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 wavelengths of the recently developed fluorescent Ca 2+ indicator fluo‐3 permit analysis of the intracellular Ca 2+ concentration, [Ca 2+ ] i , in flow cytometry with a 488‐nm argon laser. The role of [Ca 2+ ] 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 [Ca 2+ ] 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 Ca 2+ mobilization were distinguished in two patients suffering from glycogen storage disease type lb. In normal neutrophils, a different time course of Ca 2+ 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 [Ca 2+ ] i , while the other did not. These results indicate heterogeneity in the neutrophil signal transduction process involved in Ca 2+ mobilization. Therefore, flow cytometric analyses can resolve changes in single‐cell [Ca 2+ ] i distribution patterns, which is important for the understanding of [Ca 2+ ] i in neutrophil heterogeneous activation processes.