Selective incorporation of (15S)-hydroxyeicosatetraenoic acid in phosphatidylinositol of human neutrophils: agonist-induced deacylation and transformation of stored hydroxyeicosanoids.

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RESUMO

The uptake and mobilization of (15S)-hydroxy-5,8,11-cis-13-trans-eicosatetraenoic acid (15-HETE), a major product of arachidonic acid metabolism, was examined with human neutrophils (polymorphonuclear leukocytes; PMNs). Upon exposure to labeled 15-HETE, PMNs rapidly (15 sec to 20 min) incorporated approximately 20% of the label into phosphatidylinositol, while less than 4% was associated with other phospholipid classes and neutral lipids. This pattern was distinct from that of either labeled arachidonate or labeled(5S)-hydroxy-8,11,14-cis-6-trans-eicosatetraenoic acid (5-HETE), which within 20 min were predominantly associated with triglycerides and phosphatidylcholine. After reversed-phase HPLC, greater than 98% of the label in phosphatidylinositol, isolated from PMNs, was released with phospholipase A2. Upon exposure to either chemotactic peptide (FMLP), phorbol 12-myristate 13-acetate, or an ionophore (A23187), 15-HETE-labeled PMNs released 15-HETE from phosphatidylinositol and displayed an impaired ability to generate leukotriene B4 (LTB4), 20-OH-LTB4, and 20-COOH-LTB4. Deacylated [3H]15-HETE was converted to (5S,15S)-dihydroxy-6,13-trans-8,11-cis-eicosatetraenoic acid (5,15-DHETE), lipoxin A4, and lipoxin B4, each carrying 3H label. PMNs labeled with 5-HETE also released and transformed this HETE when stimulated. However, the profile of labeled products differed between PMNs with either esterified 15-HETE or 5-HETE. When activated, 5-HETE-labeled PMNs generated both 5,20-DHETE and 5,15-DHETE but not labeled lipoxins. Threshold aggregation induced by FMLP with 15-HETE-labeled PMNs was inhibited (approximately 2 orders of magnitude), while the threshold response was relatively unimpaired with either A23187 or phorbol 12-myristate 13-acetate-induced aggregation. Results indicate that 15-HETE is rapidly esterified into phosphatidylinositol of PMNs, which can be mobilized and transformed upon exposure of the cells to a second signal. Moreover, they suggest that eicosanoid intermediates other than arachidonic acid can be stored by cells, released via signal transduction, and oxygenated to generate alternative profiles of eicosanoids.

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