Jenkinson William, Jenkinson Eric, Anderson Graham
MRC Center for Immune Regulation, University of Birmingham.
J Vis Exp. 2008 Aug 28(18):906. doi: 10.3791/906.
In the thymus, interactions between developing T-cell precursors and stromal cells that include cortical and medullary epithelial cells are known to play a key role in the development of a functionally competent T-cell pool. However, the complexity of T-cell development in the thymus in vivo can limit analysis of individual cellular components and particular stages of development. In vitro culture systems provide a readily accessible means to study multiple complex cellular processes. Thymus organ culture systems represent a widely used approach to study intrathymic development of T-cells under defined conditions in vitro. Here we describe a system in which mouse embryonic thymus lobes can be depleted of endogenous haemopoeitic elements by prior organ culture in 2-deoxyguanosine, a compound that is selectively toxic to haemopoeitic cells. As well as providing a readily accessible source of thymic stromal cells to investigate the role of thymic microenvironments in the development and selection of T-cells, this technique also underpins further experimental approaches that include the reconstitution of alymphoid thymus lobes in vitro with defined haemopoietic elements, the transplantation of alymphoid thymuses into recipient mice, and the formation of reaggregate thymus organ cultures [corrected].
在胸腺中，发育中的T细胞前体与包括皮质和髓质上皮细胞在内的基质细胞之间的相互作用，在功能健全的T细胞库的发育中起着关键作用。然而，体内胸腺中T细胞发育的复杂性会限制对单个细胞成分和特定发育阶段的分析。体外培养系统提供了一种易于研究多种复杂细胞过程的方法。胸腺器官培养系统是一种广泛使用的方法，用于在体外特定条件下研究T细胞的胸腺内发育。在此，我们描述了一种系统，通过在2-脱氧鸟苷中进行预先的器官培养，可以使小鼠胚胎胸腺叶中的内源性造血成分缺失，2-脱氧鸟苷是一种对造血细胞具有选择性毒性的化合物。该技术不仅提供了一种易于获取的胸腺基质细胞来源，以研究胸腺微环境在T细胞发育和选择中的作用，还为进一步的实验方法奠定了基础，这些方法包括用特定的造血成分在体外重建无淋巴细胞的胸腺叶、将无淋巴细胞的胸腺移植到受体小鼠体内，以及形成重组胸腺器官培养物[已修正]。
