Ligand-operated synthesis of 4-series and 5-series leukotrienes in human neutrophils: critical dependence on exogenous free fatty acid supply.
The influence of exogenously supplied free arachidonic acid (AA) and eicosapentaenoic acid (EPA) on the 5-lipoxygenase metabolism in human neutrophils (PMN) was investigated. Simultaneous application of A23187 with incremental concentrations of free AA caused a dose-dependent augmentation of the ionophore-elicited eicosanoid generation [release of leukotriene B4 and its omega-oxidation products, nonenzymatic hydrolysis products of leukotriene A4, and 5-hydroxyeicosatetraeneoic acid (5-HETE)]. A23187 challenge in the presence of free EPA resulted in the dose-dependent appearance of corresponding n - 3-derived metabolites, parallelled by a decrease in 4-series leukotrienes and 5-HETE. The inflammatory ligands formyl-methionyl-leucyl-phenylalanine and platelet-activating factor evoked no substantial eicosanoid generation in the absence of exogenously supplied polyunsaturated fatty acids (PUFAs). Addition of free AA or EPA in parallel with the ligand challenge evoked exclusive and dose-dependent generation of the respective leukotrienes and 5-HETE or 5-hydroxyeicosapentaenoic acid. Total amounts of 5-lipoxygenase products elicited under these conditions approached those in ionophore-stimulated PMN, with platelet-activating factor challenge surpassing the formyl-methionyl-leucylphenylalanine-evoked effect by approximately 50%. Two thirds of the maximum effect was obtained in the presence of only 10 microM free PUFA. Use of labeled fatty acids suggested exclusive origin of the eicosanoids from the exogenously provided precursor PUFA. Critical dependence on timing was noted; maximum response occurred upon simultaneous application of PUFA and ligand, and only 5 min of delay between AA or EPA addition and ligand challenge sufficed to reduce the formation of respective metabolites to less than 20%. EPA competed with AA and was noted to be the preferred substrate for ligand-evoked eicosanoid synthesis. In contrast to the simultaneous addition of free PUFAs, preloading of PMN with AA or EPA for 60 min revealed only very moderate or even no influence on ionophore- or ligand-evoked eicosanoid synthesis. We conclude that inflammatory ligands induce marked stimulation of PMN eicosanoid synthesis, with critical dependence on the presence of free precursor PUFAs. Preference of EPA over AA is observed under these conditions.
Grimminger
F
F
Department of Internal Medicine, Justus Liebig University, Giessen, FRG.
Dürr
U
U
Seeger
W
W
D000001:Calcimycin / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D005230:Fatty Acids, Nonesterified / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D006893:Hydroxyeicosatetraenoic Acids / Q000378:metabolism; D066298:In Vitro Techniques; D015289:Leukotrienes / Q000096:biosynthesis* / Q000737:chemistry; D008024:Ligands; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D010084:Oxidation-Reduction; D010972:Platelet Activating Factor / Q000494:pharmacology
D005230:Fatty Acids, Nonesterified; D006893:Hydroxyeicosatetraenoic Acids; D015289:Leukotrienes; D008024:Ligands; D010972:Platelet Activating Factor; D000001:Calcimycin; C022022:5-hydroxy-6,8,11,14-eicosatetraenoic acid; D009240:N-Formylmethionine Leucyl-Phenylalanine