IDENTIFICATION OF MOUSE CYP2C INVOLVED IN ARACHIDONIC ACID

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AIMS: Little is known about the endogenous function of the CYP2Cs. We postulate that these enzymes have important roles in the metabolism of physiologically important endogenous substrates such as arachidonic acid and retinoic acid (Vitamin A). Our goal has been to identify all CYP2Cs in the mouse, determine their contribution to the production of particular eicosanoid metabolites and to identify their physiological role in these tissues. The P450-derived AA metabolites are known to possess potent biological actions in numerous tissues. These effects depend on the regio- and stereochemistry of the products. We are attempting to determine the organ and cell-specific localization of these isoforms. A long-range goal is to produce knockout mice to study the physiological importance of this class of enzymes, and to overexpress these enzymes in cell lines to further establish their function. ACCOMPLISHMENTS We have identified five (potentially six) CYP2Cs in the mouse. We expressed five of these in cDNA expression systems examined their regio and stereospecificity in the formation of epoxyeicosatrienoic acids (EETs) and hydroxyeicosatetraenoic acids (HETES). They were found to produce very specific regiospecific and stereospecific products. CYP2C29 is the predominant hepatic form and produces 14,15-EET. Western blotting and RT-PCR indicted that lung contained relatively large amounts of CYP2C29. CYP2C40 was found in high amounts in colon, cecum, gut, and was also found in kidney and heart. CYP2C40 was found to metabolize arachidonic acid (AA) to 16R- HETE (66%) and 16S-HETE (34%). This is the first P450 found to produce this HETE as a primary product. 16R-HETE is thought to be important in processes such as renal vasodilation and inhibiting neutrophil aggregation and adhesion. Kidney contained CYP2C40 and a new unidentified CYP2C isoform. Endothelial cells of arteries contained CYP2C29 and some CYP2C40. - endogenous compound, arachidonic acid, retinoic acid, Vitamin A, epoxyeicosatrienoic acid