Upon stimulation by various ligands, freshly isolated human peripheral neutrophils (PMN) respond in a variety of ways, such as Superoxide (O-2) generation, phagocytosis, enzyme release, migration etc. Chemotactic peptide formylmethionyl-leucyl-phenylalanine (FMLP) and opsonized zymo- san activate neutrophils by a receptor-mediated mechanism, while phorbol myristate acetate and dioctanoylglycerol activate the cells by a mechanism involving Ca2+- and phospholipid-dependent protein kinase (PKC). Receptor-mediated but not PKC-mediated O-2 generation in PMN was enhanced by the priming of recombinant human granulocyte colony stimulating factor (G-CSF). FMLP-dependent luminol chemiluminescence was also enhanced by G-CSF. However, no appreciable enhancement was observed in FMLP-induced intracellular calcium ion concentration ([Ca2+]i). Enhancement of FMLP- induced generation of O-2 by G-CSF was inhibited by genistein or α-cyano-3-ethoxy-4-hydroxy-5- phenylthiomethylcinnamamide (ST 638), inhibitors of tyrosine kinase (TK), and was stimulated by staurosporine and 1-(5-isoquinolinesulfonyl)-3-methyl-piperazine (H-7), inhibitors of PKC. The ED50 values of genistein and ST 638 for the inhibition of the FMLP-induced O-2 generation from G-CSF were 0.5 and 5 muM, respectively. In contrast, O-2 generation by PKC activation without G-CSF priming was inhibited by stauroporine and H-7, but was stimulated by genisten and ST 638. These results suggested that the enhancing effect of G-CSF on receptor-mediated generation of the O-2 might be regulated by protein kinases, such as TK and PKC and that the TK inhibitor selectively inhibited the G-CSF-primed receptor-mediated O-2 generation of neutrophils.
ASJC Scopus subject areas