The regulation of neutrophil phospholipase A2 by granulocyte-macrophage colony-stimulating factor and its role in priming superoxide production.
Experiments were performed to investigate the relative role of phospholipase A2 (PLA2) in the activation and cytokine-mediated priming of neutrophil superoxide production. PLA2 activity was measured with a radiometric assay which discriminates between PLA2 and the downstream enzyme, 5-lipoxygenase. In cells that had not been primed by prior incubation with granulocyte-macrophage colony stimulating factor (GM-CSF), PLA2 and NADPH oxidase were differentially stimulated by the chemotactic peptide N-formyl-met-leu-phe (FMLP), calcium ionophore, or phorbol ester. In addition, inhibition of PLA2 by mepacrine (0-100 micromol/l) did not concomitantly inhibit FMLP-stimulated superoxide production. These findings suggest that the activity of PLA2 and NADPH oxidase may be uncoupled in the unprimed cell. In cells preincubated with GM-CSF, time- and dose-dependent priming of FMLP-stimulated PLA2 responses were observed and inhibition of PLA2 by mepacrine was accompanied by the inhibition of FMLP-stimulated superoxide production down to the level of unprimed cells. The effect of mepacrine was not due to inhibition of FMLP receptor expression. These data suggest that a mepacrine-sensitive PLA2 may have a role in the GM-CSF mediated priming of superoxide production. Using ionophore-stimulated PLA2 activity as a model, we showed that Bordatella pertussis toxin did not inhibit GM-CSF mediated priming, demonstrating that a pertussis-sensitive GTP-binding protein does not mediate signal transduction from the GM-CSF receptor to PLA2. The tyrosin kinase inhibitor, genestein, selectively inhibited GM-CSF primed but not unprimed PLA2 activity, demonstrating that GM-CSF-mediated priming requires tyrosine kinase activity.
Roberts
P J
PJ
Department of Haematology, University College London Medical School, London.
Williams
S L
SL
Linch
D C
DC
D016718:Arachidonic Acid / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology*; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism; D019255:NADPH Oxidases; D009504:Neutrophils / Q000201:enzymology*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D011796:Quinacrine / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism*
D013481:Superoxides; D016718:Arachidonic Acid; D000001:Calcimycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D010741:Phospholipases A; D054467:Phospholipases A2; D011796:Quinacrine