Arachidonic acid is metabolized by mammalian cells through the """"""""Arachidonate Cascade"""""""" to a variety of products with important biological activities and which have been implicated in the pathophysiology of many disease states. Our group recently described a role for the microsomal cytochrome P-450 system as a new member of the """"""""arachidonate Cascade."""""""" We have provided evidence indicating the invivo formation of some of the novel metabolites produced by this enzyme system and shown that the epoxyeicosatrienoic acid derivatives are potent in vitro mediators for the release of several peptide hormones such as for example insulin and glucagon from isolated pancreatic islets; somatostatin from median eminence and luteinizing hormone from anterior pituitary cells. By the use of stable- and radioisotopes and established extraction, HPLC and GC/MS procedures we propose to 1) study the in vivo formation of epoxy- and dihydroxyeicosatrienoic acid drivatives; 2) complete studies of the NADPH dependent, liver microsomal metabolism of arachidonic acid; 3) study the NADPH dependent and independent secondary metabolism of selected metabolites; 4) study the primary and secondary metabolism of arachidonic acid by purified components of the microsomal monooxygenase coupled to epoxide hydrolases and GSH S-transferases; 5) utilize isolated hepatocytes as an integrated model for the metabolism of arachidonic acid. During the course of these studies, novel metabolites will be submitted for biological testing and if active, their synthesis attempted. It is expected that the knowledge gained from these studies will lead to a better understanding of the basic biochemistry of this novel pathway in liver cells and will allow us to make useful predictions and pose relevant questions with regard to the nature and functions of the products formed by this system in extrahepatic tissues, where the concentrations of the enzymes involved are far lower. The latter will allow is to establish the generality of the pathway and to delineate its possible role in the control of organ and cell homeostasis. This work should help to establish a possible role of cytochrome P-450 in the metabolism of endogenous unsaturated fatty acids and in the formation of eicosanoids with biological activity.
