Differences in the expression of protein kinase C isoforms and its translocation after stimulation with phorbol ester between young-adult and middle-aged ventricular cardiomyocytes isolated from Fischer 344 rats.

Abstract

It is known that the tolerance against ischemia-reperfusion and the effects of preconditioning decrease in aged hearts, but the mechanisms responsible for this diminished ischemic tolerance and reduced efficacy of preconditioning remain unknown. To determine the age-related changes in these mechanisms, protein kinase C (PKC) isoform expression and its translocation by phorbol ester were analyzed because PKC is believed to be involved in preconditioning. Immunoblotting and immunostaining analysis were performed with isoform-specific PKC antibodies using cardiomyocytes isolated from young-adult (12-week-old: 12W) and middle-aged (50-week-old: 50W) Fischer 344 rats. There was significantly greater PKC-delta expression in both the cytosolic and membrane fractions of 12W cardiomyocytes than in 50W ones. Exposure of cardiomyocytes to 100 nmol/L 4-beta-phorbol 12-myristate 13-acetate (PMA) caused translocation of PKC-delta from the cytosol to the membrane in the 12W group, whereas in the 50W group, the translocation was attenuated. Immunostaining confirmed the PKC-delta translocation in the 12W cardiomyocytes. Oil pellet examination showed that the translocation of PKC-delta induced by preconditioning was associated with cell protection from ischemic injury in the 12W group only. Age-related changes in PKC isoform expression and activation in cardiomyocytes might be responsible for the reduced ischemic tolerance and less efficient preconditioning that accompanies aging.