Regulation of prostaglandin biosynthesis by nitric oxide is revealed by targeted deletion of inducible nitric-oxide synthase

Abstract

We investigated the effects of targeted deletion of the inducible NO synthase (iNOS) gene on the formation of prostaglandins in vivo and ex vivo. Peritoneal macrophages were obtained from control and iNOS-deficient mice, and prostaglandin E2 (PGE2) was quantified after stimulation with γ- interferon and lipopolysaccharide to induce COX-2. Total nitrate and nitrite production was completely abolished in cells from iNOS-deficient animals compared with control cells. PGE2 formation by cells from iNOS-deficient animals was decreased compared with cells from control animals 80% at 12 h (0.85 ± 0.90 ng/106 cells versus 15.4 ± 2.1 ng/106 cells, p < 0.01) and 74% at 24 h (9.4 ± 4.3 ng/106 cells versus 36.8 ± 4.1 ng/106 cells, p < 0.01). COX-2 protein expression was not significantly different in cells from control or knockout animals. Levels of PGE2 in the urine of iNOS-deficient mice were decreased 78% (0.24 ± 0.14 ng/mg of creatinine versus 1.09 ± 0.66 ng/mg of creatinine, p < 0.01) compared with control animals. In addition, the levels of urinary F2-isoprostanes, an index of endogenous oxidant stress, were significantly decreased in iNOS-deficient animals. In contrast, the levels of thromboxane B2 derived from platelets allowed to aggregate ex vivo were significantly increased in iNOS-deficient mice compared with wild- type mice. These studies support the hypothesis that NO and/or NO-derived species modulate cyclooxygenase activity and eicosanoid production in vivo.