Heterogeneity in the mobilization of cytoplasmic calcium by human polymorphonuclear leukocytes in response to fMLP, C5a and IL-8/NAP-1.
The visible excitation and emission wave-lengths of the recently developed fluorescent Ca2+ indicator fluo-3 permit analysis of the intracellular Ca2+ concentration, [Ca2+]i, in flow cytometry with a 488-nm argon laser. The role of [Ca2+]i in human polymorphonuclear leukocyte heterogeneity was investigated in response to formyl-methionyl-leucyl-phenylalanine (fMLP), C5a, and interleukin 8/neutrophil attractant/activation protein 1 (IL-8/NAP-1) by flow cytometry. The [Ca2+]i changes in different subpopulations within a heterogeneous cell suspension were resolved upon stimulation with fMLP. Using an anti-CD16 phycoerythrin-conjugated antibody and fluo-3 simultaneously, neutrophils affected and nonaffected in Ca2+ mobilization were distinguished in two patients suffering from glycogen storage disease type 1b. In normal neutrophils, a different time course of Ca2+ mobilization of neutrophil subpopulations immediately after stimulation with fMLP was detected. In addition, after stimulation with a low concentration of IL-8/NAP-1 (10(-10) M) two subsets of neutrophils appeared; one of them showed an increase in [Ca2+]i, while the other did not. These results indicate heterogeneity in the neutrophil signal transduction process involved in Ca2+ mobilization. Therefore, flow cytometric analyses can resolve changes in single-cell [Ca2+]i distribution patterns, which is important for the understanding of [Ca2+]i in neutrophil heterogeneous activation processes.
Elsner
J
J
Department of Immunobiology, Fraunhofer Institut ITA, Hannover, Germany.
Kaever
V
V
Emmendörffer
A
A
Breidenbach
T
T
Lohmann-Matthes
M L
ML
Roesler
J
J
D000814:Aniline Compounds; D002118:Calcium / Q000378:metabolism*; D015936:Complement C5a / Q000494:pharmacology*; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D016209:Interleukin-8 / Q000494:pharmacology*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014966:Xanthenes
D000814:Aniline Compounds; D016209:Interleukin-8; D014966:Xanthenes; C059715:Fluo-3; D009240:N-Formylmethionine Leucyl-Phenylalanine; D015936:Complement C5a; D002118:Calcium