NA-phenotype-dependent differences in neutrophil Fc gamma RIIIb expression cause differences in plasma levels of soluble Fc gamma RIII.
Soluble Fc gamma RIII in plasma is primarily derived from neutrophils and is a measure of the total body neutrophil mass. We have developed a new, sensitive 'sandwich' ELISA to measure soluble Fc gamma RIII in plasma and released Fc gamma RIII in cell supernatants. Both sFc gamma RIIIa, derived from NK cells and sFc gamma RIIIb, derived from neutrophils are detected in the assay. However, plasma analysis of Fc gamma RIIIB gene-deficient donors suggested that sFc gamma RIIIa contributes only marginally to the total amount measured in healthy individuals. Furthermore, we observed that plasma of homozygous NA1-positive donors contained lower amounts of sFc gamma RIII than plasma of homozygous NA2-positive donors. Heterozygous donors were found to have intermediate levels of sFc gamma RIII in their plasma. Hemizygous Fc gamma RIIIB gene-deficient donors were found to have half the amount of sFc gamma RIII in their plasma compared to donors with two Fc gamma RIIIB alleles. These NA phenotype-dependent differences in plasma sFc gamma RIII could not be contributed to either an assay artefact or NA-dependent differences in shedding of Fc gamma RIIIb upon neutrophil activation. Calibration curves constructed with plasma of homozygous donors did nor reveal NA-dependent differences in antibody affinity. Measurement of released Fc gamma RIIIb in supernatants of neutrophils stimulated with PMA, and inhibition of this signal with human IgG revealed no NA-dependent differences. However, NA-dependent differences in neutrophil Fc gamma RIIIb expression were present, comparable to the differences found in plasma levels of sFc gamma RIII. Differences in the amounts of released Fc gamma RIII in supernatants of NA-typed apoptotic neutrophils were similar to initial differences in Fc gamma RIIIb expression, again being lower in NA1-positive than in heterozygous and NA2-positive donors. In conclusion, NA-dependent differences in plasma levels of soluble Fc gamma RIII seem to be caused by differences in expression of the receptor on the neutrophil membrane.
Koene
H R
HR
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, University of Amsterdam, The Netherlands.
de Haas
M
M
Kleijer
M
M
Roos
D
D
von dem Borne
A E
AE
D000941:Antigens / Q000276:immunology*; D017209:Apoptosis / Q000502:physiology; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D009504:Neutrophils / Q000276:immunology* / Q000502:physiology; D010641:Phenotype; D017452:Receptors, IgG / Q000378:metabolism*
D000941:Antigens; D017452:Receptors, IgG