Oxygen deprivation but not a combination of oxygen, glucose, and serum deprivation induces DNA degradation in mouse cortical neurons in vitro: attenuation by transgenic overexpression of CuZn-superoxide dismutase.
The present work was designed to study the possible implication of apoptosis in ischemic neuronal death, a phenomenon that has been suggested to be involved in neurodegeneration following focal as well as global ischemia. In this study, mouse cortical neurons in primary culture were subjected to oxygen deprivation or oxygen, glucose, and serum deprivation to simulate hypoxia and "ischemia-like" conditions; also, cellular viability as well as DNA degradation were investigated. The results showed that DNA degradation occurred in neurons subjected to oxygen deprivation but not to oxygen and substrate deprivation together. This DNA degradation, resulting in a laddering by agarose gel electrophoresis, could be prevented by cycloheximide and actinomycin-D treatments, although these inhibitors were unable to reduce neuronal death. To investigate if DNA degradation could be elicited by an intracellular free radical generation during reoxygenation, transgenic neurons overexpressing copper-zinc superoxide dismutase were subjected to 9 h of oxygen deprivation and analyzed after 24 h of reoxygenation. The results showed a significant attenuation of DNA degradation in these cells and confirmed a possible relationship between reactive oxygen species and neuronal apoptosis. This study opens the way to further investigations regarding the involvement of an apoptotic process in necrotic neuronal death, and provides some new insights into the mechanisms underlying selective sensitivity of neuronal cells to oxygen and glucose deprivation.
Copin
J C
JC
Department of Neurological Surgery, University of California, School of Medicine, San Francisco 94143-0651, USA.
Reola
L F
LF
Chan
T Y
TY
Li
Y
Y
Epstein
C J
CJ
Chan
P H
PH
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D015687:Cell Hypoxia / Q000502:physiology*; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D016895:Culture Media, Serum-Free; D004249:DNA Damage / Q000502:physiology*; D053938:DNA Fragmentation; D004587:Electrophoresis, Agar Gel; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D005947:Glucose / Q000502:physiology*; D007150:Immunohistochemistry; D051379:Mice; D008822:Mice, Transgenic; D016900:Neurofilament Proteins / Q000378:metabolism; D009474:Neurons / Q000201:enzymology / Q000378:metabolism*; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D013482:Superoxide Dismutase / Q000096:biosynthesis* / Q000235:genetics
D016895:Culture Media, Serum-Free; D016900:Neurofilament Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D017382:Reactive Oxygen Species; D013482:Superoxide Dismutase; D005947:Glucose