REGULATION OF ENDOTHELIAL CONSTITUTIVE NITRIC-OXIDE SYNTHASE BY PROTEIN-KINASE-C

Protein kinase C (PKC) plays a key role in a variety of signal transdu ction processes. The promoter region of the endothelial constitutive n itric oxide synthase (ecNOS) gene contains a transcriptional factor AP -1 binding element. In the present study, we sought to determine the e ffect of PKC inhibition on the expression of ecNOS in cultured bovine aortic endothelial cells (BAEC). The PKC inhibitor staurosporine (10 t o 100 nmol/L) increased the expression of ecNOS mRNA, assessed by Nort hern analysis, in a dose-dependent manner. A newly developed, more spe cific PKC inhibitor, chelerythrine (1 to 3 mu mol/L), also increased t he level of ecNOS mRNA. Incubation of BAEC with phorbol 12-myristate 1 3-acetate (100 nmol/L) for 24 hours, which downregulates PKC, increase d ecNOS mRNA expression. The protein content of ecNOS, assessed by Wes tern analysis, was also increased in staurosporine-treated or cheleryt hrine-treated BAEC. The release of nitrogen oxides from staurosporine- treated or chelerythrine-treated cells both under basal conditions and in response to calcium ionophore A23187 was significantly increased ( P<.05). In conclusion, the present study suggests that regulation of e cNOS is mediated by PKC. The increased release of nitric oxide induced by PKC inhibition may play a protective role against atherogenic proc ess.