Regulation of Contractile Function by cMLC2v Phosphorylation in Heart Failure

cMLC2v 磷酸化对心力衰竭收缩功能的调节

• 批准号: 8908835
• 负责人: Margaret Louise Novak
• 金额: $ 5.42万
• 依托单位: LOYOLA UNIVERSITY CHICAGO
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2017-07-13
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8908835
• 关键词: ATP phosphohydrolase; Adrenergic Agents; Adrenergic beta-Antagonists; Adverse effects; Affect; American; Animal Model; Cardiac; Cardiac Myocytes; Cardiac Myosins; Clinical; Contractile Proteins; Diagnosis; Disease Progression; Diuretics; Functional disorder; Future; Goals; Head; Health; Heart; Heart Atrium; Heart failure; Human; In Situ; Kinetics; Length; Longevity; Measures; Methods; Microfilaments; Mus; Myosin ATPase; Myosin Heavy Chains; Myosin Light Chain Kinase; Myosin Regulatory Light Chains; Oryctolagus cuniculus; Patients; Performance; Phosphoric Monoester Hydrolases; Phosphorylation; Phosphorylation Site; Preparation; Production; Protein Isoforms; Proteins; Pump; Quality of life; Rattus; Recombinants; Regulation; Reporting; Role; Skin; Testing; Thick Filament; Thin Filament; Transfection; Ventricular; Work; adrenergic; gene therapy; genetic regulatory protein; heart function; improved; interest; mutant; myosin phosphatase; new therapeutic target; novel; novel therapeutics; outcome forecast; overexpression; palliative; pressure; restoration; targeted treatment; therapeutic target

 DESCRIPTION (provided by applicant): Heart failure (HF) is a major clinical problem with extremely poor prognosis. Though altered phosphorylation of thin filament proteins is known to contribute to HF progression, much less is known about phosphorylation of the thick filament protein cardiac myosin regulatory light chain (cMLC2v). cMLC2v modulates force production by altering myosin head orientation. Its phosphorylation status is controlled by myosin light chain kinase (cMLCK) and phosphatase (MLCP). Decreased cMLC2v phosphorylation and cMLCK expression are observed during HF in patients and animal models, though increased phosphorylation has also been reported and may be an adaptive mechanism. In mice, cMLCK overexpression ameliorates HF; conversely, cMLCK disruption exacerbates HF. However, mice have two cMLC2v phosphorylation sites, each of which may have a separate function, whereas human cMLC2v has only one phosphorylation site. Thus, though cMLC2v phosphorylation is a key regulator of contraction, its role in human HF is largely unknown. Defining the role of cMLC2v phosphorylation in human health and HF will open new therapeutic avenues. Current HF therapies, such as beta-blockers and diuretics, are non-specific and largely palliative; in contrast, therapies targeting cMLC2v would be cardiac-specific and could improve heart function at the fundamental level of the contractile proteins. We hypothesize that cMLC2v phosphorylation is a critical regulator of human cardiomyocyte function, and manipulation of cMLC2v phosphorylation may be a novel therapy for treatment of HF. In Aim 1, we will use skinned cardiac preparations to study regulation of contractile function by cMLC2v phosphorylation in failing and non-failing human heart. We predict that cMLC2v phosphorylation will be decreased during HF, and that experimentally increasing cMLC2v phosphorylation will improve contractile function, especially in failing versus non-failing hearts. Because cMLC2v may also impact Ca2+ transients and may preferentially alter dynamic and loaded contractions, Aim 2 will investigate regulation of dynamic contractile function and Ca2+ handling by cMLC2v phosphorylation in intact electrically stimulated cardiomyocytes from failing and non-failing rabbit hearts (which have high cMLC2v homology with human and identical phosphorylation site). We expect that increasing cMLC2v phosphorylation by cMLCK transfection will improve contractile function as assessed by simulated PV loops, and will increase amplitude but decrease duration of the Ca2+ transient; we expect that cMLCP overexpression will have the opposite effect. In summary, we hypothesize that reduced cMLC2v phosphorylation is a major mechanism underlying contractile dysfunction in human HF, and restoration of cMLC2v phosphorylation may be a novel therapeutic target. Future studies will use pharmacological and/or gene therapy to treat HF in animal models and eventually in patients.

 描述（由申请人提供）：心力衰竭（HF）是一个预后极差的主要临床问题，尽管已知细丝蛋白磷酸化的改变会导致心力衰竭的进展，但对粗丝蛋白心肌肌球蛋白的磷酸化知之甚少。调节轻链 (cMLC2v) 通过改变肌球蛋白头部方向来调节力的产生，其磷酸化状态由肌球蛋白轻链激酶控制。在患者和动物模型中观察到 cMLC2v 磷酸化和 cMLCK 表达降低，但也有报道称 cMLCK 过度表达可改善 HF，这可能是一种适应性机制。然而，小鼠有两个 cMLC2v 磷酸化位点，每个位点可能具有单独的功能，而人类 cMLC2v 只有一个磷酸化位点，因此，尽管 cMLC2v 磷酸化是收缩的关键调节因子，但其在人类心力衰竭中的作用在很大程度上尚不清楚，确定 cMLC2v 磷酸化在人类健康和心力衰竭中的作用将为当前的心力衰竭治疗开辟新的途径。 β-受体阻滞剂和利尿剂等非特异性且很大程度上是姑息性的；相反，针对 cMLC2v 的治疗具有心脏特异性，可以改善病情；我们发现 cMLC2v 磷酸化是人类心肌细胞功能的关键调节因子，并且控制 cMLC2v 磷酸化可能是治疗心力衰竭的一种新疗法。在目标 1 中，我们将使用带皮心脏制剂。研究衰竭和非衰竭人类心脏中 cMLC2v 磷酸化对收缩功​​能的调节，我们预测 cMLC2v 磷酸化在心力衰竭期间会降低。并且通过实验增加 cMLC2v 磷酸化将改善收缩功能，特别是在衰竭心脏与非衰竭心脏中，因为 cMLC2v 也可能影响 Ca2+ 瞬变，并且可能优先改变动态和负荷收缩，因此目标 2 将研究 cMLC2v 对动态收缩功能和 Ca2+ 处理的调节。来自衰竭和非衰竭兔心脏的完整电刺激心肌细胞中的磷酸化（与人类和相同的 cMLC2v 具有高度同源性）我们预计通过 cMLCK 转染增加 cMLC2v 磷酸化将改善通过模拟 PV 环评估的收缩功能，并且会增加振幅但减少 Ca2+ 瞬变的持续时间；我们预计 cMLCP 过度表达将产生相反的效果。 cMLC2v 磷酸化是人类心力衰竭收缩功能障碍的主要机制，恢复 cMLC2v 磷酸化可能是一个新的治疗靶点。未来的研究将使用药物和/或基因疗法来治疗动物模型中的心力衰竭，并最终治疗患者。