Brain mitochondrial DNA is not damaged by prolonged cardiac arrest or reperfusion.
Postischemic reperfusion is known to cause iron-mediated peroxidation of polyunsaturated fatty acids in membranes, including mitochondrial membranes, in the brain cortex. Consequently, we tested the hypothesis that this radical-mediated damage would extend to DNA. Mitochondrial DNA (mtDNA) was chosen because of its presence at a known site of free radical formation, its sensitivity and ease of assay, and its known lack of any repair systems. In model experiments we utilized endonuclease III or piperidine to amplify topological form conversions in mtDNA damaged by in vitro reactions with hydroxyl radical. We then applied the amplified detection assays to dog brain mtDNA isolated after 2 or 8 h of reperfusion following a 20-min cardiac arrest. We found that ischemia and reperfusion caused no topological form conversions in mtDNA. Similarly, nucleotide incorporation by a gap-filling reaction showed no sensitivity to digestion of the mtDNA by exonuclease III, an enzyme known to remove blocked 3' termini at the site of radical-generated nicks. Furthermore, the recovery of mtDNA was similar in all experimental groups, suggesting that putatively damaged forms had not been removed by rapid degradation. Thus, despite mitochondrial membrane damage, brain mtDNA does not accumulate oxygen radical damage during postischemic brain reperfusion.
White
B C
BC
Department of Emergency Medicine, Wayne State University School of Medicine, Detroit, Michigan 48201.
Tribhuwan
R C
RC
Vander Laan
D J
DJ
DeGracia
D J
DJ
Krause
G S
GS
Grossman
L I
LI
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002545:Brain Ischemia / Q000378:metabolism*; D004249:DNA Damage; D004272:DNA, Mitochondrial / Q000378:metabolism*; D004285:Dogs; D005090:Exodeoxyribonucleases / Q000378:metabolism; D005609:Free Radicals; D006324:Heart Arrest, Induced; D015424:Reperfusion; D013997:Time Factors
D004272:DNA, Mitochondrial; D005609:Free Radicals; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III