Protection by inhibition of poly (ADP-ribose) synthetase against oxidant injury in cardiac myoblasts In vitro.
Peroxynitrite and hydroxyl radical are reactive oxidants produced during myocardial reperfusion injury. In various cell types, including macrophages and smooth muscle cells, peroxynitrite and hydrogen peroxide cause DNA single strand breakage, which triggers the activation of the nuclear enzyme poly (ADP-ribose) synthetase (PARS), resulting in cytotoxicity. Using 3-aminobenzamide and nicotinamide, inhibitors of PARS, we investigated the role of PARS in the pathogenesis of myocardial oxidant injury in H9c2 cardiac myoblasts in vitro. Peroxynitrite (100-1000 microM), hydrogen peroxide (0.3-10 microM) and the NO donor compounds S-nitroso-N-accetyl-DL-penicillamine (SNAP) and diethyltriamine NONOate all caused a dose-dependent reduction of the mitochondrial respiration of the cells, as measured by the mitochondrial-dependent conversion of MTT to formazan. Peroxynitrite and hydrogen peroxide, but not the NO donors caused activation of cellular PARS activity. The suppression of mitochondrial respiration by peroxynitrite and hydrogen peroxide, but not by the NO donors, was ameliorated by pharmacological inhibition of PARS. The protection by the PARS inhibitors diminished at extremely high concentrations of the oxidants. Hypoxia (1 h) followed by reoxygenation (1-24 h) also resulted in a significant activation of PARS, and caused a suppression of mitochondrial respiration, which was prevented by inhibition of PARS. Similar to the results obtained with the pharmacological inhibitors of PARS, a fibroblast cell line which derives from the PARS knockout mouse was protected against the suppression of mitochondrial respiration in response to peroxynitrite and reoxygenation, but not to NO donors, when compared to the result of cells derived from wild-type animals. Based on our data, we suggest that activation of PARS plays a role in the myocardial oxidant injury.
Gilad
E
E
Division of Critical Care, Children's Hospital Medical Center, 3333 Burnet Avenue, Cincinnati, Ohio 45229, USA.
Zingarelli
B
B
Salzman
A L
AL
Szabó
C
C
D000818:Animals; D015687:Cell Hypoxia; D002478:Cells, Cultured; D004789:Enzyme Activation / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000494:pharmacology; D051379:Mice; D008929:Mitochondria, Heart / Q000187:drug effects / Q000378:metabolism; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D009566:Nitrates / Q000494:pharmacology; D009569:Nitric Oxide / Q000494:pharmacology; D016877:Oxidants / Q000494:pharmacology*; D018384:Oxidative Stress; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; D051381:Rats
D009566:Nitrates; D016877:Oxidants; D000067856:Poly(ADP-ribose) Polymerase Inhibitors; C039642:peroxynitric acid; D009569:Nitric Oxide; D006861:Hydrogen Peroxide