Measurement of nitric oxide and peroxynitrite generation in the postischemic heart. Evidence for peroxynitrite-mediated reperfusion injury.
Altered nitric oxide (NO.) production is a critical factor in tissue reperfusion injury; however, controversy remains regarding these alterations and how they cause injury. Since superoxide (O-2) generation is triggered during the early period of reperfusion the cytotoxic oxidant peroxynitrite (ONOO-) could be formed, but it is not known if this occurs. Therefore electron paramagnetic resonance and chemiluminescence studies were performed of the magnitude and time course of NO., O-2, and ONOO- formation in the postischemic heart. Isolated rat hearts were subjected either to normal perfusion or to reperfusion after 30 min of ischemia in the presence of the NO. trap Fe2+-N-methyl-D-glucamine dithiocarbamate with electron paramagnetic resonance measurements performed on the effluent. Although only trace signals were present prior to ischemia, prominent NO. adduct signals were seen during the first 2 min of reflow which were abolished by nitric oxide synthase (NOS) inhibition. Similar studies with the O-2 trap 5, 5-dimethyl-1-pyrroline N-oxide demonstrated a burst of O-2 generation over the first 2 min of reflow. Chemiluminescence measurements using 5-amino-2,3-dihydro-1,4-phthalazinedione (luminol) demonstrated a similar marked increase in ONOO- which was blocked by NOS inhibitors or superoxide dismutase. NOS inhibition or superoxide dismutase greatly enhanced the recovery of contractile function in postischemic hearts. Immunohistology demonstrated that the ONOO--mediated nitration product nitrotyrosine was formed in postischemic hearts but not in normally perfused controls. Thus, NO. formation is increased during the early period of reflow and reacts with O-2 to form ONOO-, which results in amino acid nitration and cellular injury.
Department of Medicine, Division of Cardiology, Johns Hopkins Medical Institutions, Johns Hopkins Bayview Medical Center, Baltimore, Maryland 21224, USA.
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Zweier
D000818:Animals; D004578:Electron Spin Resonance Spectroscopy; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D007150:Immunohistochemistry; D009200:Myocardial Contraction; D017202:Myocardial Ischemia / Q000378:metabolism* / Q000503:physiopathology; D015428:Myocardial Reperfusion Injury / Q000209:etiology / Q000378:metabolism* / Q000503:physiopathology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009566:Nitrates / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species; D013482:Superoxide Dismutase / Q000378:metabolism
D004791:Enzyme Inhibitors; D009566:Nitrates; D017382:Reactive Oxygen Species; C039642:peroxynitric acid; D009569:Nitric Oxide; D013482:Superoxide Dismutase; D019331:NG-Nitroarginine Methyl Ester