The normal immune function of leukocytes depends upon intercellular adhesion. However, under pathologic conditions such as ischemic disease and enhanced complement activation extensive neutrophil aggregation has been shown to result in leukostasis, leukopenia, and subsequent tissue damage. In the absence of degranulation neutrophil adherence and other responses to soluble, monovalent stimuli are transient. Quantitative and molecular information about this transient adherence is limited. First, details about the kinetics of aggregate formation and the distribution of aggregate size are not available from light scatter (""""""""aggregometry"""""""") and are difficult to obtain from microscopic studies. Second, the balance of nonspecific repulsive forces, hydrodynamic shear force, and the number of molecular adhesive bridges which govern adhesive interaction among neutrophils is not defined. Third, the molecular mechanisms which proceed via surface receptors and ultimately lead to adhesion between activated neutrophils are only partially understood. We have begun to investigate the quantitative and molecular details of homotypic neutrophil aggregation using novel flow cyotmetric techniques. A mathematical model based on particle geometry and rates of aggregate formation and breakup is currently used to analyze the macroscopic process. This approach has enabled us to begin to examine the impact of the number and lifetime of adhesive receptors on the time course of aggregation in isolated neutrophils and whole blood. The methodology provides a unique foundation from which to integrate the quantitative macroscopic description of the adhesion events with the molecular underpinnings which determine aggregation. The current objectives are to: 1) define, analyze, and model the quantitative aspects of aggregate size, distribution, and lifetime in a hydrodynamic environment in which the fluid shear stress and cell-cell encounter frequency are controlled; 2) gain insight to the molecular requirements of the adhesion threshold in terms of the dynamic balance between the rate and lifetime of bond formation, and electrostatic surface repulsion. By varying adhesive sites by titration with neutralizing antibodies to adhesion molecules, aggregation will be quantitated as a function of the receptor/counter-receptor number and critical separation stress; 3) verify the macroscopic interpretations by developing cytometric and imaging technologies which will allow analysis of the molecular anatomy of fluorescently labeled antibodies to specific adhesive epitopes.
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| Simon, S I; Rochon, Y P; Lynam, E B et al. (1993) Beta 2-integrin and L-selectin are obligatory receptors in neutrophil aggregation. Blood 82:1097-106 |
| Simon, S I; Chambers, J D; Butcher, E et al. (1992) Neutrophil aggregation is beta 2-integrin- and L-selectin-dependent in blood and isolated cells. J Immunol 149:2765-71 |
