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

描述（由申请方提供）：平滑肌细胞增殖的控制在几种病理状态中起关键作用，包括动脉粥样硬化、动脉瘤、移植血管病变、血管腔内手术支架植入术后再狭窄和静脉移植物失效。虽然过度的血管平滑肌细胞（VSMC）增殖参与新生内膜增生，并且VSMC的凋亡参与动脉瘤形成和动脉粥样硬化斑块破裂，但是关于SMC增殖如何在体内被控制以及所述控制如何在疾病过程中失败的许多信息还有待了解。这种知识上的差距阻碍了旨在纠正SMC功能障碍的新治疗或预防措施的开发。本实验室最近发现，白细胞介素-2（IL-2）有助于维持体内VSMCs。该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导致人类病理学。本提案的目的是开始确定IL-2对VSMC生物学的影响。中心假设是IL-2促进血管平滑肌细胞中分化表型的存活和维持。这一假设将通过以下具体目标进行检验：目的I：确定单体和多聚体IL-2如何影响VSMC功能。这一目标将通过测试IL-2对VSMC功能的几种标准测定的影响来实现。目的二：探讨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.这一目的将通过使用transwell测定来实现，其中VSMC与动脉碎片分离。将评估IL-2的释放，并使用乙酰肝素酶抑制剂来确定该酶的作用。目的III：建立IL-2对体内动脉瘤形成的影响。这一目标将通过对人动脉瘤组织标本中存在的IL- 2形式进行蛋白质印迹分析，并通过确定IL-2是否可以逆转IL-2缺陷小鼠中的动脉瘤形成来实现。拟议的工作是创新的，因为它利用了新的发现，即IL-2与大动脉和小动脉内存在的硫酸乙酰肝素寡糖相关，并且在这些位置硫酸乙酰肝素结合的IL-2影响VSMC表型和存活。完成拟议的研究将提供一个增强的了解血管细胞的稳态是如何通过一个迄今为止未知的途径启动的IL-2的调节。这一贡献是重要的，因为血管细胞稳态的调节是多种血管病理的基础，但靶向平滑肌细胞的治疗干预非常有限。由于心血管疾病是美国的头号死亡原因，这项工作解决了NIH的使命，以治疗人类疾病。从拟议的和未来的研究中获得的知识将支持建立新的治疗方法，药物递送系统，甚至靶向或利用平滑肌细胞的组织结构。 公共卫生相关性：血管平滑肌细胞的凋亡是许多病理状态的基础，包括动脉粥样硬化、动脉瘤以及静脉移植物和移植器官的衰竭。拟议的研究涉及更好地了解在正常和疾病状态下血管平滑肌细胞的增殖和死亡是如何调节的。鉴于美国心血管疾病的流行，这项拟议中的研究对公共卫生具有重要意义。

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.