Oxidation of pyridine nucleotides during Fas- and ceramide-induced apoptosis in Jurkat cells: correlation with changes in mitochondria, glutathione depletion, intracellular acidification and caspase 3 activation

Jurkat T cells showed a major, early decrease in blue auto-fluorescence in response to Fas/Apo-1/CD95 cross-linking or stimulation with cell-permeant ceramide. This indicates the oxidation/depletion of NADH or NADPH before th e onset of apoptosis, Kinetic studies, cytofluorimetric multiparameter anal yses and cell sorting experiments indicated a close temporal relationship b etween NAD(P)H oxidation/depletion and the dissipation of the mitochondrial transmembrane potential (Delta Psi (m)). In contrast. NAD(P)H depletion wa s detected well before several other changes associated with late apoptosis , including enhanced superoxide generation, phosphatidylserine exposure on the cell surface, loss of cytosolic K+, decreased cytoplasmic pH, nuclear D NA fragmentation, cell shrinkage, loss of viability and the appearance of t he mitochondrial antigen APO2.7. Full activation of caspase 9 and caspase 3 appeared to be correlated with the appearance of superoxide anions in the mitochondria, and followed the drop in NADPH. Overexpression of the apoptos is-inhibitory proto-oncogene Bcl-2, which encodes an inhibitor of the mitoc hondrial permeability transition (PT) pore, delayed both the Delta Psi (m) disruption and the depletion of NAD(P)H. Similar effects were observed with the pharmacological PT pore inhibitors, bongkrekic acid and cyclosporin A. Thus there appears to be a close functional relationship between mitochond rial and cellular redox changes during early apoptosis; events that are inh ibited by Bcl-2.