Background

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.

Objective

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.

Methods

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.

Results

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.

Conclusion

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.

中文翻译：

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培养具有肥大细胞表型和功能的细胞：外周血和骨髓作为来源的比较。

 背景

由于难以分离足够数量的这些组织驻留细胞，阻碍了对控制肥大细胞（MC）功能的机制的研究。因此，许多研究小组使用从祖细胞中获得的培养人类 MC。然而，这些培养方法在主要来源材料、培养持续时间和条件方面存在显着差异。因此，最终获得的细胞可能表现出形态、表型和/或功能异质性。

 客观的

比较疑似克隆性 MC 病患者的外周血和骨髓祖细胞培养的细胞的表型和功能。这些细胞分别称为 PBCMC 和 BMCMC。

 方法

比较了二十对从 CD34 ⁺祖细胞开始的 PBCMC 和 BMCMC 培养物。细胞培养4周。表型分析包括吉姆萨和 CD117 染色以及 CD117、CD203c、FcεRI、MRGPRX2、CD300a、CD32、CD63 和 CD25 的流式细胞术染色。功能评估包括测量 IgE 高亲和力受体 (FcεRI) 与抗 FcεRI 交联以及 MRGPRX2 与 P 物质连接后 CD63 的上调。

 结果

PBCMC 和 BMCMC 在表型上具有可比性。在功能上，用抗 FcεRI 和 P 物质激活后，PBCMC 和 BMCMC 表现出类似的溶酶体脱粒标记物 CD63 上调。然而，PBCMC 的产量高于 BMCM，并且外周血培养物比骨髓培养物更纯。

 结论

PBCMC 是较难获得的 BMCMC 的一种有吸引力的替代品，可用于探索控制 IgE 和 MRGPRX2 依赖性 MC 激活和脱颗粒的复杂机制。与 BMCMC 不同，PBCMC 易于访问并可进行重复分析。