Glutathione depletion causes cytochrome c release even in the absence of cell commitment to apoptosis.

We demonstrate here that the release of mature cytochrome c from mitochondria is a cellular response to the depletion of glutathione, the main intracellular antioxidant, independently from the destiny of the cells, i.e., apoptosis or survival. On the one hand, cytosolic cytochrome c was detected in cells where the inhibition of glutathione synthesis led to glutathione depletion without impairing viability or in tight concomitance with glutathione depletion prior to puromycin-induced apoptosis. Removal of the apoptogenic agent prior to apoptosis, but after glutathione extrusion and cytochrome c release, led to recovery of preapoptotic cells, which resume healthy features, i.e., restoration of normal glutathione levels and disappearance of cytosolic cytochrome c. On the other hand, in an example of apoptosis occurring without glutathione depletion, no translocation of cytochrome c from mitochondria to cytosol was detected. Unlike the other instances of apoptosis, in this case caspase 3 was not activated, thus suggesting the following oxidant-related apoptotic pathway: glutathione depletion, cytochrome c release, and caspase 3 activation. These results show that cytochrome c release is not a terminal event leading cells to apoptosis, but rather is the consequence of a redox disequilibrium that, under some circumstances, may be associated with apoptosis.