Release of azurophilic granule contents in fMLP-stimulated neutrophils requires two activation signals, one of which is a rise in cytosolic free Ca2+.
We have used a continuous spectrofluorimetric method to analyse the role of cytosolic free Ca2+ ([Ca2+]i) in the lysosomal enzyme release from the azurophilic granules in human neutrophils stimulated with f-Met-Leu-Phe (fMLP) in the presence of cytochalasin B. Measurements were performed with the beta-glucuronidase substrate 4-methylumbelliferyl-beta-D-glucuronide. We found that the transient rise in [Ca2+]i induced by fMLP is a necessary signal to obtain maximal degranulation. When this Ca2+ transient is prevented by the Ca2+ chelator BAPTA, degranulation can still be induced by a stimulated Ca2+ influx, albeit to a lower extent. We also studied the degranulation process in the neutrophils of a patient with a generalized chemotactic defect. Release of beta-glucuronidase from the patient's neutrophils could not be induced despite the occurrence of a normal Ca2+ response and normal degranulation of specific granules. We conclude that, besides an increase in [Ca2+]i, an additional signal is required for the fusion of azurophilic granules with the plasma membrane in human neutrophils.
Niessen
H W
HW
Central Laboratory, The Netherlands Red Cross Blood Transfusion Service, Amsterdam.
Kuijpers
T W
TW
Roos
D
D
Verhoeven
A J
AJ
D002118:Calcium / Q000378:metabolism*; D015550:Cell Degranulation; D003571:Cytochalasin B / Q000494:pharmacology; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000201:enzymology; D003600:Cytosol / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005966:Glucuronidase / Q000378:metabolism*; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D014155:Transcobalamins / Q000378:metabolism
D014155:Transcobalamins; D003571:Cytochalasin B; D004533:Egtazic Acid; D009240:N-Formylmethionine Leucyl-Phenylalanine; D005966:Glucuronidase; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium