Synthesis of 4- and 5-series leukotrienes in the lung microvasculature challenged with Escherichia coli hemolysin: critical dependence on exogenous free fatty acid supply.

Escherichia coli hemolysin (HlyA) has been identified as a potent inductor of phosphoinositide hydrolysis and related metabolic responses in neutrophils (Grimminger and colleagues, 1991, J. Clin. Invest. 88:1531-1539). In isolated perfused rabbit lungs, which harbor a large number of entrapped microvascular leukocytes, we investigated the effect of a low dose of HlyA on lipoxygenase product formation in the presence of exogenous free arachidonic acid (AA), eicosapentaenoic acid (EPA), or both precursor fatty acids. Leukotrienes (LT) and hydroxyeicosatetra(penta)enoic acids (HET[P]E) in the recirculating perfusate were quantified using high-performance liquid chromatography techniques. In the absence of exogenous precursor fatty acid supply, 0.02 hemolytic units/ml HlyA elicited only minor amounts of LTs and 5-HETE. AA, 10 microM, provoked the generation of limited quantities of LTB4, LTE4, and 5-HETE. Combined application of HlyA and AA caused a manifold amplification of 4-series LT and 5-HETE generation, with predominance of cysteinyl-LTs. EPA, 10 microM, elicited the synthesis of 5-series LTs accompanied by marked quantities of 5-HEPE. Dual stimulation with HlyA and EPA provoked exclusive generation of excessive quantities of all 5-series 5-lipoxygenase products. When HlyA was administered in the presence of both AA (10 microM) and EPA (10 microM), the n-3 fatty acid clearly turned out to be the preferred substrate, with ratios of the various 5-series to 4-series products ranging between 1.8 and 14.5. Moreover, the absolute quantities of AA-derived metabolites and the total sum of all 5-lipoxygenase products was markedly reduced under these conditions. We conclude that the HlyA-evoked 5-lipoxygenase product formation in the pulmonary vasculature of the rabbit is critically dependent on the presence of free precursor fatty acids. The profile of LTs suggests neutrophil (PMN)-related transcellular eicosanoid synthesis as a major underlying metabolic pathway. EPA represents the preferred substrate as compared with AA, resulting in a marked suppression of AA metabolite formation. Therapeutic attempts to provide n-3 fatty acids via the intravenous route may have a major impact on lipid mediator profiles in PMN-related inflammatory events.