Oxidative neurotoxicity in rat cerebral cortex neurons: synergistic effects of H2O2 and NO on apoptosis involving activation of p38 mitogen-activated protein kinase and caspase-3.
Oxidative stress in the brain has been increasingly associated with the development of numerous human neurological diseases. Microglia, activated upon neuronal injury or inflammatory stimulation, are known to release superoxide anion (*O(2) (-)), hydrogen peroxide (H(2)O(2)), and nitric oxide (NO), thereby further contributing to oxidative neurotoxicity. The reaction of NO and *O(2) (-), forming the toxic peroxynitrite (ONOO(-)), has been proposed to play a pathogenic role in neuronal injury. However, the interactions between H(2)O(2) and NO during oxidative stress, which may promote or diminish cell death, is less clear. In this study, we explored oxidative neurotoxicity induced by H(2)O(2) plus NO in primary cultures of rat cerebral cortex neurons. As the mechanisms may involve reactions between H(2)O(2) and NO, we monitored the production of ONOO(-)and reactive oxygen species (ROS) throughout the experiments. Results indicated that the NO donor S-nitroso-N-acetyl-D, L-penicillamine (SNAP) and H(2)O(2) by themselves elicited neuronal death in a concentration- and time-dependent manner. Sublytic concentrations of H(2)O(2) plus SNAP were sufficient to induce neuronal apoptosis as determined by DNA laddering and fluorescent staining of apoptotic nuclei. Transient ONOO(-)increase was accompanied by rapid H(2)O(2) decay and NO production, whereas ROS slowly decreased following treatment. Furthermore, p38 mitogen-activated protein kinase (MAPK) activation and the cleavage of caspase-3 were observed. Conversely, inhibition of p38 MAPK and caspase-3 significantly reduced apoptotic death induced by H(2)O(2) plus SNAP. These data suggest that H(2)O(2) and NO act synergistically to induce neuronal death through apoptosis in which activation of p38 MAPK and caspase-3 is involved.
Wang
Jiz-Yuh
JY
Graduate Institute of Life Sciences, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Shum
Andrew Y C
AY
Ho
Yi-Jung
YJ
Wang
Jia-Yi
JY
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000378:metabolism / Q000473:pathology; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005110:Extracellular Space / Q000737:chemistry; D005333:Fetus; D006861:Hydrogen Peroxide / Q000032:analysis / Q000378:metabolism* / Q000494:pharmacology; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling; D020928:Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology*; D009569:Nitric Oxide / Q000032:analysis / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000502:physiology*; D030421:Peroxynitrous Acid / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000032:analysis; D026423:S-Nitroso-N-Acetylpenicillamine / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases
D004791:Enzyme Inhibitors; D020030:Nitric Oxide Donors; D016877:Oxidants; D017382:Reactive Oxygen Species; D030421:Peroxynitrous Acid; D009569:Nitric Oxide; D026423:S-Nitroso-N-Acetylpenicillamine; D006861:Hydrogen Peroxide; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases