Heart Function Decline and Aging

心脏功能衰退与衰老

• 批准号: 10265347
• 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 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10265347
• 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）是老年人口的重大健康问题，包括退伍军人。基础 旧心脏（哦，19-22个月）的功能降低，而发展HF的倾向增加只是 未完全探索和治疗不佳。初步结果表明，在OH心脏功能中， 心肌细胞（CM）收缩较差，线粒体（MITO）呼吸功能降低。我们的 初步结果还显示了新发现的结果，即OH与年轻心（YH，2-3个月）中的CM 线粒体钙Uniporter（MCU）复合物（MCUC）显着降低了钙（Ca2+）电导 导致MITO矩阵freeca2+浓度（[Ca2+] m）降低，导致MITO呼吸降低， 心脏代谢和收缩功能。特定的MCUC成员蛋白显示OH CM的水平降低。 OH CM中，必需的MCU调节器（EMRE）的水平降低了70％。在没有EMRE的情况下 MCUC CA2+电导丢失。此外，MCU显示OH CM下降30％。原则 假设是可以通过恢复EMRE和MCU来显着提高OH的性能 使用腺相关的病毒载体（AAV）表达的OH CM（OH+EMRE+MCU）的水平（OH+EMRE+MCU） 编码EMRE和MCU的转基因（TGES）或对MITO Ca2+出口的药理抑制作用。测试这个 假设我们追求三个紧密相关的目标。在AIM 1中，我们通过还原来增强MCUC Ca2+电导 EMRE和MCU朝向YH范围，并确定其对MITO和胞质CA2+处理的影响， MITO呼吸道，心脏代谢和收缩功能以及动物存活。我们还抑制丝菌钠 Ca2+交换器（MNCLX）和Mito Ca2+用药理学化合物导出，将[Ca2+] M返回YH 水平和改善CM收缩。在AIM 2中，我们确定是否在 OH+EMRE+MCU，特别是增加CM死亡并增加心肌梗塞大小（MI） 缺血/再灌注（I/R）。初步结果表明，OH+EMRE+MCU与OH的MI大小减少。 主要负（DN）DNEMRE和DNMCU表达对Mito Ca2+处理和CM收缩的影响 在YH和OH中进行了调查。我们比较OH+EMRE+MCU与SERCA2 TGE的表达中的救援效应 在OH（哦+SERCA2）中。初步结果表明基础和最大离体心脏功能 OH+EMRE+MCU和OH+SERCA2。在AIM 3中，我们探索了介导标记的分子机制 OH CM中EMRE蛋白水平的降低。初步结果指出miRNA 215的相互作用，即 OH CM的3.4倍增加，EMR MRNA的5'UTR抑制翻译起始。我们还探索是否 染色质重塑为更开放的状态发生在OH+EMRE+MCU vs OH中，DNA可及性增加 可以增加基因转录，包括SERCA2基因。染色质重塑涉及染色质 需要ATP水解的重塑电动机。在YH，OH和OH+EMRE+MCU的研究中进行了研究 弗吉尼亚州圣地亚哥和UCSD的合作者，他们拥有专业知识和设备来执行以下操作 实验。我们将测定法用于带有深度测序（ATAC-SEQ）和 染色质免疫沉淀测序（CHIP-SEQ）具有聚合酶II抗体以探测DNA可及性 用于基因转录。 ChIP-Seq途径富集和RNA测序（RNA-Seq）用于确定 基因表达的变化。为了建立与升高ATP级别的链接，我们使用OH+EMRE+MCU的CM和 OH有或没有protonophore FCCP，该FCCP迅速消散质子梯度并抑制ATP 形成。有限的初步结果，需要从ATAC-SEQ和CHIP-SEQ进行确认和扩展 表明在OH+EMRE+MCU与OH CM中的DNA可及性提高。 CHIP-SEQ途径富集分析 RNA-seq数据表明基因表达谱的转移向编码与心脏相关的蛋白质的mRNA OH+EMRE+MCU与OH中的肌肉收缩和肌肉结构的发展。紧密联系的目标将 导致新知识，并可能导致新颖的治疗方法。