15(S)-hydroxyeicosatetraenoic acid-induced angiogenesis requires Src-mediated Egr-1-dependent rapid induction of FGF-2 expression

To understand the mechanisms underlying 15(S)-hydroxyeicosatetraenoic acid [15(S)-HETE]-induced angiogenesis, we studied the role of Egr-1. 15(S)-HETE induced Egr-1 expression in a time-dependent manner in human dermal microvascular endothelial cells (HDMVECs). Blockade of Egr-1 via forced expression of its dominant-negative mutant attenuated 15(S)-HETE -induced HDMVEC migration and tube formation as well as Matrigel plug angiogenesis. 15(S)-HETE induced Egr-1 expression requires Src activation. In addition, adenovirus-mediated expression of dominant-negative mutant of Src blocked 15(S)-HETE's effects on migration and tube formation of HDMVECs and Matrigel plug angiogenesis. 15(S)-HETE induced fibroblast growth factor-2 (FGF-2) expression rapidly via Src-mediated production of Egr-1. Cloning and mutational analysis of FGF-2 promoter revealed that Egr-1 binding site proximal to transcription start site is required for 15(S)-HETE induced FGF-2 expression. Neutralizing antibody-mediated suppression of FGF-2 function also attenuated the effects of 15(S)-HETE on HDMVEC migration and tube formation as well as Matrigel plug angiogenesis. Furthermore, in contrast to wild-type mice, 12/15-LOX-/- mice exhibited decreased Matrigel plug angiogenesis in response to AA, which was rescued by 15(S)-HETE. On the basis of these observations, we conclude that 15(S)-HETE-induced angiogenesis requires Src-mediated Egr-1-dependent rapid induction of FGF-2. These findings may suggest that 15(S)-HETE could be a potential endogenous regulator of pathologic angiogenesis associated with atherosclerosis and restenosis. (Blood. 2010; 115: 2105-2116)