Zinc Protection Against Ischemia-Reperfusion Injury in Heart

锌可预防心脏缺血再灌注损伤

• 批准号: 10652915
• 负责人: ZUI PAN
• 金额: $ 44.3万
• 依托单位: UNIVERSITY OF TEXAS ARLINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10652915
• 关键词: Acute; Address; Antioxidants; Apoptosis; Area; Award; Biochemistry; CRISPR screen; CRISPR/Cas technology; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cardiovascular Physiology; Cell Death; Cell Survival; Cell membrane; Cells; Clinical Trials; Confocal Microscopy; Data; Doctor of Philosophy; EFRAC; Echocardiography; Formulation; Genes; Genetic; Genetic Transcription; Goals; Growth; Health; Heart; Heart failure; Hispanic; Homeostasis; Human; Hypoxia; Individual; Infarction; Inflammation; Injury; Ionophores; Ischemia; Laboratories; Left ventricular structure; Link; Mammalian Cell; Mediating; Membrane; Metabolism; Metallothionein; Methods; Minority; Molecular; Monitor; Mus; Muscle; Muscle Cells; Myocardial Ischemia; Myocardial dysfunction; Natural regeneration; Necrosis; Nutrient; Osmolar Concentration; Oxidative Stress; Perfusion; Phase; Phlorhizin; Proteins; Quantitative Reverse Transcriptase PCR; Reactive Oxygen Species; Regimen; Regulation; Reperfusion Injury; Reperfusion Therapy; Reporting; Research; Resources; Risk; Role; Sarcolemma; Signal Pathway; Sodium; Stroke; Students; Techniques; Testing; Texas; Therapeutic; Tissues; Toxic effect; Training; United States National Institutes of Health; Validation; Western Blotting; Zinc; Zinc supplementation; bone; candidate validation; cardioprotection; career; cell growth regulation; college; conditioning; doctoral student; dosage; genetic manipulation; genome-wide; graduate student; heart damage; in vivo; inhibitor; innovation; knock-down; member; myoinositol; overexpression; pharmacologic; post stroke; prevent; programs; protective effect; reverse genetics; screening; side effect; temporal measurement; undergraduate student; whole genome; zinc-binding protein

Project Summary This project is aimed to provide NIH R15 REAP research opportunities for our underrepresented graduate students at UT Arlington, a Hispanic-Serving Institute in North Texas. During acute ischemia-reperfusion (I/R), reactive oxygen species (ROS) is generated at the reperfusion phase and results in catastrophic damage to heart. Thus, the administration of antioxidant agents to prevent or ameliorate ROS detrimental effects is an active research area. The nutrient, zinc, has multifaceted antioxidant effects and has been shown to protect against I/R injury in heart. But a safe formulation, dosage, and delivery mode of zinc have not been established. Previously, zinc dosages yielded toxic effects (e.g., halted growth, cell death), and the delivery mode of zinc as an ionophore across plasma membrane produced many side-effects (e.g., membrane damage, cell death, etc.). Our long-term goal is to establish a therapeutic zinc regimen to protect heart against I/R injury. To progress to such human clinical trials, we must initially address two fundamental questions: What entity besides zinc transporters regulates zinc influx in muscle cells? Can enhancement of such intrinsic entity, without using side effect-prone zinc ionophore, protect the cardiomyocytes from I/R injury? Toward addressing the two questions, we performed unbiased genome-scale CRISPR/Cas9-based screening to search for such new entity. Unexpectedly, a gene on the top validated candidate list was SLC5A3, also called SMIT1, a sodium myo-inositol transporter. SLC5A3 has never been linked to zinc metabolism. Our preliminary data showed that knockdown of SLC5A3 negated zinc-induced protection in HL- 1 cardiomyocytes in hypoxia /reoxygenation (H/R) injury. Interestingly, hypoxic post-conditioning (PostC) enhanced HL-1 cell survival upon H/R injury, and its beneficial effect was diminished in SLC5A3-knockdown cells. Moreover, PostC with zinc perfusion protect the heart (in terms of reduced apoptosis, maintaining left ventricle ejection fraction in echocardiography study) from I/R induced cardiac dysfunction in mice. The protective function of zinc in both cultured cardiomyocytes and whole heart was blunted by a SLC5A3 inhibitor phloridzin. Based on these data, we hypothesize that SLC5A3 mediates zinc influx in cardiomyocytes, which can be enhanced to protect heart from I/R injury. Two specific aims are formulated to test this hypothesis: 1) to define the molecular mechanisms by which SLC5A3 mediates zinc influx in cardiomyocytes; 2) to conduct proof-of-concept study by targeting SLC5A3-mediated zinc influx to protect heart from I/R injury. The innovation includes: (i) the first study to show an unrecognized role of SLC5A3 in regulating zinc homeostasis; (ii) the identification of the therapeutic potential of SLC5A3- mediated zinc influx for heart I/R injury. As one of six founding members in Bone-Muscle Research Center at UTA, the PI’s laboratory has required expertise, excellent resources to train PhD students in cardiovascular physiology.

项目摘要 该项目旨在为我们代表性不足的研究生提供NIH R15 REAP研究机会 德州大学阿灵顿分校的学生，这是一所位于德克萨斯州北部的西班牙裔服务学院。在急性缺血再灌注期间 在缺血再灌注（I/R）中，活性氧（ROS）在再灌注阶段产生，并导致灾难性的缺血再灌注。 对心脏的伤害。因此，施用抗氧化剂以预防或改善有害的ROS， 影响是一个活跃的研究领域。营养素锌具有多方面的抗氧化作用， 保护心脏免受I/R损伤。但安全的配方，剂量和锌的输送方式， 尚未建立。以前，锌剂量产生毒性作用（例如，停止生长，细胞死亡）， 锌作为离子载体穿过质膜的递送模式产生许多副作用（例如， 膜损伤、细胞死亡等）。我们的长期目标是建立一个治疗锌方案，以保护 心脏对抗I/R损伤为了进行这样的人体临床试验，我们必须首先解决两个基本问题。 问题：除了锌转运蛋白外，还有什么物质调节肌肉细胞中锌的流入？可以增强 这样的内在实体，不使用副作用倾向的锌离子载体，保护心肌细胞免受I/R 受伤？为了解决这两个问题，我们进行了基于CRISPR/Cas9的公正基因组规模的研究 筛选以寻找这种新实体。出乎意料的是，在最有效的候选名单上的一个基因， SLC5A3，也称为SMIT1，是一种钠肌醇转运蛋白。SLC5A3从未与锌有关 新陈代谢.我们的初步数据表明，SLC5A3的敲低否定了锌诱导的HL-60细胞保护作用。 1心肌细胞缺氧/复氧（H/R）损伤。有趣的是，低氧后处理（PostC） 增强HL-1细胞在H/R损伤后的存活，并且其有益作用在SLC5A3敲低时减弱 细胞此外，PostC与锌灌注保护心脏（在减少细胞凋亡，维持左心室功能方面）。 超声心动图研究中的心室射血分数）从I/R诱导的小鼠心功能不全中获得。的 锌在培养的心肌细胞和整个心脏中的保护功能被SLC 5A3减弱 抑制剂根皮苷。基于这些数据，我们假设SLC5A3介导锌内流， 心肌细胞，其可以增强以保护心脏免受I/R损伤。两个具体目标是 为了验证这一假设，我们制定了以下方法：1）确定SLC5A3介导锌的分子机制 2）通过靶向SLC5A3介导的锌内流进行概念验证研究， 保护心脏免受I/R损伤。创新包括：（一）第一项研究表明， SLC5A3在调节锌稳态中的作用;（ii）鉴定SLC5A3的治疗潜力， 介导的锌内流对心脏I/R损伤的影响。作为骨骼肌肉研究中心的六个创始成员之一， UTA，PI的实验室需要专业知识，优秀的资源来培养心血管领域的博士生 physiology.