Uniform field controlled magnetic cell targeting to stents

描述(申请人提供)：这项拟议的研究将探索这样一个假设，即磁引导下内皮细胞(EC)靶向支架动脉可以减少支架血管成形术后的血管再闭塞(再狭窄)。以再内皮化为导向的策略有可能预防再狭窄，同时避免目前使用的治疗的副作用。然而，为了充分实现这一潜力，需要显著提高细胞递送效率。这项研究将探索两个新的想法：1)应用 磁性纳米颗粒(MNP)用于为EC提供足够的磁响应性，使其能够靶向输送到支架动脉；以及2)使用统一的磁场控制的EC磁靶向可逆磁化支架，以增强再内皮化和抑制支架植入后的再狭窄。目前的研究将针对以下具体目标：目标1：关于其物理性质、细胞相容性、摄取和降解动力学的表征。生物可降解的荧光标记的MNP将使用聚合物沉淀法的改进而制成。MNP的大小、组成和磁性将被表征，它们的细胞相容性、内在化和降解动力学将是Aim 1实验的主要终点。用交流梯度磁强计测量了MNP和经MNP处理的EC的磁响应性。动力学研究将在培养细胞中使用荧光法和全球FQrster共振能量转移测量进行。MNP的细胞相容性将通过荧光细胞毒性分析来确定。目的2：EC靶向和再内皮化研究。磁性与非磁性给药后动脉的定位和组织分布 在大鼠颈动脉支架模型中，将分别采用体内生物发光法和发光法对稳定表达萤火虫荧光素酶的MNP-EC进行研究。治疗后1天、7天和28天将比较支架靶向细胞的局部动力学特征，并用定量免疫组织化学方法确定内皮化程度。目的3：靶向EC的抗再狭窄作用。在磁性条件下，MNP负载EC的动物再狭窄程度将通过计算机形态计量学来确定，并在分娩后4周比较非磁性输送和只植入支架的对照。

DESCRIPTION (provided by applicant): The proposed research will explore the hypothesis that magnetically guided targeting of endothelial cells (EC) to stented arteries can reduce vessel reocclusion (restenosis) following stent angioplasty. Re-endothelialization oriented strategies have potential to prevent restenosis while avoiding the side effects of the currently used therapies. However, in order to fully realize this potential a significant improvement in cell delivery efficiency is required. This study will investigate two novel ideas: 1) the application of magnetic nanoparticles (MNP) for providing EC with magnetic responsiveness adequate for their targeted delivery to stented arteries; and 2) using a uniform field-controlled magnetic targeting of EC to reversibly magnetizable stents for enhanced reendothelialization and inhibition of restenosis post stenting. The present studies will address the following specific aims: AIM 1: MNP characterization with respect to their physical properties, cell compatibility, uptake and degradation kinetics. Biodegradable fluorescent-labeled MNP will be formulated using a modification of the polymer precipitation approach. The size, composition and magnetic properties of MNP will be characterized, and their cell compatibility, internalization and degradation kinetics will be the main endpoints of the Aim 1 experiments. The magnetic responsiveness of MNP and EC treated with MNP will be measured by alternating gradient magnetometer. The kinetic studies will be performed in cultured cells using fluorimetry and global FQrster Resonance Energy Transfer measurements. Cell compatibility of MNP will be determined using fluorimetric cell toxicity assays. AIM 2: EC targeting and reendothelialization studies. Arterial localization and tissue distribution after magnetic vs. non- magnetic delivery of MNP-impregnated EC stably expressing firefly luciferase will be studied by in vivo bioluminescence and luminometry, respectively, in the rat carotid stenting model. The local kinetic profiles of stent-targeted cells will be compared 1, 7 and 28 days post treatment with endothelialization determined using a quantitative immunohistochemical strategy. AIM 3: antirestenotic efficacy of targeted EC. The extent of restenosis in animals treated under magnetic conditions with MNP-loaded EC will be determined by computerized morphometry and compared that in non-magnetic delivery and 'stenting only' controls four weeks post delivery.