Shear and time-dependent changes in Mac-1, LFA-1, and ICAM-3 binding regulate neutrophil homotypic adhesion.
We examined the relative contributions of LFA-1, Mac-1, and ICAM-3 to homotypic neutrophil adhesion over the time course of formyl peptide stimulation at shear rates ranging from 100 to 800 s-1. Isolated human neutrophils were sheared in a cone-plate viscometer and the kinetics of aggregate formation was measured by flow cytometry. The efficiency of cell adhesion was computed by fitting the aggregate formation rates with a model based on two-body collision theory. Neutrophil homotypic adhesion kinetics varied with shear rate and was most efficient at 800 s-1, where approximately 40% of the collisions resulted in adhesion. A panel of blocking Abs to LFA-1, Mac-1, and ICAM-3 was added to assess the relative contributions of these molecules. We report that 1) LFA-1 binds ICAM-3 as its primary ligand supporting homotypic adhesion, although the possibility of other ligands was also detected. 2) Mac-1 binding to an unidentified ligand supports homotypic adhesion with an efficiency comparable to LFA-1 at low shear rates of approximately 100 s-1. Above 300 s-1, however, Mac-1 and not LFA-1 were the predominant molecules supporting cell adhesion. This is in contrast to neutrophil adhesion to ICAM-1-transfected cells, where LFA-1 binds with a higher avidity than Mac-1 to ICAM-1. 3) Following stimulation, the capacity of LFA-1 to support aggregate formation decreases with time at a rate approximately 3-fold faster than that of Mac-1. The results suggest that the relative contributions of beta2 integrins and ICAM-3 to neutrophil adhesion is regulated by the magnitude of fluid shear and time of stimulus over a range of blood flow conditions typical of the venular microcirculation.
Neelamegham
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Department of Chemical Engineering, State University of New York, Buffalo, NY 14260, USA.
Taylor
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AD
Shankaran
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Smith
C W
CW
Simon
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SI
D015703:Antigens, CD; D000943:Antigens, Differentiation; D001809:Blood Viscosity / Q000276:immunology*; D002448:Cell Adhesion / Q000276:immunology; D015815:Cell Adhesion Molecules / Q000097:blood*; D002449:Cell Aggregation / Q000276:immunology; D002450:Cell Communication / Q000276:immunology; D002465:Cell Movement / Q000276:immunology*; D002634:Chemotaxis, Leukocyte / Q000276:immunology; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D018056:Hemorheology; D006801:Humans; D008024:Ligands; D016169:Lymphocyte Function-Associated Antigen-1 / Q000097:blood*; D016177:Macrophage-1 Antigen / Q000097:blood*; D009504:Neutrophils / Q000276:immunology*; D009930:Organic Chemicals; D011485:Protein Binding / Q000276:immunology; D013997:Time Factors
D015703:Antigens, CD; D000943:Antigens, Differentiation; D015815:Cell Adhesion Molecules; D005456:Fluorescent Dyes; C483616:ICAM3 protein, human; D008024:Ligands; D016169:Lymphocyte Function-Associated Antigen-1; D016177:Macrophage-1 Antigen; D009930:Organic Chemicals; C057718:6-(Dimethylamino)-2-((1E,3E)-4-(4-(dimethylamino)phenyl)-1,3-butadien-1-yl)-1-ethylquinolinium perchlorate; D016650:Fluorescein-5-isothiocyanate