title: deltaPKC inhibition or varepsilonPKC activation repairs endothelial vascular dysfunction by regulating eNOS post-translational modification.
abstract: The balance between endothelial nitric oxide synthase (eNOS)-derived nitric oxide (NO) and reactive oxygen species (ROS) production determines endothelial-mediated vascular homeostasis. Activation of protein kinase C (PKC) has been linked to imbalance of the eNOS/ROS system, which leads to endothelial dysfunction. We previously found that selective inhibition of delta PKC (deltaPKC) or selective activation of epsilon PKC (varepsilonPKC) reduces oxidative damage in the heart following myocardial infarction. In this study we determined the effect of these PKC isozymes in the survival of coronary endothelial cells (CVEC). We demonstrate here that serum deprivation of CVEC increased eNOS-mediated ROS levels, activated caspase-3, reduced Akt phosphorylation and cell number. Treatment with either the deltaPKC inhibitor, deltaV1-1, or the varepsilonPKC activator, psivarepsilonRACK, inhibited these effects, restoring cell survival through inhibition of eNOS activity. The decrease in eNOS activity coincided with specific de-phosphorylation of eNOS at Ser1179, and eNOS phosphorylation at Thr497 and Ser116. Furthermore, deltaV1-1 or psivarepsilonRACK induced physical association of eNOS with caveolin-1, an additional marker of eNOS inhibition, and restored Akt activation by inhibiting its nitration. Together our data demonstrate that (1) in endothelial dysfunction, ROS and reactive nitrogen species (RNS) formation result from uncontrolled eNOS activity mediated by activation of deltaPKC or inhibition of varepsilonPKC; (2) inhibition of deltaPKC or activation of varepsilonPKC corrects the perturbed phosphorylation state of eNOS, thus increasing cell survival. Since endothelial health ensures better tissue perfusion and oxygenation, treatment with a deltaPKC inhibitor and/or an varepsilonPKC activator in diseases of endothelial dysfunction should be considered.
authors.affiliation: Department of Molecular Biology, University of Siena, Via Aldo Moro, 2, 53100, Siena, Italy.
authors.forename: Martina
authors.initials: M
authors.lastname: Monti
authors.forename: Sandra
authors.initials: S
authors.lastname: Donnini
authors.forename: Antonio
authors.initials: A
authors.lastname: Giachetti
authors.forename: Daria
authors.initials: D
authors.lastname: Mochly-Rosen
authors.forename: Marina
authors.initials: M
authors.lastname: Ziche
mesh_terms: D000818:Animals; D017209:Apoptosis; D002417:Cattle; D051242:Caveolin 1 / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism / Q000473:pathology*; D004789:Enzyme Activation; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D010100:Oxygen / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors*; D051744:Protein Kinase C-epsilon / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D026361:Reactive Nitrogen Species
chemical_list: D051242:Caveolin 1; D026361:Reactive Nitrogen Species; D052250:Nitric Oxide Synthase Type III; D051745:Protein Kinase C-delta; D051744:Protein Kinase C-epsilon; D006152:Cyclic GMP; D010100:Oxygen