INHIBITION BY MANOALIDE OF FMLP-STIMULATED ELASTASE RELEASE FROM HUMAN NEUTROPHILS

Abstract Incubation of human polymorphonuclear leukocytes (PMNLs) with the chemotactic factor N -formyl- l -methionyl- l -leucyl- l -phenylalanine (fMLP) resulted in a concentration-dependent release of the neutral protease elastase. This response was inhibited by pretreatment of the PMNLs with manoalide ( ic 50 approximately 0.08 μM). To understand the mechanism of this inhibition, we examined the effect of manoalide on the signal-transduction pathway believed to mediate fMLP stimulation. We observed in fura-2 loaded cells that pretreatment with manoalide blocked fMLP-induced increases in cytosolic free-calcium ( ic 50 approximately 0.15 μM). However, manoalide had no effect on inositol 1,4,5-trisphosphate (IP 3 ) production at concentrations which completely inhibited the Ca 2+ signal. Furthermore, manoalide was approximately 50-fold less potent as an inhibitor of phospholipase C activity in membrane preparations of PMNLs than as an inhibitor of calcium mobilization in whole cells. These data indicate that manoalide can block stimulation of human PMNLs through inhibition of Ca 2+ mobilization, but that this occurs at a site beyond phospholipase C activation and inositol phosphate turnover.