A Real Time Chemotaxis Assay Unveils Unique Migratory Profiles amongst Different Primary Murine Macrophages

Chemotaxis assays are an invaluable tool for studying the biological activity of inflammatory mediators such as CC chemokines, which have been implicated in a wide range of chronic inflammatory diseases. Conventional chemotaxis systems such as the modified Boyden chamber are limited in terms of the...

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Veröffentlicht in:PloS one Jg. 8; H. 3; S. e58744
Hauptverfasser: Iqbal, Asif J., Regan-Komito, Daniel, Christou, Ivy, White, Gemma E., McNeill, Eileen, Kenyon, Amy, Taylor, Lewis, Kapellos, Theodore S., Fisher, Edward A., Channon, Keith M., Greaves, David R.
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States Public Library of Science 14.03.2013
Public Library of Science (PLoS)
Schlagworte:
Actin
Analysis
Animals
Antigens
Assaying
Biological activity
Biology
Bone marrow
Bone Marrow Cells - cytology
Boyden chamber
CC chemokines
CD14 antigen
Cell migration
Chemokines
Chemokines, CC - pharmacology
Chemokinesis
Chemotaxis
Chemotaxis - drug effects
Cytochalasin D
Cytokines
Cytoskeleton
Cytoskeleton - drug effects
Cytoskeleton - metabolism
Ethics
Gene expression
GTP-Binding Protein alpha Subunits, Gi-Go - metabolism
Humans
Inflammation
Inflammatory diseases
Laboratories
Macrophages
Macrophages - cytology
Male
Medicine
Mice
Mice, Inbred C57BL
Monocyte chemoattractant protein 1
Monocytes
Monocytes - cytology
Monocytes - drug effects
Pathology
Penicillin
Pertussis
Pertussis toxin
Populations
Proteins
Real time
Signal transduction
Signal Transduction - drug effects
Signaling
Time Factors
Toxins
New York
United States > US
ISSN:1932-6203, 1932-6203
Online-Zugang:Volltext
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Beschreibung
Zusammenfassung:Chemotaxis assays are an invaluable tool for studying the biological activity of inflammatory mediators such as CC chemokines, which have been implicated in a wide range of chronic inflammatory diseases. Conventional chemotaxis systems such as the modified Boyden chamber are limited in terms of the data captured given that the assays are analysed at a single time-point. We report the optimisation and validation of a label-free, real-time cell migration assay based on electrical cell impedance to measure chemotaxis of different primary murine macrophage populations in response to a range of CC chemokines and other chemoattractant signalling molecules. We clearly demonstrate key differences in the migratory behavior of different murine macrophage populations and show that this dynamic system measures true macrophage chemotaxis rather than chemokinesis or fugetaxis. We highlight an absolute requirement for Gαi signaling and actin cytoskeletal rearrangement as demonstrated by Pertussis toxin and cytochalasin D inhibition. We also studied the chemotaxis of CD14(+) human monocytes and demonstrate distinct chemotactic profiles amongst different monocyte donors to CCL2. This real-time chemotaxis assay will allow a detailed analysis of factors that regulate macrophage responses to chemoattractant cytokines and inflammatory mediators.
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Competing Interests: The authors have declared that no competing interests exist.
Conceived and designed the experiments: AJI GEW EAF KMC DRG. Performed the experiments: AJI DRK IC EM AK LT TSK DRG. Analyzed the data: AJI IC DRG. Contributed reagents/materials/analysis tools: AJI GEW EM KMC DRG. Wrote the paper: AJI GEW EAF DRG.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0058744