Modulating HSC-niche interactions to understand aging and improve transplantation

项目总结/摘要 该建议的目标是：（1）阐明造血（血液形成） 干细胞（HSC）及其微环境中的因子（即，小生境）导致血液中与年龄相关的变化 形成;和（2）确定相关的HSC和生态位机制，增强HSC植入后， 移植更具体地说，我们的目标是实现这些目标，分别：（1）通过操纵Lnk（Sh 2b 3） 在HSC中的表达，以鉴定HSC老化和骨髓旁路分化途径的内在调节因子 (see下文）;和（2）通过鉴定小生境中的“支持者”细胞，以及它们相应的分子机制， 使HSC能够植入。该项目将提供一个机制的基础，制定战略，以治疗年龄- 本发明涉及造血干细胞移植（HSCT）的方法，从而解决两个问题： 重要的医疗需求。HSCT是一种治疗多种血液病（包括 免疫缺陷，自身免疫和白血病），但仍然是危险的，部分原因是必要的预HSCT 骨髓预处理，通常是放疗或化疗。这样的条件作用是必要的，使“空间” 或清除骨髓中供供体HSC移植的小生境，但也会导致显著的发病率。与 总体目标是提高我们对HSC-生态位相互作用的理解，本研究计划：（1）将确定 在衰老过程中维持HSC功能的新策略， 血液系统疾病;和（2）改善HSC移植，可能通过减少对毒性预处理的需要， 条件反射目的1是基于以下发现：（i）Lnk缺陷的HSC不表现出典型的年龄相关的HSC分化。 野生型HSC的骨髓偏好;（ii）在免疫表型定义的HSC区室中， 不同的自我更新的骨髓限制性祖细胞（MyRPs），直接从HSC产生， 骨髓旁路途径;和（iii）MyRPs随年龄增长而增加。通过对LNK缺乏的分析， HSC在MyRP生成方面存在缺陷，我们将确定调节骨髓旁路的机制。 分化通过转录组学分析，我们将鉴定MyRP产生的新的假定调节因子， HSC老化，并通过功能测定证实这些。目的2是基于发现Lnk缺陷的HSC 在HSCT中，即使在没有预处理的情况下，也比野生型HSC具有更大的移植潜力。我们 将鉴定增强HSC植入的非HSC骨髓细胞类型。我们阐明了细胞机制 使用2D和3D显微镜追踪骨髓定植， 移植后。总之，本研究计划将使用LNK缺陷作为一个系统， 了解HSC老化的基本机制，并确定改善HSC移植的策略 植活率。这项研究项目将建立在改善临床移植的长期目标之上 通过更好地理解干细胞功能和干细胞生态位，

PROJECT SUMMARY/ABSTRACT The goals of this proposal are: (1) to elucidate specific mechanisms by which hematopoietic (blood-forming) stem cells (HSCs) and factors within their microenvironment (i.e., niche) lead to age-associated changes in blood formation; and (2) to identify related HSC and niche mechanisms that enhance HSC engraftment after transplantation. More specifically, we aim to achieve these goals, respectively: (1) by manipulating Lnk (Sh2b3) expression in HSCs to identify intrinsic regulators of HSC aging and the myeloid-bypass differentiation pathway (see below); and (2) by identifying “supporter” cells in the niche, and their corresponding molecular mechanisms, that enable HSC engraftment. This project will provide a mechanistic basis to devise strategies to treat age- associated hematologic disorders and improve methods of HSC transplantation (HSCT), thereby addressing two important medical needs. HSCT is a curative therapy for a number of hematological diseases (including immunodeficiencies, autoimmunities, and leukemias) but is still risky, in part due to the necessary pre-HSCT bone marrow conditioning, usually irradiation or chemotherapy. Such conditioning is necessary to make “space” or clear niches in the bone marrow for donor HSCs to engraft, but also causes significant morbidity. With the overall goal of improving our understanding of HSC-niche interactions, this research program: (1) will identify novel strategies to sustain HSC function during aging, potentially preventing or treating age-associated hematologic disorders; and (2) to improve HSC transplantation, potentially by reducing the need for toxic pre- conditioning. Aim 1 is based on findings that: (i) Lnk-deficient HSCs do not exhibit the typical age-associated myeloid bias of wild-type HSCs; (ii) within the immunophenotypically-defined HSC compartment are functionally distinct self-renewing myeloid-restricted progenitor cells (MyRPs) that are directly generated from HSCs through a myeloid-bypass pathway; and (iii) MyRPs increase in frequency with age. Through analysis of Lnk-deficient HSCs, which are defective in MyRP generation, we will identify mechanisms regulating myeloid-bypass differentiation. From transcriptomics analysis, we will identify new putative regulators of MyRP production and HSC aging, and confirm these through functional assays. Aim 2 is based on the finding that Lnk-deficient HSCs have greater engraftment potential than wild-type HSCs in HSCT, even in the absence of preconditioning. We will identify non-HSC bone marrow cell types that enhance HSC engraftment. We elucidate cellular mechanisms that contribute to this “supporter cell” function using 2D and 3D microscopy to trace bone marrow colonization following transplantation. In summary, this research proposal will use Lnk-deficiency as a system to both understand fundamental mechanisms of HSC aging and identify strategies to improve HSC transplantation engraftment rates. This research project will build on the long-term objectives of improving clinical transplantation and healthy aging through better understanding of stem cell function and the stem cell niche.