Peripheral blood cultured mast cells: Phenotypic and functional outcomes of different culture protocols

Jessy Elst; Vito Sabato; Marie-Line M van der Poorten; Margaretha Faber; Athina L Van Gasse; Leander P De Puysseleyr; Chris H Bridts; Christel Mertens; Michel Van Houdt; Marcus Maurer; Margo M Hagendorens; Didier G Ebo

doi:10.1016/j.jim.2021.113003

Peripheral blood cultured mast cells: Phenotypic and functional outcomes of different culture protocols

J Immunol Methods. 2021 May:492:113003. doi: 10.1016/j.jim.2021.113003. Epub 2021 Feb 26.

Affiliations

¹ University of Antwerp, Faculty of Medicine and Health Sciences, Department of Immunology, Allergology, Rheumatology and the Infla-Med Centre of Excellence, Antwerp (Belgium) and Immunology, Allergology, Rheumatology, Antwerp University Hospital, Antwerp, Belgium.
² University of Antwerp, Faculty of Medicine and Health Sciences, Department of Immunology, Allergology, Rheumatology and the Infla-Med Centre of Excellence, Antwerp (Belgium) and Immunology, Allergology, Rheumatology, Antwerp University Hospital, Antwerp, Belgium; Department of Immunology and Allergology, AZ Jan Palfijn Gent, Ghent, Belgium.
³ University of Antwerp, Faculty of Medicine and Health Sciences, Department of Immunology, Allergology, Rheumatology and the Infla-Med Centre of Excellence, Antwerp (Belgium) and Immunology, Allergology, Rheumatology, Antwerp University Hospital, Antwerp, Belgium; University of Antwerp, Faculty of Medicine and Health Sciences, Department of Paediatrics and the Infla-Med Centre of Excellence, Antwerp (Belgium) and Paediatrics, Antwerp University Hospital, Antwerp, Belgium.
⁴ Dermatological Allergology, Allergie-Centrum-Charité, Department of Dermatology and Allergy, Charité - Universitätsmedizin Berlin, Germany.
⁵ University of Antwerp, Faculty of Medicine and Health Sciences, Department of Immunology, Allergology, Rheumatology and the Infla-Med Centre of Excellence, Antwerp (Belgium) and Immunology, Allergology, Rheumatology, Antwerp University Hospital, Antwerp, Belgium; Department of Immunology and Allergology, AZ Jan Palfijn Gent, Ghent, Belgium. Electronic address: immuno@uantwerpen.be.

PMID: 33647250
DOI: 10.1016/j.jim.2021.113003

Abstract

Background: Mast cells (MCs) play a pivotal role in innate and adaptive immune responses. However, MCs are also involved in different pathologic conditions. Studies on the mechanisms that govern human MC functions are impeded by their limited and difficult recovery. Therefore, several research groups have developed protocols to culture human MCs from progenitor cells. These protocols vary with respect to culture duration and used maturation cytokines. How MCs obtained by different protocols differ in phenotype and functionality is currently unknown.

Objective: To compare different protocols for the generation of human MCs from peripheral blood progenitors.

Methods: Thirteen paired human MC cultures were investigated. MCs were cultured form CD34⁺ progenitors cells for 4 or 8 weeks and with or without the addition of IL-6. Phenotyping comprised staining for CD117, CD203c, FcεRI, MRGPRX2, CD300a and CD32. Functional studies included measurements of the up-regulation of CD63 and CD203c after allergen-specific cross-linking of sIgE/FcεRI complexes or ligation of MRGPRX2 with substance P and different drugs.

Results: Cell cultures for 4 weeks in the presence of IL-6 consistently yielded the highest numbers of MCs. MCs cultured for 8 weeks with IL-6 showed more autofluorescence significantly impeding correct analyses of FcεRI and CD32. The density of FcεRI and CD32 was comparable between the different culture conditions. MRGPRX2 expression was significantly higher in the 8 week cultures. The density of CD300a was increased in the cultures with IL-6. Cells cultured for 8 weeks were more responsive to MRGPRX2 activation. In contrast, the 4-weeks cultures with IL-6 showed significantly higher allergen-specific activation.

Conclusion: Four weeks of culture with IL-6 are sufficient to generate sizeable numbers of human mast cells from blood progenitors, thereby enabling simultaneous exploration of allergen-specific sIgE/FcεRI cross-linking and non-specific activation via MRGPRX2.

Keywords: Flow cytometry; Human mast cell; IgE; IgE-dependent activation; MRGPRX2; Mast cell activation.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Cells, Cultured / immunology
Cells, Cultured / metabolism
Culture Media / metabolism
Flow Cytometry
Healthy Volunteers
Humans
Immunophenotyping
Interleukin-6 / metabolism*
Mast Cells / immunology*
Mast Cells / metabolism
Nerve Tissue Proteins / metabolism
Primary Cell Culture / methods*
Receptors, G-Protein-Coupled / metabolism
Receptors, Neuropeptide / metabolism
Time Factors

Substances

Culture Media
IL6 protein, human
Interleukin-6
MRGPRX2 protein, human
Nerve Tissue Proteins
Receptors, G-Protein-Coupled
Receptors, Neuropeptide