Phenotypic and functional characterization of in vitro cultured human mast cells.

Abstract

Mast cell progenitor cells, derived from CD34+ hematopoietic stem cells, enter the circulation and subsequently mucosal or connective tissues where they mature to mast cells. Upon activation, mast cells increase the expression of activation markers, e.g. CD63, and release histamine amongst other mediators. Traditionally, release of these mediators is quantified using assays measuring their extracellular concentration in the supernatant of stimulated cells. Human mast cells (HuMC) were cultured from peripheral blood, phenotypically characterized, passively sensitized with allogenic IgE antibodies and finally stimulated by anti-IgE that crosslinks IgE/FcεRI complexes. Alterations in the number of cells positive for CD63 and release of histamine were quantified simultaneously by flow cytometry. In culture, two distinct CD45+ cell populations were identified: CD117+ CD203c+hi and CD117- CD203c+low cells. Both populations showed positivity for FcεRI, tryptase and chymase, and contained histamine. Activation resulted in a significant increase of cells positive for CD63+ up to 21% (range: 11-39) for CD117+ CD203c+hi cells (P = 0.005), and 27% (18-55) CD63+ for CD117- CD203c+low cells (P = 0.02). Baseline histamine content was higher for CD117+ CD203c+hi cells than for CD117- CD203c+low cells, respectively 994 (695-6815) Molecules of Equivalent Specific Fluorochrome V500 per cell (MESF-V500/cell) and 797 (629-4978) MESF-V500/cell (P = 0.02). After activation, CD117+ CD203c+hi cells showed significant histamine release of 578 (366-1521) MESF-V500/cell, whilst CD117- CD203c+low cells resulted in 310 (217-366) MESF-V500/cell histamine release. This study discloses that culturing HuMC from CD34+ progenitors yields 2 phenotypically distinct cell populations that display a greatly similar response upon cross-linking of IgE/FcεRI complexes. © 2016 International Clinical Cytometry Society.