Abstract
Mast cells (MCs) are a versatile cell type playing key roles in tissue morphogenesis and host defence against bacteria and parasites. Furthermore, they can enhance immunological danger signals and are implicated in inflammatory disorders like fibrosis. This granulated cell type originates from the myeloid lineage and has similarities to basophilic granulocytes, both containing large quantities of histamine and heparin. Immature murine mast cells mature in their destination tissue and adopt either the connective tissue (CTMC) or mucosal (MMC) type. Some effector functions are executed by activation/degranulation of MCs which lead to secretion of a typical set of MC proteases (MCPT) and of the preformed or newly synthesized mediators from its granules into the local microenvironment. Due to the potential accumulation of mutations in key signalling pathway components of corresponding MC cell-lines, primary cultured MCs are an attractive mean to study general features of MC biology and aspects of MC functions relevant to human disease. Here, we describe a simple protocol for the simultaneous isolation of mature CTMC-like murine MCs from the peritoneum (PMCs) and immature MC precursors from the bone marrow (BM). The latter are differentiated in vitro to yield BM-derived MCs (BMMC). These cells display the typical morphological and phenotypic features of MCs, express the typical MC surface markers, and can be propagated and kept in culture for several weeks. The provided protocol allows simple amplification of large quantities of homogenous, non-transformed MCs from the peritoneum and bone marrow-derived mast cells for cell- and tissue-based biomedical research.
Highlights
Mast cells (MCs) are tissue-resident cells that are linked to the innate immune system
The proposed protocols of these pioneering studies are used in many laboratories to isolate immature BM-derived MCs (BMMC) or mature peritoneal MCs (PMCs)
Some of the biological attributes of MCs described were only established in immortalized cell lines resulting in findings that must be interpreted cautiously
Summary
Mast cells (MCs) are tissue-resident cells that are linked to the innate immune system. The proposed protocols of these pioneering studies are used in many laboratories to isolate immature BMMCs or mature PMCs. In principle, MCs can be derived from multipotent progenitor cells that are matured in specialized culture media, or directly isolated as functional MC from diverse tissues that are classified as tissue MC. Murine tissue MCs with a phenotype that is more consistent with connective tissue MCs can be isolated from the peritoneum and to a lesser amount from mucosa or skin [10] Both MC entities may significantly differ from each other in functional terms and each by itself might have limited functional explanatory power in estimating general properties of MCs. As a consequence, the simultaneous availability and comparative analysis of culture-maturated and tissue-derived MCs from one biological source would be helpful to conduct experiments with more functional significance. The resulting cells are highly homogenous and show the typical MC phenotype and expression of marker genes and can be propagated in culture
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