Caspase activation accompanying cytochrome c release from mitochondria is possibly involved in nitric oxide-induced neuronal apoptosis in SH-SY5Y cells.
It is well known that caspases are produced as proforms, which are proteolytically cleaved and activated during apoptosis or programmed cell death. We report here that caspases are activated during apoptosis by treatment with NOC18, a nitric oxide (NO) donor. Our present experiments have examined the way in which NO induces neuronal cell death, using a new type of NO donor that spontaneously releases only NO without enzymatic metabolism. NOC18 induced apoptosis in human neuroblastoma SH-SY5Y cells in a concentration- and time-dependent manner as estimated by DNA fragmentation assay, FACScan analysis, and nuclear morphology. Oxyhemoglobin, an NO trapper, suppressed NOC18-triggered DNA fragmentation, indicating that NO from NOC18 is a real activator in this study. Upon the induction of apoptosis, an increase in caspase-3-like protease activity, but not caspase-1, was observed. Procaspase-2 protein, an inactive form of caspase-2, decreased dramatically. In addition, NOC18 also resulted in poly (ADP-ribose) polymerase (PARP) cleavage, yielding an 85-kDa fragment typical of caspase activity. Oxyhemoglobin blocked the decrease of procaspase-2 and the cleavage of PARP by NOC18 in a concentration-dependent manner. Moreover, NO elicited the release of cytochrome c into the cytosol during apoptosis. These results suggest that both stimulation of caspase activity and cytochrome c release are partly involved in NO-induced neuronal apoptosis.
Uehara
T
T
Department of Pharmacology, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan.
Kikuchi
Y
Y
Nomura
Y
Y
D017209:Apoptosis / Q000502:physiology*; D020170:Caspase 1 / Q000235:genetics / Q000378:metabolism*; D053143:Caspase 2; D053148:Caspase 3; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D003374:Coumarins / Q000494:pharmacology; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D003574:Cytochrome c Group / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D008928:Mitochondria / Q000201:enzymology; D009474:Neurons / Q000166:cytology* / Q000201:enzymology; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009603:Nitroso Compounds / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology
C112287:Ac-aspartyl-glutamyl-valyl-aspartyl-aminomethylcoumarin; D003374:Coumarins; D015853:Cysteine Proteinase Inhibitors; D003574:Cytochrome c Group; C093285:NOC 18; D020030:Nitric Oxide Donors; D009603:Nitroso Compounds; D009842:Oligopeptides; D009569:Nitric Oxide; C505407:CASP3 protein, human; D053143:Caspase 2; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1