Ischaemic preconditioning improves proteasomal activity and increases the degradation of δPKC during reperfusion

Aims The response of the myocardium to an ischemic insult is regulated by two highly homologous protein kinase C (PKC) isozymes, δ and ϵPKC. Here we determined the spatial and temporal relationships between these two isozymes in the context of ischemia/reperfusion (I/R) and ischemic preconditioning (IPC) to better understand their roles in cardioprotection. Methods and results Using an ex vivo rat model of myocardial infarction, we found that short bouts of ischemia and reperfusion prior to the prolonged ischemic event (IPC) diminished δPKC translocation by 3.8 fold and increased ϵPKC accumulation at mitochondria by 16 fold during reperfusion. In addition, total cellular levels of δPKC decreased by 60±2.7% in response to IPC, whereas the levels of ϵPKC did not significantly change. Prolonged ischemia induced a 48±11% decline in the ATP-dependent proteasomal activity and increased the accumulation of misfolded proteins during reperfusion by 192±32%; both of these events were completely prevented by IPC. Pharmacological inhibition of the proteasome or selective inhibition of ϵPKC during IPC restored δPKC levels at the mitochondria while decreasing ϵPKC levels, resulting in a loss of IPC-induced protection from I/R. Importantly, increased myocardial injury was the result, in part, of restoring a δPKC-mediated I/R pro-apoptotic phenotype by decreasing pro-survival signaling and increasing cytochrome c release into the cytosol. Conclusion Taken together, our findings indicate that IPC prevents I/R injury at reperfusion by protecting ATP-dependent 26S proteasomal function. This decreases the accumulation of the pro-apoptotic kinase, δPKC, at cardiac mitochondria, resulting in the accumulation of the pro-survival kinase, ϵPKC.