title: A real time chemotaxis assay unveils unique migratory profiles amongst different primary murine macrophages.
abstract: 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.
authors.affiliation: Sir William Dunn School of Pathology, University of Oxford, Oxford, United Kingdom. asif.iqbal@path.ox.ac.uk
authors.forename: Asif J
authors.initials: AJ
authors.lastname: Iqbal
authors.forename: Daniel
authors.initials: D
authors.lastname: Regan-Komito
authors.forename: Ivy
authors.initials: I
authors.lastname: Christou
authors.forename: Gemma E
authors.initials: GE
authors.lastname: White
authors.forename: Eileen
authors.initials: E
authors.lastname: McNeill
authors.forename: Amy
authors.initials: A
authors.lastname: Kenyon
authors.forename: Lewis
authors.initials: L
authors.lastname: Taylor
authors.forename: Theodore S
authors.initials: TS
authors.lastname: Kapellos
authors.forename: Edward A
authors.initials: EA
authors.lastname: Fisher
authors.forename: Keith M
authors.initials: KM
authors.lastname: Channon
authors.forename: David R
authors.initials: DR
authors.lastname: Greaves
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D019742:Chemokines, CC / Q000494:pharmacology; D002633:Chemotaxis / Q000187:drug effects; D003599:Cytoskeleton / Q000187:drug effects / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000166:cytology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009000:Monocytes / Q000166:cytology / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
chemical_list: D019742:Chemokines, CC; D019206:GTP-Binding Protein alpha Subunits, Gi-Go