Leukotriene Formation by Eosinophil Leukocytes. Analysis with Ion-Pair High Pressure Liquid Chromatography and Effect of the Respiratory Burst

Abstract

Ion-pair reverse-phase high pressure liquid chromatography, a novel, high-resolution method for the separation of leukotrienes was described. Using this technique, the production and release of leukotrienes from purified horse eosinophil leukocytes was studied following stimulation with the ionophore A23187. At least 11 different metabolites with spectroscopical characteristics of leukotrienes were resolved. Four of them exhibited the biological activity of the slow-reacting substance and LTC4/LTD4 spectra with absorption maxima at 278/281 nm. The other metabolites were virtually devoid of slow-reacting substance activity and exhibited LTB4 spectra with maxima at 269/271 nm. Since ionophore A23187, under experimental conditions, induced a respiratory burst in the eosinophils, 2 modifiers of this response, 2-deoxyglucose, an inhibitor of glucose metabolism and catalase, a scavenger of H2O2, were used to investigate the influence of the burst on leukotriene generation. An overall inhibition of leukotriene release was induced by 2-deoxyglucose. Catalase strongly decreased the formation of LTB4 and its stereoisomers and correspondingly enhanced the formation of LTC4 and LTD4. The increase of glucose in the medium augmented the production of B4-type leukotrienes while decreasing that of LTC4 and LTD4. The respiratory burst apparently is involved in leukotriene synthesis by eosinophil leukocytes.