Comparing the efficacy of pharmacological preconditioning with myristic acid-conjugated, TAT- conjugated and native protein kinase C epsilon peptide activator in myocardial ischemia/reperfusion (MI/R) Models

<div class="line" id="line-21"> <span style='color: rgb(80, 80, 80); font-family: "Open Sans", Arial, sans-serif; font-size: 12px;'> Protein kinase C epsilon (PKC&varepsilon;) activation is thought to be central in mediating cardioprotection conferred by myocardial ischemic preconditioning. PKC&varepsilon; activation via PKC&varepsilon; activator peptide (PKC&varepsilon;+, HDAPIGYD, MW= 887 g/mol) as pretreatment (PT) is a pharmacological means to mimic preconditioning. However, unconjugated or native PKC&varepsilon;+ requires the use of cell permeabilization methods for efficacious intracellular targeting to improve cardiac function and reduce necrotic cell death. Therefore, conjugating PKC&varepsilon;+ to known intracellular delivery moieties may be a method to effectively activate PKC&varepsilon; for cardioprotection. To test this hypothesis, we subjected isolated perfused rat hearts to PT with PKC&varepsilon;+ conjugated to either myristic acid (Myr-HDAPIGYD, MW= 1097 g/mol, 10mM, n=5) or transactivating (TAT) carrier peptide (YGRKKRRQRRR-CC-HDAPIGYD, MW= 2632 g/mol, 10mM, n=6) to evaluate the efficacy of these intracellularly targeted peptide analogs in attenuating contractile dysfunction and infarct size after MI (30 min)/R (90 min) in comparison with native PKC&varepsilon;+ PT (10mM, n=5) and untreated control hearts (n=6). We found that compared to control hearts, PT with Myr-PKC&varepsilon;+ and TAT-PKC&varepsilon;+ significantly reduced infarct size from 41&plusmn;3% to 29 &plusmn;1% and 28&plusmn;2% (p&lt;0.05) respectively compared to control hearts assessed by 1% triphenyltetrazolium chloride staining of heart tissue. By contrast, hearts PT with native PKC&varepsilon;+ showed almost no change in infarct size compared to controls, 41&plusmn;3% versus 35&plusmn;2%. Both myristic acid- and TAT-conjugated PKC&varepsilon;+ PT hearts restored post-reperfused left ventricular developed pressure (LVDP) to 52&plusmn;5% and 50&plusmn;10% compared to both native PKC&varepsilon;+ PT and control hearts which only recovered to 33&plusmn;9% and 38&plusmn;4% of initial baseline values respectively, but this improvement was not statistically significant. These preliminary results indicate that increasing cellular permeability of PKC&varepsilon;+ via conjugation to either myristic acid or TAT significantly improved its efficaciousness in attenuating infarct size when given before ischemia as a pharmacologic mimic of ischemic preconditioning. These results suggest that Myr- or TAT-conjugated PKC&varepsilon;+ may be an effective treatment to attenuate cell death in coronary bypass or organ transplantation settings. </span></div>