Apelin is an endogenous peptide found in various tissues and organs included the cardiovascular system, where it displays inotropic and vasodilator effects and protects myocardium against ischaemia-reperfusion injury. Depending on the number of aminoacids, different forms of Apelin have been classified. Out of the various forms, apelin-13 has been found to be the most active on the cardiovascular system. The mechanism of the myocardial protection exerted by Apelin is still controversial. An intervention of a phosphatidylinositol 3-kinase (PI3K) as a signal transducer has not yet been fully confirmed. The present research aims at investigating whether PI3K leads to protection via nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway. Twentyeight anaesthetised (ketamine (90 mg/kg) and xylazine (10 mg/kg)) rats were killed by decapitation. The hearts were excised and perfused with oxygenated buffer. A 30 min global ischaemia was followed by a 120 min reperfusion. Left ventricular pressure was measured and infarct size was assessed by nitro-blue tetrazolium test (Group I; n=5). Group II hearts were perfused with 0.5 μM apelin-13 during the first 20 min of reperfusion (n=5); in addition to Apelin, Group III-VI hearts received the following compounds 5 min before and 25 min after ischaemia: the PI3K inhibitor LY 294002 (15 μM) (n=4) or the NO-synthase inhibitor L-nitro-n-arginine (L-NNA) (100 μM) (n=5) or the guanylyl-cyclase (GC) inhibitor 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (10 μM) (n=4) or the blocker of mitochondrial ATP sensitive K+ channels (mitoATP K+) 5-hydroxydecanoate (5-HD) (100 μM ) (n=5), respectively. Data are expressed as mean±sem and were statistically analyzed by 1-way ANOVA. After ischaemia and reperfusion, the infarction was extended to the 54±3% of the left ventricular mass. Moreover, starting from the late phase of ischaemia, the left ventricular end diastolic pressure (LVEDP) increased from 6±1 to 40±9 mmHg (p<0.001), without any significant change of the left ventricular systolic pressure (LVSP). The increase of LVEDP, which was taken as an index of myocardial contracture, was responsible for about 70% reduction of the left ventricular developed pressure (LVDevP ). Apelin-13 significantly (p<0.001) reduced the infarct size to 26±4%. It also reduced LVEDP by about 50% (p<0.001) thus limiting the fall of LVDevP to about 20% (p<0.001). The inhibition of PI3K, NO-synthase and GC and the blockade of mitoATP K+ channels prevented the effect of Apelin-13 on infarct size and contracture. Our results indicate that Apelin-induced protection is mediated by NO-cGMP cascade which is likely to be elicited by PI3K-AKT pathway as suggested by its suppression by PI3K inhibition. The protection includes not only a limitation of the infarct size, but also a reduction of myocardial contracture.
Involvement in NO-cGMP pathway in cardiac protection by Apelin-13 in the rat
RASTALDO, Raffaella;CAPPELLO, SANDRA;FOLINO, Anna;LOSANO, Giovanni;PAGLIARO, Pasquale
2010-01-01
Abstract
Apelin is an endogenous peptide found in various tissues and organs included the cardiovascular system, where it displays inotropic and vasodilator effects and protects myocardium against ischaemia-reperfusion injury. Depending on the number of aminoacids, different forms of Apelin have been classified. Out of the various forms, apelin-13 has been found to be the most active on the cardiovascular system. The mechanism of the myocardial protection exerted by Apelin is still controversial. An intervention of a phosphatidylinositol 3-kinase (PI3K) as a signal transducer has not yet been fully confirmed. The present research aims at investigating whether PI3K leads to protection via nitric oxide (NO)-cyclic guanosine monophosphate (cGMP) pathway. Twentyeight anaesthetised (ketamine (90 mg/kg) and xylazine (10 mg/kg)) rats were killed by decapitation. The hearts were excised and perfused with oxygenated buffer. A 30 min global ischaemia was followed by a 120 min reperfusion. Left ventricular pressure was measured and infarct size was assessed by nitro-blue tetrazolium test (Group I; n=5). Group II hearts were perfused with 0.5 μM apelin-13 during the first 20 min of reperfusion (n=5); in addition to Apelin, Group III-VI hearts received the following compounds 5 min before and 25 min after ischaemia: the PI3K inhibitor LY 294002 (15 μM) (n=4) or the NO-synthase inhibitor L-nitro-n-arginine (L-NNA) (100 μM) (n=5) or the guanylyl-cyclase (GC) inhibitor 1H-[1,2,4]Oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) (10 μM) (n=4) or the blocker of mitochondrial ATP sensitive K+ channels (mitoATP K+) 5-hydroxydecanoate (5-HD) (100 μM ) (n=5), respectively. Data are expressed as mean±sem and were statistically analyzed by 1-way ANOVA. After ischaemia and reperfusion, the infarction was extended to the 54±3% of the left ventricular mass. Moreover, starting from the late phase of ischaemia, the left ventricular end diastolic pressure (LVEDP) increased from 6±1 to 40±9 mmHg (p<0.001), without any significant change of the left ventricular systolic pressure (LVSP). The increase of LVEDP, which was taken as an index of myocardial contracture, was responsible for about 70% reduction of the left ventricular developed pressure (LVDevP ). Apelin-13 significantly (p<0.001) reduced the infarct size to 26±4%. It also reduced LVEDP by about 50% (p<0.001) thus limiting the fall of LVDevP to about 20% (p<0.001). The inhibition of PI3K, NO-synthase and GC and the blockade of mitoATP K+ channels prevented the effect of Apelin-13 on infarct size and contracture. Our results indicate that Apelin-induced protection is mediated by NO-cGMP cascade which is likely to be elicited by PI3K-AKT pathway as suggested by its suppression by PI3K inhibition. The protection includes not only a limitation of the infarct size, but also a reduction of myocardial contracture.
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