Biological role of EET in induction of vascular endotberial growth factor underr hypoxia

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Cytochrome P450 (P450) catalyze the NADPH-dependent oxidation of arachidonic acid to several unique eicosanoids such as epoxyeicosatrienoic acid (EET) which has four isomers, 5,6-EET, 8,9-EET, 11,12-EET, and 14,15-EET. A particular interest in the epoxygenase reaction has developed because of the potent biological activities (modulation of vascular tone and anti-inflammatory activity etc.) attributed to the EETs. I focused on a new biological function of EETs which induce vascular endotherial growth factor (VEGF) under hypoxia. In this study, human hepatoma cell, Hep3B and human umbilical artery endotherial cell (HUAEC) were used under normoxic and hypoxic conditions. An inhibitor for phospholipase A2 which supplies arachidonic acid from plasma membrane, MAFP, inhibited the VEGF induction in HUAEC under hypoxia. NADPH-P450 reductase (NPR) supplies electrons to P450 and is necessary for P450 to oxidize arachidonic acid. An inhibitor for NPR, DPIC, inhibited the VEGF induction. These results suggest that arachidonic acid and P450 are important for the VEGF induction in HUAEC under hypoxia. The same effects on Hep3B cells were also observed. In Hep3B cells, inhibitors for P450s, sulfaphenazol (an inhibitor for CYP2C8 and CYP2C9) and ketoconazole (an inhibitor for CYP3A) inhibited hypoxia response (erythropoietin induction) efficiently. Addition of 11,12-EET or 14,15-EET induced VEGF in HUAEC under hypoxia but hydration products, 11,12-DHETE and 14,15-DHETE did not induce VEGF. Hypoxia-inducible factor (HIF-1) is necessary for induction of VEGF under hypoxia. 11,12-EET and 14,15-EET did not change the levels of HIF-1. In conclusion, EETs produced by P450s play an important role in hypoxic response of cells. EETs induced VEGF independent on HIF-1 which is an important factor of hypoxic response.