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 该项目的目的是为我们代表性不足的研究生提供NIH R15收获研究机会 北德克萨斯州西班牙裔服务学院UT阿灵顿的学生。在急性缺血再灌注期间 （I/R）在再灌注阶段产生活性氧（ROS），并导致灾难性 伤害心脏。那就是抗氧化剂以预防或改善ROS确定的施用 效果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，一直以来 显示可防止心脏中的I/R伤害。但是锌的安全配方，剂量和递送模式具有 未建立。以前，锌剂量产生有毒作用（例如，停止生长，细胞死亡），并且 锌作为离子机跨质膜的递送模式产生了许多副作用（例如， 膜损伤，细胞死亡等）。我们的长期目标是建立一种治疗性锌疗法以保护 对I/R伤害的心脏。为了进行此类人类临床试验，我们必须最初解决两个基本问题 问题：除锌转运蛋白外，哪些实体调节锌细胞中的锌影响？可以增强 这种固有的实体无需使用副作用易于副作用的锌离子载体，可以保护心肌细胞免受I/R的影响 受伤？为了解决这两个问题，我们进行了公正的基因组规模CRISPR/CAS9 筛选以搜索此类新实体。出乎意料的是，最高验证的候选人名单上的基因是 SLC5A3，也称为SMIT1，一种肌醇钠肌醇转运蛋白。 SLC5A3从未与锌链接 代谢。我们的初步数据表明，SLC5A3的敲低否定了锌诱导的HL-的保护 1个缺氧 /重氧（H /R）损伤中的1个心肌细胞。有趣的是，缺氧后调节（Postc） H/R损伤时HL-1细胞的生存增强，其有益效果在SLC5A3-KNOCKDOWN中降低 细胞。此外，带有锌灌注的Postc保护心脏（就凋亡减少而言，保持左侧 超声心动图研究中的心室射血分数）来自I/R诱导的小鼠心脏功能障碍。这 锌在培养的心肌细胞和整个心脏中的保护功能被SLC5A3钝化 抑制剂菲洛津。基于这些数据，我们假设SLC5A3介导了锌的影响 心肌细胞可以增强以保护心脏免受I/R损伤。两个具体目标是 伪造来检验此假设：1）定义SLC5A3介质锌的分子机制 心肌细胞的影响； 2）通过针对SLC5A3介导的锌的影响进行概念证明研究 保护心脏免受I/R伤害。该创新包括：（i）首次展示未认识到的作用的研究 SLC5A3在调节锌稳态方面； （ii）鉴定SLC5A3-的治疗潜力 介导的锌影响心脏I/R损伤。作为骨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，出色的资源来培训医生心血管的学生 生理。