Abstract 12257: Selective Inhibition of δPkc-mediated Pyruvate Dehydrogenase Kinase Phosphorylation Alone is Sufficient to Induce Protection From Ischemia and Reperfusion Injury
Rationale: Delta-PKC, whose activation induces cardiac injury following ischemia and reperfusion (I/R) injury, phosphorylates a number of substrates. To determine which of these phosphorylation events is critical to induce cardiac injury, we developed a novel separation-of-function inhibitor that selectively inhibits the phosphorylation of one substrate. We showed that following I/R, delta-PKC-mediated phosphorylaton of pyruvate dehydrogenase kinase (PDK) increased PDK-induced pyruvate dehydrogease phosphorylation and inactivation and inhibition of ATP regeneration. We therefore determined if selective inhibition of delta-PKC-mediated PDK phosphorylation is sufficient to induce cardioprotection.
Methods: We designed a short peptide inhibitor, called ψPDK, to selectively inhibit delta-PKC binding to and phosphorylation of PDK. Using an ex vivo model of I/R, isolated rat hearts (6 per group) were subjected to 30 min ischemia followed by 60 min reperfusion, and the tested peptides (1 µM each) were delivered at reperfusion for 20 minutes. I/R-induced injury was determined using TTC staining, CPK release and activation of stress-induced response.
Results: ψPDK treatment caused a selective inhibition of I/R-induced PDK phosphorylation which resulted in increased ATP levels and decreased H2O2 release. However, the phosphorylaton of two other delta-PKC substrates, the cytoskeletal protein, MARCKS, and the mitochondrial fission protein, dynamin-related protein 1, was not inhibited. Further, ψPDK treatment led to a 50% reduction in cardiac infarction, in CPK release and in JNK phosphorylation, all markers of cardiac injury. DeltaV1-1, a peptide inhibitor of all delta-PKC functions, caused a similar reduction in these markers.
Conclusion: Of the several delta-PKC-mediated phosphorylation events following I/R, delta-PKC-induced PDK phosphorylation is sufficient to cause cardiac injury, as measured in an ex vivo model of myocardial infarction.
- © 2012 by American Heart Association, Inc.