Genetic Mechanisms of Resistance against Cardiac Preconditioning

心脏预适应抵抗的遗传机制

• 批准号: 8141871
• 负责人: Matthias L. Riess
• 金额: --
• 依托单位: CLEMENT J. ZABLOCKI VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8141871
• 关键词: ATP Synthesis Pathway; Affect; African American; Aging; Agonist; Attenuated; Binding Proteins; Blood Vessels; Calcium; Candidate Disease Gene; Cardiac; Cardiac Myocytes; Cardiovascular system; Cell physiology; Chromosomes; Chromosomes, Human, Pair 6; Complement; Complex; Coronary; DNA; Dahl Hypertensive Rats; Disease; Exhibits; Failure; Genes; Genetic; Genetic Models; Genetic Predisposition to Disease; Genotype; Goals; Heart; Individual; Infarction; Injury; Institution; Investigation; Ischemia; Ischemic Preconditioning; K-Series Research Career Programs; Measurement; Measures; Mechanics; Mediating; Medical; Membrane Potentials; Metabolic; Mitochondria; Modeling; Morbidity - disease rate; Muscle Cells; Myocardial; Myocardial Infarction; Myocardial Ischemia; Myocardial tissue; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase; Norway; Oxidation-Reduction; Oxidative Stress; PPAR gamma; Patients; Peroxisome Proliferator-Activated Receptors; Peroxisome Proliferators; Phenotype; Play; Population; Production; Rattus; Rattus norvegicus; Reactive Oxygen Species; Reperfusion Injury; Reperfusion Therapy; Research; Research Project Grants; Resistance; Role; Signal Pathway; Superoxides; System; Techniques; Testing; Tissue Viability; Variant; Ventricular; Veterans; Wisconsin; base; clinical practice; consomic; functional outcomes; helix-loop-helix protein differentiation inhibitor; human NOS3 protein; inhibitor/antagonist; medical schools; mortality; oxidation; preconditioning; prevent; protein expression; receptor; research study; resistance mechanism; resistant strain; salt sensitive; success; tool

DESCRIPTION (provided by applicant): As a VA anesthesiologist and physiologist I study protection against myocardial infarction, a significant medical problem in aging Veterans. In Ischemic Preconditioning (IPC), for example, several shorter ischemic periods before sustained ischemia/reperfusion (IR) attenuate infarction. Various triggering mechanisms have been described for IPC, including nitric oxide (NO) and superoxide formation. Genetic predisposition, however, may be an important confounding factor when trying to transfer it into clinical practice; while some patients may benefit others do not. Profound differences in ischemic tolerance exist not just among but also within certain species. For example, Dahl Salt Sensitive (SS) rats are more susceptible to IR injury than Brown Norways (BN), making them an ideal model to study the genotype of diseases phenotypically similar to African-American patients. In a unique chromosomal substitution (consomic) model constructed at the Medical College of Wisconsin introgression of BN chromosome 6 into SS renders the resulting SS6BN consomic more IR resistant while narrowing the genetic difference to a single chromosome. This offers an excellent starting point to study the genetic basis of cardioprotective strategies like IPC. Although NO has been implicated in decreasing infarct size in BN vs SS without IPC, it is yet unknown if SS can respond to IPC and if resistance to IPC is related to differences in endothelial NO synthase (eNOS) activity possibly modulated by the DNA-binding protein inhibitor Id2 and the peroxisome proliferator-activated receptor 3 (PPAR3). eNOS can produce NO or superoxide. Both can modulate mitochondrial function, which in turn can function as a trigger and effector of IPC. Thus, my overall hypothesis is that failure/success of cardioprotection by IPC is mediated by genes regulating NO production and mitochondrial function. I therefore propose to use this consomic model to study two specific aims: I) Determine if a different genetic background is responsible for differential cellular and mitochondrial protection by IPC, and II) if eNOS and/or its upstream modulators Id2 and PPAR3 are candidate genes responsible for this differential protection. Four hypotheses are tested: 1) Genes on BN chromosome 6 are necessary for IPC as evidenced by better cardiac function and less infarction in BN & SS6BN vs SS. 2) Genes on BN chromosome 6 are necessary for IPC as evidenced by more efficient mitochondrial function in BN & SS6BN vs SS. 3) Protection of cellular and mitochondrial function by IPC in intact hearts depends on the signaling pathway Id2->PPAR3->eNOS->NO modulated by genes on rat chromosome 6. 4) Differential protection of cellular and mitochondrial function in isolated cardiomyocytes depends on NO availability. Approaches for 1 & 2: Various cardiac and mitochondrial functions, NO production and infarct size are measured in intact, beating hearts to assess quantity and quality of genetically determined protection by IPC. Approaches for 3 & 4: Additional beating heart experiments are conducted in the presence of NOS inhibitors, an NO-donor, or a PPAR3 agonist or antagonist, and differences in Id2, PPAR3, and eNOS expression and NO levels and localization are determined. In addition, different mitochondrial functions in the absence or presence of NO are measured in isolated myocytes to determine the role and origin of NO in mediating differential cellular and mitochondrial protection by IPC in this genetic model. Genetic tools are essential to study the role of genetic predisposition. Correlating functional outcomes, infarct size, and mitochondrial functions with IPC, NO levels, protein expressions and genotype will allow us to define the signaling pathway and role of NO and mitochondrial function in cardioprotection and to delineate the subcellular and mitochondrial phenotypes associated with the cardioprotective genotype variation. This CDA marks the indispensable basis for further investigations on the role of genetics in cardioprotection and will be a critical milestone to achieve investigative independence in cardiovascular research at the VA. PUBLIC HEALTH RELEVANCE: As a VA anesthesiologist and cardiac physiologist I study protective strategies against myocardial infarction, a significant medical problem in Veterans. One such strategy is "Preconditioning". Genetic predisposition, however, may be a confounding factor when trying to transfer its use into clinical practice. Profound differences in tolerance to ischemia exist among and even within certain species. Utilizing a unique genetic rat model it could be shown that transfer of one chromosome from a more resistant strain into a more sensitive strain renders the resulting "consomic" more resistant against infarction while narrowing the genetic difference to a single chromosome compared to the sensitive parental strain. This offers an excellent starting point to study the genetic basis and mechanisms of cardioprotection, from intact hearts to isolated mitochondria. This proposal is the indispensable basis for me to further investigate the role of genetics in cardioprotection and represents a critical milestone to achieve independence in cardiovascular research in the VA system.

描述（由申请人提供）： 作为一名退伍军人管理局麻醉师和生理学家，我研究预防心肌梗塞，这是老年退伍军人面临的一个重大医疗问题。例如，在缺血预适应（IPC）中，持续缺血/再灌注（IR）之前的几个较短的缺血期可以减轻梗塞。已经描述了 IPC 的各种触发机制，包括一氧化氮 (NO) 和超氧化物的形成。然而，当试图将其应用于临床实践时，遗传倾向可能是一个重要的混杂因素。虽然有些患者可能会受益，但另一些患者则不会。缺血耐受性不仅存在于某些物种之间，而且存在于某些物种内部。例如，达尔盐敏感 (SS) 大鼠比棕色挪威 (BN) 大鼠更容易受到红外线损伤，这使它们成为研究与非裔美国患者表型相似的疾病基因型的理想模型。在威斯康星医学院构建的独特染色体替代（康体）模型中，BN 6 号染色体渗入 SS 中，使所得的 SS6BN 康体更具 IR 抗性，同时将遗传差异缩小到单个染色体。这为研究 IPC 等心脏保护策略的遗传基础提供了一个极好的起点。尽管 BN 与无 IPC 的 SS 相比，NO 与减少梗塞面积有关，但尚不清楚 SS 是否能够对 IPC 做出反应，以及对 IPC 的抵抗是否与可能受 DNA 结合调节的内皮 NO 合酶 (eNOS) 活性差异有关。蛋白质抑制剂 Id2 和过氧化物酶体增殖物激活受体 3 (PPAR3)。 eNOS 可以产生 NO 或超氧化物。两者都可以调节线粒体功能，而线粒体功能又可以作为 IPC 的触发器和效应器。因此，我的总体假设是 IPC 心脏保护的失败/成功是由调节 NO 产生和线粒体功能的基因介导的。因此，我建议使用这个群体模型来研究两个具体目标：I) 确定不同的遗传背景是否导致 IPC 的差异细胞和线粒体保护，以及 II) eNOS 和/或其上游调节剂 Id2 和 PPAR3 是否是候选基因负责这种差动保护。测试了四个假设：1) BN 6 号染色体上的基因对于 IPC 是必需的，BN 和 SS6BN 与 SS 相比具有更好的心脏功能和更少的梗塞即可证明这一点。 2) BN 6 号染色体上的基因对于 IPC 是必需的，BN 和 SS6BN 比 SS 的线粒体功能更有效就证明了这一点。 3) IPC 对完整心脏中细胞和线粒体功能的保护取决于大鼠 6 号染色体上基因调节的信号通路 Id2->PPAR3->eNOS->NO。 4) 离体心肌细胞中细胞和线粒体功能的差异保护取决于无空房。方法 1 和 2：在完整、跳动的心脏中测量各种心脏和线粒体功能、NO 产生和梗塞大小，以评估 IPC 遗传确定的保护的数量和质量。方法 3 和 4：在 NOS 抑制剂、NO 供体或 PPAR3 激动剂或拮抗剂存在的情况下进行额外的心脏跳动实验，并确定 Id2、PPAR3 和 eNOS 表达以及 NO 水平和定位的差异。此外，在分离的肌细胞中测量NO不存在或存在时的不同线粒体功能，以确定NO在该遗传模型中通过IPC介导差异细胞和线粒体保护中的作用和起源。遗传工具对于研究遗传倾向的作用至关重要。将功能结果、梗死面积和线粒体功能与 IPC、NO 水平、蛋白质表达和基因型相关联，将使我们能够定义信号通路以及 NO 和线粒体功能在心脏保护中的作用，并描绘与心脏保护基因型相关的亚细胞和线粒体表型变化。该 CDA 标志着进一步研究遗传学在心脏保护中的作用不可或缺的基础，并将成为 VA 心血管研究实现研究独立性的一个重要里程碑。 公共卫生相关性： 作为一名退伍军人管理局麻醉师和心脏生理学家，我研究心肌梗塞的保护策略，这是退伍军人中一个重要的医学问题。其中一种策略是“预处理”。然而，当试图将其应用转移到临床实践时，遗传倾向可能是一个混杂因素。在某些物种之间甚至内部，对缺血的耐受性存在巨大差异。利用独特的遗传大鼠模型，可以证明，将一条染色体从更具抵抗力的品系转移到更敏感的品系中，可以使所得的“体”对梗塞具有更强的抵抗力，同时与敏感的亲本品系相比，将遗传差异缩小到单条染色体。这为研究心脏保护的遗传基础和机制（从完整的心脏到分离的线粒体）提供了一个极好的起点。该提案是我进一步研究遗传学在心脏保护中的作用不可或缺的基础，也是 VA 系统心血管研究实现独立的一个重要里程碑。