Enhanced mtDNA repair capacity protects pulmonary artery endothelial cells from oxidant-mediated death.
In rat cultured pulmonary arterial (PA), microvascular, and venous endothelial cells (ECs), the rate of mitochondrial (mt) DNA repair is predictive of the severity of xanthine oxidase (XO)-induced mtDNA damage and the sensitivity to XO-mediated cell death. To examine the importance of mtDNA damage and repair more directly, we determined the impact of mitochondrial overexpression of the DNA repair enzyme, Ogg1, on XO-induced mtDNA damage and cell death in PAECs. PAECs were transiently transfected with an Ogg1-mitochondrial targeting sequence construct. Mitochondria-selective overexpression of the transgene product was confirmed microscopically by the observation that immunoreactive Ogg1 colocalized with a mitochondria-specific tracer and, with an oligonucleotide cleavage assay, by a selective enhancement of mitochondrial Ogg1 activity. Overexpression of Ogg1 protected against both XO-induced mtDNA damage, determined by quantitative Southern analysis, and cell death as assessed by trypan blue exclusion and MTS assays. These findings show that mtDNA damage is a direct cause of cell death in XO-treated PAECs.
Dobson
Allison W
AW
Department of Cell Biology and Neuroscience, College of Medicine, University of South Alabama, Mobile, Alabama 36688, USA.
Grishko
Valentina
V
LeDoux
Susan P
SP
Kelley
Mark R
MR
Wilson
Glenn L
GL
Gillespie
Mark N
MN
D000818:Animals; D016923:Cell Death / Q000187:drug effects / Q000235:genetics*; D002478:Cells, Cultured; D004260:DNA Repair / Q000502:physiology*; D004272:DNA, Mitochondrial / Q000378:metabolism*; D043382:DNA-Formamidopyrimidine Glycosylase; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D008928:Mitochondria / Q000201:enzymology / Q000235:genetics; D009699:N-Glycosyl Hydrolases / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D016877:Oxidants / Q000378:metabolism; D010641:Phenotype; D011651:Pulmonary Artery / Q000166:cytology / Q000378:metabolism*; D011667:Pulmonary Veins / Q000166:cytology / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection; D014969:Xanthine Oxidase / Q000378:metabolism
D004272:DNA, Mitochondrial; D016877:Oxidants; D014969:Xanthine Oxidase; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase