Arachidonate 12-Lipoxygenase and Cyclooxygenase:PGE Isomerase are Predominant Pathways for Oxygenation in Bovine Tracheal Epithelial Cells

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Abstract
The capacity of bovine tracheal epithelial cells to convert arachidonic acid to oxygenation products with potential biologic activity was studied in homogeneous preparations of isolated cells. Purified epithelial cell suspensions were incubated with radiolabeled arachidonic acid, and oxygenated metabolites were identified using high-pressure liquid chromatography and gas chromatography—mass spectrometry. The cells released predominantly two products during incubation with 0.3 to 150 µM arachidonic acid for 1 to 60 min at 37° C: prostaglandin E2 (PGE2) and 12-hydroxyeicosatetraenoic acid (12-HETE). Concentration-response curves for the two products yielded half-maximal effects at 2 and 45 µM arachidonic acid, respectively. Stereochemical analysis by chiral-phase high-pressure liquid chromatography demonstrated that the epithelial 12-HETE consisted exclusively of the 12(S) isomer, providing supporting evidence that it was derived from an arachidonate 12-lipoxygenase. Epithelial cells prelabeled with arachidonic acid and incubated with 5 µM A23187 to stimulate endogenous arachidonic acid metabolism also released two predominant products with the chromatographic properties of PGE2 and 12-HETE. The findings demonstrate that bovine tracheal epithelial cells express both a cyclooxygenase:PGE isomerase and a 12-lipoxygenase pathway and therefore implicate this pathway as a new source of epithelial cell mediators.

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