Neutrophil-activating polypeptides IL-8 and NAP-2 induce identical signal transduction pathways in the regulation of lysosomal enzyme release.
Interleukin-8 (IL-8/NAP-1), the neutrophil-activating peptide 2 (NAP-2), and formyl-peptides (fMLP) have been described as potent stimulators for human neutrophils (PMN). We have compared the mechanism of signal transduction induced by these factors during neutrophil activation (elastase release), by using activators and inhibitors and by direct measurement of the enzymatic activity of kinases. Moreover, costimulation kinetics of the combined factors were analyzed. Our results show that each of these stimulators induces elevated levels of cAMP, indicating the activation of adenylate-cyclase. Further results obtained with the kinase inhibitor H-7 and the cAMP analogue dibutyryl-cAMP (dbcAMP) gave evidence that cAMP-dependent kinases are involved in the down-regulation of the activation process. Degranulation could not be prevented by the inhibitor W-7, nor did the treatment of cells with calcium ionophore (A23187) lead to elevation of intracellular calcium levels. Both phenomena exclude the participation of calcium calmodulin-dependent kinases. Further results obtained with the novel protein kinase C (PKC) inhibitor BM 41440 as well as by direct measurement of PKC enzyme activity demonstrated the involvement of PKC in fMLP-mediated stimulation but not with IL-8/NAP-1 or NAP-2. Analysis of costimulation experiments conducted with these three factors and TPA confirmed these results and gave evidence that fMLP activates two different signaling pathways, one of which is PKC dependent, while the other is not. Moreover, our data indicate that NAP-2, IL-8/NAP-1, and fMLP use identical PKC-independent transduction mechanisms.
Petersen
F
F
Department of Immunology and Cell Biology, Forschungsinstitut Borstel, Federal Republic of Germany.
Van Damme
J
J
Flad
H D
HD
Brandt
E
E
D000595:Amino Acid Sequence; D002118:Calcium / Q000502:physiology; D002147:Calmodulin / Q000502:physiology; D000242:Cyclic AMP / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects; D016209:Interleukin-8 / Q000494:pharmacology*; D007700:Kinetics; D008247:Lysosomes / Q000187:drug effects* / Q000201:enzymology; D008969:Molecular Sequence Data; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects* / Q000201:enzymology; D010196:Pancreatic Elastase / Q000097:blood / Q000187:drug effects; D010455:Peptides / Q000494:pharmacology*; D011493:Protein Kinase C / Q000502:physiology; D011494:Protein Kinases / Q000502:physiology; D015398:Signal Transduction / Q000187:drug effects*; D001620:beta-Thromboglobulin
D002147:Calmodulin; D016209:Interleukin-8; C061615:PPBP protein, human; D010455:Peptides; D001620:beta-Thromboglobulin; D009240:N-Formylmethionine Leucyl-Phenylalanine; C009862:connective tissue-activating peptide; D000242:Cyclic AMP; D011494:Protein Kinases; D011493:Protein Kinase C; D010196:Pancreatic Elastase; D002118:Calcium