Building the Hematopoietic Stem Cell Niche

描述(由申请人提供)： 各种细胞类型的层次化和严密控制的组织是正常组织和器官的标志，支持这一提议的假设是，预先定义3D组织结构中单个细胞的特定位置和由此产生的细胞-细胞相互作用将允许人们创建高功能组织，其中细胞-细胞相互作用对细胞表型的作用可以被精确描绘。这一概念将通过开发人类造血的3D模型来探索，在该模型中，将探索骨祖细胞和血管细胞在定义造血干细胞(HSC)生态位中的作用。其具体目标包括(1)开发微流控技术，使单细胞能够大规模包裹在高度定义的细胞外基质模拟物中，从而确定基质信号如何在单细胞水平调控间充质干细胞的分化；(2)创建混合集成电路/微流控电路系统，使人们能够将包含单个细胞的皮升液滴和合成的ECM组装成具有预先定义的结构和组织的3D组件；以及(3)确定适当的HSC和代表骨髓HSC的细胞的体外组装是否能够产生具有体外重建造血功能的造血组织。该项目的成功将导致创造一套新的工具，使3D组织的形成具有精确定义的细胞放置以及同型和异型细胞-细胞相互作用。这些工具很可能广泛用于建立新的组织发育和药物筛选的体外模型，以及来自各种细胞类型的体内组织替代。由于干细胞对环境信号特别敏感，不适当的细胞-细胞和细胞-基质相互作用可能导致在培养中发现的干细胞分化命运的不可逆和不受欢迎的改变。该项目开发的系统将使我们能够研究血管细胞和成骨细胞/成骨细胞在维持人类HSC生态位中的具体作用，这在体内是一个难以解决的问题。更好地定义和创建利基模型，以了解正常的造血和涉及血细胞的病理，并在各种治疗场所按需进行造血，这是至关重要的。到目前为止，关于这一主题的关键研究都依赖于啮齿动物模型，许多发现与人类生物学的相关性目前尚不清楚。 (评论者评论结束)

DESCRIPTION (provided by applicant): A hierarchal and tightly controlled organization of various cell types is the hallmark of normal tissues and organs, and the hypothesis underlying this proposal is that pre-defining the specific location and resultant cell- cell interactions of individual cells within a 3D tissue construct will allow one to create highly functional tissues in which the role of cell-cell interactions on cell phenotype can be precisely delineated. This concept will be explored by developing a 3D model of human hematopoiesis, in which osteoprogenitors and vascular cells will be probed for their roles in defining the hematopoietic stem cell (HSC) niche. The specific aims include (1) the development of microfluidic techniques to allow large-scale encapsulation of single cells in highly defined extracellular matrix mimics in order to determine how matrix cues regulate mesenchymal stem cell differentiation at the single cell level, (2) the creation of hybrid integrated circuit/microfluidic circuit systems to enable one to assemble picoliter drops containing individual cells and synthetic ECM into 3D assemblies with pre-defined structure and organization, and (3) determining whether appropriate in vitro assembly of HSCs and cells representative of the bone marrow HSC niche can yield functional hematopoietic tissues capable of recreating hematopoiesis in vitro. Success in this project will lead to the creation of a new set of tools that will enable formation of 3D tissues with precisely defined cell placement, and homotypic and heterotypic cell-cell interactions. These tools are likely to be broadly useful to the creation of new in vitro models of tissue development and drug screening, and in vivo tissue replacements from a variety of cell types. As stem cells are particularly sensitive to environmental cues, inappropriate cell-cell and cell-matrix interactions likely lead to the irreversible and undesirable alterations in stem cell differentiation fate found in culture. The systems developed in this project will allow us to investigate the specific role of vascular cells and osteoprogenitors/osteoblasts in maintaining the human HSC niche, which is a difficult question to address in vivo. It is crucial to better define and create models of the niche to understand normal hematopoiesis and pathologies involving blood cells, and to enable hematopoiesis on demand in various therapeutic venues. The key studies to date on this topic have relied on rodent models, and the relevance of many findings to human biology is currently unclear. (End of Reviewers' Comment)