Quantitative In Vivo Optical Imaging of Vascular Response to Hind Limb Ischemia

描述（由申请人提供）：外周动脉疾病（PAD）估计影响多达20%的65岁以上的美国成年人，并导致下肢功能和整体生活质量下降。治疗性血管生成，刺激新血管以取代功能障碍的患病动脉的功能，代表了一种有前途的相对较新的方法，可以恢复外周循环和PAD患者的生活质量。小鼠后肢缺血模型是一种常用的临床前系统，用于测试新的治疗性血管生成策略，包括生长因子递送、基因治疗和细胞治疗。后肢缺血模型也为研究体内新血管形成的生物学机制提供了一个明确的和先前表征的平台。然而，由于一个或多个缺点，包括缺乏提供的定量数据，用户的主观性，空间分辨率不足，缺乏纵向体内成像能力，以及需要采购多组小鼠来分析血管结构和功能，目前缺乏用于表征该模型系统中的血管生成反应的分析技术。当前提案的中心目标是开发一种光学活体成像技术，该技术可用于采集关于血管形态、血流和氧合的共配准3D图像数据。所获得的定量数据集将提供可从单剂量麻醉下的单只小鼠获取的互补结构-功能数据量的显著改善。此外，由于可以在一只小鼠中进行重复成像，因此应加快临床前和基础研究的通量，并降低实验成本。提出了两个目标，即（1）技术开发和（2）相对于传统的“金标准”技术的技术验证。所提出的技术有可能为血管生成研究人员提供具有高度影响力的创新性新工具，并加速开发用于治疗PAD重大临床问题的新治疗策略。 公共卫生相关性：外周肢体缺血的小动物模型在理解缺血组织的新血管形成的基本机制和促血管生成疗法的临床前测试中起着不可或缺的作用。然而，一个显着的缺点存在于可用性的定量，高分辨率的方法，纵向评估血管形态和功能的活动物。在这里，我们建议开发一种新的光学成像方法，能够同时评估血管形态，血流和氧合在活的小鼠，而不需要外源性造影剂管理。该技术将能够对血管结构/功能进行更可靠的评估，预计将加速血管生成机制控制和治疗性血运重建技术开发方面的新发现。

DESCRIPTION (provided by applicant): Peripheral arterial disease (PAD) is estimated to affect as many as 20% American adults over 65 years of age and results decreased lower extremity function and overall quality of life. Therapeutic angiogenesis, stimulation of new blood vessels to replace the function of dysfunctional, diseased arteries, represents a promising, relatively new approach that could restore peripheral circulation and quality of life for PAD patients. The mouse hind limb ischemia model is a commonly used preclinical system for testing new therapeutic angiogenesis strategies including growth factor delivery, gene therapy, and cell therapy. The hind limb ischemia model also provides a well-defined and previously-characterized platform for studying the biological mechanisms of neovascularization in vivo. However, analytical techniques for characterization of the angiogenic response in this model system are at present lacking due to one or more shortcomings including lack of quantitative data provided, user subjectivity, inadequate spatial resolution, lack of longitudinal in vivo imaging capability, and the requirement for procuring multiple sets of mice for analysis of both vessel structure and function. The central goal of the current proposal is to develop an optical, intravital imaging technique that can be used to acquire co-registered, 3D image data on vessel morphology, blood flow, and oxygenation. The acquired quantitative data sets will provide a significant improvement in the amount of complementary structure-function data that can be taken from a single mouse under a single dose of anesthesia. Furthermore, throughput of preclinical and basic studies should be accelerated and experimental costs decreased because repeated imaging can be carried out in a single mouse. Two aims are proposed that span (1) technique development and (2) technique validation relative to traditional "gold standard" techniques. The proposed technique has the potential to provide innovative, new tools of high impact to angiogenesis researchers and to accelerate development of new therapeutic strategies for treatment of significant clinical problem of PAD. PUBLIC HEALTH RELEVANCE: Small animal models of peripheral limb ischemia play an integral role in understanding the basic mechanisms of neovascularization of ischemic tissues and preclinical testing of pro-angiogenic therapies. However, a significant shortcoming exists in the availability of quantitative, high resolution methodologies for longitudinal assessment of blood vessel morphology and function within living animals. Here, we propose to develop a novel optical imaging approach capable of simultaneous assessment of vascular morphology, blood flow, and oxygenation in living mice without the need for exogenous contrast agent administration. This technique will enable more robust assessments of vascular structure/function and is anticipated to accelerate new discoveries on the mechanistic control of angiogenesis and the development of therapeutic revascularization technologies.