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 autofluorescence 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 the onset of apoptosis. Kinetic studies, cytofluorimetric multiparameter analyses and cell sorting experiments indicated a close temporal relationship between NAD(P)H oxidation/depletion and the dissipation of the mitochondrial transmembrane potential (DeltaPsi(m)). In contrast, NAD(P)H depletion was 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 DNA fragmentation, cell shrinkage, loss of viability and the appearance of the 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 apoptosis-inhibitory proto-oncogene Bcl-2, which encodes an inhibitor of the mitochondrial permeability transition (PT) pore, delayed both the DeltaPsi(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 mitochondrial and cellular redox changes during early apoptosis; events that are inhibited by Bcl-2.
Gendron
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Institut Cochin de Génétique Moléculaire, INSERM U129, CHU Cochin Port-Royal, 24 rue du Faubourg Saint-Jacques, F-75014 Paris, France. pxpetit@icgm.cochin.inserm.fr
Schrantz
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Métivier
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Kroemer
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Maciorowska
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Sureau
F
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Koester
S
S
Petit
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PX
D048868:Adaptor Proteins, Signal Transducing; D000227:Adenine Nucleotides / Q000737:chemistry / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000032:analysis; D000255:Adenosine Triphosphate / Q000032:analysis; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins / Q000494:pharmacology; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002518:Ceramides / Q000494:pharmacology; D004789:Enzyme Activation; D005978:Glutathione / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019169:Jurkat Cells; D018715:Microscopy, Video; D008928:Mitochondria / Q000378:metabolism; D009249:NADP / Q000032:analysis / Q000737:chemistry; D010084:Oxidation-Reduction; D011188:Potassium / Q000378:metabolism; D000090063:Proto-Oncogene Mas; D017382:Reactive Oxygen Species / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D000227:Adenine Nucleotides; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D002518:Ceramides; C490780:FAF1 protein, human; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D017382:Reactive Oxygen Species; D009249:NADP; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D005978:Glutathione; D011188:Potassium