Interactions between the ADORA2b/Sphk1axis and the AE1-Hb switch in red blood cell aging in vivo and in vitro

ADORA2b/Sphk1axis 和 AE1-Hb 开关在体内和体外红细胞老化中的相互作用

• 批准号: 10369002
• 负责人: Angelo D'Alessandro
• 金额: $ 63.65万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-15 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10369002
• 关键词: Acclimatization; Adenosine; Adenosine A2B Receptor; Affect; Affinity; Age; Aging; Aldehyde-Lyases; Altitude; American; Animal Model; Anions; Antigens; Antioxidants; Binding; Biochemical; Blood; Blood Banks; Blood Circulation; Blood Transfusion; Blood donor; Cell Aging; Cell Nucleus; Cell Survival; Cells; Cellular Metabolic Process; Cellular biology; Chlorides; Complex; Cytolysis; Cytosol; Data; Disease; Docking; Energy Metabolism; Enzymes; Erythrocyte Transfusion; Erythrocytes; Exposure to; Face; Functional disorder; Gene Expression Regulation; Glucose; Glucosephosphate Dehydrogenase; Glucosephosphate Dehydrogenase Deficiency; Glyceraldehyde-3-Phosphate Dehydrogenases; Glycolysis; Health; Hemoglobin; Hospitals; Hour; Human; Human body; Hypoxia; In Vitro; Individual; Intervention; Iron; Life; Logistics; Longevity; Lung; Medical; Membrane; Metabolic; Metabolic stress; Metabolism; Modification; Morphology; Mus; N-terminal; NADH; NADP; Organelles; Oxidation-Reduction; Oxidative Stress; Oxygen; Pathologic; Pathology; Pathway interactions; Patients; Pentosephosphate Pathway; Peripheral; Persons; Plasma; Play; Predisposition; Procedures; Proteins; Proteomics; Reaction; Role; SPHK1 enzyme; Savings; Sea; Series; Sickle Cell Anemia; Signal Transduction; Site; Technology; Tissues; Translations; Vaccination; age related; base; cell age; cofactor; deoxyhemoglobin; diphosphoglycerate; gene product; in vivo; mouse model; normoxia; novel; oxidant stress; oxidative damage; repaired; response; senescence; sensor; tool

ABSTRACT Red Blood Cells (RBCs) represent ~83% of the total human cells in the body. RBC hemoglobin (Hb), which is critical for their function to carry and deliver oxygen to peripheral tissues, constitutes ~90% of the total protein content of a mature RBCs. During their lifespan of 120 days in the bloodstream, RBCs are constantly exposed to oxidant stress, which mostly arises from Fenton and Haber-Weiss reactions triggered by Hb-Iron in the presence of oxygen. However, RBCs lack nuclei and organelle and, as such, they cannot synthesize new proteins to replace oxidatively damaged components. Therefore, in order to cope with oxidant stress, RBCs have evolved unique mechanisms that rely on signaling axes and their capacity to trigger metabolic reprogramming to favor antioxidant defenses (the Pentose Phosphate Pathway – PPP) over energy metabolism (glycolysis). One such mechanism relies on the two most abundant proteins in RBC cytosols and membranes: Hb and anion exchanger 1 (AE1), respectively. Owing to its capacity to “sense” oxygen, in response to hypoxia, deoxygenated Hb migrates to the membrane, where it binds the N-terminus of AE1. This mechanism releases a series of glycolytic enzymes, which are inhibited under high-oxygen tensions owing to their binding to the same region of AE1 with high affinity for deoxygenated Hb. This phenomenon favors energy metabolism under low oxygen tensions (e.g., high-altitude hypoxia), while it creates a metabolic bottleneck in energy metabolism to promote a critical antioxidant pathway when oxidant stress is high: the Pentose Phosphate Pathway (PPP). This mechanism is referred to as the AE1-Hb switch in this proposal. Of note, glucose 6-phosphate dehydrogenase (G6PD) is not only the rate-limiting enzyme of the PPP, but also the target of the most common enzymopathy in humans, G6PD deficiency, which affects ~400 million people. While RBCs from G6PD-deficient subjects are perfectly healthy in the absence of oxidant stress, RBCs from these individuals are characterized by a shorter lifespan and susceptibility to lysis following oxidative insults. Oxidative stress to RBCs is not only relevant within the context of RBC senescence. Significant oxidant stress arises during RBC storage under blood bank conditions for blood transfusion purposes, a common in hospital medical procedure and a life-saving intervention for ~3-4 million Americans every year. However, little is known about the impact of the AE1-Hb switch and G6PD deficiency in the context of RBC aging in vitro (blood bank). In parallel, when studying human acclimatization to high-altitude hypoxia, we discovered a novel axis, the ADORA2b/Sphk1 axis, that interplays with the AE1-Hb switch to favor oxygen off-loading in healthy individuals as they climb to high-altitude, where oxygen is limited in comparison to sea level. By leveraging a combination of state-of-the art omics technologies (fluxomics and Xlinking proteomics) and a mix of well-established and novel animal models (exclusively developed for this proposal – e.g., G6PD-def mice) we will investigate the interplay of the AE1-Hb switch and ADORA2b/Sphk1 in the context of oxidant stress and hypoxia during RBC aging in vivo (senescence) and in vitro (blood bank).

摘要 红细胞（RBC）占人体总细胞的约83%。RBC血红蛋白（Hb）， 对于它们携带氧气并将氧气输送到外周组织的功能至关重要，占总蛋白质的约90 成熟红细胞的含量。在血液中120天的生命周期中，红细胞不断暴露于 氧化应激，这主要是由于芬顿和Haber-Weiss反应引发的Hb-铁在 氧气的存在。然而，红细胞缺乏细胞核和细胞器，因此，它们不能合成新的 蛋白质，以取代氧化损伤的成分。因此，为了应对氧化应激，红细胞 进化出了依赖于信号轴的独特机制及其触发代谢重编程的能力 有利于抗氧化防御（戊糖磷酸途径-PPP）超过能量代谢（糖酵解）。 其中一种机制依赖于红细胞胞浆和膜中最丰富的两种蛋白质：血红蛋白和阴离子 交换器1（AE1）。由于其能够"感知"氧气，在缺氧时， Hb迁移到膜上，在那里它结合AE1的N-末端。这种机制释放出一系列 糖酵解酶，这是抑制下高氧张力，由于它们的结合相同的区域， AE1对脱氧血红蛋白具有高亲和力。这种现象有利于低氧条件下的能量代谢 张力（例如，高海拔缺氧），同时它在能量代谢中产生代谢瓶颈，以促进 氧化应激高时的关键抗氧化途径：戊糖磷酸途径（PPP）。这 该机制在本建议中被称为AE1-Hb开关。值得注意的是，葡萄糖6-磷酸脱氢酶 葡萄糖-6-磷酸脱氢酶（G6PD）不仅是PPP的限速酶，也是PPP最常见的酶病的靶标， G6PD缺乏症，影响约4亿人。虽然来自G6PD缺陷受试者的RBC 在没有氧化应激的情况下完全健康，来自这些个体的RBC的特征在于较短的 寿命和氧化损伤后对溶解的敏感性。红细胞的氧化应激不仅与 RBC衰老的背景。在血库下储存红细胞期间出现显著的氧化应激 输血的条件，一种常见的医院医疗程序和挽救生命的干预措施 每年约有3 - 4百万美国人。然而，关于AE1-Hb开关和G6PD的影响知之甚少。 在体外RBC老化的背景下缺乏（血库）。与此同时，当研究人类对 在高原缺氧中，我们发现了一个新的轴，ADORA2b/Sphk1轴，它与AE1-Hb相互作用， 当健康人爬到高海拔时，他们会转向有利于氧气的卸载，因为那里的氧气是有限的。 与海平面相比。通过利用最先进的组学技术（通量组学和 Xlinking蛋白质组学）和成熟的新型动物模型（专门为此开发）的混合 建议-例如，G6PD-def小鼠），我们将研究AE1-Hb开关和ADORA2b/Sphk1的相互作用， 体内（衰老）和体外（血库）RBC老化期间氧化应激和缺氧的背景。