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Originally published In Press as doi:10.1074/jbc.M201143200 on March 18, 2002

J. Biol. Chem., Vol. 277, Issue 21, 18670-18676, May 24, 2002
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Eicosanoid Regulation of Vascular Endothelial Growth Factor Expression and Angiogenesis in Microvessel Endothelial Cells*

Alexander MezentsevDagger , Francesca SetaDagger , Michael W. DunnDagger , Naoya Ono§, John R. Falck§, and Michal Laniado-SchwartzmanDagger ||

From the Dagger  Department of Pharmacology, New York Medical College, Valhalla, New York 10595 and Departments of § Biochemistry and  Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas 75235

12(R)-Hydroxy-5,8,14-eicosatrienoic acid (HETrE) is a potent inflammatory and angiogenic eicosanoid in ocular and dermal tissues. Previous studies suggested that 12(R)-HETrE activates microvessel endothelial cells via a high affinity binding site; however, the cellular mechanisms underlying 12(R)-HETrE angiogenic activity are unexplored. Because the synthesis of 12(R)-HETrE is induced in response to hypoxic injury, we examined its interactions with vascular endothelial growth factor (VEGF) in rabbit limbal microvessel endothelial cells. Addition of 12(R)-HETrE (0.1 nM) to the cells increased VEGF mRNA levels with maximum 5-fold increase at 45 min. The increase in VEGF mRNA was followed by an increase in immunoreactive VEGF protein. 12(R)-HETrE (0.1 nM) rapidly activated the extracellular signal-regulated kinases (ERKs) ERK1 and ERK2. Moreover, preincubation of cells with PD98059, a selective inhibitor of MEK-1, inhibited 12(R)-HETrE-induced VEGF mRNA. Addition of VEGF antibody to cells grown in Matrigel-coated culture plates inhibited 12(R)-HETrE-induced capillary tube-like formation, suggesting that VEGF mediates, at least in part, the angiogenic response to 12(R)-HETrE. The results indicate that in microvessel endothelial cells, 12(R)-HETrE induces VEGF expression via activation of ERK1/2 and that VEGF mediates, at least in part, the angiogenic activity of 12(R)-HETrE. Given the fact that both VEGF and 12(R)-HETrE are produced in the cornea after hypoxic injury, their interaction may be an important determinant in the development of neovascularized tissues.

* This work was supported by Grants EY06513 and GM31278 from the National Institutes of Health and by the Robert A. Welch Foundation.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

|| To whom correspondence should be addressed: Dept. of Pharmacology, New York Medical College, Basic Science Bldg. 530, Valhalla, NY 10595. Tel.: 914-594-4153; Fax: 914-594-4303; E-mail: michal_schwartzman@nymc.edu.

Copyright © 2002 by The American Society for Biochemistry and Molecular Biology, Inc.
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