Abstract
Oxidative/nitrative stress caused by peroxynitrite, the reaction product of superoxide () and nitric oxide (NO), is the primary cause of myocardial ischemia/reperfusion injury. The present study determined whether INO-4885 [5,10,15,20-tetra[N-(benzyl-4′-carboxylate)-2-pyridinium]-21H,23H-porphine iron(III) chloride], a new peroxynitrite decomposition catalyst, may provide cellular protection and protect heart from myocardial ischemia/reperfusion injury. Adult male mice were subjected to 30 min of ischemia and 3 or 24 h of reperfusion. Mice were randomized to receive vehicle, INO-4885 without catalytic moiety, or INO-4885 (3-300 μg/kg i.p.) 10 min before reperfusion. Infarct size, apoptosis, nitrotyrosine content, NO/ production, and inducible nitric-oxide synthase (iNOS)/NADPH oxidase expression were determined. INO-4885 treatment reduced ischemia/reperfusion-induced protein nitration and caspase 3 activation in a dose-dependent fashion in the range of 3 to 100 μg/kg. However, doses exceeding 100 μg/kg produced nonspecific effects and attenuated its protective ability. At the optimal dose (30 μg/kg), INO-4885 significantly reduced infarct size (p < 0.01), decreased apoptosis (p < 0.01), and reduced tissue nitrotyrosine content (p < 0.01). As expected, INO-4885 had no effect on ischemia/reperfusion-induced iNOS expression and NO overproduction. To our surprise, this compound significantly reduced superoxide production and partially blocked NADPH oxidase overexpression in the ischemic/reperfused cardiac tissue. Additional experiments demonstrated that INO-4885 provided better cardioprotection than N-(3-(aminomethyl)benzyl)acetamidine (1400W, a selective iNOS inhibitor), apocynin (an NADPH oxidase inhibitor), or Tiron (a cell-permeable superoxide scavenger). Taken together, our data demonstrated that INO-4885 is a cardioprotective molecule that attenuates myocardial reperfusion injury by facilitating peroxynitrite decomposition and inhibiting NADPH oxidase-derived production.
Footnotes
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This work was supported by the American Diabetes Association [Grants 7-06-JF59 and 7-08-RA-98]; the National Institutes of Health [Grant 2R01HL-63828]; and the American Heart Association [Grant GIA0855554D].
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doi:10.1124/jpet.108.144352.
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ABBREVIATIONS:, superoxide; TTC, triphenyltetrazolium chloride; ONOO-, peroxynitrite; NO, nitric oxide; I/R, ischemia/reperfusion; ONOO-, peroxynitrite; INO-4885, 5,10,15,20-tetra[N-(benzyl-4′-carboxylate)-2-pyridinium]-21H,23H-porphine iron(III) chloride; MI, myocardial ischemia; INO-C, INO-4885 without catalytic moiety; iNOS, inducible nitric-oxide synthase; MI/R, myocardial ischemia/reperfusion; AAR, area at risk; PBS, phosphate-buffered saline; DAPI, 4,6-diamidino-2-phenylindole; TUNEL, terminal deoxynucleotidyl transferase dUTP nick-end labeling; pNA, p-nitroanilide; 1400W, N-(3-(aminomethyl)benzyl)acetamidine.
- Received August 1, 2008.
- Accepted November 21, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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