Cardiopulmonary bypass with cardioplegic arrest activates protein kinase C in the human myocardium

Abstract

Background: The protein kinase C (PKC) family consists of 12 isoforms, 6 of which have been found in human myocardium (PKC alpha, beta I/beta II, delta, epsilon, eta, and lambda/iota). These kinases function in regulation of contractility, ion channels, and in cellular protection or damage during ischemia-reperfusion injury. This study investigated the effects of controlled ischemia-reperfusion injury through cardioplegic arrest on PKC activity in patients undergoing cardiac surgery.

Study design: Myocardium and skeletal muscle were harvested from patients before and after cardiopulmonary bypass and cardioplegic arrest. Total PKC (n=12) was isolated and specifically, the PKC delta (n=8) and PKC epsilon (n=8) isoforms were immunoprecipitated for use in functional kinase assays. Cellular fractions (n=4) were separated by differential ultracentrifugation and analyzed by Western blotting for membrane translocation (an indirect indicator of increased activity). Immunofluorescent staining for PKC delta and PKC epsilon was performed on myocardial sections. An in vitro assay of hypoxic cardioplegic arrest followed by reoxygenation was performed using isolated cardiomyocytes, and apoptosis was assessed.

Results: Cardioplegic arrest was associated with a 24.4% +/- 4.6% increase (p < 0.05) in total PKC activity, a 26.7% +/- 4.9% increase (p < 0.05) in PKC delta activity, and a 35.3% +/- 14% increase (p < 0.05) in PKC epsilon activity in myocardium. Cardioplegic arrest induced migration of PKC delta and epsilon to the z-line of the cardiomyocyte. Inhibition of PKC delta in the in vitro studies demonstrated a considerable reduction in apoptotic cells.

Conclusions: PKC delta and epsilon have previously been shown to mediate and protect, respectively, from ischemia-reperfusion injury after myocardial ischemia. Demonstration of increases in their activity after cardioplegic arrest provides support for their possible role in myocardial function after cardiac surgery. Isoform-specific modulators may be of potential therapeutic value in treating postoperative myocardial dysfunction.