Thrombin triggers the de novo expression of an inducible NO synthase in porcine aortic valve endothelial cells

The nitric oxide (NO) production by porcine aortic valve endothelial cells was estimated in cusps incubated at 37°C by measuring their cyclic GMP content and the nitrite levels of the incubation medium. After a stabilization period, incubations for 5 min with acetylcholine, bradykinin, ADP and bovine thrombin resulted in a receptor-mediated increase in cyclic GMP which could be blocked by EGTA, $\text{N-ω-}\text{nitro}\text{-}\text{L}\text{-}\text{arginine}$ methyl ester (L-NAME) and N^G-monomethyl-L-arginine (L-NMMA). Incubation with lipopolysaccharide (endotoxin) from E. coli O111:B4 or bovine thrombin for 5 h, dose-dependently increased nitrite production as well as cyclic GMP content. The elevated nitrite production was completely abolished in the presence of the protein synthesis inhibitor cycloheximide, was reduced by more than 50% by dexamethasone but was not affected by EGTA. L-NMMA dose-dependently reduced the increased nitrite production and cyclic GMP content. These results suggest that besides the presence of a constitutive NO synthase in porcine aortic valve endothelial cells thrombin, like lipopolysaccharide, triggers the de novo expression of an inducible Ca²⁺-independent NO synthase.