The Metabolic Regulation of Hematopoietic Stem Cell Function

项目摘要/摘要 一个根本的问题是，体内代谢物水平的生理变化是否会影响干细胞的功能 或者组织再生。由于技术上的原因，我们对躯体干细胞代谢的了解仍然有限 与研究体内稀有细胞代谢相关的挑战。为了解决这个问题，我们优化了 代谢组学方法的敏感性，使稀有细胞群体的分析成为可能。我们使用这种方法来 比较造血干细胞(HSCs)和多种限制性干细胞之间的代谢产物水平 小鼠骨髓中分离的造血祖细胞。我们发现每个造血干细胞和 祖细胞群体具有明显的新陈代谢特征。人和小鼠的造血干细胞区别于 大多数其他造血细胞都含有异常高水平的抗坏血酸(维生素C)。抗坏血酸在小鼠体内的耗竭， 在5%的美国人中观察到的水平，增加了HSC的频率和功能，部分是通过减少 TET2，一种抑制白血病发展的胞嘧啶去甲基酶。抗坏血酸消耗，比如TET2缺失， 与Flt3ITD合作，促进骨髓生成和白血病生成。抗坏血酸以细胞自主方式作用于 主要通过TET2依赖机制负向调节HSC功能和骨髓生成。这些 观察结果可能与公共健康有关，因为美国人的血浆抗坏血酸水平差异很大。 这在很大程度上是由于饮食差异。在任何时候，13%的美国人都被认为是抗坏血酸缺乏。在AIM 1，我们建议测试抗坏血酸水平是否影响稳态造血或在 造血压力。在目标2中，我们建议测试抗坏血酸耗竭是否促进克隆 在稳定状态下或对造血压力作出反应时的造血作用。克隆性造血术 最近发现，不确定的潜势在健康的老年人和患者中都很常见。 患有再生障碍性贫血、实体癌和接受过造血移植的患者。.的存在 克隆性造血与包括心血管疾病在内的不良健康后果有关。克隆性 造血通常由TET2的一个等位基因丢失或DNMT3A功能丧失突变引起。我们 抗坏血酸耗竭可通过以下途径促进克隆性造血的发生和发展 降低TET2功能。在目标3中，我们将评估抗坏血酸消耗和 TET2缺乏对HSC功能和骨髓生成有调节作用。我们希望这些研究能扩大我们的理解 代谢如何调节HSC表观基因组、HSC功能、正常造血和克隆 造血术。

PROJECT SUMMARY/ABSTRACT A fundamental question is whether physiological variations in metabolite levels in vivo influence stem cell function or tissue regeneration. Our understanding of somatic stem cell metabolism remains limited due to the technical challenges associated with studying metabolism in rare cells in vivo. To address this issue we optimized the sensitivity of metabolomics methods to enable the analysis of rare cell populations. We used this approach to compare metabolite levels between hematopoietic stem cells (HSCs) and a wide range of restricted hematopoietic progenitors isolated from mouse bone marrow. We found that each hematopoietic stem and progenitor cell population has a distinct metabolic signature. Human and mouse HSCs are distinguished from most other hematopoietic cells by unusually high levels of ascorbate (vitamin C). Ascorbate depletion in mice, to a level observed in 5% of Americans, increases HSC frequency and function, partly by reducing the activity of Tet2, a cytosine demethylase that suppresses leukemia development. Ascorbate depletion, like Tet2 deletion, cooperates with Flt3ITD to promote myelopoiesis and leukemogenesis. Ascorbate acts cell-autonomously to negatively regulate HSC function and myelopoiesis mainly through Tet2-dependent mechanisms. These observations are likely relevant to public health as plasma ascorbate levels vary widely among Americans, largely due to dietary differences. At any one time, 13% of Americans are considered ascorbate deficient. In Aim 1, we propose to test whether ascorbate levels influence steady-state hematopoiesis or regeneration after hematopoietic stresses. In Aim 2, we propose to test whether ascorbate depletion promotes clonal hematopoiesis under steady state conditions or in response to hematopoietic stresses. Clonal hematopoiesis of indeterminate potential has recently been shown to be common in healthy older people as well as in patients with aplastic anemia, solid cancers, and patients who have received hematopoietic transplants. The presence of clonal hematopoiesis is associated with adverse health outcomes including cardiovascular disease. Clonal hematopoiesis is usually caused by the loss of one allele of Tet2 or by loss-of-function mutations in Dnmt3a. We hypothesize that ascorbate depletion can promote the development and progression of clonal hematopoiesis by reducing Tet2 function. In Aim 3 we will assess the molecular mechanisms by which ascorbate depletion and Tet2 deficiency regulate HSC function and myelopoiesis. We expect these studies to expand our understanding of how metabolism regulates the HSC epigenome, HSC function, normal hematopoiesis, and clonal hematopoiesis.