Development of an In Vivo Stem Cell Tracking Method Using Ultrasound Imaging

描述（由申请人提供）：本申请涉及广泛的挑战领域（14）干细胞和特定的挑战主题14-GB-102：成像干细胞迁移和分化。在西方国家，由心肌梗死和随后的不良左心室重构引起的充血性心力衰竭是发病率、死亡率和医疗保健支出的主要原因。尽管在心力衰竭的治疗方面取得了进展，但目前的药理学和机械疗法不能解决主要的潜在问题，即心肌细胞的死亡。心脏自身有限的自我修复能力进一步加剧了当代疗法的局限性。这种困境激发了人们对心脏修复治疗的探索，并引发了对干细胞向受损心肌输送治疗的探索。在急性心肌梗死的动物研究中，细胞制剂如未分级成人骨髓或骨髓源性间充质干细胞（MSC）的递送改善了灌注和心室功能指数。有益效果的机制尚未完全了解，部分原因是方法有限，用于连续跟踪交付后的干细胞命运。最近的数据支持这样的假设，即MSC的旁分泌功能占主导地位，以促进心肌愈合，血管生成，内源性自我修复和减轻炎症。令人鼓舞的临床前结果迅速导致了急性梗死细胞治疗的临床试验，这就迫切需要安全，非侵入性，易于获得的方法来跟踪和量化交付后的干细胞。事实上，治疗方案的优化和对治疗性细胞递送的机械见解的获得与连续跟踪干细胞体内生物分布和活力的能力有关。具有可视化干细胞的临床潜力的当前成像方法利用用于磁共振或核成像的探针。这些技术在辐射暴露、空间分辨率、干细胞遗传修饰的要求、技术复杂性以及对连续成像的阻碍方面具有局限性。因此，为了解决成像技术中的这一差距，目前的研究应用将开发一种基于超声的技术，用于在体内可视化外源递送的干细胞。声学活性气体填充的聚合物微泡将被开发为用于在治疗递送之前离体标记MSC的造影剂。待测试的总体假设是MSC摄取微泡将使得能够使用临床可用的超声扫描仪在体内可视化干细胞运输。该项目将开始微泡化学的体外验证和优化，声学参数和细胞毒性的测试;进行小动物体内概念验证;然后将成像技术应用于MSC治疗的临床相关大型动物模型。这些研究将最终开发出一种安全、非侵入性、便携式、高分辨率、体内、真实的时间超声方法，用于可视化治疗性递送的MSC。预计干细胞的对比增强超声成像将在评价用于改善人群中缺血性心脏病和充血性心力衰竭负担的基于细胞的疗法中发挥关键作用。 公共卫生相关性：基于细胞的治疗，如间充质干细胞（MSC）的管理，是一种有前途的新方法来修复心肌梗死后的心脏。这一策略的成功实施需要更深入地了解MSC赋予治疗益处的机制，这反过来又需要了解局部或全身递送后MSC的分布和命运。该提案旨在开发一种基于超声的方法，用于可视化可用于患者的MSC。这种新的成像技术可能有助于MSC治疗的临床应用，最终可能减少心肌梗死和充血性心力衰竭的死亡和残疾。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (14) Stem Cells, and specific Challenge Topic 14-GB-102: Imaging Stem Cell Migration and Differentiation. Congestive heart failure resulting from myocardial infarction and subsequent adverse left ventricular remodeling is a major cause of morbidity, mortality and health care spending in Western nations. Despite advances in the treatment of heart failure, current pharmacologic and mechanical therapies do not resolve the principal underlying issue, which is the death of cardiomyocytes. The limitations of contemporary therapies are further exacerbated by the heart's own limited ability for self-repair. This dilemma has motivated a search for reparative treatments for the heart, and led to the exploration of therapeutic delivery of stem cells to injured myocardium. In animal studies of acute myocardial infarction, delivery of cell preparations such as unfractionated adult bone marrow or marrow-derived mesenchymal stem cells (MSCs) has improved perfusion and indices of ventricular function. The mechanisms of beneficial effect are incompletely understood, partly because methods are limited for serial tracking of stem cell fate after delivery. Recent data favor the hypothesis that paracrine functions of MSCs predominate to facilitate myocardial healing, angiogenesis, endogenous self-repair, and mitigate inflammation. Encouraging pre-clinical results have rapidly led to clinical trials of cell therapy in acute infarction, which has created a critical need for safe, non-invasive, readily available methods for tracking and quantifying stem cells after delivery. Indeed, the optimization of treatment protocols and acquisition of mechanistic insights into therapeutic cell delivery, are linked to the ability to serially track stem cell biodistribution and viability in vivo. Current imaging methods with clinical potential for visualizing stem cells utilize probes for magnetic resonance or nuclear imaging. These have limitations in terms of radiation exposure, spatial resolution, the requirement for genetic modification of stem cells, technical complexity, and resulting impediments to serial imaging. Accordingly, to address this gap in imaging technology, the current research application will develop an ultrasound-based technology for visualizing exogenously delivered stem cells in vivo. Acoustically active gas-filled polymer microbubbles will be developed as contrast agents for ex vivo labeling of MSCs prior to therapeutic delivery. The overall hypothesis to be tested is that MSC uptake of the microbubbles will enable in vivo visualization of stem cell trafficking using clinically available ultrasound scanners. The project will begin with in vitro validation and optimization of microbubble chemistry, testing of acoustic parameters and cytotoxicity; proceed to in vivo proof of concept in small animals; and then apply the imaging technology to a clinically relevant large animal model of MSC therapy. These studies will culminate in the development of a safe, non-invasive, portable, high resolution, in vivo, real time ultrasound method for visualizing therapeutically delivered MSCs. It is anticipated that contrast- enhanced ultrasound imaging of stem cells will play a pivotal role in the evaluation of cell-based therapies for ameliorating the burden of ischemic heart disease and congestive heart failure in human populations. PUBLIC HEALTH RELEVANCE: Cell-based therapies, such as administration of mesenchymal stem cells (MSCs), are a promising new approach to repair the heart after myocardial infarction. Successful implementation of this strategy requires a greater insight into the mechanism by which MSCs confer therapeutic benefit, which in turn requires an understanding of the distribution and fate of MSCs after local or systemic delivery. This proposal aims to develop an ultrasound-based approach for visualizing MSCs that can be used in patients. This new imaging technology may facilitate the clinical application of MSC therapy, which may ultimately decrease death and disability from myocardial infarction and congestive heart failure.