Dynamic Single Cell Imaging of Coronary Microvascular Dysfunction in the Failing Heart

尽管几十年来心脏病的临床诊断和治疗取得了进展，但心脏病的流行仍在不断上升。 全球老龄化人口中的心力衰竭。从历史上看，很多注意力都集中在动脉粥样硬化、急性 心肌梗死，以及大血管冠心病的治疗。然而，人们对此知之甚少 机械层面关于心脏中巨大的细小动脉、毛细血管和静脉网络 冠状动脉、冠脉微循环和越来越多的证据表明冠状动脉功能障碍 微循环对包括心力衰竭在内的各种形式的心脏病的影响。体外实验，组织学 分析和计算模型提供了关于冠状动脉微循环是如何 在衰竭的心脏中调节和重塑，但以前的研究一直无法直接评估这个复合体 活体心脏细胞水平的微循环生理学。活体光学显微镜正被用于 神经科学和肿瘤生物学在体内破译动态血管生理学，但这些技术 由于收缩运动造成的严重成像限制，该技术不适用于心脏。我们 最近开创了活体成像的先河，以执行无运动伪影的细胞分辨率 心脏跳动中的显微镜。在这项工作的基础上，这项提案试图调查 应用活体显微镜定量标测冠脉微血管功能障碍的研究 在心脏病动物模型中，微循环向下延伸到毛细血管水平。研究将在小鼠身上进行 比较健康对照组的微循环功能，在运动性肥厚模型中， 在压力超负荷导致病理性肥大和心力衰竭的模型中。此外， 微血管周细胞的特殊fic作用将作为毛细血管血fl的主要调节因子而被研究。 活体共聚焦显微镜和双光子显微镜、多路fl荧光记者和细胞种fic光遗传学 将使用方法来记录和操作微血管flow，并关联异常flow模式 与心肌细胞代谢，在fl炎症反应，和fi的BROSIS。这项工作将带来新的理解 在单细胞水平上对衰竭心脏的微循环病理生理学的研究和对 临床治疗学。

Despite decades of advances in clinical diagnosis and treatment of heart disease, there is a rising epidemic of heart failure in an aging population worldwide. Historically, much focus has been on atherosclerosis, acute myocardial infarction, and treatment of large vessel coronary disease. However, relatively little is known at a mechanistic level about the vast network of small arteries, capillaries and veins in the heart beyond the large coronary arteries, the coronary microcirculation, and increasing evidence links dysfunction of the microcirculation to various forms of heart disease including heart failure. In vitro experiments, histologic analysis, and computational modeling have provided insight about how the coronary microcirculation is regulated and remodels in the failing heart, but prior studies have been unable to directly evaluate the complex microcirculatory physiology of the heart at the cellular level in vivo. Intravital optical microscopy is being used in the neurosciences and tumor biology to decipher dynamic vascular physiology in vivo, but these techniques have not been applicable in the heart due to severe imaging limitations imposed by contractile motion. We have recently pioneered intravital imaging methods to perform motion-artifact free, cellular resolution microscopy in the beating heart. Building on this work, this proposal seeks to investigate mechanisms of coronary microvascular dysfunction by utilizing intravital microscopy to quantitatively map flow in the coronary microcirculation down to the capillary level in animal models of heart disease. Studies will be performed in mice comparing microcirculatory function in healthy controls, in a model of physiologic hypertrophy due to exercise, and in a model of pressure overload leading to pathologic hypertrophy and heart failure. In addition, the specific role of microvascular pericytes will be investigated as a master regulator of capillary blood flow. Intravital confocal and two-photon microscopy, multiplexed fluorescent reporters, and cell-specific optogenetics approaches will be used to record and manipulate microvascular flow, and to correlate abnormal flow patterns with cardiomyocyte metabolism, inflammatory response, and fibrosis. This work will lead to new understanding of microcirculatory pathophysiology in the failing heart at the single cell level and promising new insights for clinical therapeutics.