δPKC interaction with the d subunit of F₁Fo ATP synthase impairs energetics and exacerbates ischemia/reperfusion injury in isolated rat hearts

Previously, we demonstrated protection against hypoxic injury in neonatal cardiac myocytes and reduced release of cardiac troponin I from perfused rat hearts by a novel peptide inhibitor [NH₂-YGRKKRRQRRRMLATRALSLIGKRAISTSVCAGRKLALKTIDWVSFDYKDDDDK-] of the delta protein kinase C (δPKC) interaction with the "d" subunit of mitochondrial F₁Fo ATP synthase (dF₁Fo). This peptide was developed in our laboratory and contains: an HIV-Tat protein transduction domain; a mitochondrial targeting motif; the δPKC-dF₁Fo inhibitor sequence; and a FLAG epitope. In the present study the δPKC-dF₁Fo inhibitor attenuated co-immunoprecipitation of δPKC with dF₁Fo, improved recovery of contractility, diminished levels of tissue t-carbonyls and 4-hydroxy-2-nonenal (HNE), and reduced 2,3,5-triphenyltetrazolium chloride-monitored infarct size following simulated global ischemia/reperfusion (IR) exposures. Perfusion of hearts with this peptide prior to IR enhanced ATP levels 2.1-fold, improved ADP (state 3)- and FCCP (maximal)-stimulated respiration in mitochondrial oxygen consumption assays, and attenuated Ca⁺⁺-induced mitochondrial swelling following ischemic injury. Mitochondrial membrane potential (assessed by JC-1) was also improved 1.6-fold by the inhibitor in hearts subsequently exposed to IR injury. Brief IR exposures did not cause mitochondrial loss of cytochrome c in the presence or absence of the inhibitor. Additionally, the inhibitor did not modify accumulation of the autophagy marker LC3_II after brief IR injury. Our results support the potential for this first-in-class peptide as a translational agent for combating cardiac IR injury.