Heart Function Decline and Aging

心脏功能衰退与衰老

• 批准号: 10427227
• 负责人: Wolfgang H Dillmann
• 金额: --
• 依托单位: VA SAN DIEGO HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-01-01 至 2022-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10427227
• 关键词: 3' Untranslated Regions; 5' Untranslated Regions; ATP Hydrolysis; Age; Aging; Animals; Antibodies; Biological Assay; Calcium; Cardiac; Cardiac Muscle Contraction; Cardiac Myocytes; Cessation of life; ChIP-seq; Chromatin; Collaborations; Complex; Consumption; Contracts; DNA; DNA Probes; Data; Development; Dominant-Negative Mutation; Elderly; Equipment; Gene Expression; Gene Expression Profile; General Population; Genes; Genetic Transcription; Health; Healthcare; Heart; Heart Mitochondria; Heart failure; Impairment; Ischemia; Knowledge; Lead; Link; Mediating; Messenger RNA; Metabolic; Metabolism; MicroRNAs; Mitochondria; Mitochondrial Matrix; Modeling; Molecular; Motor; Mus; Mutate; Normal Range; Pathway interactions; Performance; Pharmacology; Polymerase; Population; Production; Proteins; Protons; Reperfusion Injury; Reperfusion Therapy; Research; Research Personnel; Respiratory physiology; Role; Sodium; Testing; Transgenes; Translation Initiation; Transposase; Untranslated Regions; Veterans; Workload; adeno-associated viral vector; base; blood pump; calcium uniporter; chromatin immunoprecipitation; chromatin remodeling; deep sequencing; experimental study; gain of function; genome-wide; heart function; improved; improved functioning; in vivo; knock-down; link protein; loss of function; member; muscular structure; myocardial infarct sizing; new therapeutic target; novel; novel strategies; novel therapeutic intervention; pressure; respiratory; response; restoration; transcriptome sequencing; transgene expression

Heart failure (HF) is a significant health problem in the elderly population, including in Veterans. The basis for the diminished function of the old heart (OH, 19-22 month) and increased propensity to develop HF is only incompletely explored and poorly treated. Preliminary results show that in OH cardiac function is diminished, cardiac myocytes (CM) contract poorly, and decreased Mitochondrial (Mito) respiratory function occurs. Our preliminary results also show novel findings that in CM from OH versus young heart (YH, 2-3 month) the Mitochondrial Calcium Uniporter (MCU) Complex (MCUC) has markedly decreased calcium (Ca2+) conductance resulting in decreased Mito matrix freeCa2+ concentration ([Ca2+]m) which leads to decreased Mito respiratory, cardiac metabolic, and contractile function. Specific MCUC member proteins show decreased levels in OH CM. The level of the Essential MCU Regulator (EMRE) is decreased by 70% in OH CM. In the absence of EMRE, MCUC Ca2+ conductance is lost. In addition MCU shows a more modest 30% decline in OH CM. The principle hypothesis is that the performance of the OH can be markedly improved by restoring EMRE and MCU levels in OH CM (OH+EMRE+MCU) using adeno-associated viral vector (AAV)-based expression of transgenes (tges) encoding EMRE and MCU or pharmacological inhibition of Mito Ca2+ export. To test this hypothesis we pursue three closely linked Aims. In Aim 1 we enhance MCUC Ca2+ conductance by restoring EMRE and MCU in OH towards the YH range and determine its influence on Mito and cytosolic Ca2+ handling, Mito respiratory, cardiac metabolic and contractile function, and animal survival. We also inhibit the Mito sodium- Ca2+ exchanger (mNCLX) and Mito Ca2+ export with a pharmacological compound, returning [Ca2+]m to the YH level and improving CM contraction. In Aim 2 we determine if maladaptive consequences occur in OH+EMRE+MCU, especially increased CM death and increased myocardial infarct size (MI) with ischemia/reperfusion (I/R). Preliminary results show a decrease in MI size in OH+EMRE+MCU versus (vs) OH. Influences of dominant negative (dn) dnEMRE and dnMCU expression on Mito Ca2+ handling and CM contraction in YH and OH are investigated. We compare rescue effects in OH+EMRE+MCU vs expression of a SERCa2 tge in OH (OH+SERCa2). Preliminary results show similar basal and maximal ex vivo cardiac function in OH+EMRE+MCU and OH+SERCa2. In Aim 3 we explore molecular mechanisms mediating the marked decrease in EMRE protein levels in OH CM. Preliminary results point to an interaction of miRNA 215, which is 3.4-fold increased in OH CM, with the 5' UTR of EMRE mRNA inhibiting translation initiation. We also explore if chromatin remodeling to a more open state occurs in OH+EMRE+MCU vs OH with increased DNA accessibility enabling increased gene transcription, including of the SERCa2 gene. Chromatin remodeling involves chromatin remodeling motors which require ATP hydrolysis. Studies in YH, OH, and OH+EMRE+MCU are conducted with collaborators at San Diego VA and UCSD who have the expertise and equipment to perform the following experiments. We use the Assay for Transposase-Accessible Chromatin with deep Sequencing (ATAC-Seq) and Chromatin Immunoprecipitation Sequencing (Chip-Seq) with a Polymerase II antibody to probe DNA accessibility for gene transcription. Chip-Seq pathway enrichment and RNA Sequencing (RNA-Seq) are used to determine changes in gene expression. To establish a link to increased ATP levels we use CM from OH+EMRE+MCU and OH with or without the protonophore FCCP which rapidly dissipates the proton gradient and inhibits ATP formation. Limited preliminary results, which need to be confirmed and extended, from ATAC-Seq and Chip-Seq indicate increased DNA accessibility in OH+EMRE+MCU vs OH CM. Chip-Seq pathway enrichment analysis and RNA-Seq data indicate a shift in the gene expression profile to mRNAs encoding proteins linked to cardiac muscle contraction and muscle structure development in OH+EMRE+MCU vs OH. The closely linked Aims will lead to new knowledge and may result in novel therapeutic approaches.

心力衰竭（HF）是老年人群（包括退伍军人）的重要健康问题。的基础 老年心脏（OH，19 - 22个月）的功能减弱和发生HF的倾向增加仅是 未完全探索和治疗不善。初步结果显示，在OH心脏功能减弱， 心肌细胞（CM）收缩不良，并且发生线粒体（Mito）呼吸功能降低。我们 初步结果还显示了新的发现，在OH与年轻心脏（YH，2 - 3个月）的CM中， 线粒体钙单向转运体（MCU）复合物（MCUC）可显著降低钙（Ca 2+）电导 导致减少的Mito基质游离Ca2+浓度（[Ca2 +] m），这导致减少的Mito呼吸， 心脏代谢和收缩功能。特定MCUC成员蛋白在OH CM中显示降低的水平。 在OH CM中，基本MCU调节器（EMRE）的水平降低了70%。在没有电磁辐射的情况下， MCUC Ca2+电导丢失。此外，MCU显示OH CM下降了30%。原则 假设通过恢复EMRE和MCU可以显著改善OH的性能 使用基于腺相关病毒载体（AAV）的表达的OH CM（OH + EMRE + MCU）中的水平， 编码EMRE和MCU的转基因（tges）或Mito Ca2+输出的药理学抑制。为了验证这一 假设我们追求三个紧密相连的目标。在目的1中，我们通过恢复MCUC的Ca2+电导来增强MCUC的Ca2+电导。 EMRE和MCU在OH朝向YH范围，并确定其对Mito和细胞溶质Ca2+处理的影响， 线粒体呼吸、心脏代谢和收缩功能与动物存活。我们也抑制水户钠- Ca2+交换器（mNCLX）和Mito Ca2+输出与药理学化合物，返回[Ca2 +] m到YH 水平和改善CM收缩。在目标2中，我们确定适应不良的后果是否发生在 OH + EMRE + MCU，尤其是CM死亡增加和心肌梗死面积（MI）增加， 缺血/再灌注（I/R）。初步结果显示，与OH相比，OH + EMRE + MCU中MI尺寸减小。 显性负性（dn）dnEMRE和dnMCU表达对Mito Ca2+处理和CM收缩的影响 在YH和OH中的反应。我们比较了OH + EMRE + MCU与SERCa2 tge表达的拯救效果， 在OH（OH + SERCa 2）中。初步结果显示，与对照组相比， OH + EMRE + MCU和OH + SERCa 2。在目标3中，我们探索了介导标记的 OH CM中EMRE蛋白水平降低。初步结果表明，miRNA 215的相互作用， 3.4-在OH CM中，EMRE mRNA的5 'UTR抑制翻译起始。我们还探讨了如果 OH + EMRE + MCU与OH相比，染色质重塑为更开放的状态，DNA可及性增加 从而增加基因转录，包括SERCa2基因。染色质重塑涉及染色质 重塑需要ATP水解的马达在YH、OH和OH + EMRE + MCU中进行了研究， 合作者在圣地亚哥弗吉尼亚州和加州大学圣地亚哥分校谁拥有专业知识和设备，以执行以下 实验我们使用深度测序转座酶可降解染色质测定法（ATAC-Seq）， 染色质免疫沉淀测序（Chip-Seq），使用聚合酶II抗体探测DNA可及性 for gene基因transcription转录.使用芯片-Seq途径富集和RNA测序（RNA-Seq）来确定 基因表达的变化。为了建立与ATP水平增加的联系，我们使用来自OH + EMRE + MCU的CM， OH，有或没有质子载体FCCP，可快速消散质子梯度并抑制ATP 阵ATAC-Seq和Chip-Seq的初步结果有限，需要确认和扩展 表明OH + EMRE + MCU中DNA可及性相对于OH CM增加。Chip-Seq途径富集分析 和RNA-Seq数据表明，基因表达谱向编码与心脏相关蛋白质的mRNA转移， OH + EMRE + MCU与OH的肌肉收缩和肌肉结构发育。紧密相连的目标将 导致新知识并可能导致新的治疗方法。