Chronic exposure of HepG2 cells to excess copper results in depletion of glutathione and induction of metallothionein

Metallothionein (MT) and reduced glutathione (GSH) play an important role in the intracellular handling of copper by preventing the generation and favouring the removal of copper-derived free radicals. The present study addressed the changes in MT and GSH that follow chronic (2 or 5 weeks) exposure of human hepatoblastoma cells (HepG2) to excess copper. Copper treatment (100 muM, 2 weeks) led to a 28-fold elevation in intracellular copper. Concomitantly, cells exhibited a seven-fold increase in total MT and an increment in its saturation with copper from 45 to 86%. Around 38% of copper in the cytosolic fraction could be accounted for by MT. GSH equivalents were substantially lowered (to 37% of basal levels) in treated cells, with only part or it being accounted for by an increase in GSSG. Copper-treatment induced no changes in catalase or GSH-peroxidase activities but it was associated with a small reduction in SOD (20%) and GSH-reductase (26%) activities, Copper-loaded cells did not differ from controls in their basal oxidative tone, however, when exposed to tert-butylhydroperoxide they exhibited a markedly greater susceptibility to undergo both oxidative stress and cell lysis. It is proposed that chronic exposure of HepG2 cells to excess copper is accompanied by "adaptive changes" in GSH and MT metabolism that would render cells substantially more susceptibility to undergo oxidative stress-related cytotoxicity. (C) 2002 Elsevier Science Ltd. All rights reserved.