Generation, Isolation, and Maintenance of Human Mast Cells and Mast Cell Lines Derived from Peripheral Blood or Cord Blood

Abstract

Antigen-mediated mast cell activation is a pivotal step in the initiation of allergic disorders including anaphylaxis and atopy. To date, studies aimed at investigating the mechanisms regulating these responses, and studies designed to identifying potential ways to prevent them, have primarily been conducted in rodent mast cells. However, to understand how these responses pertain to human disease, and to investigate and develop novel therapies for the treatment of human mast cell-driven disease, human mast cell models may have greater relevance. Recently, a number of systems have been developed which allow investigators to readily obtain sufficient quantities of human mast cells to conduct these studies. These mast cells release the appropriate suite of inflammatory mediators in response to known mast cell activators including antigen. These systems have also been employed to examine the signaling events regulating these responses. Proof of principle studies have also demonstrated utility of these systems for the identification of potential inhibitors of mast cell activation and growth. In this unit, we describe techniques for the development and culture of human mast cells from their progenitors and the culture of human mast cell lines. The relative merits and drawbacks of each model are also described.

PMX-53 as a Dual CD88 Antagonist and an Agonist for Mas-Related Gene 2 (MrgX2) in Human Mast Cells

“…Half the cell culture medium was replaced weekly with media containing 100 ng/ml rhSCF and 100 ng/ml rhIL-6. Cells were used for experiments after 7 to 10 weeks in culture (Venkatesha et al, 2005;Rådinger et al, 2010). LAD2 cells were maintained in StemPro-34 medium containing nutrient supplements (Invitrogen) supplemented with 2 mM L-glutamine, 100 IU/ml penicillin, 100 g/ml streptomycin, and 100 ng/ml rhSCF (Kirshenbaum et al, 2003;Rådinger et al, 2010).…”

Mas-related Gene X2 (MrgX2) Is a Novel G Protein-coupled Receptor for the Antimicrobial Peptide LL-37 in Human Mast Cells

“…Hemidepletions were performed weekly with media containing rhSCF (100 ng/ml) and rhIL-6 (100 ng/ml) (30). Cells were used for experiments after 7-10 weeks in culture.…”

A Phenotypic Screening Approach in Cord Blood–Derived Mast Cells to Identify Anti-Inflammatory Compounds

“…We therefore used a surrogate sample-mast cells differentiated over 8 to 10 weeks from a CBDMC preparation-as precedented in the literature. 7,8,[16][17][18] The differentiation and maintenance of the culture were critical to ensure the robustness of the ultimate screening assay and carefully controlled, with cultures maintained at strict densities, regular cell counting and feeding, further QC checks by granule staining for mast cell identification, and batch testing for sensitivity to IgE-driven degranulation before the small-molecule screening assays.…”