Effect of elevated dATP on contractile function and the Frank-Starling relationship in models of dilated cardiomyopathy

dATP 升高对扩张型心肌病模型收缩功能和 Frank-Starling 关系的影响

• 批准号: 10391887
• 负责人: Farid Moussavi-Harami
• 金额: $ 4.46万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-05-01 至 2022-01-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10391887
• 关键词: Address; Adult; Anabolism; Animal Model; Beryllium; Binding; Calcium; Cardiac; Cardiac Myocytes; Data; Deoxyadenosines; Dextrans; Dilated Cardiomyopathy; Enzymes; Epidemic; Fluorides; Genetic Models; Heart; Heart failure; Human; Impairment; Infarction; Kinetics; Length; Medical; Modeling; Mus; Mutation; Myocardium; Patients; Process; Prognosis; Rattus; Recombinants; Relaxation; Ribonucleotide Reductase; Rodent Model; Sarcomeres; Sturnus vulgaris; Systolic heart failure; Testing; Thin Filament; Tissues; Transgenic Organisms; Workload; adeno-associated viral vector; alpha Tropomyosin; improved; inorganic phosphate; machine learning method; mortality; novel; novel therapeutics; overexpression; recruit; response; tripolyphosphate

Abstract Dilated cardiomyopathy (DCM) is the most common form of systolic heart failure with poor prognosis and no specific treatment to address the underlying contractile deficit. In addition to the loss of contractility, there is also impairment in the Frank-Starling relationship, an adaptive process that is described as the increase in contractile force in response to increased preload. We are proposing to investigate a novel therapy for DCM that addresses both loss of contractility and impairment in the Frank-Starling relationship. This novel therapy is achieved by increasing intracellular levels of 2-deoxy ATP (dATP) in cardiomyocytes via increasing the expression of the enzyme ribonucleotide reductase (R1R2), the rate-limiting step in de novo dNTP biosynthesis. Increased intracellular levels dATP in cardiomyocytes increases contraction by enhancing cross-bridge binding and cycling kinetics and improving allosteric activation of contraction. Recent data suggests that in addition to improved contraction, increased dATP level also enhances the Frank Starling relationship. I will propose specific aims to assess the effect of increased dATP on the Frank Starling relationship and contraction in two animal models of DCM. In the first aim, I will use a genetic model of DCM with a D230N mutation in alpha-tropomyosin (D230N Tm). Using recombinant D230N Tm, I will test the hypothesis that this mutation reduces thin filament activation and, as a consequence, the kinetics of contractile activation and relaxation. I will then determine the effect of increasing dATP content on these deficits. Using tissue from transgenic D230N Tm mice, I will test the hypothesis that this mutation decreases calcium sensitivity of force and length-dependent activation. I will correct these functional deficits by increasing dATP content. Using AAV6-cR1R2, an adeno-associated viral vector that restricts R1R2 over-expression to cardiac myocytes, I will test the hypothesis that increasing dATP levels improves contractility of isolated cardiomyocytes, and improves systolic function in both adult and young D230N Tm mice. Similarly, I will use post-infarct model of DCM in rats to evaluate the effect of cross-bridge activation on length dependent activation (LDA)-the Frank Starling relationship at the sarcomere level. Next, I will use alternative agents to increase (dextran, EMD 50733) or decrease (BDM, beryllium fluoride, high inorganic phosphate) cross-bridge recruitment and evaluate their effect on LDA in demembranated post- infarct DCM rat trabecula. I will complete this aim by testing the hypothesis that cross-bridge activation augments LDA in human myocardium from patients with DCM. Extension Project: Use data from above studies in D230N Tm DCM rodent model to simulate realistic cardiac twitches and use machine-learning methods for parameter optimization.

抽象的 扩张的心肌病（DCM）是收缩性心力衰竭的最常见形式，预后不良 并且没有具体的治疗方法来解决潜在的收缩赤字。除了失去收缩性外， 坦率的宣传关系也有损害，这是一种适应性过程，被描述为 收缩力增加，以响应提前加载。我们建议调查小说 DCM的治疗既解决坦率的关系中的收缩力和损害的损失。 通过增加心肌细胞中的2-脱氧ATP（DATP）的细胞内水平来实现这种新型疗法 通过增加酶核糖核苷酸还原酶（R1R2）的表达，DE的速率限制步骤 Novo DNTP生物合成。心肌细胞中DATP的细胞内水平升高可通过 增强跨桥结合和循环动力学并改善收缩的变构激活。 最近的数据表明，除了提高收缩外，DATP级别还提高了 弗兰克·斯塔琳的关系。我将提出特定的目的，以评估增加Datp对Frank的影响 DCM的两个动物模型中的Starling关系和收缩。在第一个目标中，我将使用遗传 DCM模型，具有D230N突变（D230N TM）中的D230N突变。使用重组D230N TM， 我将检验以下假设：该突变减少了细丝的激活，因此 收缩激活和放松的动力学。然后，我将确定增加DATP内容的效果 在这些缺陷上。使用转基因D230N TM小鼠的组织，我将测试这种突变的假设 降低力和长度依赖性激活的钙敏感性。我会纠正这些功能 通过增加DATP内容来赤字。使用AAV6-CR1R2，这是一种限制腺相关的病毒载体 R1R2过表达对心肌细胞，我将测试以下假设，即增加DATP水平有所改善 孤立的心肌细胞的收缩力，并改善成人和年轻D​​230N TM的收缩功能 老鼠。同样，我将使用大鼠DCM的Infarct模型来评估跨桥激活的效果 在依赖性激活（LDA）上 - 肉皮级别的弗兰克·史塔琳（Frank Starling）关系。接下来，我会的 使用替代剂增加（Dextran，EMD 50733）或减少（BDM，氟化铍，高 无机磷酸盐）跨桥的募集，并评估其对LDA的影响 梗死DCM大鼠小梁。我将通过测试跨桥激活的假设来完成这一目标 DCM患者的人体心肌增强LDA。 扩展项目：使用D230N TM DCM啮齿动物模型的上述研究数据模拟现实 心脏抽搐并使用机器学习方法进行参数优化。