Leukotriene formation by eosinophil leukocytes. Analysis with ion-pair high pressure liquid chromatography and effect of the respiratory burst.

Abstract

In this paper, we describe ion-pair reverse-phase high pressure liquid chromatography, a novel, high-resolution method for the separation of leukotrienes. Using this technique, we have studied the production and release of leukotrienes from purified horse eosinophil leukocytes 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 an LTC 4/LTD 4 spectra with absorption maxima at 278/281 nm. The other metabolites were virtually devoid of slow-reacting substance activity and exhibited LTB 4 spectra with maxima at 269/271 nm. Since ionophore A23187, under our experimental conditions, induced a respiratory burst in the eosinophils, two modifies of this response, 2-deoxyglucose, an inhibitor of glucose metabolism and catalase, a scavenger of hydrogen peroxide, 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 LTB 4 and its stereoisomers and correspondingly enhanced the formation of LTC 4 and LTD 4. On the other hand, the increase of glucose in the medium augmented the production of B4-type leukotrienes while decreasing that of LTC 4 and LTD 4. These results indicate that the respiratory burst is involved in leukotriene synthesis by eosinophil leukocytes.