Magnetic Targeting of Stents with Endothelial Cells for Prevention of Restenosis

描述（由申请人提供）：在过去的十年里，支架成形术的出现和最近药物洗脱支架的使用导致了血管疾病治疗的范式转变。血管介入的有害后遗症是血管壁不可避免的机械损伤的结果。内皮单层的破坏暴露了底层介质，并诱导了一系列细胞和生物事件，导致血管壁功能异常。增强损伤后血管壁内皮细胞数量的策略可以通过重建管腔屏障和细胞分泌旁分泌因子来限制血栓形成、血管痉挛和新内膜形成等并发症。先前快速“内皮化”植入装置的策略并没有显示出临床应用所需的有效性、安全性和易用性。在此，我们提出了一种将内皮细胞磁性靶向支架的方法，该方法基于使用适度均匀的磁场来最大限度地磁化负载磁性纳米粒子的内皮前体细胞，并在钢支架钢丝网内产生大的局部磁场梯度。这种机制将允许实现最大的磁力，这将导致内皮细胞在血管损伤部位的有效定位。我们在大鼠颈动脉支架模型中使用牛主动脉内皮细胞的初步数据表明了这种方法的可行性。在这个项目中，我们计划研究更多与治疗相关的细胞，这些细胞能够分化为内皮表型细胞（即同种异体血液生长内皮细胞，BOEC和内皮祖细胞，EPC），这些细胞来自同一物种（大鼠），将接受磁细胞治疗。拟议研究的具体目标1将侧重于开发用于进一步磁靶向研究的内皮前体细胞的分离、培养和表征方案。特异性目标2将集中于生物可降解磁性纳米颗粒细胞负载方案的优化，评估细胞负载对细胞形态、生长和功能完整性保存的影响。这一目标也将解决一些机械方面的磁负载和操纵EC细胞，评估其基因表达谱，粘附性和血栓形成性。特异性目的3将致力于利用合适的动物模型定量评估磁性细胞在体内靶向的效率，研究脱靶内皮细胞的器官生物分布，以及检查磁性定位异体内皮细胞对损伤血管壁的有益治疗效果。长期治疗效果也将被评估。我们真诚地相信，所得到的数据，加上同时进行的临床前磁性植入物开发工作，将为磁性细胞靶向血管和其他应用提供广泛实施的策略。

DESCRIPTION (provided by applicant): For the past decade the advent of stent angioplasty and the even more recent use of drug eluting stents have resulted in a paradigm shift in the care of vascular disease. Deleterious sequelae of vascular interventions are the result of unavoidable mechanical damage to the vessel wall. Disruption of the endothelial monolayer exposes the underlying media and induces a cascade of cellular and biological events, resulting in abnormal vascular wall function. Strategies that enhance the number of endothelial cells in the vessel wall following injury may limit complications such as thrombosis, vasospasm, and neointimal formation, through reconstitution of a luminal barrier and cellular secretion of paracrine factors. Previous strategies to rapidly "endothelialize" implanted devices did not show desired efficacy, safety, and ease of use required for clinical applications. Herein we propose a method for magnetic targeting of endothelial cells to stents based on use of a modest uniform magnetic field to both maximally magnetize the magnetic nanoparticle-loaded endothelial precursor cells and produce large local magnetic field gradients within the steel stent wire network. This mechanism will allow achieving maximized magnetic force that will result in efficient localization of endothelial cells at the blood vessel injured site. Our preliminary data using bovine aortic endothelial cells in the rat carotid-stenting model indicate on feasibility of this approach. In this project we plan to study more therapeutically relevant cells that are capable of differentiating into cells with endothelial phenotype (i.e. allogeneic blood outgrowth endothelial cells, BOEC and endothelial progenitor cells, EPC) derived from same species (rats) that will receive magnetic cell therapy. Specific Aim 1 of the proposed research will focus on the development of protocols for isolation, culture and characterization of endothelial precursor cells to be used for further magnetic targeting studies. Specific Aim 2 will concentrate on the optimization of protocols for cell loading by biodegradable magnetic nanoparticles, evaluating the effects of cell loading on cell's morphology, growth, and preservation of functional integrity. This aim will also address some mechanistic aspects of magnetically loaded and manipulated EC cells, evaluating their gene expression profiles, adhesion and thrombogenicity. Specific Aim 3 will be dedicated to quantitatively evaluate the efficiency of magnetic cell targeting in vivo using appropriate animal model, to investigate the organ biodistribution of the off-targeted endothelial cells as well as to examine the beneficial therapeutic effect on the injured vessel wall by magnetically localized allogeneic endothelial cells. The long term therapeutic effect will also be assessed. We sincerely believe that resulting data, coupled with concurrent pre-clinical work on magnetic implant development will provide a widely implementable strategy of magnetic cell targeting in vascular and other applications.