Mitochondrial and Nuclear Mechanisms of Estrogen Receptor-Mediated Cardioprotecti

雌激素受体介导的心脏保护的线粒体和核机制

• 批准号: 7842217
• 负责人: DONNA HOPE KORZICK
• 金额: $ 18.97万
• 依托单位: PENNSYLVANIA STATE UNIVERSITY, THE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-15 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7842217
• 关键词: 1-Phosphatidylinositol 3-Kinase; Acute; Address; Adult; Age; Aging; Agonist; American; Amino Acid Sequence; Animal Model; Antioxidants; Apoptosis; Apoptotic; Arts; Attenuated; Binding; Biochemical; Biological Assay; Cardiac; Cardiovascular Diseases; Cardiovascular system; Cause of Death; Cell Death; Cell Survival; Centrifugation; Cessation of life; Chronic; Clinical Research; Clinical Trials; Computer software; Conflict (Psychology); Coronary; Data; Demographic Aging; Development; Diagnosis; Economic Burden; Electron Spin Resonance Spectroscopy; Enzymes; Epidemiology; Estrogen Nuclear Receptor; Estrogen Receptors; Estrogen Replacement Therapy; Estrogens; Event; Female; Gene Expression; Genetic Transcription; Genomics; Glycogen Synthase Kinases; Health Care Costs; Heart; Heart Diseases; Heart failure; Hospitals; Image; In Vitro; Infarction; Ischemia; Label; Ligation; Mediating; Mediator of activation protein; Medical; Membrane; Mitochondria; Mitochondrial Proteins; Modeling; Molecular; Myocardial Infarction; Necrosis; Nitric Oxide Synthase; Nuclear; Nuclear Protein; Nuclear Proteins; Ontology; Operative Surgical Procedures; Outcome; Oxidative Stress; Pathway interactions; Peptide Sequence Determination; Peptides; Persons; Phenotype; Phosphorylation; Physiological; Population; Post-Translational Protein Processing; Postmenopause; Production; Protein Kinase; Proteins; Proteomics; Rattus; Receptor Activation; Receptor Gene; Receptor Signaling; Reperfusion Injury; Reperfusion Therapy; Resources; Risk Factors; Role; Scaffolding Protein; Signal Pathway; Signal Transduction; Testing; Therapeutic; Time; Up-Regulation; Western Blotting; Woman; acute coronary syndrome; age effect; age related; aged; aging population; cell growth regulation; estrophilin; improved; in vivo; juvenile animal; men; middle age; mimetics; mortality; multiple reaction monitoring; new therapeutic target; non-genomic; novel; oxidant stress; public health relevance; receptor expression; research study; senescence

DESCRIPTION (provided by applicant): Post-menopausal women are especially vulnerable to ischemic insult, and ~80,000 more cardiovascular deaths occur yearly in aged women than in aged men. Conflicting data exist on the role of estrogen (E2) in modulating ischemic tolerance (IT), due in part to the failed efficacy of E2 replacement to improve IT. Protective signaling pathways in the heart known to be influenced by E2 action include (PI3K)-Akt, eNOS and GSK3-2. Collectively, these signals impact mitochondrial events critical to cell survival to limit necrotic and apoptotic cell death. Here, we propose to test the novel paradigm that maladaptive estrogen receptor (ER) expression in the aged heart leads to a vicious cycle of increased oxidative stress (ROS) and ischemia/reperfusion (I/R injury), while selective engagement of non-genomic ER pathways can rescue this phenotype. We further propose that non-genomic ER signaling results in protein kinase C5 (PKC5) activation, a well-known mediator of cardioprotection. The following specific aims will be tested: 1) determine which ER subtype(s) reduce IT in aged, gonadectomized female rats, 2) determine the molecular mechanism(s) by which ERs alter ROS (ONOO- and O-2) through eNOS uncoupling, post-translational modification (PTM) or the PI3- Akt-GSK-32 axis, 3) determine the role of PKC5 in rapid ER signaling, and 4) identify novel mitochondrial PKC5 binding targets following non-genomic ER activation. Using a model of global ischemia, we will employ specific ER and PKC5 agonists and EPR spectroscopy to assess ROS. Apoptosis will be assessed through in vivo coronary ligation, western blotting for cellular ER subtype targeting and downstream signals, qRT- PCR for ER- mediated gene expression, and multiple reaction monitoring to assess PTMs of ERs. A targeted, state-of-the- art proteomic iTRAQ 8plex approach and LC MS/MS will be used to identify novel cardiac mitochondrial PKC5 protein partners and phosphorylation. We propose that non-genomic activation of ERs can compensate for E2 deficiency in aged female hearts, resulting in improved IT. The confirmation of our hypothesis will strongly support new targets for the treatment of acute coronary syndrome in the aging population and address the potential use of ER/PKC5 therapeutics as viable approaches to improve IT in post-menopausal women. PUBLIC HEALTH RELEVANCE: The leading cause of death among persons aged 75 years or older is heart disease, and deaths due to heart disease have actually increased by 60% from 1970 to 2002 among persons aged 80 years or older. Age is also the leading risk factor for the development of heart failure (CHF) and more Americans aged 65 and older are treated and discharged from hospitals with CHF than with any other diagnosis. Interestingly, mortality rates due to CHF are significantly greater in aged women vs aged men. CVD rates and associated mortality are also significantly greater in aged women when compared to aged men, and data from large scale clinical trials suggest that estrogen replacement therapy is ineffective in reducing, and may actually increase CVD outcomes. Collectively, this expanding aged demographic will undoubtedly require increased allocation of medical resources, raising the economic burden of health care costs for all Americans. The cellular mechanisms by which estrogen exerts effects in the aged heart, however, are poorly understood. The experiments associated with this proposal are expected to identify novel protein targets previously unstudied with estrogen deficiency. If our hypothesis on estrogen receptors is correct, new therapeutic targets for the treatment of ischemic cardiovascular disease in the aged population will be realized, i.e. use of estrogen receptor therapeutics as viable approaches to improve ischemic tolerance in aged women.

描述（由申请人提供）：绝经后女性特别容易受到缺血性损伤的影响，老年女性每年发生的心血管死亡人数比老年男性多约80，000人。关于雌激素（E2）在调节缺血耐受（IT）中的作用存在明确的数据，部分原因是E2替代改善IT的无效功效。已知心脏中受E2作用影响的保护性信号传导途径包括（PI 3 K）-Akt、eNOS和GSK 3 -2。总的来说，这些信号影响对细胞存活至关重要的线粒体事件，以限制坏死和凋亡性细胞死亡。在这里，我们建议测试新的范例，即在老年心脏中的适应不良雌激素受体（ER）表达导致氧化应激（ROS）和缺血/再灌注（I/R损伤）增加的恶性循环，而非基因组ER途径的选择性参与可以挽救这种表型。我们进一步提出，非基因组ER信号转导导致蛋白激酶C5（PKC 5）激活，这是一种众所周知的心脏保护介质。将测试以下具体目标：1）确定哪种ER亚型降低老年性腺切除雌性大鼠的IT，2）确定ER改变ROS的分子机制（ONOO-和O-2）通过eNOS解偶联、翻译后修饰（PTM）或PI 3- Akt-GSK-32轴，3）确定PKC 5在快速ER信号传导中的作用，和4）在非基因组ER活化后鉴定新的线粒体PKC 5结合靶。使用全脑缺血模型，我们将采用特定的ER和PKC 5激动剂和EPR光谱来评估ROS。将通过体内冠状动脉结扎、细胞ER亚型靶向和下游信号的蛋白质印迹、ER介导的基因表达的qRT-PCR和多反应监测来评估细胞凋亡，以评估ER的PTM。将使用靶向、最先进的蛋白质组学iTRAQ 8 plex方法和LC MS/MS鉴定新型心脏线粒体PKC 5蛋白伴侣和磷酸化。我们提出，ER的非基因组激活可以弥补老年女性心脏E2缺乏，从而改善IT。我们的假设的确认将有力地支持新的目标，为治疗急性冠状动脉综合征在老龄化人口和解决潜在的使用ER/PKC 5治疗作为可行的方法，以改善IT在绝经后妇女。公共卫生关系：75岁或以上的人的主要死因是心脏病，1970年至2002年，80岁或以上的人因心脏病死亡的人数实际上增加了60%。年龄也是心力衰竭（CHF）发展的主要风险因素，65岁及以上的美国人接受CHF治疗和出院的人数多于任何其他诊断。有趣的是，老年女性CHF的死亡率明显高于老年男性。与老年男性相比，老年女性的心血管疾病发病率和相关死亡率也明显更高，来自大规模临床试验的数据表明，雌激素替代疗法在减少心血管疾病方面无效，实际上可能会增加心血管疾病的结局。总的来说，这种不断扩大的老年人口无疑需要增加医疗资源的分配，增加所有美国人的医疗保健费用的经济负担。然而，雌激素在老年心脏中发挥作用的细胞机制却知之甚少。与该建议相关的实验有望确定以前未研究过的雌激素缺乏症的新蛋白质靶点。如果我们对雌激素受体的假设是正确的，将实现治疗老年人群缺血性心血管疾病的新的治疗靶点，即使用雌激素受体治疗作为改善老年女性缺血耐受性的可行方法。