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Culturing cells with mast cell phenotype and function: Comparison of peripheral blood and bone marrow as a source.
- Jessy Elst, Didier G. Ebo, Margaretha A. Faber, Athina L. Van Gasse, Ine I. Decuyper, Marie-Line M. van der Poorten, Chris H. Bridts, Leander P. De Puysseleyr, Christel Mertens, Margo M. Hagendorens, Luc S. De Clerck, Mark Walschot, Anke Verlinden, Daniela Berger, Peter Valent, Vito Sabato
- Journal of immunological methods 2021 v.495 pp. 113061
- blood, bone marrow, cell structures, crosslinking, flow cytometry, humans, mast cells, phenotype, substance P
- Studies on the mechanisms that govern mast cell (MC) functions are hindered by the difficulties in isolating sufficient numbers of these tissue-resident cells. Therefore, many research groups use cultured human MCs obtained out of progenitor cells. However, these culture methods significantly differ regarding primary source material, culture durations and conditions. Consequently, the finally obtained cells are likely to exhibit morphological, phenotypical and/or functional heterogeneity.To compare the phenotype and functionality of cells cultured from peripheral blood and bone marrow progenitor cells from patients with suspected clonal MC disease. These cells are designated as PBCMCs and BMCMCs, respectively.Twenty paired PBCMCs and BMCMCs cultures starting from CD34⁺ progenitor cells were compared. Cells were cultured for 4 weeks. Phenotyping included Giemsa and CD117 staining and flow cytometric staining for CD117, CD203c, FcεRI, MRGPRX2, CD300a, CD32, CD63 and CD25. Functional assessment included measurement of the up-regulation of CD63 after cross-linking of the high affinity receptor for IgE (FcεRI) with anti-FcεRI and ligation of MRGPRX2 with substance P.PBCMCs and BMCMCs are phenotypically comparable. Functionally, after activation with anti-FcεRI and substance P, PBCMCs and BMCMCs show similar up-regulation of the lysosomal degranulation marker CD63. However, the yield of PBCMCs is higher than BMCMs and peripheral blood cultures are purer than bone marrow cultures.PBCMCs are an attractive alternative to the more difficult to obtain BMCMCs for the exploration of the complex mechanisms that govern IgE- and MRGPRX2-dependent MC activation and degranulation. Unlike BMCMCs, PBCMCs are easily accessible and enable repetitive analyses.