Obestatin affords cardioprotection to the ischemic-reperfused isolated rat heart and inhibits apoptosis in cultures of similarly stressed cardiomyocytes

Abstract

Obestatin, a newly discovered peptide encoded by the ghrelin gene, induces the expression of genes regulating pancreatic β-cell differentiation, insulin biosynthesis, and glucose metabolism. It also activates antiapoptotic signaling pathways such as phosphoinositide 3-kinase (PI3K) and ERK1/2 in pancreatic β-cells and human islets. Since these kinases have been shown to protect against myocardial injury, we sought to investigate whether obestatin would exert cardioprotective effects. Both isolated perfused rat heart and cultured cardiomyocyte models of ischemia-reperfusion (I/R) were used to measure infarct size and cell apoptosis as end points of injury. The presence of specific obestatin receptors on cardiac cells as well as the signaling pathways underlying the obestatin effect were also studied. In the isolated heart, the addition of rat obestatin-(1–23) before ischemia reduced infarct size and contractile dysfunction in a concentration-dependent manner, whereas obestatin-(23–1), a synthetic analog with an inverse aminoacid sequence, was ineffective. The cardioprotective effect of obestatin-(1–23) was observed at concentrations of 10–50 nmol/l and was abolished by inhibiting PI3K or PKC by the addition of wortmannin (100 nmol/l) or chelerythrine, (5 μmol/l), respectively. In rat H9c2 cardiac cells or isolated ventricular myocytes subjected to I/R, 50 nmol/l obestatin-(1–23) reduced cardiomyocyte apoptosis and reduced caspase-3 activation; the antiapoptotic effect was blocked by the inhibition of PKC, PI3K, or ERK1/2 pathways. In keeping with these functional findings, radioreceptor binding results revealed the presence of specific high-affinity obestatin-binding sites, mainly localized on membranes of the ventricular myocardium and cardiomyocytes. Our data suggest that, by acting on specific receptors, obestatin-(1–23) activates PI3K, PKC-ε, PKC-δ, and ERK1/2 signaling and protects cardiac cells against myocardial injury and apoptosis induced by I/R.