The Roles of Lipid Metabolism in the Maintenance of Hematopoietic Stem Cells

摘要 我们研究的一个主要目标是确定指导造血干细胞的关键代谢途径。 (HSC)命运决定。虽然早期的发现表明HSCs主要依赖于糖酵解，但新的证据 来自我们实验室和其他实验室的研究表明，线粒体代谢，特别是脂肪酸氧化，是 对HSC命运的决定至关重要。我们假设线粒体的新陈代谢在启动时被重塑。 的命运选择过程，以满足不断变化的需求，适当的HSC功能。然而，我们对此的理解 HSC自我更新和脂质代谢之间的关系仍然有限。为了鉴定代谢物- 依赖途径，我们已经使用了一种适应的面向基因表达的生物信息学工具和我们自己的 代谢组学分析。我们还建立了一种生物传感器来评估脂肪酸的氧化活性。 活细胞，以确定与肝星状细胞受控平衡相关的代谢模式。量化 实时成像和我们的单细胞方法将阐明对称或不对称的过程 HSC分裂过程中线粒体的分离。我们创新的本地移植系统将使我们能够 监测单个HSCs在体内的迁移和细胞分裂，以及我们建立的图像引导技术 直接从活体动物的骨髓中提取单个细胞的微吸管将使后续 单细胞分析。对结果数据的分析将对造血干细胞的命运决定过程产生新的见解， 并促进开发新的治疗策略，将HSCs的分裂平衡转向自我 通过新陈代谢操作进行更新。这项提议的目标有三个：(1)在目标1中，我们将诱导 选择性消耗线粒体中的代谢物以识别脂肪酸的代谢靶点 影响HSC命运的代谢；(2)在目标2中，我们将使用关键基因的药物或遗传调节 影响脂肪酸氧化或其下游靶点以定义线粒体之间的代谢串扰 以及(3)在目标3中，我们将评估产生HSC分裂对称性的协调过程 以及对分裂平衡的分析，将为深入了解体内脂肪酸代谢与 HSC命运选择。如果成功，拟议的研究将通过提供更深层次的 理解协调HSC命运决定的代谢线索，并将提出潜在的方法 通过代谢调控使HSCs分裂平衡向自我更新方向转变以改善临床 移植后的结果。

ABSTRACT A leading goal of our research is identification of the key metabolic pathways directing hematopoietic stem cell (HSC) fate decisions. While early findings suggested that HSCs depend mainly on glycolysis, emerging evidence from our lab and others has shown that mitochondrial metabolism, and particularly fatty acid oxidation, is essential to HSC fate determination. We hypothesize that mitochondrial metabolism is remodeled at the initiation of the fate choice process to meet the changing needs of proper HSC function. However, our understanding of the relationship between HSC self-renewal and lipid metabolism remains limited. To identify metabolite- dependent pathways, we have used an adapted gene expression-oriented bioinformatics tool and our own metabolomics analyses. We have also established a biosensor for assessment of fatty acid oxidation activity in live cells to determine the metabolic modes which are relevant to the controlled equilibrium of HSCs. Quantitative live imaging and our single cell approaches will illuminate the processes of symmetric or asymmetric mitochondrial segregation during HSC division. Our innovative local transplantation system will allow us to monitor the migration and cell divisions of single HSCs in vivo, and our established image-guided technique of micropipette aspiration of individual cells directly from the bone marrow of live animals will enable subsequent single-cell assay. Analysis of the resulting data will yield new insights into the fate decision process of HSCs, and facilitate the development of new therapeutic strategies for shifting the division balance of HSCs toward self- renewal through metabolic manipulation. The goals of this proposal are three-fold: (1) In Aim 1, we will induce the selective consumption of metabolites localized in the mitochondria to identify metabolic targets of fatty acid metabolism that affect HSC fate; (2) in Aim 2, we will use pharmacological or genetic modulation of key genes impacting fatty acid oxidation or its downstream targets to define the metabolic crosstalk between mitochondria and the cytosol; and (3) in Aim 3 we will evaluate the coordinated process that yields HSC division symmetry in vivo, and analysis of division balance will provide insights into the in vivo relevance of fatty acid metabolisms to HSC fate choice. If successful, the proposed research will positively impact the field by providing a deeper understanding of the metabolic cues coordinating HSC fate decisions, and will suggest potential methods of shifting the division balance of HSCs toward self-renewal through metabolic manipulation to improve clinical outcomes after transplantation.