Advertisement

Mast Cells pp 163-177 | Cite as

Isolation and Characterization of Human Intestinal Mast Cells

  • Axel LorentzEmail author
  • Gernot Sellge
  • Stephan C. Bischoff
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1220)

Abstract

Mast cells are granulated immune cells typically located at barrier sites of the body, such as the skin and the mucosa of the respiratory, urogenital, and gastrointestinal tract. They are well known for their capacity to participate in the orchestration of inflammatory and immune responses by releasing a broad array of mediators as a consequence of IgE-dependent and IgE-independent activation. Mast cells derive from myeloid progenitors, but in contrast to other myeloid cells, they leave the bone marrow in an immature state; therefore, mast cells are not visible in the blood under normal conditions. For full maturation, the tissue environment is necessary. Thus, mature mast cells can be only isolated from tissue such as skin or mucosal sites, which makes mast cell isolation complicated. This chapter describes methods to isolate, purify, and culture mast cells from the human intestinal mucosa. Human mucosal mast cells can be used to characterize their mediators and to study the mechanisms of human mast cell activation, signal transduction, and exocytosis in response to specific stimuli.

Key words

Mast cells Human Intestinal Gut Bowel Cell isolation Cell culture Cell activation Mediator release assay 

Notes

Acknowledgments

The authors thank all former and current colleagues and in particular C. A. Dahinden, K. Wordelmann, S. Schwengberg, C. T. Mierke, G. Weier, T. Gebhardt, L. E. Sander, S. P. Frank, A. Mrasori, K. Feuser, and Y. Soltow who were involved in establishing the methods described here.

References

  1. 1.
    Bischoff SC (2007) Role of mast cells in allergic and non-allergic immune responses: comparison of human and murine data. Nat Rev Immunol 7:93–104PubMedCrossRefGoogle Scholar
  2. 2.
    Kalesnikoff J, Galli SJ (2008) New developments in mast cell biology. Nat Immunol 9:1215–1223PubMedCentralPubMedCrossRefGoogle Scholar
  3. 3.
    Galli SJ, Grimbaldeston M, Tsai M (2008) Immunomodulatory mast cells: negative, as well as positive, regulators of innate and acquired immunity. Nat Rev Immunol 8:478–486PubMedCentralPubMedCrossRefGoogle Scholar
  4. 4.
    Bischoff SC (2009) Physiological and pathophysiological functions of intestinal mast cells. Semin Immunopathol 31:185–205PubMedCrossRefGoogle Scholar
  5. 5.
    Marshall JS (2004) Mast cell responses to pathogens. Nat Rev Immunol 4:787–799PubMedCrossRefGoogle Scholar
  6. 6.
    Rivera J, Gilfillan AM (2006) Molecular regulation of mast cell activation. J Allergy Clin Immunol 117:1214–1225PubMedCrossRefGoogle Scholar
  7. 7.
    Bischoff SC, Dahinden CA (1992) c-Kit ligand: a unique potentiator of mediator release by human lung mast cells. J Exp Med 175:237–244PubMedCrossRefGoogle Scholar
  8. 8.
    Bischoff SC, Sellge G, Lorentz A, Sebald W, Raab R, Manns MP (1999) IL-4 enhances proliferation and mediator release in mature human mast cells. Proc Natl Acad Sci U S A 96:8080–8085PubMedCentralPubMedCrossRefGoogle Scholar
  9. 9.
    Lorentz A, Schwengberg S, Sellge G, Manns MP, Bischoff SC (2000) Human intestinal mast cells are capable of producing different cytokine profiles: role of IgE receptor cross-linking and IL-4. J Immunol 164:43–48PubMedCrossRefGoogle Scholar
  10. 10.
    Babina M, Guhl S, Starke A, Kirchhof L, Zuberbier T, Henz BM (2004) Comparative cytokine profile of human skin mast cells from two compartments strong resemblance with monocytes at baseline but induction of IL-5 by IL-4 priming. J Leukoc Biol 75:244–252PubMedCrossRefGoogle Scholar
  11. 11.
    Hundley TR, Gilfillan AM, Tkaczyk C, Andrade MV, Metcalfe DD, Beaven MA (2004) Kit and FcεRI mediate unique and convergent signals for release of inflammatory mediators from human mast cells. Blood 104:2410–2417PubMedCrossRefGoogle Scholar
  12. 12.
    Lorentz A, Wilke M, Sellge G, Worthmann H, Klempnauer J, Manns MP, Bischoff SC (2005) IL-4 induced priming of human intestinal mast cells for enhanced survival and Th2 cytokine generation is reversible and associated with an increased activity of ERK1/2 and c-Fos. J Immunol 174:6751–6756PubMedCrossRefGoogle Scholar
  13. 13.
    Feuser K, Feilhauer K, Staib L, Bischoff SC, Lorentz A (2011) Akt crosslinks IL-4 priming, stem cell factor signaling, and IgE dependent activation in mature human mast cells. Mol Immunol 48:546–552PubMedCrossRefGoogle Scholar
  14. 14.
    Lorentz A, Schwengberg S, Mierke C, Manns MP, Bischoff SC (1999) Human intestinal mast cells produce IL-5 in vitro upon IgE receptor cross-linking and in vivo in the course of intestinal inflammatory disease. Eur J Immunol 29:1496–1503PubMedCrossRefGoogle Scholar
  15. 15.
    He SH (2004) Key role of mast cells and their major secretory products in inflammatory bowel disease. World J Gastroenterol 10:309–318PubMedGoogle Scholar
  16. 16.
    Guhl S, Babina M, Neou A, Zuberbier T, Artuc M (2010) Mast cell lines HMC-1 and LAD2 in comparison with mature human skin mast cells—drastically reduced levels of tryptase and chymase in mast cell lines. Exp Dermatol 19:845–847PubMedCrossRefGoogle Scholar
  17. 17.
    Kovarova M, Latour AM, Chason KD, Tilley SL, Koller BH (2010) Human embryonic stem cells: a source of mast cells for the study of allergic and inflammatory diseases. Blood 115:3695–3703PubMedCentralPubMedCrossRefGoogle Scholar
  18. 18.
    Rådinger M, Jensen BM, Kuehn HS, Kirshenbaum A, Gilfillan AM (2010) Generation, isolation, and maintenance of human mast cells and mast cell lines derived from peripheral blood or cord blood. Curr Protoc Immunol Chapter 7:Unit 7.37Google Scholar
  19. 19.
    Schulman ES, Macglashan DW, Peters SP, Schleimer RP, Newball HH, Lichtenstein LM (1982) Human lung mast cells: purification and characterization. J Immunol 129:2662–2667PubMedGoogle Scholar
  20. 20.
    Gibbs BF, Wierecky J, Welker P, Henz BM, Wolff HH, Grabbe J (2001) Human skin mast cells rapidly release preformed and newly generated TNF-alpha and IL-8 following stimulation with anti-IgE and other secretagogues. Exp Dermatol 10:312–320PubMedCrossRefGoogle Scholar
  21. 21.
    Kulka M, Metcalfe DD (2010) Isolation of tissue mast cells. Curr Protoc Immunol Chapter 7:Unit 7.25Google Scholar
  22. 22.
    Befus AD, Dyck N, Goodacre R, Bienenstock J (1987) Mast-cells from the human intestinal lamina propria—isolation, histochemical subtypes, and functional-characterization. J Immunol 138:2604–2610PubMedGoogle Scholar
  23. 23.
    Lowman MA, Rees PH, Benyon RC, Church MK (1988) Human mast cell heterogeneity: histamine release from mast cells dispersed from skin, lung, adenoids, tonsils, and colon in response to IgE-dependent and nonimmunologic stimuli. J Allergy Clin Immunol 81:590–597PubMedCrossRefGoogle Scholar
  24. 24.
    Bischoff SC, Schwengberg S, Raab R, Manns MP (1997) Functional properties of human intestinal mast cells cultured in a new culture system: enhancement of IgE receptor-dependent mediator release and response to stem cell factor. J Immunol 159:5560–5567PubMedGoogle Scholar
  25. 25.
    Bischoff SC, Sellge G, Schwengberg S, Lorentz A, Manns MP (1999) Stem cell factor-dependent survival, proliferation and enhanced releasability of purified mature mast cells isolated from human intestinal tissue. Int Arch Allergy Immunol 118:104–107PubMedCrossRefGoogle Scholar
  26. 26.
    Gebhardt T, Sellge G, Lorentz A, Raab R, Manns MP, Bischoff SC (2002) Cultured human intestinal mast cells express functional IL-3 receptors and respond to IL-3 by enhancing growth and IgE receptor-dependent mediator release. Eur J Immunol 32:2308–2316PubMedCrossRefGoogle Scholar
  27. 27.
    Sander LE, Frank SP, Bolat S, Blank U, Galli T, Bigalke H, Bischoff SC, Lorentz A (2008) Vesicle associated membrane protein (VAMP)-7 and VAMP-8, but not VAMP-2 or VAMP-3, are required for activation-induced degranulation of mature human mast cells. Eur J Immunol 38:855–863PubMedCrossRefGoogle Scholar
  28. 28.
    Frank SP, Thon KP, Bischoff SC, Lorentz A (2011) SNAP-23 and syntaxin-3 are required for chemokine release by mature human mast cells. Mol Immunol 49(1–2):353–358PubMedCrossRefGoogle Scholar
  29. 29.
    Walev I, Bhakdi SC, Hofmann F, Djonder N, Valeva A, Aktories K, Bhakdi S (2001) Delivery of proteins into living cells by reversible membrane permeabilization with streptolysin-O. Proc Natl Acad Sci U S A 98:3185–3190PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Axel Lorentz
    • 1
    Email author
  • Gernot Sellge
    • 2
  • Stephan C. Bischoff
    • 1
  1. 1.Department of Nutritional MedicineUniversity of HohenheimStuttgartGermany
  2. 2.Department of Medicine III, University Hospital (UKA)University of Aachen (RWTH)AachenGermany

Personalised recommendations