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Static and Dynamic Assays of Cell Adhesion Relevant to the Vasculature

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1430))

Abstract

Methods are described for analyzing adhesion of isolated cells (such as leukocytes, tumor cells, or precursor cells) to purified adhesion receptors or cultured endothelial cells. “Static” assays (where cells are allowed to settle on the adhesive substrates) and flow-based assays (where cells are perfused over the substrates) are compared. Direct observations of the time course of adhesion and migration can be made when purified proteins or endothelial cells are cultured in plates, after cells are allowed to settle onto them for a desired period. In the flow-based assay, cells are perfused through coated glass capillaries, flow-channels incorporating coated plates, or commercially available preformed channels. Again, direct video-microscopic observations are made. In this assay various stages of capture, immobilization, and migration can be followed. In general, the static systems have higher throughput and greatest ease of use, but yield less detailed information, while the flow-based assay is most difficult to set up but is most physiologically relevant if one is interested in the dynamics of adhesion in the vasculature.

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References

  1. McGettrick HM, Butler LM, Nash GB (2005) Analysis of leukocyte migration through monolayers of cultured endothelial cells. Methods Mol Biol 370:37–54

    Article  Google Scholar 

  2. Nash GB (2004) Adhesion between platelets and leukocytes or endothelial cells. Methods Mol Biol 272:199–214

    PubMed  Google Scholar 

  3. Springer TA (1995) Traffic signals on endothelium for lymphocyte recirculation and leukocyte emigration. Annu Rev Physiol 57:827–872

    Article  CAS  PubMed  Google Scholar 

  4. Bahra P, Rainger GE, Wautier JL, Luu N-T, Nash GB (1998) Each step during transendothelial migration of flowing neutrophils is regulated by the stimulatory concentration of tumour necrosis factor-alpha. Cell Ad Commun 6:491–501

    Article  CAS  Google Scholar 

  5. Luu NT, Rainger GE, Nash GB (1999) Kinetics of the different steps during neutrophil migration through cultured endothelial monolayers treated with tumour necrosis factor-alpha. J Vasc Res 36:477–485

    Article  CAS  PubMed  Google Scholar 

  6. Smith CW, Kishimoto TK, Abbassi O, Hughes B, Rothlein R, McIntire LV, Butcher E, Anderson DC, Abbass O (1991) Chemotactic factors regulate lectin adhesion molecule 1 (LECAM-1)-dependent neutrophil adhesion to cytokine-stimulated endothelial cells in vitro. J Clin Invest 87:609–618

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Butler LM, Jeffery HC, Wheat RL, Rae PC, Townsend K, Alkharsah KR, Schulz TF, Nash GB, Blackbourn DJ (2011) Kaposi’s sarcoma-associated herpesvirus infection of endothelial cells inhibits neutrophil recruitment through an interleukin-6-dependent mechanism: a new paradigm for viral immune evasion. J Virol 85:7321–7332

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Cinamon G, Shinder V, Alon R (2001) Shear forces promote lymphocyte migration across vascular endothelium bearing apical chemokines. Nature Immunol 2:515–522

    Article  CAS  Google Scholar 

  9. Konya V, Ullen A, Kampitsch N, Theiler A, Philipose S, Parzmair GP, Marsche G, Peskar BA, Schuligoi R, Sattler W, Heinemann A (2013) Endothelial E-type prostanoid 4 receptors promote barrier function and inhibit neutrophil trafficking. J Allergy Clin Immunol 131:532–540

    Article  CAS  PubMed  Google Scholar 

  10. Warren KJ, Iwami D, Harris DG, Bromberg JS, Burrell BE (2014) Laminins affect T cell trafficking and allograft fate. J Clin Invest 124:2204–2218

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Cooke BM, Usami S, Perry I, Nash GB (1993) A simplified method for culture of endothelial cells and analysis of adhesion of blood cells under conditions of flow. Microvasc Res 45:33–45

    Article  CAS  PubMed  Google Scholar 

  12. Butler LM, Rainger GE, Rahman M, Nash GB (2005) Prolonged culture of endothelial cells and deposition of basement membrane modify the recruitment of neutrophils. Exp Cell Res 310:22–32

    Article  CAS  PubMed  Google Scholar 

  13. Sheikh S, Gale Z, Rainger GE, Nash GB (2004) Methods for exposing multiple cultures of endothelial cells to different fluid shear stresses and to cytokines, for subsequent analysis of inflammatory function. J Immunol Methods 288:35–46

    Article  CAS  PubMed  Google Scholar 

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Correspondence to Gerard B. Nash Ph.D. .

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Butler, L.M., McGettrick, H.M., Nash, G.B. (2016). Static and Dynamic Assays of Cell Adhesion Relevant to the Vasculature. In: Martin, S., Hewett, P. (eds) Angiogenesis Protocols. Methods in Molecular Biology, vol 1430. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3628-1_16

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  • DOI: https://doi.org/10.1007/978-1-4939-3628-1_16

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3626-7

  • Online ISBN: 978-1-4939-3628-1

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