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Am J Physiol Gastrointest Liver Physiol 283: G709-G718, 2002. First published May 15, 2002; doi:10.1152/ajpgi.00013.2002
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Vol. 283, Issue 3, G709-G718, September 2002

Resistance to Fas-induced apoptosis in hepatocytes: role of GSH depletion by cell isolation and culture

Lina Musallam1,2, Chantal Éthier1, Pierre Selim Haddad2, Francine Denizeau3, and Marc Bilodeau1

1 Centre de recherche, Centre hospitalier de l'Université de Montréal-Hôpital Saint-Luc, Montréal, H2X 1P1; 2 Département de Pharmacologie, Université de Montréal, Montréal, H3C 3T4; and 3 Département de Chimie/Biochimie, Université du Québec à Montréal, Montréal, Canada H3C 3P8

The involvement of reduction/oxidation (redox) state in cell sensitivity to apoptosis has been suggested by several studies in which induction of apoptosis was shown to require oxidative stress or GSH extrusion. On the other hand, biochemical studies of caspases revealed that their activation necessitates a reduced cysteine in their active site. This is ensured by maintaining intact intracellular glutathione status during apoptotic induction as reported by in vivo studies. Therefore, we investigated the relationship between intracellular glutathione levels and the sensitivity of mouse hepatocytes in culture to Fas-induced apoptosis as well as potential mechanisms responsible for this sensitivity. We found that total and reduced glutathione levels are decreased by one-half after cell isolation procedure and further decline by 25% during cell culture for 2 h in normal Williams' E medium. Cell culture in medium supplemented with cysteine and methionine maintains glutathione at a level similar to that measured just after cell isolation. Results show that the capacity of Fas to activate caspase-8 and to induce apoptosis requires important intracellular glutathione levels and high GSH/total glutathione ratio. In conclusion, the present study shows that intracellular glutathione plays an important role in maintaining the apoptotic machinery functional and is thus capable of transmitting the apoptotic signal.

glutathione; redox; Bid; BCL-xl; caspase-8; apoptosis


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