Metabolism of arachidonic acid by rat adrenal glomerulosa cells: Synthesis of hydroxyeicosatetraenoic acids and epoxyeicosatrienoic acids

Metabolites of arachidonic acid have been implicated in the regulation of aldosterone release. To form a basis for further investigations in this area, the present study has isolated and identified the metabolites formed from exogenous arachidonic acid by adrenal zona glomerulosa cells and characterized the effects of several inhibitors on the synthesis of these eicosanoids. Rat adrenal glomerulosa cells metabolized exogenous [¹⁴C]arachidonic acid to products comigrating with the prostaglandins (PGs), hydroxyeicosatetraenoic acids (HETEs) and epoxyeicosatrienoic acids (EETs). The metabolites were found in the cells and the incubation media; however, none of the metabolites were found esterified to cellular lipids. The major metabolites were identified as 6-keto PGF_1α, PGE₂, PGF_2α PGD₂, 12(S)-HETE, 15(S)-HETE, 14, 15-EET, 11, 12-EET, 8, 9-EET, and 5, 6-EET. The identities of the HETEs and EETs were confirmed by gas chromatography/mass spectrometry. There was no evidence for the synthesis of leukotrienes. The cyclooxygenase inhibitor, indomethacin, the lipoxygenase inhibitors, nordihydroguaiaretic acid, baicalein and AA861, and the combined cyclooxygenase/lipoxygenase inhibitors, BW755C and eicosatetrayenoic acid, inhibited the formation of the [¹⁴C]PGs, the [¹⁴C] HETEs, and the [¹⁴C]EETs. Metyrapone and clotrimazole, inhibitors of cytochrome P₄₅₀, increased the synthesis of [¹⁴C]PGs and [¹⁴C]HETEs and reduced the synthesis of [¹⁴C] EETs. Superoxide dismutase did not alter arachidonic acid metabolism. In contrast, arachidonic acid metabolism was increased in cells pretreated with catalase. These data indicate that adrenal glomerulosa cells metabolize exogenous arachidonic acid to a number of oxygenated metabolites including PGs, HETEs, and EETs. From studies with inhibitors, the EETs appear to be synthesized by a cytochrome P₄₅₀epoxygenase and the HETEs by lipoxygenases.