FLOW CYTOMETRIC ANALYSIS/MODELING OF LEUKOCYTE ADHESION

  • Simon, Scott I, (PI)

Project: Research project

Description

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.
StatusFinished
Effective start/end date7/1/924/30/98

Funding

  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health: $22,973.00
  • National Institutes of Health
  • National Institutes of Health

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Adhesives
Neutrophils
Leukocytes
Hydrodynamics
Leukostasis
Complement Activation
Leukopenia
Neutralizing Antibodies
Static Electricity
Epitopes
Technology
Light
Antibodies

Keywords

  • Medicine(all)
  • Immunology and Microbiology(all)