Abstract T P83: The CYP Epoxygenase Metabolite of Arachidonic Acid 14,15- Epoxyeicosatrienoic Acid: a Novel Therapeutic Target in Neuroprotection

Background: Neuronal cells rely on their metabolic coupling with astrocytes, which release factors that affect phenotype of brain cell types. Both astrocytes and neuronal cells express the CYP epoxygenases that metabolize arachidonic acid to four regioisomeric epoxyeicosatrienoic acids (EETs), including 14,15-EET. Circadian rhythms are evident in daily variations of cardiovascular function and in stroke incidence. Ischemic stroke is associated with increased production of H2O2 that causes cellular oxidation and neuronal death. The purpose of this study is to investigate neuroprotective effects of 14,15-EET against oxidative damage. Methods: We investigated the circadian pattern of production and neuroprotective effects of 14,15-EET against H2O2-induced oxidative damage in co-cultures of astrocytes and neurons. Results: We found significant 24h rhythms in 14,15 EET production in astrocytes. Stimulation of neuronal cells with H2O2 (0.1 and 1 mM, 1h) elicited lower cell viability compared to vehicle treated group. Co-culturing of neuronal cells with astrocytes prevented neuronal death induced by H2O2 (0.1 and 1 mM, 1h) application. Pretreating neuronal cells with 14,15-EET (0.1-30 μM, 30 min) before stimulation with H2O2 increased neuronal cell viability. Preincubation of the co-cultured cells with the soluble epoxide hydrolase (sEH) inhibitor 12-(3-adamantan-1-yl-ureido)-dodecanoic acid (AUDA) (8-800 nM, 1h) increased endogenous 14,15-EET in the media and protected against H2O2 (1 mM, 1h) induced damage. In contrast inhibition of endogenous EETs production by miconazole (1-20 μM, 1h) before stimulation of cells with H2O2 (1 mM, 1h) elicited decreased cell viability and increased neuronal death. Treatment with 14,15-EET increased formation of the prosurvival phospho-Akt in co-cultures of astrocytes and neurons. Conclusions: These findings indicate that 14,15-EET is released from astrocytes in a circadian manner and is neuroprotective against oxidant damage that could prevail in incidences of ischemic stroke. The findings further suggest that activation of phospho-Akt could be the underlying mechanism for the neuroprotective effects of 14,15-EET, and a novel therapeutic target for stroke related neuronal disorders.