Differential effects of nitric oxide on the activity of prostaglandin endoperoxide H synthase-1 and-2 in vascular endothelial cells

A number of studies have demonstrated that prostacyclin and nitric oxide (N O) regulate blood pressure, blood flow and platelet aggregation, In this pa per, we have examined the possible relationship between NO and prostaglandi n endoperoxide H synthase (PGHS)-1 and -2 activities in cultured bovine aor tic endothelial cells. In the non-activated condition endothelial cells exp ressed PGHS-1 activity alone. When these cells were pretreated with aspirin to inactivate their PGHS-1 and then activated by serum and phorbol ester ( TPA) for 6 h, the cells expressed PGHS-2 activity alone. The PGHS activity was assessed by the generation of 6-ketoprostaglandin F-1 alpha (6-ketoPGF( 1 alpha)), a stable metabolite of prostacyclin, after the treatment of thes e cells with arachidonic acid. The simultaneous addition of NOC-7, a NO don or, with arachidonic acid did not affect the production of 6-ketoPGF(1 alph a) in PGHS-1 expressed cells, but attenuated it in PGHS-2-expressed cells. The inhibitory effect of NOC-7 on PGHS-2 activity was dose dependent, and t he different effects of NOC-7 on the activities of PGHS isozymes were also observed in other NO donors. To confirm the different effect of NO on PGHS isozymes demonstrated in the cultured endothelial cells, we carried out an ex vivo perfusion assay in aorta isolated from normal and lipopolysaccharid e (LPS)-treated rats. In the aortae isolated from normal rats, where domina nt expression of PGHS-1 was expected, the NO donor did not affect the PGHS activity, while in aortae isolated from LPS-treated rats, where PGHS-2 was dominantly expressed, the NO donor dramatically inhibited the PGHS activity , suggesting that NO suppressed PGHS-2 activity alone. The inhibitory effec t of NO on PGHS-2 activity was not mediated by cyclic GMP (cGMP), since (a) methylene blue, an inhibitor of soluble guanylate cyclase did not abolish the inhibitory effect of the NO donor on PGHS-2 activity, and (b) 8-Br-cGMP , a permeable cGMP analogue, failed to mimic the effect of NO donors. These data suggest that the effect of NO on prostacyclin production in endotheli al cells was dependent on the expression rate of PGHS-1 and PGHS-2 in the c ells. (C) 2000 Harcourt Publishers Ltd.