Mechanisms and treatment of impaired erythrocyte ATP release in older adults

老年人红细胞 ATP 释放受损的机制和治疗

• 批准号: 8979328
• 负责人: Matthew Racine
• 金额: $ 3.4万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8979328
• 关键词: Activities of Daily Living; Adenine Nucleotides; Adenosine Triphosphate; Adult; Age; Aging; Ascorbic Acid; Binding; Biology; Blood; Blood Circulation; Blood Gas Analysis; Blood Vessels; Blood flow; Blood gas; Cardiovascular system; Cells; Cilostazol; Clinical Research; Cyclic AMP; Diamide; Disease; Doppler Ultrasonography; Doppler Ultrasound; Elderly; Electrolytes; Endothelium; Erythrocyte Membrane; Erythrocytes; Exercise; Fatigue; Forearm; Goals; Hemoglobin; Human; Hypertension; Hypoxia; Impairment; In Vitro; Intervention; Lead; Link; Luciferases; Measures; Membrane; Methodology; Mission; Muscle; Outcome; Oxidative Stress; Oxygen; Pathway interactions; Pharmaceutical Preparations; Physiological; Plasma; Property; Purinoceptor; Quality of life; Regulation; Research Priority; Saline; Science; Signal Transduction; Skeletal Muscle; Stimulus; Structure; Techniques; Testing; Time; Tissues; Training; Translating; Vasodilation; cardiovascular disorder risk; cardiovascular risk factor; fasudil; improved; in vivo; indexing; insight; luciferin; new therapeutic target; novel; public health relevance; response; stressor; young adult

 DESCRIPTION (provided by applicant): The adenine nucleotide ATP is released from erythrocytes (RBCs) into the circulation in response to hemoglobin (Hb) deoxygenation and cell deformation, and binds directly to purinergic receptors along the endothelium to evoke vasodilation and increase tissue blood flow and oxygen delivery. Isolated RBC ATP release during Hb deoxygenation, and plasma [ATP] and forearm blood flow (FBF) responses during exercise and systemic hypoxia, are impaired in healthy older vs. young adult humans. However, the underlying cause of these age-associated impairments remains unclear. Thus, the overall aim of this proposal is to identify changes within the RBC that contribute to impaired ATP release and vascular control with advancing age. Specific Aim 1 will test the hypothesis that age-associated changes in RBC structure and function - specifically, membrane deformability, oxidative stress, and cyclic AMP signaling - are mechanistically-linked to impaired ATP release during Hb deoxygenation in isolated RBCs from older adults. To test this, isolated RBC ATP release in response to normoxic (pO2 ~110-120mmHg) and hypoxic (pO2 ~15-25mmHg) stimuli will be quantified in RBCs from young and older adults using the luciferin-luciferase technique after incubation with either saline (control) or pharmacological agents to modify membrane deformability (Y-27632/ diamide), oxidative stress (ascorbic acid), or intracellular [cAMP] (cilostazol). Specific Aim 2 will test the hypothesis that in vivo treatment of impaired RBC ATP release with age will improve isolated RBC ATP release during Hb deoxygenation and in vivo vascular and plasma ATP responsiveness to systemic hypoxia and graded-intensity handgrip exercise in older adults, and that this will be associated with improved exercise capacity. To test this, the treatment from Specific Aim 1 that elicits the greatest improvement in Hb deoxygenation-induced ATP release in RBCs from older adults will be identified, and the corresponding pathway will be targeted using in vivo, systemic administration of either fasudil (deformability), ascorbic acid (oxidative stress), or cilostazol (cAMP signaling), in young and older adults. When peak plasma concentration of the drug is achieved, RBCs will be isolated and Hb deoxygenation-induced ATP release will be quantified as in Specific Aim 1; in conjunction, FBF responses to systemic isocapnic hypoxia (80% SpO2) and graded-intensity handgrip exercise (5%, 15%, 25% MVC) will be quantified using Doppler ultrasound and plasma [ATP] will be measured using the luciferin-luciferase technique. Submaximal rhythmic handgrip exercise time-to-fatigue will be assessed as an index of exercise capacity. The expected outcomes will provide novel insight into the impairments in RBC function and subsequent in vivo vascular control that occur with normal human aging; this insight is relevant to NHLBI's mission and research priorities, particularly those of the Vascular Biology and Hypertension Branch in the Division of Cardiovascular Sciences, given that impaired vascular control with advancing age leads to an increased risk of cardiovascular/ischemic disease and a decline in functional capacity and overall quality of life.

 描述（由适用提供）：腺嘌呤核丁基ATP从红细胞（RBC）释放到循环中，以响应血红蛋白（Hb）脱氧和细胞变形，并直接与沿着内皮的嘌呤能受体结合，以使血管降低和增加组织血液流量和氧气流量。在健康老年人与年轻人中，孤立的RBC ATP释放在HB脱氧过程中，运动和全身性缺氧期间的血浆[ATP]和前臂血流（FBF）反应受损。但是，这些与年龄相关的障碍的根本原因尚不清楚。这是该提案的总体目的是确定RBC内的变化，这些变化会导致ATP释放受损和随着年龄的增长的影响。具体目标1将检验以下假设：RBC结构和功能的年龄相关变化 - 具体来说，膜缺陷性，氧化应激和环状AMP信号传导 - 在来自老年人的分离RBC中的HB脱氧过程中机械地链接以损害ATP释放。为了测试这一测试，将对年轻人和老年人使用荧光素 - 荧光素酶技术在RBC中量化孤立的RBC ATP释放，以响应正常氧（PO2〜110-120mmHg）和低氧（PO2〜15-25mmHg）刺激。应力（抗坏血酸）或细胞内[CAMP]（Cilostazol）。具体目标2将检验以下假设：在HB脱氧过程中，体内治疗RBC ATP释放受损将改善孤立的RBC ATP释放，体内血管和血浆ATP ATP对系统性低氧和对全身性缺氧的反应能力以及老年人的分级强度锻炼，这将与运动能力提高相关。测试 这是从特定目的1的治疗方法中确定的，从老年人中引起的HB脱氧诱导的ATP释放的最大改善将得到确定，并且将使用体内使用Fasudil（可变形性），氧化应激（氧化压力），或Camp signal signalsing（Camp ock and Yound offers），年轻人和年轻人，对相应的途径进行靶向。当达到药物的峰血浆浓度时，将分离RBC，并将Hb脱氧诱导的ATP释放量化为特定目标1中的ATP释放；结合使用Doppler Ultrasound和Clasma [ATP]，将对FBF对全身性异位症缺氧（80％SPO2）和分级强度手式运动（5％，15％，25％MVC）的反应进行定量。次最大的有节奏的手夹锻炼时间将作为运动能力的指数评估。预期的结果将为RBC功能的损伤以及随后在正常人类衰老而发生的体内血管控制中提供新的见解。这种见解与NHLBI的使命和研究重点有关，尤其是心血管科学划分的血管生物学和高血压分支的洞察力，因为鉴于血管控制受损而增长的血管控制受损会导致心血管/缺血性疾病的风险增加，而且功能能力和整体质量和生活质量的下降。