Endothelial cytochrome P450 (CYP) 2C epoxygenases regulate vascular tone and homeostasis by the generation of epoxyeicosatrienoic acid (EET). EETs have been implicated in the regulation of endothelial cell proliferation and angiogenesis. As the expression of CYP 2C is regulated by different stimuli, including hypoxia, we investigated whether EETs play a role in hypoxia-induced angiogenesis. In human endothelial cells, hypoxia (1% O2) enhanced the activity of the CYP 2C9 promoter and increased the expression of CYP 2C mRNA and protein. Additionally, elevated levels of 11,12-EET and 11,12-DHET were detected. These responses were even more pronounced in bovine retina endothelial cells. The CYP inhibitor MS-PPOH and the EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (EEZE) as well as CYP 2C antisense oligonucleotide treatment prevented hypoxia-induced cell migration and tube formation in vitro. In vivo EEZE inhibited the hypoxia-induced angiogenesis in the chick chorioallantoic membrane assay. These data indicate that CYP 2C expression and EET formation are increased by hypoxia and that CYP 2C-derived EETs are implicated in the hypoxia-induced migration and angiogenesis.