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)是老年人群中的一个重要健康问题，包括退伍军人。的基础 陈旧心脏(OH，19-22个月)的功能减弱和发生心力衰竭的倾向增加只是 不完全探索和不好的对待。初步结果显示，在OH中，心脏功能减弱， 心肌细胞收缩不良，线粒体(Mito)呼吸功能下降。我们的 初步结果还显示，在来自OH和年轻心脏(YH，2-3个月)的CM中，新的发现 线粒体钙统一转运体(MCU)复合体(MCUC)显著降低钙(Ca~(2+))电导 导致线粒体基质游离钙浓度([Ca~(2+)]m)降低，从而导致线粒体呼吸减少， 心脏代谢和收缩功能。特定的MCUC成员蛋白在OH CM中的水平降低。 在OH CM中，基本MCU调节器(EMRE)的水平降低了70%。在埃姆雷不在的情况下 MCUC钙电导丧失。此外，MCU的OH CM下降幅度较小，为30%。原则性 假设通过恢复EMRE和MCU可以显著提高OH的性能 腺相关病毒载体(AAV)在OH CM(OH+EmRE+MCU)中的表达水平 编码EmRE和MCU的转基因(TGE)或药物抑制线粒体钙输出。为了测试这一点 假设我们追求三个紧密相连的目标。在目标1中，我们通过恢复 EMRE和MCU在OH中朝向YH范围，并确定其对线粒体和胞浆钙离子处理的影响， 线粒体呼吸、心脏代谢和收缩功能，以及动物存活。我们还抑制了Mito钠- Ca~(2+)交换器(MNCLX)和Mito Ca~(2+)与一种药理化合物一起输出，使[Ca~(2+)]m返回YH 水平和改善CM收缩。在目标2中，我们确定不适应后果是否发生在 OH+EmRE+MCU，尤其是增加CM死亡率和增加心肌梗死面积(MI) 缺血/再灌流(I/R)。初步结果表明，与(VS)OH相比，OH+EmRE+MCU的MI尺寸减小。 显性负性(DN)dnEMRE和dnMCU表达对线粒体钙处理和心肌收缩的影响 In_(YH)和In_(OH)的研究。我们比较了在OH+EmRE+MCU中的救援效果与表达SERCA2 TGE的效果 在OH(OH+SerCa2)中。初步结果显示，基础和最大体外心功能相似 OH+EmRE+MCU和OH+SERCA2。在目标3中，我们探索了调节被标记的分子机制。 OHCM中EmRE蛋白水平降低。初步结果表明miRNA215之间存在相互作用 在OH CM中增加了3.4倍，Emre mRNA的5‘UTR区抑制了翻译起始。我们还探讨了如果 在OH+EmRE+MCU和OH中，染色质重塑到更开放的状态，DNA可及性增加 使基因转录增加，包括SERCA2基因。染色质重塑涉及染色质 改造需要三磷酸腺苷水解的马达。在YH、OH和OH+EMRE+MCU中进行了研究 圣地亚哥退伍军人管理局和加州大学圣地亚哥分校的合作者拥有执行以下任务的专业知识和设备 实验。我们使用转座酶可及染色质深度测序分析(ATAC-SEQ)和 用多聚酶II抗体进行染色质免疫沉淀测序(ChIP-SEQ)检测DNA可及性 用于基因转录。ChIP-Seq途径浓缩和RNA测序(RNA-Seq)用于确定 基因表达的变化。为了建立到增加的ATP水平的链接，我们使用来自OH+EmRE+MCU的CM和 无论有没有质子载体FCCP，它都能迅速消散质子梯度并抑制ATP 队形。ATAC-Seq和Chip-Seq的初步结果有限，需要确认和推广 表明与OH CM相比，OH+EmRE+MCU中的DNA可获得性更高。ChIP-Seq途径富集化分析 RNA-Seq数据表明，基因表达谱向编码与心脏相关的蛋白质的mRNAs转移 OH+EmRE+MCU VS OH组的肌肉收缩和肌肉结构发育。紧密相连的目标将 带来新的知识，并可能导致新的治疗方法。