title: Ischemic post-conditioning reduces infarct size of the in vivo rat heart: role of PI3-K, mTOR, GSK-3beta, and apoptosis.
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-3beta) 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-3beta 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-3beta 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-3beta, 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.
authors.affiliation: Department of Medicine and Cardiology, Heart Center Dresden University Hospital, University of Technology Dresden, P.O. Box 95, Fetscherstr. 76, Dresden 01307, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Wagner
authors.forename: Diana
authors.initials: D
authors.lastname: Tillack
authors.forename: Gregor
authors.initials: G
authors.lastname: Simonis
authors.forename: Ruth H
authors.initials: RH
authors.lastname: Strasser
authors.forename: Christof
authors.initials: C
authors.lastname: Weinbrenner
mesh_terms: D000818:Animals; D017209:Apoptosis; D015153:Blotting, Western; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006321:Heart / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D007511:Ischemia / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D008297:Male; D015428:Myocardial Reperfusion Injury / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D058570:TOR Serine-Threonine Kinases
chemical_list: D038362:Glycogen Synthase Kinase 3; D000071679:Glycogen Synthase Kinase 3 beta; D047908:Intracellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases; D017346:Protein Serine-Threonine Kinases; D058570:TOR Serine-Threonine Kinases; C546845:mTOR protein, rat; C000605508:Gsk3b protein, rat