Regulation of skin microvasculature angiogenesis, cell migration, and permeability by a specific inhibitor of PKCα

Activation of protein kinase C (PKC) induces phenotypic changes in the morphology of microvascular endothelial cells that affect major functions of the microvasculature. These functions include the first stages of sprouting in angiogenesis, cell migration following wounding, and vascular permeability. The specific isoform(s) of PKC responsible for each of these changes has not been previously identified. In this study, we used two inflammatory agents, IL-1 beta and phorbol myristic acetate, to activate PKC isozymes and specific inhibitors of PKC alpha (Go6976) and PKC beta (hispidin) to distinguish how each of these isoform(s) controls angiogenesis, wound healing, and permeability. In all cases, only inhibition of PKC a inhibited each of these functions when compared to the inhibition of PKC beta. Additional analysis of the mechanism of action of Go6976 (RT-PCR, Western blots, and immunohistochemistry) of the changes in the phosphorylated and nonphosphorylated forms of PKC alpha in the cell membrane and cytoplasm confirmed the specificity of PKC alpha inhibition by Go6976. These studies therefore indicate a specific and a regulatory role of the PKC a isoform in three major endothelial cell functions that are important in the maintenance of microvascular homeostasis.