Identification of functional domains in the AGM region linked to the hierarchical organisation of the developing haematopoietic stem cell lineage

血液系统的基础是被称为造血干细胞(HSCs)的血液干细胞，它能产生所有类型的血细胞。尽管可以活跃地产生成熟的血细胞，但造血干细胞在生命周期中并不会耗尽，因为每次分裂时，它们至少会产生一份自身的副本。造血干细胞是研究最多的干细胞类型，为其他干细胞类型的分析提供了一个模型。这些强大的不朽细胞在生物体中的重要性引起了科学界和普通公众的极大关注。尽管在这一领域取得了重大进展，但造血干细胞在胚胎发育过程中产生的确切来源和机制仍然知之甚少。腹主动脉-性腺-中肾(AGM)区是胚胎发育中最早出现造血干细胞的重要器官。我们最近开发了一种强大的技术，使我们能够在培养的发育中的AGM区域复制大量的HSCs(在四天的培养期间，HSCs的数量大约增加了150倍，Taoudi等人，Cell Stem Cell，2008)。造血干细胞的发展是一个多步骤的过程，依赖于与周围细胞的相互作用。这个培养系统第一次使我们能够研究单个细胞群体在HSC发育中的作用。在这里，我们建议将AGM区域解构为适合操作和分析的“积木”。我们将确定那些具有促进胚胎前体细胞(这里称为前HSCs)发育为最终HSCs的功能细胞。我们的初步实验表明，在发育过程中，前HSCs经历了不同的成熟阶段。为了有效地实现这一目标，我们不仅将从AGM区域纯化不同类型的细胞，而且还将使用一种特殊的转基因小鼠品系来获得不同类型的永生细胞系的文库。这样的细胞系将在我们的实验中用作可再生的标准材料。我们已经确定了一种细胞系，能够促进一种类型的PF前HSCs发育为特定的HSCs。因此，该项目的重要理论基础是，HSC发育的不同阶段与年度大会区域内不同的微环境有关。使用上述策略，我们将能够将HSC发育的某些阶段映射到AGM区域内的特定形态结构域。此外，基于所获得的知识，我们的目标是仅使用上述分析中确定的必要“构件”来重新设计年度股东大会区域。通过这一点，我们将设计一个功能完善的年度股东大会培养系统，以便进行进一步的深入分析。

At the foundation of the blood system lie blood stem cells called haematopoietic stem cells (HSCs) which give rise to all types of blood cells. In spite of active production of mature blood cells, HSCs are not exhausted during the lifespan, since every time they divide they produce at least one copy of themselves. HSCs are the best studied stem cell type which serves a model for analysis of other stem cell types. The importance of these potent 'immortal' cells in the organism attracts considerable attention both from scientific community and general public. Despite significant progress in this field the exact origin and mechanisms whereby HSCs emerge during embryo development remain poorly understood. The aorta-gonad-mesonephros (AGM) region is an important organ in the developing embryo in which HSCs first appear. We have recently developed a powerful technique which allows us to reproduce massive generation of HSCs in cultured developing AGM regions (approximately 150-fold increase in HSCs was achieved during a four day culture period, Taoudi et al., Cell Stem Cell, 2008). Development of HSCs is a multi-step process dependent on interaction with surrounding cells. This culture system for the first time allowed us to investigate the role of individual cell populations in HSC development. Here we propose to deconstruct the AGM region into 'building blocks' suitable for manipulation and analysis. We will identify those which have functional capacity to promote development of embryonic precursors (called here pre-HSCs) into definitive HSCs. Our preliminary experiments show that during development, pre-HSCs go through distinct stages of maturation. To effectively pursue this goal we will not only purify different cell types from the AGM region but also derive a library of immortal cell lines of different types using a special transgenic mouse strain. Such cell lines will then be used as renewable standard material in our experiments. We have already identified one cell line which is capable of promoting development of one type pf pre-HSCs into definitive HSCs. Thus, important rationale for this project is that distinct stages of HSC development are associated with different microenvironments within the AGM region. Using the above strategy, we will be able to map certain stages of HSC development to specific morphological domains within the AGM region. In addition, based on the knowledge obtained, we aim to re-design the AGM region using only essential 'building blocks' identified in the analysis described above. By this we will engineer a well characterised functional AGM culture system accessible for further in-depth analysis.