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Enhanced aggregation of human neutrophils by MnCl 2 or DTT differentiates the roles of L‐selectin and β 2 ‐integrins
Author(s) -
Lynam Eric B.,
Rogelj Snezna,
Edwards Bruce S.,
Sklar and Larry A.
Publication year - 1996
Publication title -
journal of leukocyte biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.819
H-Index - 191
eISSN - 1938-3673
pISSN - 0741-5400
DOI - 10.1002/jlb.60.3.356
Subject(s) - integrin , dithiothreitol , selectin , adhesion , flow cytometry , microbiology and biotechnology , biophysics , intracellular , cell adhesion , biology , biochemistry , chemistry , cell adhesion molecule , receptor , cell , enzyme , organic chemistry
MnCl 2 and dithiothreitol (DTT) enhance the adhesive functions of β 2 ‐integrins. We have used these agents and flow cytometry to distinguish the contributions of β 2 ‐integrins and L‐selectin to neutrophil aggregation. Although neither compound induced aggregation, they prolonged N ‐formylmethionyl‐leucyl‐phenylalanine‐induced aggregation and produced larger aggregates. Because activated polymorphonuclear granulocytes (PMN) shed L‐selectin in the presence of MnCl 2 , but not DTT, we could evaluate the role of L‐selectin in the early and late stages of aggregation. Blocking L‐selectin sites with DREG200 Fab and/or β 2 ‐integrin sites with IB4 Fab indicated that aggregation under all conditions remained β 2 ‐integrin‐ and L‐selectin‐dependent. Disaggregation was integrin‐dependent whether L‐selectin was present or shed. The disaggregation kinetics suggested that integrin bonds turned over at a slower rate in MnCl 2 ‐treated cells. Enhanced aggregation due to DTT and MnCl 2 required sustained energy output, suggesting intracellular rather than strictly conformational control. These results provide evidence that PMN aggregation, like leukocyte‐endothelial cell adhesion, utilizes L‐selectin to form intercellular contacts that are maintained through activated integrins. J. Leukoc. Biol . 60: 356–364; 1996.