High resolution 3D functional imaging of cerebrovascular perfusion in mice

描述(由申请人提供)：非侵入性血流成像技术--向下至毛细血管水平的分辨率--对于研究和诊断有血管病因或受累的疾病是至关重要的。目前的光学成像技术能够达到这一分辨率，但大多数通常局限于二维，如激光散斑和光学固有信号成像技术，而其他技术，如共焦显微镜，成像深度有限(&lt；300？m)。然而，通常需要以毛细血管水平的分辨率对微循环床内深层的血管血液灌流进行三维(3D)可视化，以揭示微血管网络的详细结构，从而可以量化组织的体积流变学和灌流状态。我们建议开发、优化和表征3D光学微血管成像(OMAG)，这是一种新的、非侵入性的光学成像方法，可以用于评估毛细血管水平分辨率的血管灌注，在微循环床深处。OMAG包括将近红外光照射到活样本上，然后使用新的算法分析后向散射光，以并行获得体积微结构结构和血液灌流图像。为了更好地理解OMAG如何感知血流，我们将开发源自组织样本的背向散射信号的严格数学和实验模型。我们将开发OMAG算法来定量评估组织中的动态血液灌流。我们将利用一个小鼠模型(在正常或病理生理条件下)来帮助发展OMAG。我们将获得小鼠脑血流的经颅图像，并将使用这些数据来改进和验证OMAG系统。之所以选择这种小鼠模型，是因为它被发现是量化单个血管和组织中的脑血流量的有用工具，以便更好地了解人类脑血管疾病的机制和治疗干预措施。我们将通过诱导小鼠急性血栓性缺血性中风(ATIS)来操纵皮质血流。我们将通过OMAG获得的血流图像与模型的预期结果(实验终点)相关联。我们还将通过证实溶栓酶和组织型纤溶酶原激活剂在小鼠ATIS期间加速再通和/或防止血栓形成进展，来验证OMAG对研究实验性中风的有用性和药物干预的效果。该项目的一个令人兴奋的方面是，在初步研究中，我们一直获得高质量的OMAG图像，通过小鼠的脑血管树一直到毛细血管水平的血液灌流。这些图像是通过小鼠完整的头骨获得的，不需要注射染料、造影剂或外科开颅手术。使用我们初步的OMAG系统，我们已经能够捕捉到实验性缺血性中风期间局灶性脑血流灌注停止，并随后可视化脑血管闭塞随时间的空间进展。这项研究的直接结果有两个：a)一个新的成像工具，将使研究人员和临床医生更好地了解组织灌流和缺血组织损伤的病理生理学；以及b)具体地说，关于皮质脑损伤和缺血组织灌流，将在建立良好的小鼠模型中获得重要的新信息。一旦以这种方式验证，我们预计，随着新问题的出现和新治疗策略的开发，OMAG系统将在未来的中风和其他疾病的研究中发挥相当大的优势。我们打算在拟议的研究中开发的OMAG系统将是紧凑、快速、光学稳定的，并且易于实施，可在研究实验室和临床环境中使用。

DESCRIPTION (provided by applicant): Non-invasive techniques for imaging blood flow - down to capillary-level resolution - are of paramount importance in order to research and diagnose diseases that have vascular etiology or involvement. Current optical imaging techniques are able to achieve this resolution, but most are typically confined to two dimensions, such as laser-speckle and optical intrinsic-signal imaging techniques, while other techniques, like confocal microscopy, has limited imaging depth (<300¿m). However, a three-dimensional (3D) visualization of vascular blood perfusion deep within microcirculatory beds at capillary-level resolution is often required to reveal the detailed architecture of the perfused microvascular network so that the volumetric rheology and perfusion status of the tissue can be quantified. We propose to develop, optimize, and characterize 3D optical micro-angiography (OMAG), a novel, non-invasive optical imaging method that can be used to assess vascular perfusion at capillary-level resolution, deep within microcirculation beds. OMAG involves shining a near infrared light onto a living sample and then analyzing the backscattered light using novel algorithms to obtain, in parallel, volumetric microstructural architecture and blood perfusion images. We will develop rigorous mathematical and experimental models of the backscattering signals originating from a tissue sample in order to better understand how OMAG senses blood flow. We will develop OMAG algorithms to quantitatively assess dynamic blood perfusion in tissue. We will utilize a mouse model (under either normal or pathophysiologic conditions) to assist in developing OMAG. We will obtain transcranial images of cerebral blood flow in the mice and will use the data to improve and validate the OMAG system. This mouse model was chosen because it has been found to be a useful tool in quantifying cerebral blood flow in individual vessels and tissues, in order to better understand mechanisms of human cerebrovascular disease and therapeutic interventions. We will manipulate cortical blood flow by inducing acute thrombotic ischemic stroke (ATIS) in the mice. We will correlate the blood flow images obtained by OMAG with the expected outcomes (experimental end points) of the models. We will also verify the usefulness of OMAG for studying experimental stroke and the effects of pharmacological interventions by confirming that thrombolytic enzyme, tissue plasminogen activator accelerates reperfusion and/or prevents thrombosis progression during ATIS in mice. An exciting aspect of this project is that in the preliminary studies, we have consistently achieved high-quality OMAG images of blood perfusion through the cerebrovascular tree down to the capillary level in mice. These images were obtained through the intact skull of the mouse without the need for dye injections, contrast agents, or surgical craniotomy. Using our preliminary OMAG system, we have been able to capture focal cerebral perfusion cessation during experimental ischemic stroke, with subsequent visualization of the spatial progression of cerebrovascular occlusions over time. The immediate outcomes of this research are twofold: a) a new imaging tool that will allow researchers and clinicians to better understand the pathophysiology of tissue perfusion and ischemic tissue injury, and b) specifically, with respect to cortical brain injury and ischemic tissue perfusion important new information will be obtained in a well-established mouse model. Once validated in this way, we anticipate that the OMAG system will be used to considerable advantage in future studies of stroke and other disorders as new questions arise and as new therapeutic strategies are developed. The OMAG system that we intend to develop in the proposed research will be compact, fast, optically stable, and easily implemented and adaptable in both research laboratories and clinical environments.