Related Experiment Video
Updated: Aug 13, 2026
13:22
Adhesion Frequency Assay for In Situ Kinetics Analysis of Cross-Junctional Molecular Interactions at the Cell-Cell Interface
Published on: November 2, 2011
Shear and time-dependent changes in Mac-1, LFA-1, and ICAM-3 binding regulate neutrophil homotypic adhesion
S Neelamegham1, A D Taylor, H Shankaran
1Department of Chemical Engineering, State University of New York, Buffalo, NY 14260, USA.
Journal of Immunology (Baltimore, Md. : 1950)
|March 22, 2000
Summary
Neutrophil adhesion involves LFA-1 binding ICAM-3, while Mac-1 dominates at higher shear rates. Molecule contributions change over time and with fluid shear, impacting venular microcirculation.
Area of Science:
- Immunology
- Cell Biology
- Biophysics
Background:
- Leukocyte adhesion molecules, including integrins like LFA-1 and Mac-1, are crucial for immune cell trafficking.
- Homotypic neutrophil adhesion, where neutrophils bind to each other, plays a role in inflammatory responses.
- Understanding the dynamics of neutrophil adhesion under varying shear stress is vital for comprehending microcirculation events.
Purpose of the Study:
- To investigate the roles of LFA-1, Mac-1, and ICAM-3 in homotypic neutrophil adhesion.
- To determine how shear rate and stimulation time influence the contributions of these adhesion molecules.
- To elucidate the kinetics and efficiency of neutrophil aggregate formation under different flow conditions.
Main Methods:
- Isolated human neutrophils were subjected to controlled shear rates (100–800 s-1) using a cone-plate viscometer.
- Neutrophil aggregate formation was quantified over time using flow cytometry.
- Blocking antibodies against LFA-1, Mac-1, and ICAM-3 were employed to assess molecular contributions.
- Adhesion efficiency was calculated by fitting aggregate formation rates to a two-body collision theory model.
Main Results:
- Neutrophil homotypic adhesion efficiency was highest at 800 s-1, with ~40% of collisions resulting in adhesion.
- LFA-1 primarily binds ICAM-3 for homotypic adhesion, though other ligands may exist.
- Mac-1 supports adhesion comparably to LFA-1 at low shear (100 s-1) but predominates above 300 s-1.
- The adhesive capacity of LFA-1 decreased ~3-fold faster than Mac-1 post-stimulation.
Conclusions:
- The relative importance of beta2 integrins (LFA-1, Mac-1) and ICAM-3 in neutrophil adhesion is dynamically regulated by fluid shear magnitude and stimulation duration.
- These findings are relevant to blood flow conditions in the venular microcirculation.
- Adhesion molecule function is shear-dependent and time-sensitive, offering insights into inflammatory processes.
Related Concept Videos
01:15Activation of Integrins
Integrins bind ligands and transmit information from outside the cell to inside or vice-versa through an "outside-in signaling" or "inside-out signaling."
In "outside-in signaling," external factors in the extracellular space bind to exposed ligand binding sites on integrins. This causes the inactive protein to undergo a conformational change to become active. Integrins are often clustered on the cell membrane. Repetitive and regularly spaced ligand binding events provide an effective stimulus.
In "outside-in signaling," external factors in the extracellular space bind to exposed ligand binding sites on integrins. This causes the inactive protein to undergo a conformational change to become active. Integrins are often clustered on the cell membrane. Repetitive and regularly spaced ligand binding events provide an effective stimulus.
01:17Intracellular Signaling Affects Focal Adhesions
Integrins act both as extracellular input receivers and as intracellular processing activators. As their name suggests, integrins are entirely integrated into the membrane structure. Their hydrophobic membrane-spanning regions interact with the phospholipid bilayer's hydrophobic region. These membrane receptors provide extracellular attachment sites for effectors like hormones and growth factors. They activate intracellular response cascades when their effectors are bound and active.
Some...
Some...
01:24Adherens Junctions
Strong contact points between adjacent cells anchor them to each other, forming tissues. Such anchoring junctions are of two types – adherens junctions and desmosomes. Adherens junctions are abundant in tissues such as epithelium and endothelium, forming a continuous zone of adhesion called the adhesion belt. In other tissues, such as heart muscle, they appear as clusters, linking the cells to produce coordinated heart muscle contraction.
Adherens Junctions are Dynamic
The endothelial cells...
Adherens Junctions are Dynamic
The endothelial cells...
01:20Cell Adhesion Molecules - Types and Functions
Cell adhesion molecules (CAMs) are pivotal to multicellularity and the coordinated functioning of tissues and organ systems. They enable physical interactions between cells and provide mechanical strength to tissues. They also function as receptors for signal transmission across the plasma membrane. The CAMs are broadly classified into four families - integrins, cadherins, selectins, and immunoglobulin-like CAMs (IgCAMs).
CAM Families
The Integrin family of proteins is primarily involved in a...
CAM Families
The Integrin family of proteins is primarily involved in a...
01:31Immunoglobulin-like Cell Adhesion Molecules
Immunoglobulin-like cell adhesion molecules or Ig-CAMs are a versatile group of cell surface glycoproteins belonging to the immunoglobulin protein superfamily. Ig-CAMs possess the characteristic immunoglobulin protein domains and other domains such as the fibronectin type III domain. The Ig domains are glycosylated to varying degrees in different Ig-CAMs.
Ig-CAMs exhibit either homophilic binding (to other Ig-CAMs) or heterophilic binding (to other ligands such as integrins). While most Ig-CAMs...
Ig-CAMs exhibit either homophilic binding (to other Ig-CAMs) or heterophilic binding (to other ligands such as integrins). While most Ig-CAMs...
01:26Tension Response at Adherens Junctions
The adherens junctions that anchor cells together are multi-protein complexes that dynamically adapt to mechanical stimuli such as tensile forces and shear stress. Mechanosensory proteins in these junctions can sense such mechanical stimuli and undergo a shift in their conformation, resulting in an altered function — a process called mechanotransduction.
α-Catenin as a Mechanosensory Protein
The α-catenin of adherens junctions is an allosteric protein with three VH (vinculin homology) domains...
α-Catenin as a Mechanosensory Protein
The α-catenin of adherens junctions is an allosteric protein with three VH (vinculin homology) domains...