The Role of Interleukin-2 in Vascular Smooth Muscle Cell Homeostasis

描述(由申请人提供)：对平滑肌细胞增殖的控制在动脉粥样硬化、动脉瘤、移植血管病变、血管内支架术后再狭窄和静脉移植物失败等几种病理情况中起着关键作用。尽管血管平滑肌细胞(VSMC)的过度增殖参与了新生内膜的增殖，VSMC的凋亡也参与了动脉瘤的形成和动脉粥样硬化斑块的破裂，但关于SMC的增殖是如何在体内被控制的以及这种控制在疾病过程中是如何失败的，我们还没有了解到很多信息。这种认识上的差距阻碍了旨在纠正SMC功能障碍的新的治疗或预防措施的发展。本实验室最近发现，白介素2(IL-2)在体内对VSMCs的维持起作用。这个实验室的总体目标是了解IL-2的调节和功能，以及两者的损伤如何在人类病理中起作用。本提案的目的是开始建立IL-2对VSMC生物学的影响。中心假说是IL-2促进血管平滑肌细胞分化表型的存活和维持。这一假说将通过以下具体目标进行检验：目的一：建立单体和多聚体IL-2如何影响VSMC功能。这一目标将通过测试IL-2对VSMC功能的几种标准检测的影响来实现。目的II：确定VSMC是否释放硫酸乙酰肝素结合的IL-2，以及释放的IL-2是否影响VSMC的功能。这一目标将通过使用Transwell检测来实现，在Transwell检测中，VSMC从动脉片段中分离出来。将评估IL-2的释放，并将使用肝素酶抑制剂来确定该酶的作用。目的：建立IL-2在体内对动脉瘤形成的影响。这一目标将通过对存在于人类动脉瘤组织样本中的IL-2形式的Western印迹分析来实现，并通过确定IL-2是否可以逆转IL-2缺乏的小鼠的动脉瘤形成来实现。这项拟议的工作是创新的，因为它利用了新的发现，即IL-2与存在于大动脉和小动脉中的硫酸乙酰肝素低聚糖有关，并且在这些位置，硫酸乙酰肝素结合的IL-2影响VSMC的表型和存活率。拟议研究的完成将提供对血管细胞内稳态如何通过IL-2启动的未知途径进行调节的更好的理解。这一贡献意义重大，因为血管细胞动态平衡的调节是多种血管病理的基础，然而针对平滑肌细胞的治疗干预极其有限。由于心血管疾病是美国头号死亡原因，这项工作致力于美国国立卫生研究院治疗人类疾病的使命。从拟议和未来的研究中获得的知识将支持建立新的治疗方法、药物输送系统，甚至是靶向或利用平滑肌细胞的组织结构。 公共卫生相关性：血管平滑肌细胞的异常是许多病理情况的基础，包括动脉粥样硬化、动脉瘤以及静脉移植物和移植器官的失败。拟议的研究涉及更好地了解血管平滑肌细胞在正常和疾病状态下的增殖和死亡是如何调节的。鉴于心血管疾病在美国的流行程度，这项拟议的研究对公共健康具有重要意义。

DESCRIPTION (provided by applicant): Control of smooth muscle cell proliferation plays a critical role in several pathologic conditions including atherosclerosis, aneurysms, transplant vasculopathy, restenosis following endo-surgical stenting of vessels, and failure of vein grafts. Although excess vascular smooth muscle cell (VSMC) proliferation is involved in neointimal hyperplasia, and apoptosis of VSMCs is involved in aneurysm formation and atherosclerotic plaque rupture, much information has yet to be learned about how SMC proliferation is controlled in vivo and how said control fails in disease processes. This gap in knowledge impedes the development of new therapeutic or preventative measures aimed at correcting SMC dysfunction. This laboratory has recently discovered that interleukin-2 (IL-2) contributes to the maintenance of VSMCs in vivo. The overall goal of this laboratory is to understand the regulation and function of IL-2, and how impairment of either contributes to human pathology. The object of this proposal is to begin to establish the impact of IL-2 on VSMC biology. The central hypothesis is that IL-2 promotes the survival and maintenance of a differentiated phenotype in vascular smooth muscle cells. This hypothesis will be tested by the following specific aims: Aim I: Establish how monomeric and multimeric IL-2 influence VSMC function. This aim will be accomplished through testing the influence of IL-2 on several standard assays of VSMC function. Aim II: Determine whether VSMC release heparan sulfate- bound IL-2 and whether the released IL-2 influences VSMC function. This aim will be achieved through the use of a transwell assay, in which VSMC are separated from pieces of artery. Release of IL-2 will be assessed and heparanase inhibitors will be used to determine the role of this enzyme. Aim III: Establish the influence of IL-2 on aneurysm formation in vivo. This aim will be accomplished by Western blot analysis of the forms of IL- 2 present in human aneurysm tissue specimens, and by determining whether IL-2 can reverse aneurysm formation in IL-2 deficient mice. The proposed work is innovative, because it capitalizes on the novel findings that IL-2 is associated with heparan sulfate oligosaccharides present within large and small arteries and that in these location heparan sulfate-bound IL-2 influences VSMC phenotypes and survival. Completion of the proposed studies will provide an enhanced understanding of how vascular cell homeostasis is regulated via a here-to-for unknown pathway initiated by IL-2. This contribution is significant because the regulation of vascular cell homeostasis underlies a multitude of vascular pathologies, yet therapeutic interventions targeting smooth muscle cells are extremely limited. Since cardiovascular disease is the number one cause of mortality in the United States, this work addresses the NIH's mission to treat human disease. Knowledge gained from both proposed and future studies will support the establishment of new therapeutics, drug delivery systems, and even tissue constructs that target or utilizes smooth muscle cells. PUBLIC HEALTH RELEVANCE: Abnormalities in vascular smooth muscle cells underlie many pathologic conditions including atherosclerosis, aneurysms, and failure of vein grafts and transplanted organs. The proposed studies concern gaining a better understanding of how proliferation and death is regulated in vascular smooth muscle cells in both normal and disease states. The proposed research is of great significance to public health, given the prevalence of cardiovascular disease in the United States.