Abstract

The effect of exogenous leukotriene B₄ (LTB₄) on opsonized zymosan-stimulated human neutrophil formation of 5-lipoxygenase products and arachidonic acid release was directly assessed using reverse-phase HPLC/tandem mass spectrometric methods for quantitation. Stable isotopically labeled LTB₄, [1,2-¹³C₂]LTB₄, caused a dose-dependent inhibition of LTB₄ production in isolated human neutrophils with significant inhibition (60 ± 7% of control levels) when 0.12 nM [¹³C₂]LTB₄ was present. Production of 5-hydroxy-6,8,11,14-eicosatetraenoic acid and release of free arachidonic acid were also dose-dependently inhibited by exogenous LTB₄. Metabolites of LTB₄, 20-hydroxy-LTB₄ and 3(S)-hydroxy-LTB₄, also significantly reduced LTB₄ production to levels as low as 10 ± 6% and 10 ± 7% of control levels, respectively, when present exogenously at 10 nM. Exogenous 5-hydroxy-6,8,11,14-eicosatetraenoic acid at concentrations as high as 10 nM produced no significant reduction in LTB₄biosynthesis during zymosan-stimulated human neutrophil production of LTB₄. The inhibitory effect of LTB₄ could be partially reversed by the LTB₄ receptor antagonist U 75302. Furthermore, an alternative stimulus, N-formyl-methionyl-leucyl-phenylalanine (100 nM), did not inhibit the production of LTB₄ in opsonized zymosan-stimulated human neutrophils. These results suggest that activation of the LTB₄ receptor on the human neutrophil during phagocytosis limits the ultimate biosynthesis of LTB₄. This autocrine effect is opposite to that observed when neutrophils have much of the signal transduction pathways bypassed when stimulated with calcium ionophore A23187 or treated with exogenous free arachidonic acid.