Inducible Nitric Oxide Synthase Mediates Prostaglandin H2 Synthase Nitration and Suppresses Eicosanoid Production

• DOI: 10.2353/ajpath.2006.050090
• Publication Date: 2007-03-16T19:36:39Z
• Authors: Ruba S. Deeb; Hao Shen; Caryn Gamss; Tatyana Gavrilova; Barbara D. Summers; Rosemary Kraemer; Gang Hao; Steven S. Gross; Muriel Lainé; Nobuyo Maeda; David P. Hajjar; Rita K. Upmacis
• Keywords: Male; 0301 basic medicine; 0303 health sciences; Blotting, Western; Myocytes, Smooth Muscle; Fluorescent Antibody Technique; Nitric Oxide Synthase Type II; Coronary Artery Disease; Blotting, Northern; Immunohistochemistry; Muscle, Smooth, Vascular; Mice; 03 medical and health sciences; Apolipoproteins E; Prostaglandin-Endoperoxide Synthases; Animals; Eicosanoids; Tyrosine; Female; Aorta; Cells, Cultured

Nitric oxide (NO) modulates the biological levels of arachidonate-derived cell signaling molecules by either enhancing or suppressing the activity of prostaglandin H2 isoforms (PGHS-1 and PGHS-2). Whether NO activates or suppresses PGHS activity is determined by alternative protein modifications mediated by NO and NO-derived species. Here, we show that inducible NO synthase (iNOS) and PGHS-1 co-localize in atherosclerotic lesions of ApoE−/− mouse aortae. Immunoblotting and immunohistochemistry revealed Tyr nitration in PGHS-1 in aortic lesions but markedly less in adjacent nonlesion tissue. PGHS-2 was also found in lesions, but 3-nitrotyrosine incorporation was not detected. 3-Nitrotyrosine formation in proteins is considered a hallmark reaction of peroxynitrite, which can form via NO-superoxide reactions in an inflammatory setting. That iNOS-derived NO is essential for 3-nitrotyrosine modification of PGHS-1 was confirmed by the absence of 3-nitrotyrosine in lesions from ApoE−/−iNOS−/− mice. Mass spectrometric studies specifically identified the active site residue Tyr385 as a 3-nitrotyrosine modification site in purified PGHS-1 exposed to peroxynitrite. PGHS-mediated eicosanoid (PGE2) synthesis was more than fivefold accelerated in cultured iNOS−/− versus iNOS-expressing mouse aortic smooth muscle cells, suggesting that iNOS-derived NO markedly suppresses PGHS activity in vascular cells. These results further suggest a regulatory role of iNOS in eicosanoid biosynthesis in human atherosclerotic lesions.