Increased 8-hydroxyguanine formation and endonuclease activity for its repair in ischemic-reperfused hearts of rats.
A type of oxidative DNA damage, 8-hydroxyguanine (8-OH-Gua) formation, and the activity for its subsequent repair, 8-OH-Gua endonuclease activity, were examined in an ischemia-reperfusion model of isolated rat hearts. The level of 8-OH-Gua in myocardial DNA was measured by a high performance liquid chromatography (HPLC) equipped with an electrochemical detector, and the 8-OH-Gua endonuclease activity was analysed by the endonuclease nicking assay using a synthetic double-stranded oligonucleotide containing an 8-OH-Gua residue as a substrate. The Langendorff-perfused rat hearts were subjected to 30 or 60 min of global ischemia, followed by reperfusion with an oxygenated or a nitrogenated Krebs-Henseleit solution. The 8-OH-Gua content in the DNA of the ischemic hearts reperfused with an oxygenated solution was three to four times higher than that of the control hearts. The levels of 8-OH-Gua did not increase either in the ischemic hearts reperfused with a nitrogenated solution or in the ischemic-reperfused hearts treated with SOD, mannitol or allopurinol. When the myocardial extract was incubated with the 8-OH-Gua-containing oligonucleotide substrate, a specific cleavage at the site of an 8-OH-Gua residue was detected. The endonuclease activity responsible for this cleavage increased two-fold in the ischemic-reperfused hearts, compared to the control. This study demonstrates that the formation of 8-OH-Gua in DNA as well as the level of its repair process, 8-OH-Gua endonuclease activity, increase in the ischemic-reperfused rat hearts in response to oxidative stress due to higher levels of oxygen free radicals.
You
H
H
Department of Pharmacology and Heart Research Institute, Seoul National University College of Medicine, Korea.
Kim
G
G
Kim
Y
Y
Chun
Y
Y
Park
J
J
Chung
M H
MH
Kim
M
M
D000818:Animals; D004247:DNA; D043382:DNA-Formamidopyrimidine Glycosylase; D006147:Guanine / Q000031:analogs & derivatives* / Q000096:biosynthesis; D006321:Heart; D008297:Male; D015425:Myocardial Reperfusion; D009206:Myocardium / Q000378:metabolism*; D009699:N-Glycosyl Hydrolases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
C024829:8-hydroxyguanine; D006147:Guanine; D004247:DNA; D009699:N-Glycosyl Hydrolases; D043382:DNA-Formamidopyrimidine Glycosylase