Protein Kinase C Inhibitors Prevent Cultured Human Endothelial Cell Stress Fiber Formation, but not Heightened Endocytosis

In order to study the signal transduction mechanism of endothelial perturbation, the effects of phorbol 12-myristate 13-acetate (PMA) and phorbol 12,13-dibutyrate (PDBu), both protein kinase C (PKC) activators, on cultured human endothelial cell (EC) hydrogen peroxide (H2O2) generation, endocytotic activity, and cytoskeletal structure have been investigated. EC were incubated with 1-100 nM PMA, or PDBu, and cellular H2O2 generation and endocytotic activity measured. PMA and PDBu exposure caused dose-dependent rises in EC H2O2 production. Likewise, EC incubated with PMA and PDBu had dose-related endocytosis increases. Cytoskeletal inspection of 10 nM PMA-perturbed EC revealed structural remodeling with stress fiber formation. Similar cellular functional changes occur in EC exposed to high low-density lipoprotein (LDL) concentrations. Protein kinase C (PKC) inhibition with 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7) prevented cytoskeletal remodeling in PMA-stimulated EC. In differenc...