Endothelial lipid droplet turnover and regulation of metabolic function

项目摘要 越来越多的证据表明，血管内皮细胞在肿瘤的发生发展中起着重要的作用。 代谢稳态的调节。因此，内皮功能障碍有助于代谢 与肥胖和胰岛素抵抗相关的紊乱。我们的实验室提供了第一个证据 内皮细胞（EC）具有在体内形成脂滴（LD）的能力，并证明LD周转 (i.e.,合成和降解）是维持EC静止和功能所必需的。具体地说， 药理学抑制LD合成途径中的关键酶，二酰基甘油乙酰转移酶（DGAT） 1和2，导致内质网（ER）应激。或者，内皮特异性脂肪缺失 甘油三酯脂肪酶（ATGL），在LD水解的限速酶，导致内皮细胞的深刻减少， 一氧化氮合酶（eNOS）蛋白水平和一氧化氮（NO）生物利用度。 事实上，这些血管扰动与全身代谢紊乱有关，但其作用 EC LD周转的影响，因为它涉及到血管和代谢功能的控制是未知的。所以在这 根据这项建议，我将定义内皮LD周转在维持血管和代谢中的具体作用。 在正常生理过程中，DIO以及对急性和慢性运动的反应。实现这些 末端，I将产生可诱导的内皮特异性ATGL敲除（iECKO）、DGAT 1-iECKO和DGAT 2-iECKO 小鼠作为工具来剖析LD周转控制血管和全身的分子机制 代谢平衡在目标1中，我将剖析LD周转中断导致的细胞机制， 在正常生理和DIO下，通过严格细致的分子和代谢 特征化在目标2中，我将解释内皮LD周转在维持胰岛素水平中的重要性。 通过全面的体内代谢研究和离体组织分析，对正常生理学和DIO的敏感性进行了研究。 最后，在目标3中，我将建立内皮LD周转在适应性调节中的作用， 使用与目标2类似的方法进行急性运动回合和慢性耐力训练。总的来说， 这些研究将阐明ECLD周转在全身代谢中迄今未知的作用。结果 这些研究将使我们更好地了解血管和代谢功能之间的相互关系 在正常生理过程中以及在肥胖和II型糖尿病的情况下。除了这些研究目标之外， 本提案描述了一个为期五年的强化指导培训计划，其目标是培养校长 研究者（PI，Nabil Boutagy，Ph.D.）成为一个独立的和高影响力的，科学家在血管领域， 生物学和代谢的监督下，他的主要导师，博士威廉Sessa，和共同导师，博士。 杰拉尔德·舒尔曼。此外，还组建了一个由世界级科学家组成的小组，负责监督和 在实现建议的研究和PI的职业发展的各个方面的指导。

Project Summary Accumulating evidence demonstrates that the vascular endothelium plays an important role in the regulation of metabolic homeostasis. In accordance, endothelial dysfunction contributes to metabolic derangements associated with obesity and insulin resistance. Our laboratory provided the first evidence that endothelial cells (ECs) have the capacity to form lipid droplets (LDs) in vivo and demonstrated that LD turnover (i.e., synthesis and degradation) is necessary for maintaining EC quiescence and function. Specifically, pharmacological inhibition of key enzymes in the LD synthesis pathway, diacylglycerol acetyl transferase (DGAT) 1 and 2, led to endoplasmic reticulum (ER) stress. Alternatively, endothelial specific deletion of adipose triglyceride lipase (ATGL), the rate-limiting enzyme in LD hydrolysis, led to a profound reduction in endothelial nitric oxide synthase (eNOS) protein levels and nitric oxide (NO) bioavailability in standard chow fed mice. Indeed, these vascular perturbations have been linked to derangements in whole-body metabolism, but the role of EC LD turnover as it relates to the control of vascular and metabolic function is unknown. Therefore, in this proposal, I will define the specific role of endothelial LD turnover in maintaining vascular and metabolic function during normal physiology, DIO and in response to acute and chronic exercise. To achieve these ends, I will generate inducible, endothelial specific ATGL knockout (iECKO), DGAT1-iECKO and DGAT2-iECKO mice as tools to dissect the molecular mechanisms by which LD turnover controls vascular and whole-body metabolic homeostasis. In aim 1, I will dissect the cellular mechanisms by which disruption in LD turnover leads to endothelial dysfunction under normal physiology and DIO via rigorous and meticulous molecular and metabolic characterization. In aim 2, I will decipher the importance of endothelial LD turnover in maintaining insulin sensitivity in normal physiology and DIO via comprehensive in vivo metabolic studies and ex vivo tissue analysis. Lastly, in aim 3, I will establish the role of endothelial LD turnover in the regulation of adaptations in response to an acute bout of exercise bout and chronic endurance training using a similar approach as aim 2. Collectively, these studies will address a heretofore unknown role of EC LD turnover in whole-body metabolism. Results from these studies will provide a better understanding of the interrelationship between vascular and metabolic function during normal physiology and in the setting of obesity and type II diabetes. In addition to these research aims, this proposal describes a five-year intensive mentored training program with the goal of developing the Principal Investigator (PI, Nabil Boutagy, Ph.D.) into an independent and high impact, scientist in the fields of vascular biology and metabolism under the supervision of his primary mentor, Dr. William Sessa, and co-mentor, Dr. Gerald Shulman. Furthermore, a team of world-class scientists has been assembled to provide oversight and guidance in achieving all aspects of the proposed research and the PI’s career development.