similar rates of MetO transamination in cytosol of both genders. Taken together, these results provide evidence for stereoselective oxidation of Met to Met-d-O under physiological conditions and suggest a major role for MetO transamination in MetO metabolism and toxicity.

Key words: analytical pharmacology/toxicology, flavin-containing monooxygenase, gender differences, glutathione homeostasis, hepatotoxicity, in vitro toxicity assays, isolated hepatocytes, metabolite kinetics, reductases