Age- and sex-dependent alterations in protein kinase C (PKC) and extracellular regulated kinase 1/2 (ERK1/2) in rat myocardium

Cardiovascular morbidity and mortality increase significantly with advancing age, with proportionally higher rates occurring in aged women when compared to aged men. The signaling alterations responsible for age-related reductions in ischemic stress reserves, particularly in aged women, are poorly understood. Accordingly, we sought to determine whether alterations in the cellular location and formation of specific protein kinase C (PKC)-extracellular regulated1/2 (ERK1/2) signaling modules (SMS) might provide insight into known age- and sex-related differences in cardiovascular disease outcomes. Cytosolic (Cyto), mitochondrial (Mito) and nuclear (Nuc) fractions were isolated from left ventricles of male (M) and female (F) adult (6mo), castrated or aged (23mo) F344 rats by centrifugation. Western blotting was used to assess PKC (α, δ, ε), p-ERK1/2 and p-Bad(Ser¹¹²) levels, and immunoprecipitation to assess PKC-ERK1/2 SMS. Cyto-PKCα levels increased with age (p < 0.0001), whereas increases in cyto-PKCα-ERK1/2 SMS were only observed in aged F (60%; p < 0.01). Mito-PKCδ and Mito-PKCδ-ERK1/2 SMS increased in M and F with age (p < 0.0001); however increases in Cyto-PKCδ were only observed in aged M (80% p < 0.0001). It is important to note that Nuc- and Mito-PKCδ-ERK1/2 SMS were 3.5- and 4.8-fold greater in males versus females, respectively (p < 0001). Increases in Mito-PKCε-ERK1/2 SMS (216%) were also specific to aged M (p < 0.0001), however, Mito-p-Bad(Ser¹¹²) levels were decreased with age in both M and F. Differences in sex hormone status could not fully account for observed age-related differences in PKC. Collectively, our results provide novel evidence for age and sex-related differences in the magnitude and distribution of cardiac PKC-ERK1/2 SMS consistent with previously described pathological and protective phenotypes, respectively.