Endothelial exocytosis and the vascular dysfunction of aging

描述(申请人提供)：老化的内皮细胞的特征是内皮扩张剂NO的活性降低，ROS的产生增加和氧化应激，以及炎症和血栓活性的增加。内皮储存颗粒，包括韦贝尔-帕拉德小体(WPBs)，含有许多在调节血管运动、血栓形成和炎症过程中发挥重要作用的介质，并可通过刺激胞吐作用迅速释放。这些内皮颗粒的胞吐和释放很可能在与衰老相关的血管病理中起着重要作用。然而，以前没有研究直接评估衰老对这一基本机制的影响。在新的初步结果中，我们证明了老化血管的天然内皮细胞在刺激胞吐过程中产生和释放病理生理介质(包括内皮素-1)的能力增强。我们的总体假设是，刺激的胞吐作用增加了内皮介质的释放和生成，导致了衰老过程中的血管功能障碍。我们希望通过增加我们对衰老如何影响这一基本机制的理解，它可能会为治疗与衰老相关的疾病提供新的方法。我们将使用Fischer大鼠临床前衰老模型来检验我们关于年轻和老年血管天然内皮细胞的假设。目的1：研究衰老对内皮细胞分泌和分泌内皮素-1的影响。这些实验将集中于评估老化的内皮细胞在胞吐过程中产生内皮素-1(ET-1)的能力增强。实验将研究ET-1在年轻和衰老血管内皮细胞中的表达、加工和胞吐释放。目的2：探讨一氧化氮在衰老血管内皮细胞WPB胞吐中的作用。这些实验将研究NO信号在调节衰老内皮细胞胞吐中的明显抑制作用。实验将集中在NO调节胞吐作用的机制上。目的3：探讨内皮细胞胞吐在衰老微循环中的病理作用。这些实验将评估失调的内皮细胞胞吐在衰老中的潜在病理生理学作用。这将通过研究胞吐作用在幼年和老年动物对肠系膜循环缺血再灌注的微血管反应中的作用来实现。 与公共健康相关：约翰·霍普金斯大学的内科医生和现代医学的偶像威廉·奥斯勒爵士(1849-1919)评论说，我们的年龄和我们的动脉一样老。我们现在知道，我们的年龄可能只与我们的内皮细胞一样老。本项目旨在分析衰老对内皮功能的一个基本机制(刺激胞吐作用)的影响，并评估其在应激诱导的与器官衰老相关的微血管功能障碍中的作用。

DESCRIPTION (provided by applicant): The aging endothelium is characterized by reduced activity of the endothelial dilator NO, increased production of ROS and oxidant stress, and increased inflammatory and thrombotic activity. Endothelial storage granules, including Weibel-Palade Bodies (WPBs) contain numerous mediators that play important roles in regulating vasomotor, thrombotic and inflammatory processes, and can be rapidly released by stimulated exocytosis. It is likely that exocytosis and release of these endothelial granules play an important role in the vascular pathology associated with aging. However, no previous studies have directly assessed the effects of aging on this fundamental mechanism. In novel preliminary results, we demonstrate that native endothelium of aging blood vessels have increased ability to generate and release pathophysiological mediators (including endothelin-1) during stimulated exocytosis. Our overall hypothesis is that increased release and generation of endothelial mediators by stimulated exocytosis contributes to vascular dysfunction in aging. We hope that by increasing our understanding of how aging affects this fundamental mechanism, it may provide novel approaches to treat diseases associated with aging. We will test our hypotheses on native endothelium of young and aging blood vessels, using the Fischer rat preclinical model of aging. Three specific aims are proposed to: AIM 1: DETERMINE THE INFLUENCE OF AGING ON THE GENERATION AND EXOCYTOSIS OF ENDOTHELIN-1 FROM ENDOTHELIUM. These experiments will focus on evaluating the increased ability of aging endothelium to generate endothelin-1 (ET-1) during exocytosis. Experiments will investigate the expression, processing and exocytotic release of ET-1 from the endothelium of young and aging blood vessels. AIM 2: DETERMINE THE ROLE OF NITRIC OXIDE IN REGULATING ENDOTHELIAL WPB EXOCYTOSIS IN AGING BLOOD VESSELS. These experiments will investigate the apparent decreased inhibitory effect of NO signaling in regulating exocytosis in aging endothelium. Experiments will focus on the mechanisms underlying NO regulation of exocytosis. AIM 3: EVALUATE THE PATHOLOGICAL ROLE OF ENDOTHELIAL EXOCYTOSIS IN THE AGING MICROCIRCULATION. These experiments will evaluate the potential pathophysiological role of dysregulated endothelial exocytosis in aging. This will be achieved by investigating the role of exocytosis in the microvascular responses of young and aging animals to ischemia- reperfusion of the mesenteric circulation. PUBLIC HEALTH RELEVANCE: The Johns Hopkins physician and icon of modern medicine, Sir William Osler (1849-1919) commented that we are only as old as our arteries. We now know that we may only be as old as our endothelium. This project proposes to analyze the influence of aging on a fundamental mechanism of endothelial function (stimulated exocytosis), and to evaluate its role in stress- induced microvascular dysfunction associated with aging organs.