Glycogen Synthase Kinase-3 Inactivation Is Not Required for Ischemic Preconditioning or Postconditioning in the Mouse

Abstract

Abstract-The inactivation of glycogen synthase kinase-3␤ (GSK-3␤) is proposed as the event integrating protective pathways initiated by preconditioning and other interventions. The inactivation of GSK-3 is thought to decrease the probability of opening of the mitochondrial permeability transition pore. The aim of this study was to verify the role of GSK-3 using a targeted mouse line lacking the critical N-terminal serine within GSK-3␤ (Ser9) and the highly homologous GSK-3␣ (Ser21), which when phosphorylated results in kinase inactivation. Postconditioning with 10 cycles of 5 seconds of reperfusion/5 seconds of ischemia and preconditioning with 6 cycles of 4 minutes of ischemia/6 minutes of reperfusion, similarly reduced infarction of the isolated perfused mouse heart in response to 30 minutes of global ischemia and 120 minutes of reperfusion. Preconditioning caused noticeable inactivating phosphorylation of GSK-3. However, both preconditioning and postconditioning still protected hearts of homozygous GSK-3 double knockin mice. Moreover, direct pharmacological inhibition of GSK-3 catalytic activity with structurally diverse inhibitors before or after ischemia failed to recapitulate conditioning protection. Nonetheless, cyclosporin A, a direct mitochondrial permeability transition pore inhibitor, reduced infarction in hearts from both wild-type and homozygous GSK-3 double knockin mice. Furthermore, in adult cardiac myocytes from GSK-3 double knockin mice, insulin exposure was still as effective as cyclosporin A in delaying mitochondrial permeability transition pore opening. Our results, which include a novel genetic approach, suggest that the inhibition of GSK-3 is unlikely to be the key determinant of cardioprotective signaling in either preconditioning or postconditioning in the mouse. (Circ Res. 2008;103:307-314.)Key Words: postconditioniong Ⅲ preconditioning Ⅲ GSK-3 Ⅲ mPTP I schemic preconditioning is a powerful and established modulator of intrinsic myocardial resistance to lethal ischemia. 1 Its clinical application in patients with acute myocardial infarction is limited as a result of the inability to predict the moment of coronary artery occlusion. More recently, Zhao et al described the concept of postconditioning as a novel strategy for protecting the heart against reperfusion-injury. 2 They demonstrated that a similar regimen of brief periods of ischemia after a lethal index ischemic insult was as protective as preconditioning. This phenomenon has subsequently been confirmed in a variety of animal models, both in vivo and in isolated heart preparations, 3 as well as in preliminary clinical interventional studies. 4,5 The endogenous cellular and molecular mechanisms involved in cardioprotection against ischemia have been extensively investigated in the field of preconditioning. 6 Because pre-and postconditioning are temporally remote, the signaling pathways might be expected to differ considerably. 2,7 However, recent evidence suggests that the early reperfusion phase after lethal ischemia is crucial i...

Cardioprotective Pathways During Reperfusion: Focus on Redox Signaling and Other Modalities of Cell Signaling

“…Cross-talk between different modalities of signaling may occur and may be the basis of controversial observations. Therefore a better understanding of this crosstalk may contribute to solve the conundrum relative to the role of RISK and=or SAFE pathway in cardioprotection deriving from conflicting results obtained with different models of pre-and postconditioning (14,70,131,174,190). It will be important to understand how posttransitional modifications of different proteins are regulated in the context of cardioprotection.…”

“…Therefore, apart from cGMP=PKG, RISK, and SAFE pathways, there are redox mechanisms and events that may be considered alternative or cooperative to the classical pathways. Of note, the importance of RISK has been recently questioned, and its relevance in larger mammals seems to be severely limited; from a translational point of view these animal models may be more suitable (76,131,173). Concerns and controversies about the possibilities of translating the results of animal experiments into human clinical trials have recently been discussed by van der Worp et al (189).…”

“…Some evidence indicates a potential mitochondrial isoform of OGT (93,128,130,131). Moreover, it has been reported that augmentation of O-GlcNAcylation levels in cardiomyocytes attenuates H 2 O 2 -induced loss of mitochondrial membrane potential (23,93,128,131) and that this may be due to O-GlcNAcylation of mitochondrial proteins such as VDAC. All these studies suggest that ROS from mitochondria contribute to the regulation of O-GlcNAcylation of proteins, which could, in turn, modulate the mitochondria response to oxidative-stress.…”

A Chemical Genetic Approach Reveals That p38α MAPK Activation by Diphosphorylation Aggravates Myocardial Infarction and Is Prevented by the Direct Binding of SB203580

“…Coronary flow was measured by timed collection of perfusate. More detailed methods and exclusion and inclusion criteria were as described previously (3,9,14,15).…”

“…All analyses of infarct size were done by an investigator who was blinded with regard to the group assignments. A more detailed description of the methods is provided in previous publications (3,9,15).…”