Connexin 43: a new player in Duchenne muscular dystrophy associated cardiomyopathy

Connexin 43：杜氏肌营养不良症相关心肌病的新参与者

• 批准号: 9494823
• 负责人: Jorge Enrique Contreras
• 金额: $ 39.48万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9494823
• 关键词: 18 year old; Address; Affect; Arrhythmia; Biochemical; Blinded; Cardiac; Cardiac Myocytes; Cardiomyopathies; Cell Membrane Permeability; Cell membrane; Cells; Cessation of life; Communities; Connexin 43; Connexins; Cysteine; Data; Development; Dilated Cardiomyopathy; Duchenne cardiomyopathy; Duchenne muscular dystrophy; Dystrophin; Environment; Exhibits; Fibrosis; Functional disorder; Genetic; Glutamic Acid; Goals; Heart; Heart Diseases; Heart failure; Hela Cells; Histologic; Homeostasis; Impairment; Intercalated disc; Ions; Knock-in Mouse; Knockout Mice; Lead; Longevity; Mechanics; Mediator of activation protein; Medical; Methionine; Molecular; Molecular Target; Mus; Mutant Strains Mice; Mutation; Myocardium; Myopathy; Oxidation-Reduction; Oxidative Stress; Oxides; Paper; Pathologic; Pathology; Pathway interactions; Patients; Permeability; Pharmacologic Substance; Pharmacology; Phosphorylation; Predisposition; Prevention; Production; Proteins; Publishing; Reactive Oxygen Species; Research Proposals; Role; Serine; Signal Transduction; Skeletal Muscle; Stem cells; Stimulus; Stress; Testing; Therapeutic; Triplet Multiple Birth; Utrophin; Xenopus oocyte; cardiogenesis; cellular pathology; driving force; dystrophic cardiomyopathy; gap junction channel; heart function; improved; mdx mouse; mitochondrial metabolism; mutant; novel therapeutic intervention; novel therapeutics; oxidation; premature; prevent; small molecule; therapeutic target

SUMMARY Duchenne muscular dystrophy (DMD) is a progressive muscle disease with severe cardiac complications. Dilated cardiomyopathy is a serious condition apparent in 100% of DMD patients over 18 years of age. Over 40% (and increasing) of DMD patients are dying from heart failure. However, studies of dystrophic cardiomyopathy remain quite limited. The culprit – loss of dystrophin – is a subsarcolemmal protein that protects skeletal and cardiac muscle from mechanically induced damage and regulates a number of downstream cellular pathways. While recent strategies to restore dystrophin levels keep hopes up, the medical and scientific communities are also seeking alternative therapeutic targets downstream of dystrophin. We found key evidence suggesting a detrimental role for connexin-43 (Cx43) in DMD-cardiomyopathy. We showed that Cx43 is upregulated and remodeled in the heart of mdxand mdx:utrophin knockout mice, as well as in the heart of DMD patients. We also showed that selective pharmacological inhibition of Cx43 function or genetic reduction of Cx43 leads to a rescue of arrhythmias and prevention of premature death in stressed mdx mice, as well as protection from the development of long-term cardiomyopathy. We determined that Cx43 in mdx is hypophosphorylated in a specific triplet of serine residues (S325/328/330). Reduction of phosphorylation in this same triplet promotes Cx43 remodeling in a variety of cardiac pathologies, leading to undesired opening of Cx43 hemichannels, arrhythmic susceptibility and irreversible long-term consequences. We believe that cardiac remodeling of Cx43 in DMD contributes to cellular Ca2+ overload and oxidative stress, leading to arrhythmias, fibrosis and consequently, to the development of heart failure. We believe that the phospho-triplet S325/328/330 is the driving force for Cx43 remodeling. In this proposal we will first determine whether expression of a mutant form of Cx43 protects mdx mice from developing cardiac pathology on functional and histological (Aim 1) as well as on cellular (Aim 2) levels. To address this question, Cx43 knockin mice will be crossed with mdx mice, to incorporate the Cx43 phospho-mutation harboring glutamic acids instead of serines in the phospho-triplet thus mimicking permanent phosphorylation. In Aim 3 we will investigate molecular mechanisms responsible for enhanced opening of Cx43 hemichannels in dystrophic hearts. In particular, we will examine if excessive ROS opens the hemichannels. Rigorous blinded functional, histological and biochemical analysis will be conducted.

概括 杜氏肌营养不良症 (DMD) 是一种进行性肌肉疾病，伴有严重的心脏并发症。 扩张型心肌病是一种严重疾病，100% 18 岁以上的 DMD 患者都会出现这种情况。超过 40%（并且还在增加）的 DMD 患者死于心力衰竭。然而，营养不良症的研究 心肌病仍然相当有限。罪魁祸首——肌营养不良蛋白的损失——是一种肌膜下蛋白， 保护骨骼和心肌免受机械引起的损伤并调节许多 下游细胞通路。虽然最近恢复肌营养不良蛋白水平的策略让人们抱有希望，但医学 科学界也在寻找肌营养不良蛋白下游的替代治疗靶点。我们 发现关键证据表明连接蛋白 43 (Cx43) 在 DMD 心肌病中发挥有害作用。我们展示了 表明 Cx43 在 mdx 和 mdx:utropin 敲除小鼠的心脏中以及在 DMD 患者的心。我们还表明，选择性药理学抑制 Cx43 功能或遗传 Cx43 的减少可挽救应激 mdx 小鼠的心律失常并预防过早死亡， 以及防止长期心肌病的发展。我们确定 mdx 中的 Cx43 是 在特定的丝氨酸残基三联体 (S325/328/330) 中低磷酸化。减少磷酸化 相同的三联体在多种心脏病中促进 Cx43 重塑，导致不期望的开放 Cx43 半通道、心律失常易感性和不可逆转的长期后果。我们相信 DMD 中 Cx43 的心脏重塑会导致细胞 Ca2+ 超载和氧化应激，从而导致 心律失常、纤维化，进而发展为心力衰竭。我们相信磷酸三重态 S325/328/330是Cx43重塑的动力。在本提案中，我们将首先确定是否 Cx43 突变体的表达可保护 mdx 小鼠在功能和功能方面免于发生心脏病理学 组织学（目标 1）以及细胞（目标 2）水平。为了解决这个问题，Cx43 敲入小鼠将被 与 mdx 小鼠杂交，整合含有谷氨酸而不是丝氨酸的 Cx43 磷酸突变 在磷酸三联体中，从而模拟永久磷酸化。在目标 3 中，我们将研究分子 负责营养不良心脏中 Cx43 半通道增强开放的机制。特别是，我们 将检查过多的 ROS 是否会打开半通道。严格的盲法功能、组织学和 将进行生化分析。