REGULATION OF ENDOTHELIAL-CELL NITRIC-OXIDE SYNTHASE MESSENGER-RNA EXPRESSION BY SHEAR-STRESS

Shear stress enhances expression of Ca2+- calmodulin-sensitive endothe lial cell nitric oxide synthase (ecNOS) mRNA and protein in bovine aor tic endothelial cells (BAEC). The present studies were performed to in vestigate mechanisms responsible for regulation of ecNOS mRNA expressi on by shear stress and to determine if this induction of ecNOS mRNA is accompanied by an enhanced nitric oxide (NO) production. Shear stress es of 15 dyn/cm(2) for 3-24 h resulted in a two- to threefold increase of ecNOS mRNA content quantified by Northern analysis in BAEC. Shear stresses (1.2-15 dyn/cm(2)) for 3 h resulted in an induction of ecNOS mRNA in a dose-dependent manner. In human aortic endothelial cells, sh ear stresses of 15 dyn/cm(2) for 3 h also resulted in ecNOS mRNA induc tion. In BAEC, this induction in ecNOS mRNA was prevented by coincubat ion with actinomycin D (10 mu g/ml). The KS channel antagonist tetraet hylammonium chloride (3 mM) prevented increase in ecNOS mRNA in respon se to shear stress. The ecNOS promotor contains putative binding domai ns for AP-1 complexes, potentially responsive to activation of protein kinase C (PKC). However, selective PKC inhibitor calphostin C (100 nM ) did not inhibit ecNOS induction by shear stress. Finally, production of nitrogen oxides under both basal conditions and in response to the calcium ionophore A-23187 (1 mu M) by BAEC exposed to shear stress wa s increased approximately twofold compared with cells not exposed to s hear stress. These data suggest that ecNOS mRNA expression is regulate d by K+ channel opening, but not by activation of PKC, and that shear not only enhances ecNOS mRNA expression but increases capacity of endo thelial cells to release NO.