Developing Contrast-Enhanced Ultrasound of Thrombosis via SPIO-RL Platelets

描述(由申请人提供)：我们建议开发一种超声技术，用于在输注的血小板相互作用形成动脉血栓时进行对比增强成像。在临床静默、闭塞前阶段检测血栓形成的能力可能会在血栓变得闭塞并导致心脏病发作、中风或坏疽肢体之前识别患者。这项技术还将指导使用相关动物模型开发新的介入策略，以最大限度地减少动脉粥样硬化中的血栓并发症。目前还没有现有的临床成像方法来直接对比血小板或检测闭塞前的临床无症状血栓形成。超顺磁性氧化铁(SPIO)是近年来出现的一种新型超声造影剂，它通过锁定检测SPIO机械耦合的回声材料(细胞或血液凝块)的诱导磁移来提供暗场成像。我们假设，携带SPIO有效载荷的可再水化、冷冻干燥(RL)血小板可以通过磁动势超声(MMUS)提供与动脉血栓形成的功能对比，同时实现长期存储能力。我们的研究策略使用一个灌注室对体外猪动脉进行实时成像，作为一个灵活的平台来表征这一新的成像技术。作为血管内皮细胞反应性的模型，我们首先将血管内皮细胞脱内皮化作为损伤模型，或加入肿瘤坏死因子诱导体外猪动脉炎症。然后，作为血栓形成并发动脉粥样硬化的模型，我们将研究具有动脉粥样硬化斑块的高胆固醇血症猪的动脉。我们的第一个具体目标是确定血栓形成过程中提供MMU造影剂所需的SPIO-RL血小板的最低循环浓度，这将为临床可行性提供基线数据。作为这一目标的一部分，我们将研究流速对血栓形成的影响，以及血栓大小对MMUS对比度的影响，同时优化成像参数。这一目的的一个探索性组成部分是研究是否可以通过MMUS信号的幅度和频率依赖性来提取血栓的硬度，这可能与识别高纤维蛋白原血症等疾病有关。我们的第二个具体目标是确定SPIO-RL血小板被结合到血栓中后SPIO的命运，以便预测药代动力学，并为未来的策略，如治疗有效载荷提供信息。血小板在血栓形成中发挥的积极和重要的作用是一个理想的功能成像平台，有望提供比单功能靶向试剂(如抗体偶联SPIO)更高的灵敏度。这项拟议的工作将验证SPIO-RL血小板在血流动力学条件下检测血栓形成的有效性，并可能导致发现动脉粥样硬化斑块出血或临床前非闭塞性血栓的能力显著提高，从而证明临床干预措施可以减轻闭塞性血栓形成。

DESCRIPTION (provided by applicant): We propose to develop an ultrasound technology for contrast-enhanced imaging of infused platelets as they interact to form arterial thrombi. The ability to detect thrombosis at a clinically silent, pre-occlusive stage can potentially identify patients before the thrombi become occlusive and cause heart attacks, strokes, or gangrenous extremities. This technology will also guide the development, using relevant animal models, of new interventional strategies for minimizing thrombotic complications in atherosclerosis. There are currently no existing clinical imaging methods to directly contrast platelets or to detect pre-occlusive, clinically silent thrombosis. Superparamagnetic iron oxides (SPIOs) have recently emerged as a new contrast agent for ultrasound that offer dark-field imaging via lock-in detection of induced magnetomotion of echogenic materials (cells or blood clots) with which SPIOs are mechanically coupled. We hypothesize that rehydratable, lyophilized (RL) platelets carrying a payload of SPIOs can provide functional contrast to arterial thrombosis via magnetomotive ultrasound (MMUS), while enabling long-term storage capability. Our research strategy employs a perfusion chamber for real-time imaging of ex vivo porcine arteries as a flexible platform to characterize this new imaging technology. As a model of vascular endothelial reactivity, we will first de-endothelialize arteries as an injury model, or add TNF cytokine to induce inflammation of ex vivo porcine arteries. Then, as a model of thrombosis complicating atherosclerosis, we will study arteries from hypercholesterolemic pigs that have atherosclerotic plaques. Our first specific aim is to determine the minimum circulating concentration of SPIO-RL platelets needed to provide MMUS contrast during thrombus formation, which will provide baseline data to inform clinical feasibility. As part of this aim we will study the effects of flowrate on thrombus formation, and thrombus size on MMUS contrast, while optimizing imaging parameters. An exploratory component of this aim is proposed to investigate whether the stiffness of the thrombus can be extracted via the amplitude and frequency-dependence of the MMUS signals, which may be of relevance for identifying disorders such as hyperfibrinogenemia. Our second specific aim is to determine the fate of SPIOs after SPIO-RL platelets have been incorporated into a thrombus, in order to predict pharmacokinetics and to inform future strategies such as therapeutic payloads. The active and essential role that platelets play in thrombus formation is a desirable platform for functional imaging, which is expected to provide improved sensitivity over mono-functionally targeted agents such as antibody-conjugated SPIOs. This proposed work will validate the efficacy of SPIO-RL platelets for thrombosis detection in hemodynamic conditions, and may lead to significant improvement in the ability to reveal atherosclerotic plaque hemorrhage or preclinical, nonocclusive thrombi, justifying clinical interventions to mitigate occlusive thrombosis.