FMLP is a potent activator of guinea-pig eosinophils but its activity is dependent on the prior overnight in vitro culture of the cells (facilitation)

Abstract

The activation of guinea-pig eosinophils was studied by measuring the production of superoxide anion (O2-) and the secretion of eosinophil peroxidase (EPO). Phorbol myristate acetate (PMA), calcium ionophore, plasma-activated zymosan, concanavalin A and recombinant human anaphylatoxin C5a induced the release of varying amounts of EPO. Some of these same activators, as well as platelet-activating factor, and aggregated homologous IgG, either by themselves or after a brief priming of the cells with low concentrations of PMA, also caused the formation of O2-. Formyl-methionyl-leucyl-phenylalanine (FMLP) failed to induce either of these reactions in freshly isolated cells. It was found serendipitously, however, that cells which had been maintained in culture overnight secreted EPO upon challenge with FMLP, and, if they were primed with PMA, they also produced O2-. The conversion from unresponsive to responsive cells ('facilitation') depended on the presence of mononuclear cells or mononuclear cell-conditioned medium in the overnight cultures. Although there also was a shift in the density of the majority of the eosinophils after overnight culture to a density lower than 1.085, this shift was not dependent on the inclusion of monocytes or of monocyte-conditioned medium (MCM) in the cultures and thus was not sufficient to impart responsiveness to FMLP. Responses of eosinophils to other activators were qualitatively unchanged after overnight facilitation. Binding studies using radiolabelled FMLP revealed that, during facilitation, binding of FMLP to guinea-pig eosinophils increased about sixfold overall and suggested the expression of a high affinity receptor. This change may explain the basis for the facilitation phenomenon.