Selected isozymes of PKC contribute to augmented growth of fetal and neonatal bovine PA adventitial fibroblasts

Abstract

We sought to determine which isozymes of protein kinase C (PKC) contribute to the increased proliferation of immature bovine pulmonary artery (PA) adventitial fibroblasts. Seven were identified in lysates of neonatal PA fibroblasts by Western blot: three Ca2+ dependent (α, βI, and βII) and four Ca2+ independent (δ, ε, ζ, and μ). Four isozymes (γ η, Θ, and ι) were not detected in fibroblasts isolated at any developmental stage. Of the seven detected isozymes, only PKC-α and -βII protein levels were higher in fetal and neonatal cells compared with adult fibroblasts. Their role in the enhanced growth of immature fibroblasts was then evaluated. The isozyme nonselective PKC inhibitor Ro-31-8220 was first compared with GF-109203X, a structural analog of Ro-31-8220 with relative specificity for the Ca2+- dependent isozymes of PKC. GF-109203X selectively inhibited the growth of immature cells and was nearly as potent as Ro-31-8220. Go-6976, a more specific inhibitor of the Ca2+-dependent isozymes, mimicked the antiproliferative effect of GF-109203X. PKC downregulation with 1 μM phorbol 12-myristate 13-acetate had the same selective antiproliferative effect on immature fibroblasts as GF-109203X and Go-6976. The protein levels of PKC-α and -βII, but not of PKC-βI, were completely degraded in response to phorbol 12-myristate 13-acetate pretreatment. These results suggest that PKC- α and -βII are important in the augmented growth of immature bovine PA adventitial fibroblasts.