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

摘要 Duchenne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的疾病，在18岁以上的DMD患者中是100%明显的。完毕 40%(而且还在增加)的DMD患者死于心力衰竭。然而，营养不良的研究 心肌病仍然相当有限。罪魁祸首--肌营养不良蛋白缺失--是一种肌膜下蛋白 保护骨骼肌和心肌免受机械性损伤，并调节许多 下游的细胞通路。虽然最近恢复营养不良蛋白水平的策略保持了希望，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-心肌病中起有害作用。我们展示了 Cx43在mdx和mdx：utroin基因敲除小鼠心脏上调和重塑，以及在 DMD患者的心脏。我们还表明，选择性药物抑制Cx43的功能或遗传 Cx43的减少导致应激MDX小鼠心律失常的挽救和过早死亡的预防， 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中发生低磷酸化(S325/328/330)。在这种情况下，磷酸化的减少 相同的三联体在各种心脏病理中促进Cx43重塑，导致不希望看到的 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患者心肌Cx43重塑导致细胞钙超载和氧化应激，导致 心律失常、纤维化，从而导致心力衰竭的发展。我们认为磷三联体 S325/328/330是Cx43重塑的驱动力。在这份提案中，我们将首先确定 Cx43突变形式的表达对mdx小鼠心脏病变的保护作用 组织学(目标1)以及细胞水平(目标2)。为了解决这个问题，Cx43敲击小鼠将 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，从而模仿永久的磷酸化。在目标3中，我们将研究分子 营养不良心脏中Cx43半通道开放增强的机制。特别是，我们 会检查过多的ROS是否会打开半脑沟。严格的盲目功能，组织学和 将进行生化分析。

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.