Regulatory Cell Signals in the Bone Marrow

描述（由申请人提供）：本提案的长期目标是确定对成骨细胞控制HSC至关重要的细胞和分子介质。我们先前已证实甲状旁腺激素（PTH）激活成骨细胞以增加造血干细胞（HSC）数量，并且PTH可提高辐射损伤后HSC的存活率。这些结果为我们提供了一个模型，以确定一种新的治疗方法，通过刺激成骨细胞来增加骨髓医源性或毒性损伤后的HSC。然而，具体的成骨细胞亚群和关键的成骨细胞依赖的分子事件调控HSC是未知的。使用药理学和遗传学模型，我们已经确定Notch信号作为PTH依赖性HSC调节的潜在介质。Notch激活需要细胞结合配体与邻近细胞上的受体直接相互作用。我们证明：1）PTH或其受体的激活刺激成骨细胞中的Notch配体Jagged 1（Jag 1）; 2）在成骨细胞中具有组成性激活PTH受体的小鼠中，HSC具有增加的Notch激活; 3）PTH依赖性HSC的增加通过抑制？分泌酶活性，这是Notch激活所必需的。我们的初步研究表明，Jag 1在成骨细胞中的表达是PTH依赖性HSC扩增所必需的。总之，这些数据表明，PTH通过成骨细胞表达Jag 1来扩增HSC，然后激活邻近骨髓细胞中的Notch信号。基于我们的数据，我们假设造血干细胞的扩张需要骨髓微环境中Jag 1启动的Notch激活。为了验证这一假设，在Aim 1中，我们将定义成骨细胞亚群，其中Jag 1是必要的，足以介导HSC扩增。在Aim 2中，我们将鉴定其中需要Notch激活以实现成骨细胞依赖性HSC扩增的细胞群（HSC、成骨细胞和/或骨髓的其他组分）。最后，在Aim 3中，我们将确定Notch信号传导对PTH的骨髓保护作用的贡献，在这种临床情况下，HSC生态位操作可能是降低发病率和死亡率的新策略。我们已经建立并充分表征了微环境信号增加HSC的体内模型。现在成骨细胞Jag 1已被确定为PTH依赖性HSC扩增的关键因素，我们有前所未有的机会，使用本文提出的体内策略来定义HSC生态位的细胞和分子组成部分。因此，我们的实验目标的完成将为骨髓微环境中的HSC操作定义新的治疗靶点，这可以用于改善骨髓损伤后的存活率。 公共卫生相关性：在这项提案中，我们研究了骨髓微环境对造血干细胞（HSC）的调节。由于HSC产生所有的血细胞，这些调节机制可以在治疗上被利用来增加血细胞损伤或缺乏的特定情况下的HSC。

DESCRIPTION (provided by applicant): The long-term objective of this proposal is to define the cellular and molecular mediators crucial for osteoblastic control of HSCs. We previously established that parathyroid hormone (PTH) activates osteoblastic cells to increase hematopoietic stem cell (HSC) numbers and that PTH improves HSC survival after radiation injury. These results give us a model to define a novel therapeutic approach to increase HSCs following iatrogenic or toxic injury to the bone marrow by stimulating osteoblastic cells. However, the specific osteoblastic cell subsets and the key osteoblastic-dependent molecular events regulating HSCs are unknown. Using pharmacologic and genetic models, we have identified Notch signaling as a potential mediator of PTH-dependent HSC regulation. Notch activation requires direct interaction of cell-bound ligands with receptors on neighboring cells. We demonstrated that 1) PTH or activation of its receptor stimulate the Notch ligand Jagged1 (Jag1) in osteoblastic cells; 2) in mice with constitutively active PTH receptors in osteoblastic cells, HSCs have increased Notch activation; 3) the PTH-dependent HSC increase is blocked by inhibition of ?-secretase activity, which is required for Notch activation. Our preliminary studies now demonstrate that expression of Jag1 in osteoblastic cells is required for the PTH-dependent HSC expansion. Together, these data suggest that PTH expands HSC through osteoblastic expression of Jag1, which then activates Notch signaling in neighboring bone marrow cells. Based on our data, we hypothesize that HSC expansion by osteoblasts requires Jag1-initiated Notch activation in the bone marrow microenvironment. To test this hypothesis, in Aim1 we will define the osteoblastic cell subset in which Jag1 is necessary and sufficient to mediate HSC expansion. In Aim2, we will identify the cell population (HSC, osteoblastic cells and/or other components of the bone marrow) in which Notch activation is required to achieve osteoblastic-dependent HSC expansion. Finally in Aim3 we will determine the contribution of Notch signaling to the myeloprotective effects of PTH, a clinical scenario in which HSC niche manipulation could be a novel strategy to reduce morbidity and mortality. We have already established and fully characterized in vivo models in which microenvironmental signals increase HSCs. Now that osteoblastic Jag1 has been identified as a key element of PTH-dependent HSC expansion, we have the unprecedented opportunity of defining the cellular and molecular components of the HSC niche using the in vivo strategies proposed here. Completion of our experimental aims will thus define novel therapeutic targets for HSC manipulation in the bone marrow microenvironment, which can be exploited to improve survival after bone marrow injury. PUBLIC HEALTH RELEVANCE: In this proposal, we study the regulation of hematopoietic stem cells (HSC) by their bone marrow microenvironment. Since HSC give rise to all blood cells, these regulatory mechanisms could be therapeutically exploited to increase HSC in specific situations of blood cell injury or deficiency.