Mitogen-activated protein kinase in neutrophils and enucleate neutrophil cytoplasts - Evidence for regulation of cell-cell adhesion

We employed neutrophils and enucleate neutrophil cytoplasts to study the activation of the mitogen-activated protein kinases (MAPKs) p44(erk1) and p42(erk2) in neutrophils by inflammatory agonists that engage G protein-linked receptors. Formyl-methionyl-leucyl-phenylalanine (FMLP) rapidly and transiently activated MAPK in neutrophils and cytoplasts, consistent with a role in signaling for neutrophil functions. FMLP stimulated p21(ras) activation in neutrophils and Raf-1 translocation from cytosol to plasma membrane in cytoplasts, with kinetics consistent with events upstream of MAPK activation. Insulin, a protein tyrosine kinase receptor (PTKR) agonist, stimulated neutrophil MAPK activation, demonstrating an intact system of PTKR signaling in these post-mitotic cells. FMLP- and insulin-stimulated MAPK activation in cytoplasts were inhibited by Bt(2)cAMP, consistent with signaling through Raf-1 and suggesting a mechanism for cAMP inhibition of neutrophil function. However, Bt(2)cAMP had no effect on FMLP-stimulated MAPK activation in neutrophils. The extent of MAPK activation by various chemoattractants correlated with their capacity to stimulate neutrophil and cytoplast homotypic aggregation. Consistent with its effects on MAPK, Bt(2)cAMP inhibited FMLP-stimulated aggregation in cytoplasts but not neutrophils. Insulin had no independent effect but primed neutrophils for aggregation in response to FMLP. Our studies support a p21(ras)-, Raf-1-dependent pathway for MAPK activation in neutrophils and suggest that neutrophil adhesion may be regulated, in part, by MAPK.