Objectives: The cardioprotection against ischemia-reperfusion (I/R) injury consists in reduction of infarct size, limitation of myocardial contracture and improvement of post-ischemic mechanical recovery. Cardioprotection may be obtained with the administration in early reperfusion of the endogenous peptide apelin which acts with a mechanism triggered by the G-protein coupled receptor APJ and includes the PI3K-Akt-NO signaling pathway. PI3K-Akt activation is counteracted by the phosphatase and tensin homolog (PTEN), whose inhibition by Src has been suggested to be required for the effectiveness of the cardioprotective interventions. Since either oxidation or phosphorylation can inhibit PTEN, the present study aims to investigate whether apelin protective mechanism involves PTEN phosphorylation by Src. Materials and methods: The experiments were carried out on Langendorff-perfused rat hearts. In the control group the hearts underwent 30-min of global ischemia and 2-hours of reperfusion. In the apelin treated group, apelin-13 (0.5 μM) was infused during the first 20-min of reperfusion. In another group PP2, the specific inhibitor of Src kinase, was co-infused with apelin. Left ventricular pressure was continuously recorded. After reperfusion infarct size was measured with nitro-blue tetrazolium technique. Western blot analysis was performed to test PTEN phosphorylation. Results: Apelin significantly (p<0.001) reduced infarct size from 60±3 to 30±3% of the left ventricle taken as the risk area. The effect of apelin on infarct size was suppressed by coinfusion of PP2. The increase in diastolic pressure, taken as an index of contracture, reached about 70 mmHg during the first 10 min of reperfusion and declined to about 45 mmHg after 2 hours of reperfusion in control group. This increase was significantly (p<0.001) reduced by apelin so that it remained around 30 mmHg for the entire period of reperfusion. Also in this case the effect of apelin was suppressed by PP2. In control group, left ventricle developed pressure (LVDevP) recovered to about 35% of pre-ischemic value at the end of reperfusion. If apelin was infused, this recovery reached about 70% of the pre-ischemic value at the end of apelin administration and remained almost unchanged for the entire period of reperfusion. Also in this case the effect of apelin was suppressed by PP2. Western blot analysis revealed that apelin increased PTEN phosphorylation, an effect which was suppressed by inhibition of Src kinase with PP2. Conclusion: myocardial protection by apelin against I/R injury includes the inhibition of PTEN by a phosphorylation induced by Src kinase.
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