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Measurement of Cellular Chemotaxis with ECIS/Taxis
11:37

Measurement of Cellular Chemotaxis with ECIS/Taxis

Published on: April 1, 2012

Characterization of transendothelial chemotaxis of T lymphocytes

S J Roth1, M W Carr, S S Rose

  • 1Department of Cardiology, Children's Hospital, Boston, MA, USA.

Journal of Immunological Methods
|December 15, 1995
PubMed

Insights

This study developed an in vitro assay for lymphocyte transendothelial chemotaxis using human umbilical vein endothelial cells. Phytohemagglutinin-stimulated supernatants effectively chemoattract peripheral blood lymphocytes, highlighting key adhesion molecules involved.

Area of Science:

  • Immunology
  • Cell Biology
  • Biomedical Research

Background:

  • Lymphocyte migration into tissues is crucial for immune responses.
  • Understanding the mechanisms of lymphocyte transendothelial migration is vital for studying inflammation and immune cell trafficking.
  • Existing in vitro models may not fully replicate physiological conditions.

Purpose of the Study:

  • To establish and validate an in vitro assay for studying lymphocyte transendothelial chemotaxis.
  • To identify chemoattractant factors and adhesion molecules involved in lymphocyte migration across endothelial monolayers.
  • To provide a physiologically relevant model for lymphocyte emigration into inflamed tissues.

Main Methods:

  • Developed an in vitro chemotaxis assay using human umbilical vein endothelial cell (HUVEC) monolayers on microporous membranes.
  • Utilized supernatants from phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMCs) as a source of chemoattractant activity.
  • Employed checkerboard analysis and monoclonal antibodies against adhesion receptors (LFA-1, ICAM-1/ICAM-2, VLA-4, VCAM-1, beta 1 integrin subunit) to investigate migration mechanisms.

Main Results:

  • PHA supernatant significantly increased peripheral blood lymphocyte (PBL) transmigration compared to control.
  • Transmigration was confirmed to be directional, not random locomotion.
  • The LFA-1/ICAM-1/ICAM-2 interaction was more critical than VLA-4/VCAM-1 for chemotaxis.
  • Monoclonal antibody against the beta 1 integrin subunit showed greater inhibition than antibodies against specific VLA subunits.
  • The assay guided the purification of monocyte chemoattractant protein-1 (MCP-1) as a key chemoattractant.

Conclusions:

  • The developed HUVEC-based assay is effective for studying lymphocyte transendothelial chemotaxis in vitro.
  • Lymphocyte migration is mediated by specific adhesion molecule interactions, with LFA-1/ICAM-1/ICAM-2 playing a dominant role.
  • The assay provides a valuable, physiologically relevant model for investigating lymphocyte emigration and can aid in the purification of chemoattractant factors like MCP-1.

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