The Contribution of Melanocyte-like Cells to Atrial Function and Development

描述(申请人提供)：我们最近在人类和小鼠的肺静脉和心房中描述了一组新的黑素细胞样细胞。在发育和成熟的心脏中，黑素细胞样细胞的模式与目前已知的任何细胞谱系都不同，并且发现于经常引发临床房性心律失常的解剖区域。这些细胞表达独特的转录信号，不同于心房肌细胞或真皮黑素细胞。有趣的是，分离的小鼠黑素细胞样细胞是电可兴奋的，并产生心房肌样动作电位。我们发现，在人和小鼠的心脏黑素细胞中表达的DOPAChrome互变酶(DCT)的遗传缺失揭示了这些细胞中的一种病理状态，即动作电位延长和后去极化。此外，成熟的DCT基因敲除小鼠保留了黑素细胞样细胞，心脏结构正常，但对房性心律失常的易感性增加。虽然心脏中有黑素细胞样细胞的野生型小鼠在基线时没有增加房性心律失常，但与心脏中没有黑素细胞样细胞的c-kit突变小鼠相比，在用毒鼠碱激动剂卡巴胆碱激发时，它们确实有更多的房性心律失常。此外，DCT基因敲除小鼠在使用活性氧清除剂治疗时，房性心律失常较少。因此，黑素细胞样细胞可能参与房性心律失常对增加的应激(如自主神经刺激或活性氧)的反应，这通常会导致临床房性心律失常。尽管我们初步确定了黑素细胞样细胞在正常生理和病理生理状态下的功能，但仍不清楚。此外，尽管我们有一些证据表明黑素细胞样细胞是可兴奋的，并可能影响心律失常的发生；但这些细胞潜在的电生理学特征需要进一步研究，以了解它们对心律失常的潜在贡献。因此，我们提出了一系列使用基因工程小鼠模型的体外和体内实验，以表征这些细胞的细胞电生理学，并确定它们对房性心律失常的贡献。提出的具体目标包括：1)阐明分离的小鼠黑素细胞样细胞电兴奋性的电压依赖电流以及黑素细胞样细胞对房性心律失常触发的直接贡献，2)确定自主神经刺激对DCT阳性黑素细胞样细胞兴奋性的影响及其对房性心律失常的贡献，3)评估活性氧对DCT阳性黑素细胞样细胞兴奋性的影响及其对房性心律失常的影响，以及4)研究黑素细胞样细胞在正常心脏中的作用。我们将获得的关于黑素细胞样细胞的基本生物学的知识可能会为我们理解心房电生理学开辟新的途径，并有可能改变对房性心律失常发病机制的认识。

DESCRIPTION (provided by applicant): We recently described a novel population of melanocyte-like cells in the pulmonary veins and atria of humans and mice. In the developing and mature heart, melanocyte-like cells are found in a pattern unlike that of any currently known cell lineage and are found in anatomic regions that often give rise to clinical atrial arrhythmia triggers. These cells express a unique transcription signature that is distinct from that of atrial myocytes or dermal melanocytes. Interestingly, isolated murine melanocyte-like cells are electrically excitable and generate atrial myocyte-like action potentials. We have found that genetic deletion of the enzyme dopachrome tautomerase (Dct), which is expressed by both human and murine cardiac melanocytes, unmasks a pathological state with action potential prolongation and afterdepolarizations in these cells. Furthermore, mature Dct knockout mice retain melanocyte-like cells and have structurally normal hearts, yet display increased susceptibility to atrial arrhythmias. While wild-type mice with melanocyte-like cells in their hearts do not have increased atrial arrhythmias at baseline, they do have more atrial arrhythmias when challenged with the muscarinic agonist carbachol compared to c-kit mutant mice that lack melanocyte-like cells in their hearts. In addition, Dct knockout mice have fewer atrial arrhythmias when treated with reactive oxygen species scavengers. Hence, melanocyte-like cells may contribute to atrial arrhythmias in response to increased stresses (i.e. autonomic stimulation or reactive oxygen species) that commonly induce clinical atrial arrhythmias. Despite our initial characterization, the function of melanocyte-like cells during normal physiologic and pathophysiologic states remains obscure. Furthermore, while we have some evidence melanocyte-like cells are excitable and may influence arrhythmogenesis; the underlying electrophysiologic characteristics of these cells require further investigation to understand their potential contribution to arrhythmias. Therefore, we are proposing a series of in vitro and in vivo experiments using genetically engineered mouse models to characterize the cellular electrophysiology of these cells and determine their contribution to atrial arrhythmias. The specific aims proposed include: 1) elucidating the voltage-dependent currents underlying the electrical excitability of isolated murine melanocyte-like cells and the direct contribution of melanocyte-like to atrial arrhythmia triggers, 2) determining the effects of autonomic stimulation upon Dct-positive melanocyte-like cellular excitability and their contribution atrial arrhythmias, 3) assessing the effects of reactive oxygen species upon the excitability of Dct-positive melanocyte-like cells and their influence upon atrial arrhythmias, and 4) investigating the role of melanocyte-like cells in the normal heart. The knowledge we will gain about the basic biology of melanocyte-like cells is likely to open new avenues in our understanding of atrial electrophysiology, with the potential for paradigm shifting insights into the pathogenesis of atrial arrhythmias.