GSK-3ß at the crossroads in the signalling of heart preconditioning: implication of mTOR and Wnt pathways.

ABSTRACT

AIMS: Ischaemic preconditioning (IPC) protects the heart against prolonged lethal ischaemia through a signalling cascade involving Akt, glycogen synthase kinase-3ß (GSK-3ß), and mitochondrial ATP-sensitive potassium channels (mitoK(ATP)). We previously demonstrated the involvement of the Wnt pathway in IPC in vivo via GSK-3ß. A downstream target might be mammalian target of rapamycin (mTOR) since Wnt can impair tuberous sclerosis complex-2 (TSC2) phosphorylation by inhibiting GSK-3ß. Here, we investigate whether the mTOR pathway is involved in cardioprotection. METHODS AND RESULTS: Isolated-perfused mouse hearts were subjected to IPC via four cycles of ischaemia/reperfusion or pharmacological preconditioning (PPC) by diazoxide, a selective mitoK(ATP) activator. IPC, like PPC, induced an inhibition/phosphorylation of GSK-3ß through Akt activation. Preconditioning also induced phosphorylation of mTOR, p70S6K, and 4E-BP1 that correlated with a significant reduction in infarct size after 40-min ischaemia and 120-min reperfusion when compared with non-preconditioned controls. Preconditioning was impaired in GSK3 knock-in mice. In transgenic mice hearts overexpressing secreted frizzled protein 1 (sFRP1, a Wnt/Frz antagonist), GSK-3ß phosphorylation, mTOR activation, and cardioprotection were impaired. Cardioprotection and its signalling were also inhibited by rapamycin (an mTOR inhibitor), 5-HD (a mitoK(ATP) blocker), and N-(2-mercaptopropionyl) glycine (MPG) as a reactive oxygen species (ROS) scavenger. CONCLUSIONS: We propose that the preconditioning signalling pathway involving an amplification loop results in a downregulation of GSK-3ß and a constant opening of mitoK(ATP) with ROS generation to activate the mTOR pathway and induce cardioprotection. The disruption of the Wnt/Frz pathway by sFRP1 modulates this loop, inducing GSK-3ß activation. This study provides evidence that cardioprotection involves both a pro-survival mTOR pathway and a developmental Wnt pathway targeting GSK-3ß.