Meurer Steffen K, Neß Melanie, Weiskirchen Sabine, Kim Philipp, Tag Carmen G, Kauffmann Marlies, Huber Michael, Weiskirchen Ralf
Institute of Molecular Pathobiochemistry, Experimental Gene Therapy and Clinical Chemistry (IFMPEGKC), RWTH Aachen University, University Hospital, Aachen, Germany.
Institute of Biochemistry and Molecular Immunology, RWTH Aachen University, University Hospital, Aachen, Germany.
PLoS One. 2016 Jun 23;11(6):e0158104. doi: 10.1371/journal.pone.0158104. eCollection 2016.
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.
肥大细胞（MCs）是一种多功能细胞类型，在组织形态发生以及宿主抵御细菌和寄生虫方面发挥着关键作用。此外，它们能够增强免疫危险信号，并与诸如纤维化等炎症性疾病有关。这种颗粒状细胞类型起源于髓系谱系，与嗜碱性粒细胞有相似之处，两者都含有大量组胺和肝素。未成熟的小鼠肥大细胞在其目的地组织中成熟，并采用结缔组织（CTMC）或黏膜（MMC）类型。肥大细胞的一些效应功能是通过激活/脱颗粒来执行的，这会导致一组典型的肥大细胞蛋白酶（MCPT）以及预先形成的或新合成的介质从其颗粒分泌到局部微环境中。由于相应肥大细胞系关键信号通路成分中潜在的突变积累，原代培养的肥大细胞是研究肥大细胞生物学一般特征以及与人类疾病相关的肥大细胞功能方面的有吸引力的手段。在这里，我们描述了一种简单的方案，用于同时从腹膜中分离成熟的类似CTMC的小鼠肥大细胞（PMCs）和从骨髓中分离未成熟的肥大细胞前体。后者在体外分化产生骨髓来源的肥大细胞（BMMC）。这些细胞表现出肥大细胞典型的形态和表型特征，表达典型的肥大细胞表面标志物，并且可以在培养中繁殖并维持数周。所提供的方案允许从腹膜和骨髓来源的肥大细胞简单扩增大量同质的、未转化的肥大细胞，用于基于细胞和组织的生物医学研究。
