The role of glycolysis and glucose oxidation in hematopoiesis

糖酵解和葡萄糖氧化在造血中的作用

• 批准号: 10340134
• 负责人: Michalis Agathocleous
• 金额: $ 40.36万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10340134
• 关键词: Ablation; Acetyl Coenzyme A; Address; B-Lymphocytes; Biochemical; Bone Marrow; Bone Marrow Cells; Carbon; Cell Respiration; Cells; Cellular Metabolic Process; Cellular biology; Citric Acid Cycle; Dependence; Embryo; Enzymes; Equilibrium; Erythropoiesis; Fermentation; Frequencies; Gatekeeping; Genetic; Glucose; Glutathione; Glycolysis; Hematopoiesis; Hematopoietic; Hematopoietic System; Hematopoietic stem cells; Homeostasis; Impairment; In Vitro; Individual; Isotopes; Label; Lactate Dehydrogenase; Measures; Mediating; Metabolic; Metabolic Pathway; Metabolism; Methods; Myelogenous; Myeloid Cells; NADH; Natural regeneration; Nature; Neurons; Nutrient; Oxidation-Reduction; Pathway interactions; Positioning Attribute; Production; Protein Isoforms; Pyruvate; Role; Specificity; Stable Isotope Labeling; T-Lymphocyte; Testing; Textbooks; Thymus Gland; Tissues; Work; cell type; experimental study; flexibility; glucose metabolism; hematopoietic differentiation; in vivo; innovation; metabolome; metabolomics; mouse model; nerve stem cell; novel; nutrient metabolism; oxidation; progenitor; pyruvate dehydrogenase; stable isotope; stem cell function; stem cells; thymic regeneration

PROJECT SUMMARY: The metabolism of nutrients has been studied using unfractionated tissues, or in vitro. An unresolved question is how nutrients are metabolized by stem cells in vivo. Our understanding of stem cell metabolism has been limited by the fact that metabolomics typically requires millions of cells, while stem cells are rare. We developed methods to profile the metabolome and to trace stable isotope labeled nutrients in hematopoietic stem cells (HSCs) and other rare cell types purified from tissues. We found that T cell progenitors in the thymus are glucose avid as compared to HSCs, myeloid and B cell restricted progenitors, in contrast to the prevailing view that HSCs are more glycolytic than hematopoietic progenitors. Stable isotope tracing experiments showed that in the bone marrow but not the thymus, glycolysis and the TCA cycle are disconnected. Hematopoietic loss of pyruvate dehydrogenase (PDH), the gatekeeper enzyme that connects glycolysis to the TCA cycle, reduced the number of double positive (DP) T cell progenitors but did not affect HSCs or other hematopoietic cell types. Loss of PDH paradoxically did not impair the TCA cycle in the thymus, but caused accumulation of pyruvate and aberrant redox balance. Cells which do not oxidize glucose in the TCA cycle are classically thought to ferment glucose through glycolysis to lactate via lactate dehydrogenase (LDH). Hematopoietic loss of LDHA, one of the two LDH isoforms, impaired development of erythroid progenitors but not HSCs, T cell progenitors or other restricted hematopoietic progenitors. The cell type specificity in the requirement of LDH and PDH in the hematopoietic system raises the question of why different stem or progenitor cell types choose to use LDH-mediated fermentation or PDH-mediated oxidation in vivo. This application’s objective is to systematically dissect the role of glycolytic as compared to oxidative metabolism in HSCs and restricted progenitors. Our hypothesis is that T cell progenitors require oxidation of glucose via PDH to regulate pyruvate levels and redox homeostasis, in contrast to HSCs, myeloid and B cell progenitors which are metabolically flexible. In Aim 1 we will test the metabolic mechanisms which mediate the effects of PDH on DP cells. In Aim 2 we will determine the cellular and metabolic effects of blocking LDHA/B or PDH alone or in combination in HSCs and restricted progenitors. In Aim 3 we will investigate the role of LDHA/B and PDH in hematopoietic and thymopoietic regeneration. These experiments will identify the contribution of glucose to metabolite pools in HSCs and progenitors in vivo, systematically test the idea that HSCs are glycolytic, and identify mechanisms by which central carbon metabolism regulates hematopoietic differentiation and regeneration. More generally our experiments will address a fundamental metabolic question by testing if stem or progenitor cells in vivo switch between glucose fermentation or oxidation, as is the textbook view, or if some cell types in vivo tolerate the loss of both major glucose catabolic pathways.

项目概要： 营养物质的代谢已经使用未分级的组织或在体外进行了研究。一个悬而未决的问题 是干细胞在体内如何代谢营养物质。我们对干细胞代谢的理解 代谢组学通常需要数百万个细胞，而干细胞很少，这一事实限制了这一点。我们 开发的方法来分析代谢组和追踪稳定同位素标记的营养素在造血 干细胞（HSC）和其他从组织中纯化的稀有细胞类型。我们发现T细胞祖细胞在 与造血干细胞、骨髓和B细胞限制的祖细胞相比，胸腺是葡萄糖亲合的， 普遍认为HSC比造血祖细胞更糖酵解。稳定同位素示踪 实验表明，在骨髓而不是胸腺中，糖酵解和TCA循环是 切断了.丙酮酸脱氢酶（PDH）的造血损失， 糖酵解的TCA循环，减少了双阳性（DP）T细胞祖细胞的数量，但不影响 HSC或其他造血细胞类型。矛盾的是，PDH的缺失并没有损害胸腺中的TCA循环， 但引起丙酮酸积累和异常氧化还原平衡。不氧化葡萄糖的细胞， 传统认为TCA循环通过乳酸脱氢酶将葡萄糖通过糖酵解发酵成乳酸 （LDH）。LDHA是LDH的两种亚型之一，其造血功能丧失损害了红系细胞的发育， 造血祖细胞，但不是HSC、T细胞祖细胞或其他限制性造血祖细胞。的细胞类型 在造血系统中LDH和PDH需求的特异性提出了为什么不同的问题 干细胞或祖细胞类型选择在体内使用LDH介导的发酵或PDH介导的氧化。 本申请的目的是系统地剖析糖酵解的作用相比，氧化 HSC和限制性祖细胞的代谢。我们的假设是，T细胞祖细胞需要氧化 与HSC、骨髓和B细胞相比， 代谢灵活的祖细胞。在目标1中，我们将测试介导 PDH对DP细胞的影响。在目的2中，我们将确定阻断LDHA/B或LDHA/B抑制剂对细胞和代谢的影响。 在HSC和限制性祖细胞中单独或组合的PDH。在目标3中，我们将研究 LDHA/B和PDH在造血和胸腺再生中的作用这些实验将确定 葡萄糖对体内HSC和祖细胞中代谢物库的贡献，系统地测试了以下想法， HSC是糖酵解的，并确定了中心碳代谢调节造血的机制。 分化和再生。更一般地说，我们的实验将解决一个基本的代谢问题， 通过测试体内的干细胞或祖细胞是否在葡萄糖发酵或氧化之间切换， 教科书的观点，或者如果体内某些细胞类型耐受两种主要葡萄糖分解代谢途径的丧失。