P131	Cytochrome P450 epoxygenases 2C8/9 are implicated in hypoxia-induced endothelial cell migration and angiogenesis.
1R.Michaelis, 1B.Fisslthaler, 1E.Barbosa-Sicard, 2J.R.Falck, 1I.Fleming, 1R.Busse
1Institut für Kardiovaskuläre Physiologie, J.W.Goethe Universität, Frankfurt/Main, DE; 2Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, US.

Endothelial cytochrome P450 (CYP) epoxygenases of the 2C family regulate vascular tone and homeostasis by the generation of the different regio-isomers of epoxyeicosatrienoic acid (EET) from arachidonic acid. EETs modulate the activity of protein kinases and phosphatases and have been implicated in the regulation of endothelial cell proliferation and angiogenesis. As the expression of CYP 2C is regulated by several pharmacological and hemodynamic stimuli, including hypoxia, we investigated whether CYP 2C-derived EETs play a role in hypoxia-induced angiogenesis. In human endothelial cells, hypoxia (1% O2) enhanced the activity of the CYP 2C9 promoter, increased the expression of CYP 2C mRNA within four hours and protein expression from eight to 24 hours. As a consequence, elevated levels of 11,12-EET and 11,12-DHET were also detected. These responses were even more pronounced in bovine retina endothelial cells. Pre-exposure of human endothelial cells to hypoxia to increase CYP 2C expression, accelerated the ability of the cells to migrate in a transwell chamber as well as to invade Matrigel. These effects were not observed in cells pre-treated with CYP 2C antisense oligonucleotides or cells incubated in the presence of the CYP inhibitor MS-PPOH or the EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (14,15-EEZE). Furthermore, 14,15-EEZE attenuated tube formation by human umbilical vein endothelial cells pre-exposed to hypoxia by approximately 50 % whereas it did not affect tube formation under normoxic conditions. In contrast, in retina endothelial cells, which express appreciable amounts of CYP 2C even under normoxic conditions, EEZE attenuated tube formation by cells cultured under normoxic conditions as well as hypoxia-induced tube formation. 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.

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