Dynamics of the diradylglycerol responses of stimulated phagocytes

Abstract

The generation of diradylglycerols (sn-1,2 diacylglycerols (DAG) and 1-O-alkyl-2-acylglycerols (AAG] was investigated in human polymorphonuclear leukocytes stimulated with fMet-Leu-Phe, phorbol myristate acetate (PMA), or A23187. With each stimulus, the elevations in the mass of DAG clearly preceded that of AAG. The levels of both lipids increased over time, peaked by 15-20 min (fMet-Leu-Phe) or 45-60 min (PMA or A23187) and returned slowly toward base line thereafter. The base-line levels of DAG were some 4-fold higher than levels of AAG. On stimulation, the relative increases in AAG (approximately 4-fold, fMet-Leu-Phe; approximately 20-fold, PMA and A23187) were much greater than the corresponding relative increases in the levels of DAG (approximately 2-fold fMet-Leu-Phe; approximately 5-fold, PMA and A23187). The diradylglycerol responses were dependent upon agonist concentration. Prior treatment with cytochalasin B augmented the fMet-Leu-Phe diradylglycerol responses but did not alter unstimulated or PMA- or A23187-stimulated diradylglycerol responses. Depletion of extracellular Ca2+ blocked responses to fMet-Leu-Phe, but not to PMA. Treatment with pertussis toxin: (a) completely blocked the responses to fMet-Leu-Phe, (b) slightly suppressed the AAG but not the DAG response to PMA, and (c) did not affect the responses to A23187. Gas chromatographic/mass spectral analyses indicated that the AAG generated during cell activation consists of a mixture of species differentiated by 1-O-alkyl chains of 16:0, 18:0, 18:1 and an additional species that remains uncharacterized. Since DAG and AAG are reportedly activators and inhibitors, respectively, of protein kinase C activities, the sequential generation of these lipid messengers may provide for a system to critically control the activation of protein kinase C.