EFFECTS OF THE DIACYLGLYCEROL KINASE INHIBITOR R59022 ON SUPEROXIDE ANION GENERATION, PLASMA-MEMBRANE DEPOLARIZATION AND CA2+ FLUXES ACTIVATED BY THE CHEMOTACTIC PEPTIDE FMLP IN HUMAN NEUTROPHILS

Compound R59022, a specific inhibitor of diacylglycerol kinase I and II, has been shown to modulate agonist mediated responses in a number of cell types. In the present report we used compound R59022 as a tool to study the role of diacylglycerol and thus of protein kinase C, in the modulation of three typical responses of human neutrophils to chemotactic peptide stimulation, i.e. O2- production, plasma membrane depolarization and [Ca2+]i increases. Our results show that in the presence of micromolar concentrations of the DG kinase inhibitor, fMLP-dependent phosphatidic acid formation is drastically reduced, while DG formation is increased. Contemporarily, fMLP induced plasma membrane depolarization and superoxide anion generation are potentiated. We also demonstrate that although at high concentrations compound R59022 can cause an increase in the [Ca2+]i, this effect does not account for its stimulation of fMLP activated response. Finally, we show that Ca2+ extrusion is activated by fMLP and that this effect is increased in the presence of R59022. Taken together these results argue in favor of a primary role in neutrophils of phospholipase C generated DG in the modulation of superoxide anion production, plasma membrane potential depolarization and Ca2+ efflux.