ANALYSIS OF MECHANISMS OF EARLY HAEMATOPOIETIC STEM CELL DEVELOPMENT CONTROLLED BY FLK1/SCL REGULATORY NETWORK

动物的血液系统通过每天产生数十亿种不同类型的红细胞、粒细胞和淋巴细胞来维持。血液系统的基础是被称为造血干细胞（HSC）的血液干细胞。因此，动物体内的所有血细胞都是由造血干细胞产生的。这种细胞类型负责接受骨髓移植的患者血液系统的长期重建。由于骨髓供体数量有限，科学家和临床医生正在寻找在实验室条件下生成造血干细胞的替代途径。胚胎干细胞（ES）在再生医学，包括血液生产方面有着巨大的前景。然而，从胚胎干细胞中产生造血干细胞是一个严峻的挑战，主要是因为胚胎如何产生造血干细胞的许多基本细节都缺失。在很大程度上，进展一直很缓慢，因为直到最近才有了能够让我们复制造血干细胞发育的培养系统。这项研究计划主要是基于我们实验室开发的一种新型的强大的分析培养系统。它使我们能够识别胚胎中造血干细胞发育的特定阶段，而这些阶段是传统方法无法检测到的。我们将使用这种培养系统来研究两个基因，称为Flk1和Scl，它们是早期HSC发展的主要调节因子。这些基因在发育中的胚胎的各种细胞群中表达，这些细胞可能参与造血干细胞的发育。此外，这些基因有可能相互调节。年代的活动。我们将研究Flk1和Scl基因如何通过在我们的培养系统中使用现代重组技术删除这些基因来调节HSC的发育阶段。这种基因消融的结果将始终通过长期移植试验来验证，这是检测造血干细胞存在和数量的金标准方法。此外，我们将对HSC发育不同阶段表达的基因进行更全面的分析，并确定Flk1和Scl缺失后该基因调控网络是如何被修饰的。为此，我们将使用一种名为Solexa的现代自动化系统。这种方法将使我们能够创建一个计算机化的遗传网络，它涉及造血干细胞的早期生成。从长远来看，该模型将为测试其他基因在造血干细胞发育中的作用提供有用的资源。

The blood system in the animal is maintained through daily generation of billions of different types of erythrocytes, granulocytes and lymphocytes. At the foundation of the blood system lie blood stem cells called haematopoietic stem cells (HSC). Thus, all blood cells in the animal are generated from HSCs. This is the cell type which is responsible for the long-term reconstitution of the blood system of patients who receive bone marrow transplants. Since the number of bone marrow donors is limited, scientists and clinicians are seeking for alternative routes for generation of HSCs in the laboratory conditions. Embryonic stem (ES) cells hold a great promise for regenerative medicine, including production of blood. However, the generation of HSCs from ES cells pose a serious challenge, mainly because many essential details of how the embryo generates HSCs are missing.To a large extent, the progress has been slow since no culture system which would allow us to replicate development of HSCs was available until recently. This research proposal is mainly based on a novel powerful and analytical culture system developed in our laboratory. It allowed us to identify specific stages of development of HSCs in the embryo, at a time when these stages are concealed from scrutiny by traditional methods. We will use this culture system to investigate two genes, called Flk1 and Scl, that are, major regulators of early HSC development. These genes are expressed in various populations of cells of the developing embryo which are potentially involved in development of HSCs. In addition, these genes have a potential to regulate each other?s activity. We will study how Flk1 and Scl genes regulate stages of HSC development by deleting these genes in our culture system using modern recombination technology. Results of such gene ablation will always be verified by a long-term transplantation assay, which is the gold-standard approach for detection of the presence and numbers of HSCs. Additionally, we will carry out more comprehensive analysis of genes expressed at different stages of HSC development and determine how this genetic regulatory network is modified after deletion of Flk1 and Scl. To this end, we will use a modern automated system called Solexa. This approach will allow us to create a computerised genetic network which is involved in early generation of HSCs. In the longer term, this model will be a useful resource for testing the role of other genes in development of HSCs.