Methods for Producing T lymphocytes in vitro from Stem Cells

T淋巴细胞是一种白色血细胞，对免疫系统的功能至关重要。这些细胞的异常功能与免疫缺陷（例如AIDS）或自身免疫（例如I型糖尿病、类风湿性关节炎）相关。T淋巴细胞来源于造血干细胞，并在胸腺中完成其成熟，胸腺是一个位于心脏附近的器官，它已经进化到专门提供促进T细胞发育的环境。造血干细胞成熟为T淋巴细胞时所经历的发育阶段已经得到了相对良好的表征。这使得能够识别细胞必须通过的关键监管检查点，以便进一步发展。其中一个检查点被称为β选择。为了通过这个检查点，细胞必须产生特定的信号，引起基因表达的变化，并允许细胞分裂和分化。T细胞发育的体外模型依赖于来源于支持基质细胞的辅助因子，其中Notch-1配体是最好理解的。然而，Notch-1配体不足以在辅助细胞不存在的情况下允许T细胞发育，表明需要由基质细胞产生的额外因子。我们已经鉴定了趋化因子CXCL 12作为β-选择过程中的辅因子，这使我们能够创建允许β-选择的无辅助细胞培养系统。我们现在建议检查层粘连蛋白和Wnt等其他基质衍生成分在无细胞系统中的作用。我们还建议测试我们对胸腺细胞发育的认识是否允许T细胞从干细胞生长而不需要基质辅助细胞。这将提供一个更简单的系统来评估分化的分子过程。此外，它可以允许T细胞的扩增，用于再生医学领域中基于细胞的治疗。

T lymphocytes, so-called because they develop in the thymus, are a type of white blood cell crucial for the function of the immune system. Aberrant function of these cells is associated with immunodeficiency (e.g. AIDS) or autoimmunity (e.g. type I diabetes, rheumatoid arthritis). T lymphocytes are derived from blood stem cells and complete their maturation in the thymus, an organ located near the heart that has evolved specifically to provide an environment that promotes T cell development. The developmental stages haematopoietic stem cells pass through as they mature into T lymphocytes have been relatively well-characterised. This has allowed the identification of key regulatory checkpoints that cells must pass through in order to develop further. One of these checkpoints is called beta-selection. In order to pass through this checkpoint cells must generate specific signals which bring about changes in gene expression and allows the cells to divide and differentiate. In vitro models of T cell development rely on accessory factors derived from supporting stromal cells of which Notch-1 ligands are the best understood. However Notch-1 ligand is insufficient to allow T cell development in the absence of accessory cells indicating additional factors produced by the stromal cells are required. We have identified the chemokine CXCL12 as a co-factor in the process of beta-selection and this has enabled us to create an accessory cell free culture system which is permissive for beta-selection. We now propose to examine the effect of additional stromal derived components such as laminin and Wnt for their effect in a cell-free system. We also propose to test whether our insight into the development of thymocytes will permit the growth of T cells from stem cells without the need for stromal accessory cells. This will provide a simpler system to evaluate the molecular processes of differentiation. Furthermore, it may permit the expansion of T cells for cell-based therapy in the area of regenerative medicine.