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)占人体细胞总数的83%。红细胞血红蛋白(Hb)，这是 对它们向外周组织运送和输送氧气的功能至关重要，占总蛋白的90% 一个成熟的红细胞的含量。红细胞在血液中的寿命为120天，不断地暴露在 氧化应激，主要由Hb-Fe引发的Fenton和Haber-Weiss反应引起 氧气的存在。然而，红细胞缺乏细胞核和细胞器，因此不能合成新的 取代氧化损伤成分的蛋白质。因此，为了应对氧化应激，红细胞已经 进化出独特的机制，依赖于信号轴及其触发代谢重新编程的能力 支持抗氧化剂防御(磷酸戊糖途径-PPP)而不是能量代谢(糖酵解)。 其中一种机制依赖于红细胞胞浆和细胞膜中含量最丰富的两种蛋白质：Hb和阴离子。 交易所1(AE1)。由于它的“感觉”氧气的能力，在低氧的反应下，脱氧 Hb迁移到膜上，在那里它与AE1的N末端结合。这一机制释放了一系列 糖酵解酶，在高氧压力下被抑制，因为它们与 对脱氧Hb具有高亲和力的AE1。这种现象有利于低氧条件下的能量代谢。 紧张(例如，高海拔低氧)，虽然它在能量代谢中造成代谢瓶颈，以促进 氧化应激高时的关键抗氧化途径：磷酸戊糖途径(PPP)。这 该机制在本提案中称为AE1-HB交换机。值得注意的是，葡萄糖6-磷酸脱氢酶 (G6PD)不仅是PPP的限速酶，也是PPP中最常见的酶病靶点。 人类，G6PD缺乏症，影响到约4亿人。而来自G6PD缺陷受试者的红细胞是 在没有氧化应激的情况下非常健康，这些个体的红细胞特征是较短 寿命和对氧化侮辱后溶解的敏感性。氧化应激对红细胞的影响不仅与 红细胞衰老的背景。红细胞在血库下储存过程中会产生明显的氧化应激 输血条件，医院常见的医疗程序和救命干预措施 每年约有300-400万美国人。然而，人们对AE1-HB开关和G6PD的影响知之甚少 红细胞体外老化背景下的缺陷(血库)。同时，在研究人类适应时 在高原缺氧时，我们发现了一个新的轴，ADORA2b/Sphk1轴，它与AE1-Hb相互作用 在健康的个体攀登到高海拔时，转向支持氧气卸载，在那里氧气被限制在 与海平面相比。通过利用最先进的组学技术(通量组学和 Xlink蛋白质组学)和成熟的和新的动物模型的组合(专门为此而开发 建议--例如，G6PD-def小鼠)我们将研究AE1-Hb开关和ADORA2b/Sphk1在 体内(衰老)和体外(血库)红细胞老化过程中氧化应激和缺氧的背景。