Nitric Oxide Produced by Endothelial Cells Increases Production of Eicosanoids Through Activation of Prostaglandin H Synthase

PGHS-2 (prostaglandin H synthase-2) is induced in mammalian cells by pro-inflammatory cytokines in tandem with iNOS [high-output (‘inducible’) nitric oxide synthase], and is co-localized with iNOS and nitrotyrosine in human atheroma macrophages. Herein, murine J774.2 macrophages incubated with lipopolysaccharide and interferon γ showed induction of PGHS-2 and generated NO using iNOS that could be completely depleted by 12(S)-HPETE [12(S)-hydroperoxyeicosatetraenoic acid; 2.4 μM] or hydrogen peroxide (500 μM) (0.42±0.084 and 0.38±0.02 nmol·min−1·106 cells−1 for HPETE and H2O2 respectively). COS-7 cells transiently transfected with human PGHS-2 also showed HPETE- or H2O2-dependent NO decay (0.44±0.016 and 0.20±0.04 nmol·min−1·106 cells−1 for 2.4 μM HPETE and 500 μM H2O2 respectively). Finally, purified PGHS-2 consumed NO in the presence of HPETE or H2O2 (168 and 140 μM·min−1·μM enzyme−1 for HPETE and H2O2 respectively), in a haem-dependent manner, with 20 nM enzyme consuming up to 4 μM NO. Km (app) values for NO and 15(S)-HPETE were 1.7±0.2 and 0.45±0.16 μM respectively. These data indicate that PGHS-2 catalytically consumes NO during peroxidase turnover and that pro-inflammatory cytokines simultaneously upregulate NO synthesis and degradation pathways in murine macrophages. Catalytic NO consumption by PGHS-2 represents a novel interaction between NO and PGHS-2 that may impact on the biological effects of NO in vascular signalling and inflammation.

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Inhibition by Extracellular cAMP of Phorbol 12-Myristate 13-Acetate-induced Prostaglandin H Synthase-2 Expression in Human Pulmonary Microvascular Endothelial Cells

Journal of Biological Chemistry ◽

10.1074/jbc.275.18.13662 ◽

2000 ◽

Vol 275 (18) ◽

pp. 13662-13667 ◽

Cited By ~ 14

Author(s):

Ismaı̈l Elalamy ◽

Fatima Ait Said ◽

Monique Singer ◽

Jean-Paul Couetil ◽

Mohamed Hatmi

Keyword(s):

Endothelial Cells ◽

Microvascular Endothelial Cells ◽

Prostaglandin H Synthase ◽

Pulmonary Microvascular Endothelial Cells ◽

Microvascular Endothelial ◽

Prostaglandin H ◽

Extracellular Camp

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Ibuprofen protects human endothelial cell prostaglandin H synthase from hydrogen peroxide

AJP Cell Physiology ◽

10.1152/ajpcell.1996.271.6.c1879 ◽

1996 ◽

Vol 271 (6) ◽

pp. C1879-C1886 ◽

Cited By ~ 6

Author(s):

D. A. Wessels ◽

S. L. Hempel

Keyword(s):

Hydrogen Peroxide ◽

Endothelial Cells ◽

Arachidonic Acid ◽

Endothelial Cell ◽

Arachidonic Acid Release ◽

Human Endothelial Cells ◽

Prostaglandin H Synthase ◽

Cyclooxygenase Activity ◽

Acid Release ◽

Prostaglandin H

Human endothelial cells exposed to H2O2 demonstrate decreased prostacyclin (PGI2) synthesis due to decreased prostaglandin H synthase (PGH synthase) activity. We tested the hypothesis that PGH synthase activity could be protected from H2O2 by a reversible nonsteroidal anti-inflammatory drug. Experiments demonstrate that ibuprofen if present during H2O2 exposure, protects endothelial cell PGH synthase against the decrease in prostaglandin formation caused by H2O2. Additional studies demonstrated that decreasing arachidonic acid release from cell phospholipids during H2O2 exposure did not protect PGI2 synthesis following H2O2 exposure. In other experiments, ibuprofen did not chelate Fe2+ in a conformation that inhibited the reactivity of Fe2+. In addition, ibuprofen did not scavenge HO. However, we demonstrate that ibuprofen significantly protects purified PGH synthase cyclooxygenase activity from the effects of H2O2. The results confirm the hypothesis. These findings suggest that ibuprofen displaces oxidant species from the cyclooxygenase site of PGH synthase, thereby preventing oxidation of the functional groups important for PGH synthase activity.

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