Induction of Microglial Anandamide-Metabolizing Cytochrome P450s Leading to the Formation of a Cannabinoid Receptor 2 Agonist with Increased Biostability
CytochromeP450 (P450) enzymes are known to oxidize the endocannabinoid anandamide, thus potentially affecting the endocannabinoid system. Work from this lab has demonstrated previously that the P450s 3A4 and 4F2 oxygenate anandamide to hydroxyeicosatetraenoic acid ethanolamides (HETE-EAs) and epoxyeicosatrienoic acid ethanolamides (EET-EAs).The current study investigated this metabolic pathway and its biological significance. We have shown that one of the metabolites of anandamide formed by these P450s, 5,6-EET -EA, is a potent agonist of the cannabinoid receptor 2 (CB2) and that it inhibited the forskolin-stimulated cyclic adenosine monophosphate (cAMP) accumulation in CHO cells stably expressing the CB2 receptor. In the central nervous system, the CB2 receptor is almost exclusively expressed on microglia, a macrophage-like immune cell. When activated, microglia have an increased expression of the CB2 receptor and this receptor is a possible target for the treatment of neuroinflammation. The mouse-derived BV-2 microglia cells stimulated with low doses of interferon-gamma exhibited an increased capacity for metabolizing anandamide to 5,6-EET-EA, corresponding to increased protein expression of the P450s 4Ft3 and 3Al. We also demonstrated that 5,6-EET-EA has a higher stability than anandamide in mouse brain homogenate. Our results suggest that epoxidation of anandamide by P450s to form 5,6-EET -EA represents an endocannabinoid bioactivation pathway in the context of immune cell function.