7-(3-(3-hydroxy-4-(4--iodophenoxy)-1-butenyl)-7-oxabicyclo(2.2.1)heptan-2-yl)-5-heptenoic-acid and 3-nitrotyrosine

The thromboxane receptor (TPr) and multiple TPr ligands, including thromboxane A(2) (TxA(2)) and prostaglandin H(2), are elevated during vascular and atherothrombotic diseases. How TPr stimulation causes vascular injury remains poorly defined. This study was conducted to investigate the mechanism by which TPr stimulation leads to vascular injury.. Exposure of bovine aortic endothelial cells to either [1S-(1α,2β(5Z),3α(1E,3R),4α]-7-[3-(3-hydroxy-4-(d'-iodophenoxy)-1-butenyl)-7-oxabicyclo-[2.2.1] heptan-2-yl]-5'-heptenoic acid (IBOP) or U46619, 2 structurally related TxA(2) mimetics, for 24 hours markedly increased the release of superoxide anions (O(2)(·-)) and peroxynitrite (ONOO(-)) but reduced cyclic GMP, an index of nitric oxide bioactivity. IBOP also significantly suppressed activity of endothelial nitric oxide synthase (eNOS), increased enzyme-inactive eNOS monomers, and reduced levels of tetrahydrobiopterin, an essential eNOS cofactor. IBOP- and U46619-induced increases in O(2)(·-) were accompanied by the membrane translocation of the p67(phox) subunit of NAD(P)H oxidase. Pharmacological or genetic inhibition of either NAD(P)H oxidase or TPr abolished IBOP-induced O(2)(·-) formation. Furthermore, TPr activation significantly increased protein kinase C-ζ (PKC-ζ) in membrane fractions and PKC-ζ phosphorylation at Thr410. Consistently, PKC-ζ inhibition abolished TPr activation-induced membrane translocation of p67(phox) and O(2)(·-) production. Finally, exposure of isolated mouse aortae to IBOP markedly increased O(2)(·-) in wild-type but not in those from gp91(phox) knockout mice.. We conclude that TPr activation via PKC-ζ-mediated NAD(P)H oxidase activation increases both O(2)(·-) and ONOO(-), resulting in eNOS uncoupling in endothelial cells.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Apoptosis; Biopterins; Bridged Bicyclo Compounds, Heterocyclic; Cattle; Cell Survival; Cells, Cultured; Cyclic GMP; Cytochrome P-450 Enzyme System; Endothelial Cells; Enzyme Activation; Enzyme Inhibitors; Fatty Acids, Unsaturated; Intramolecular Oxidoreductases; Mice; Mice, Inbred C57BL; Mice, Knockout; NADPH Oxidases; Nitric Oxide; Nitric Oxide Synthase Type III; Oxidative Stress; Peroxynitrous Acid; Phosphoproteins; Phosphorylation; Protein Kinase C; Protein Processing, Post-Translational; Protein Transport; Receptors, Immunologic; Receptors, Thromboxane A2, Prostaglandin H2; RNA Interference; Signal Transduction; Superoxides; Time Factors; Tyrosine