Ca(2+)-independent caspase-3 but not Ca(2+)-dependent caspase-2 activation induced by oxidative stress leads to SH-SY5Y human neuroblastoma cell apoptosis.
Continuous and long-lasting exposure to tert-butylhydroperoxide (t-BOOH) increased the number of apoptotic SH-SY5Y human neuroblastoma cells both in the presence and in the absence of the intracellular Ca(2+) ion chelator 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA). In addition, t-BOOH exposure induced activation of CPP32, as demonstrated by poly-(ADP-ribose) polymerase (PARP) cleavage, and of ICH-1L caspases. Exposure to t-BOOH also induced a time-dependent release of cytochrome c. Interestingly, in the presence of BAPTA, CPP32 activation still occurred, whereas ICH-1L activation was blocked. Ac-DEVD-CHO, an inhibitor of CPP32 activity, prevented the appearance of apoptotic cells, whereas the inhibitor of ICH-1L activity Z-VDVAD-FMK did not. Collectively, these findings demonstrate that in SH-SY5Y neuroblastoma cells exposure to continuous and long-lasting oxidative stress induced activation of caspase-3 that was independent of intracellular Ca(2+) ion concentration ([Ca(2+)](i)) elevation but led to cell apoptosis. In contrast, caspase-2 activation was dependent on [Ca(2+)](i) increase but did not result in apoptosis.
Amoroso
Salvatore
S
Department of Pharmacology, School of Medicine, University of Ancona, Ancona, Italy. amoroso@unina.it
D'Alessio
Angela
A
Sirabella
Rossana
R
Di Renzo
Gianfranco
G
Annunziato
Lucio
L
D017209:Apoptosis / Q000187:drug effects; D015153:Blotting, Western; D002118:Calcium / Q000037:antagonists & inhibitors / Q000378:metabolism*; D053143:Caspase 2; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000378:metabolism; D005609:Free Radicals / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling; D009447:Neuroblastoma / Q000201:enzymology / Q000378:metabolism*; D009842:Oligopeptides / Q000378:metabolism; D018384:Oxidative Stress; D013997:Time Factors; D014407:Tumor Cells, Cultured; D020122:tert-Butylhydroperoxide / Q000494:pharmacology*
D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D005609:Free Radicals; D009842:Oligopeptides; C099158:acetyl-aspartyl-glutamyl-valyl-aspartal; C427308:benzoylcarbonyl-valyl-aspartyl-valyl-alanyl-aspartyl-fluoromethyl ketone; D004533:Egtazic Acid; D020122:tert-Butylhydroperoxide; C505407:CASP3 protein, human; D053143:Caspase 2; D053148:Caspase 3; D020169:Caspases; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium