Endothelial Progenitor Cell Seeded Self-Expanding Vascular Stent

描述(申请人提供)：目的：本研究的目标是开发一种将自扩张镍钛合金支架与自体外周血源性内皮祖细胞(EPC)相结合的技术，以减少支架内常见的潜在破坏性并发症-支架内再狭窄和血栓形成。镍钛支架用于治疗外周血管疾病患者的血管狭窄，到2020年，美国将有超过2000万患者受到这种疾病的影响。目的和方法：本研究的目的是优化一种从外周血中提取的内皮祖细胞植入镍钛合金支架的新技术，并评价外周血内皮祖细胞在支架表面的作用。我们计划制造一种创新的镍钛输送系统，通过在支架外鞘中激光钻孔微孔，允许在植入后几分钟内种植支架，这样当迫使含有EPC的溶液通过支架侧口时，可以利用溶剂阻力原理在支架内侧表面种植内皮祖细胞。我们还将优化支架成像和再压迫方法。在Aim I.B.中，我们将在体外流动回路中评估生理剪应力下镍钛合金支架上的细胞保留率，并测试细胞壁磷蛋白是否介导了黏附。在AIM I.C.中，我们通过使用Ionics/Siever一氧化氮分析仪定量介质样品中的亚硝酸盐和通过Western印迹分析不同流动条件下内皮祖细胞在支架表面的血栓调节蛋白表达来评估一氧化氮的产生。在目标2中，我们将在猪的动物模型中测试EPC涂层的镍钛支架。Aim 2.A.将允许我们测试我们的新方法，即在镍钛合金支架上植入荧光标记的内皮祖细胞，并在短期存活研究中将它们植入3只实验动物体内。接下来，在目标2.B中，我们将在7头猪的左右颈外静脉分别植入一枚EPC涂层支架和一枚未涂层(对照)支架。治疗和对照支架在血栓形成和再狭窄方面的差异将通过组织形态计量学半定量分析以及扫描和荧光显微镜进行评估。影响：本方案中所获得的知识将加深我们对切应力下内皮祖细胞在镍钛上的功能和黏附机制的理解。如果成功，我们将为一项新技术提供概念验证，这将显著提高自膨胀支架的性能，并可能使数百万患者受益。 公共卫生相关性：这项研究的目标是开发一种新技术，以避免镍钛支架危险和危及生命的堵塞。为了实现这一目标，我们的目标是在支架植入人体之前立即在支架上涂上内皮祖细胞。如果成功，这将使支架畅通无阻，从而使数百万患者受益。

DESCRIPTION (provided by applicant): Objectives: It is the goal of this study to develop a technology to line self-expanding Nitinol stents with autologous peripheral blood-derived endothelial progenitor cells (EPCs) in order to reduce the common and potentially devastating complications of in-stent restenosis and thrombosis. Nitinol stents are used to treat narrowing of blood vessels in patients with peripheral vascular disease, an illness that will affect over 20 million patients in the U.S. by 2020. Aims and Methods: It is our Aim I in this study to optimize a novel technology to seed Nitinol stents with EPCs derived from peripheral blood and to evaluate EPC function on the stent surface ex vivo. We plan on manufacturing an innovative Nitinol delivery system that allows seeding the stents within minutes of implantation by laser drilling micropores in the outside stent sheath so that EPCs can be seeded on the inside strut surface by using the principle of solvent drag when forcing EPC-containing solution through a stent side- port. We will also optimize stent imaging and re-compressing methods. In Aim I.B. we will assess cell retention on Nitinol stents under physiological shear stress in an ex vivo flow circuit and test whether adhesion is mediated by cell wall phosphoproteins. In Aim I.C. we evaluate nitric oxide production by quantifying nitrite in media samples with an Ionics/Sievers Nitric Oxide Analyzer and thrombomodulin expression by EPCs on the stent surface with Western blot analysis under different flow conditions. In Aim 2, we will test EPC-coated Nitinol stents in a porcine animal model. Aim 2.A. will allow us to test our novel methods of seeding Nitinol stents with fluorescently-labeled EPCs and implanting them into 3 pilot animals in a short-term survival study. Following, in Aim 2.B. we will implant one EPC-coated and one uncoated (control) stent into the right and left external jugular vein of 7 pigs. The differences in thrombosis and restenosis between treated and control stents will be evaluated by histomorphometric semiquantitative analyses, as well as scanning- and fluorescent microscopy. Impact: The knowledge gained in this proposal will deepen our understanding of the function and adhesion mechanism of EPCs on Nitinol under shear stress. If successful, we will have provided proof-of-concept for a novel technology, which will significantly improve the performance of self-expanding stents and may benefit millions of patients. PUBLIC HEALTH RELEVANCE: The goal of this research is to develop a novel technology in order to avoid the dangerous and life-threatening clogging of Nitinol stents. In order to achieve this goal we aim to coat stents with EPCs immediately before implanting them in into the human body. If successful, this will render stents free from obstructions - thus benefitting millions of patients.