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Ischemic post-conditioning reduces infarct size of the in vivo rat heart: role of PI3-K, mTOR, GSK-3β, and apoptosis

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Abstract

Post-conditioning by repetitive cycles of reperfusion/ischemia after prolonged ischemia protects the heart from infarction. The objectives of this study were: Are kinases (PI3-kinase, mTOR, and GSK-3β) involved in the signaling pathway of post-conditioning? Does post-conditioning result in a diminished necrosis or apoptosis? In open chest rats the infarct size was determined after 30 min of regional ischemia and 30 min of reperfusion using propidium iodide and microspheres. Post-conditioning was performed by three cycles of 30 s reperfusion and reocclusion each, immediately upon reperfusion. PI3-kinase and mTOR were blocked using wortmannin (0.6 mg/kg) or rapamycin (0.25 mg/kg), respectively. The phosphorylation of GSK-3β and p70S6K was determined with phospho-specific antibodies. TUNEL staining and detection of apoptosis-inducing factor (AIF) were used for the determination of apoptosis. Control hearts had an infarct size of 49 ± 3%, while post-conditioning significantly reduced it to 29 ± 3% (P < 0.01). Wortmannin as well as rapamycin completely blocked the infarct size reduction of post-conditioning (51 ± 2% and 54 ± 5%, respectively). Western blot analysis revealed that post-conditioning increased the phosphorylation of GSK-3β by 2.3 times (P < 0.01), and this increase could be blocked by wortmannin, a PI3-kinase inhibitor. Although rapamycin blocked the infarct size reduction, phosphorylation of p70S6K was not increased in post-conditioned hearts. After 2 h of reperfusion, the post-conditioned hearts had significantly fewer TUNEL-positive nuclei (35 %) compared to control hearts (53%; P < 0.001). AIF was equally reduced in post-conditioned rat hearts (P < 0.05 vs. control). Infarct size reduction by ischemic post-conditioning of the in vivo rat heart is PI3-kinase dependent and involves mTOR. Furthermore, GSK-3β, which is thought to be a regulator of the mPTP, is part of the signaling pathway of post-conditioning. Finally, apoptosis was inhibited by post-conditioning, which was shown by two independent methods. The role of apoptosis and/or autophagy in post-conditioning has to be further elucidated to find therapeutic targets to protect the heart from the consequences of acute myocardial infarction.

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Acknowledgments

Part of this study was presented at the meetings of the European Society of Cardiology, the International Society of Heart Research, and the German Society of Cardiology 2006. This study was supported by the Deutsche Forschungsgemeinschaft, Bonn (We 1955/2-2). Part of this work was sponsored by a grant from the Roland Ernst Stiftung für Gesundheitswesen. We thank Janet Lehmann for her excellent technical assistance.

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  1. Department of Medicine and Cardiology, Heart Center Dresden University Hospital, University of Technology Dresden, P.O. Box 95, Fetscherstr. 76, Dresden, 01307, Germany

    Claudia Wagner, Diana Tillack, Gregor Simonis, Ruth H. Strasser & Christof Weinbrenner

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Wagner, C., Tillack, D., Simonis, G. et al. Ischemic post-conditioning reduces infarct size of the in vivo rat heart: role of PI3-K, mTOR, GSK-3β, and apoptosis. Mol Cell Biochem 339, 135–147 (2010). https://doi.org/10.1007/s11010-009-0377-x

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