VEGF-C/VEGFR3 AND LYMPHATIC TRANSPORT OF CHOLESTEROL FROM ATHEROSCLEROTIC PLAQUE

项目总结 我们的长期目标是了解如何去除斑块中的巨噬细胞和胆固醇以促进 疾病消退。我们最近发现，淋巴管是胆固醇的通道 从动脉壁和其他组织中取出。胆固醇逆向转运(RCT)是指 胆固醇从体内被动员起来，通过粪便排泄。关于动脉粥样硬化， 从巨噬细胞中动员胆固醇以清除和排泄是最相关的。多年来，许多人 关于如何将胆固醇从巨噬细胞动员到高密度脂蛋白(HDL-C)上的细节浮出水面。 然而，关于高密度脂蛋白-胆固醇随后是如何从组织中排出的，人们知之甚少，包括 动脉壁的动脉粥样硬化斑块等部位，在进入排泄途径之前回到血浆中。 20世纪80年代初，几位医生通过临床或实验观察，提出了一种新的治疗方法--S 淋巴运输受损与动脉粥样硬化之间的联系。然而，数量上的重要性 RCT中的淋巴管还没有被检查，除了一项令人信服的研究估计Net 人体淋巴中高密度脂蛋白胆固醇的流量是相当大的。最近，我们使用了实验小鼠模型，其中 淋巴流动的通畅可通过手术或遗传方法进行调节。在皮肤方面，我们能够完全 切断淋巴流动，使我们能够证明淋巴管在数量上是 巨噬细胞RCT后胆固醇向血浆的动员。在动脉粥样硬化影响的主动脉壁中，我们还 使用一种外科方法追踪来自于 以脉搏追逐的方式形成斑块。通过外科手术将主动脉移植到受体体内，并重新吻合 抗VEGFR3单抗阻断或不阻断淋巴管系统。这种封锁极大地保留了D6- 动脉粥样硬化动脉中的胆固醇，提示淋巴管在胆固醇动员中起关键作用 从皮肤上观察到的来自大动脉的。在目标1中，我们将采取关键的下一步，改进方法，将 使我们能够更好地量化淋巴管在从主动脉中清除胆固醇中的作用，并评估 淋巴管的阻断是否真的在主动脉壁局部起作用。在初步数据中，我们表明 血管内皮生长因子受体3的配体血管内皮生长因子C治疗载脂蛋白E/-小鼠可恢复受损的淋巴转运 在高胆固醇血症后，允许我们测试假设，血管内皮生长因子-C作用于淋巴管 治疗上维持巨噬细胞胆固醇后胆固醇转运出斑块的关键途径 外流受到刺激。这一假设提出了一个尚未在该领域得到解决的根本问题： 有效的斑块消退真正依赖于斑块中胆固醇的去除，只有胆固醇的去除。 来自巨噬细胞，或者两者都不是？我们的研究设计是解决这个根本问题的理想方案，同时 同时深入挖掘支持淋巴管功能的新概念可能有助于解决 动脉粥样硬化斑块中的炎症。

PROJECT SUMMARY Our long-term goal is to understand how to remove macrophages and cholesterol from plaques to promote disease regression. We recently discovered that lymphatic vessels serve as the conduits by which cholesterol is removed from the artery wall and other tissues. Reverse cholesterol transport (RCT) is the process by which cholesterol is mobilized from the body for excretion through the feces. With respect to atherosclerosis, the mobilization of cholesterol from macrophages for removal and excretion is most relevant. Over the years, many details emerged regarding how cholesterol is mobilized from macrophages to be loaded onto HDL (HDL-C). However, little was known about how HDL-cholesterol subsequently makes its way out of tissues, including sites like atherosclerotic plaques of the artery wall, to return to plasma before entering pathways for excretion. A handful of clinical or experimental observations led a few physicians in the early 1980's to propose a connection between impaired lymphatic transport and atherosclerosis. However, the quantitative importance of lymphatic vessels in RCT had not been examined, aside from a compelling study that estimated that the net flux of HDL-C through human lymph is substantial. Recently, we utilized experimental mouse models where the patency of lymphatic flow could be modulated by surgical or genetic methods. In skin, we were able to fully abrogate lymphatic flow allowing us to demonstrate that lymphatics are quantitatively the major route for cholesterol mobilization to plasma following macrophage RCT. In atherosclerosis-affected aortic walls, we also used a surgical approach to track deuterium-labeled cholesterol ([2H] D6-cholesterol; D6-cholesterol) from plaques in a pulse-chase manner. Aortas were surgically transplanted into recipients with re-anastomosis of the lymphatic vasculature blocked or not with anti-VEGFR3 mAb. This blockade significantly retained D6- cholesterol in the atherosclerotic aorta, suggesting a key role for lymphatic vessels in cholesterol mobilization from the aorta as observed in skin. In aim 1, we will take a critical next step with refined approaches that will allow us to better quantify the role of lymphatic vessels in cholesterol removal from the aorta and to assess whether the blockade on lymphatics is truly acting locally at the aortic wall. In preliminary data, we show that treating apoE-/- mice with VEGF-C, the ligand for VEGFR3, restores impaired lymphatic transport that occurs following hypercholesterolemia, allowing us to test the hypothesis that VEGF-C acts on lymphatic vessels to therapeutically sustain a critical route for cholesterol transport out of plaques after macrophage cholesterol efflux is stimulated. This hypothesis raises a fundamental question not yet addressed in the field: does effective plaque regression truly depend upon cholesterol removal from plaques, only upon cholesterol removal from macrophages, or perhaps neither? Our study design is ideal to address this fundamental issue while simultaneously digging deeply into a new concept that supporting lymphatic vessel function may help resolve inflammation in the atherosclerotic plaque.