Ca2+-dependent production and release of IL-8 in human neutrophils.
IL-8, a potent neutrophil chemoattractant that is elevated about 200-fold in exudative neutrophils isolated from localized inflammatory sites in vivo, is thought to play a major role in recruitment of neutrophils to inflammatory sites. Incubation of peripheral blood neutrophils with thapsigargin, an inhibitor of the endoplasmic reticulum Ca2+-sequestering-ATPase, causes a dose-dependent induction of IL-8 synthesis that continues for up to 8 h. Cycloheximide inhibits the thapsigargin-induced IL-8 production, suggesting the induction of protein synthesis de novo. In addition, Northern blot analysis of mRNA isolated from neutrophils indicates that thapsigargin treatment increases IL-8 mRNA in a time- and dose-dependent manner. Thapsigargin also induces a biphasic rise in the intracellular Ca2+ concentration, [Ca2+]i, which is composed of an initial (within 15 s) EGTA-insensitive elevation in [Ca2+]i, followed by a delayed (2-min) EGTA-sensitive component. Addition of EGTA before thapsigargin inhibited the induction of IL-8 production. Experiments in which EGTA was added at various times after thapsigargin treatment indicated that a sustained Ca2+ influx was required for maximum IL-8 production. Ascomycin and cyclosporin A, inhibitors of the Ca2+-dependent phosphatase, calcineurin, also inhibited thapsigargin-induced IL-8 production. Thus, in neutrophils, a prolonged increase in [Ca2+]i stimulates IL-8 transcription and synthesis, possibly through a calcineurin-dependent pathway.
Clinical Services Program, Science Applications International Corp. Frederick, Frederick Cancer Research and Development Center, MD 21702, USA.
D B
DB
Kuhns
H A
HA
Young
E K
EK
Gallin
J I
JI
Gallin
D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D016209:Interleukin-8 / Q000276:immunology / Q000378:metabolism*; D018375:Neutrophil Activation; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction / Q000276:immunology; D019284:Thapsigargin / Q000494:pharmacology
D004791:Enzyme Inhibitors; D016209:Interleukin-8; D012333:RNA, Messenger; D019284:Thapsigargin; D000252:Calcium-Transporting ATPases; D002118:Calcium