Abstract
Harnessing endogenous cardioprotectants is a novel therapeutic strategy to combat ischemia/reperfusion (I/R) injury. Thrombin causes I/R injury, whereas exogenous adenosine prevents I/R injury. We hypothesized that blocking thrombin receptor activation with a protease-activated receptor (PAR) 4 antagonist would unmask the cardioprotective effects of endogenous adenosine. The protective role of two structurally unrelated PAR4 antagonists, trans-cinnamoyl-YPGKF-amide (tc-Y-NH2) and palmitoyl-SGRRYGHALR-amide (P4pal10), were evaluated in two rat models of myocardial I/R injury. P4pal10 (10 μg/kg) treatment before ischemia significantly decreased infarct size (IS) by 31, 21, and 19% when given before, during, and after ischemia in the in vivo model. tc-Y-NH2 (5 μM) treatment before ischemia decreased IS by 51% in the in vitro model and increased recovery of ventricular function by 26%. To assess whether the cardioprotective effects of PAR4 blockade were due to endogenous adenosine, isolated hearts were treated with a nonselective adenosine receptor blocker, 8-sulfaphenyltheophylline (8-SPT), and tc-Y-NH2 before ischemia. 8-SPT abolished the protective effects of tc-Y-NH2 but did not affect IS when given alone. Adenosine-mediated survival pathways were then explored. The cardioprotective effects of tc-Y-NH2 were abolished by inhibition of Akt (wortmannin), extracellular signal-regulated kinase 1/2 [PD98059 (2′-amino-3′-methoxyflavone)], nitric-oxide synthase [NG-monomethyl-l-arginine (l-NMA)], and KATP channels (glibenclamide). PD98059, l-NMA, and glibenclamide alone had no effect on cardioprotection in vitro. Furthermore, inhibition of mitochondrial KATP channels [5-hydroxydecanoic acid (5-HD)] and sarcolemmal KATP channels (sodium (5-(2-(5-chloro-2-methoxybenzamido)ethyl)-2-methoxyphenylsulfonyl)(methylcarbamothioyl)amide; HMR 1098) abolished P4pal10-induced cardioprotection in vivo. Thrombin receptor blockade by PAR4 inhibition provides protection against injury from myocardial I/R by unmasking adenosine receptor signaling and supports the hypothesis of a coupling between thrombin receptors and adenosine receptors.
Footnotes
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This study was supported in part by National Institutes of Health Grants HL54075 (to J.E.B.) and HL0831 (to G.J.G.). J.L.S. is supported by National Institutes of Health Training Grant HL07792 (awarded to the Medical College of Wisconsin).
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.107.133595.
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ABBREVIATIONS: I/R, ischemia/reperfusion; GPCR, G protein-coupled receptor; AR, adenosine receptor; ERK, extracellular signal-regulated kinase; NOS, nitric-oxide synthase; sarc, sarcolemmal; mito, mitochondrial; PAR, protease-activated receptor; AP, activating peptide; tc-Y-NH2, trans-cinnamoyl-YPGKF-amide; P4pal10, palmitoyl-SGRRYGHALR-amide; SD, Sprague-Dawley; PCR, polymerase chain reaction; PD98059, 2′-amino-3′-methoxyflavone; LVDP, left ventricular developed pressure; l-NMA, NG-monomethyl-l-arginine; 8-SPT, 8-sulfaphenyltheophylline; AAR, area at risk; NO, nitric oxide; 5-HD, 5-hydroxydecanoic acid; PI3K, phosphoinositide-3 kinase; HMR 1098, sodium (5-(2-(5-chloro-2-methoxybenzamido)ethyl)-2-methoxyphenylsulfonyl)(methylcarbamothioyl)amide.
- Received October 24, 2007.
- Accepted November 29, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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