Health & Environmental Research Online (HERO)


Print Feedback Export to File
6341856 
Journal Article 
Differential up-regulation of specific and azurophilic granule membrane markers in electropermeabilized neutrophils 
Niessen, HW; Verhoeven, AJ 
1992 
Yes 
Cellular Signalling
ISSN: 0898-6568 
Elsevier 
4 
5 
501-509 
English 
1419488 
WOS:A1992JR48800005 
We have developed an alternative method to study the degranulation in electropermeabilized human neutrophils by measuring the up-regulation of the specific membrane markers CD63 (residing in the azurophilic granules of resting neutrophils) and CD67 (present in specific granules). The expression of these marker proteins was measured by the binding of specific antibodies to paraformaldehyde-fixed cells and subsequent flow cytometry. We first investigated whether the changes in CD63 and CD67 expression after stimulation of intact cells were comparable with earlier measurements of neutrophil degranulation, in which the release of soluble marker proteins was measured. These experiments indicated that this new method compares favourably with earlier studies, both with respect to kinetics and stimulus dependency. Subsequently, we applied this method (which does not include centrifugation of the cells) to study degranulation in electropermeabilized neutrophils. In permeabilized neutrophils, a clear up-regulation of the specific granule marker CD67 was observed upon incubation with a free Ca2+ concentration of 1 microM, a value of the cytosolic free Ca2+ concentration occurring in formylmethionyl-leucyl-phenylalanine (FMLP)-activated neutrophils. The azurophilic granule marker CD63 required GTP-gamma-S besides 1 microM Ca2+ for a significant up-regulation. Hence, our study indicates a different requirement for intracellular signals of the two main types of granules in human neutrophils. 
NEUTROPHILS; DEGRANULATION; ELECTROPERMEABILIZATION; MEMBRANE SURFACE EXPRESSION