An optical approach to 3-dimensional micro-mechanical imaging of the extra-cellular matrix (ECM)

项目概要/摘要 本提案的目标是开发和验证激光散斑场微流变学（SLIM）技术， 组织ECM的微机械映射，横向分辨率为10 μm，轴向分辨率为60 μm， 穿透深度为5 mm。细胞所感知的ECM硬度正在成为一个突出的 一种先于发病机制并通过协调几乎所有方面来指导其进展的微观机械线索 细胞的行为。ECM的过度和不规则的微机械重塑与广泛的 一系列病理学，包括心血管疾病、纤维化疾病和癌症，它们共同构成了 造成了全球50%以上的死亡尽管如此，我们对潜在机制的理解仍然严重不足。 有限的，因为目前没有成像工具可用于ECM的微机械映射， 与细胞相关的尺度。 SLIM测量随时间变化的散斑强度波动。斑点是一种颗粒状的强度图案，形成时， 相干激光束从组织反向散射。散射粒子的布朗位移 ECM动态地调制散斑波动。这些波动反过来又与 成像组织的粘弹性。在柔顺区域中，无限制的布朗位移引起 快速波动的斑点，而在刚性区域，受约束的运动引起有限的强度变化， 斑点颗粒强度波动的像素相关分析提供了一个2D深度整合图， 组织内的机械性能。然而，该图的分辨率受限于散斑颗粒大小， 由光学器件的数值孔径（NA）设置。此外，由于光的多次散射，散斑波动 被整个照明体积内的散射粒子的布朗位移调制。作为 结果，所评估的图缺乏深度信息。因此，本提案的第一个目标是解决这些问题， 通过引入创新的SLIM平台，能够实现高分辨率、深度分辨率、大视场， 在没有物理扫描和重新聚焦在样品上的情况下，ECM的微机械映射。我们的第二 然后，我们的目标是确定ECM的微观力学特性与已知疾病标志之间的联系 进展，通过专注于乳腺癌作为模型。SLIM在微机械领域的独特能力 ECM的断层摄影能够识别病原体的关键生物力学介质。它还打开了多个 基于靶向细胞-ECM微机械相互作用的途径用于疾病的治疗管理。

Project Summary/Abstract The goal of this proposal is to develop and validate a laser SpeckLe fIeld Microrheology (SLIM) technology for micromechanical mapping of the tissue ECM, with lateral resolution of 10 μm, axial resolution of 60 μm, and a penetration depth of 5 mm penetration depth. ECM stiffness, as perceived by cells, is emerging as a prominent micro-mechanical cue that precedes pathogenesis and directs its progression by orchestrating nearly all aspects of cellular behavior. Excessive and irregular micro-mechanical remodeling of the ECM is implicated in a broad spectrum of pathologies, including cardiovascular disease, fibrotic disorders, and cancer, which together account for over 50% of death worldwide. Nevertheless, our understanding of the underlying mechanisms is severely limited as currently there are no imaging tools available for micromechanical mapping of the ECM at length scales pertinent to cells. SLIM measures the time-varying speckle intensity fluctuations. Speckle is a grainy intensity pattern, formed when a coherent laser beam is back scattered from tissue. Brownian displacements of scattering particles within the ECM dynamically modulate the speckle fluctuations. These fluctuations in turn are intimately related to the viscoelastic properties of imaged tissue. In compliant regions, unrestricted Brownian displacements provoke rapidly fluctuating speckle spots, whereas in rigid areas, restrained motions elicit limited intensity variations of speckle grains. Pixel-wise correlation analysis of intensity fluctuations provides a 2D depth-integrated map of mechanical properties within the tissue. However, the resolution of this map is limited to the speckle grain size, set by the Numerical Aperture (NA) of optics. In addition, due to multiple scattering of light, speckle fluctuations are modulated by the Brownian displacements of the scattering particles within the entire illuminated volume. As a result, the evaluated map lacks depth information. Therefore, the first goal of this proposal is to address these issues by introducing an innovative SLIM platform, capable of high resolution, depth-resolved, large FoV, micromechanical mapping of the ECM, without physical scanning and refocusing on the sample. Our second goal is then to identify the link between the micromechanical properties of ECM and known hallmarks of disease progression, by focusing on breast cancer as a model. The unique capability of SLIM for micro-mechanical tomography of ECM enables identifying the key biomechanical mediators of pathogenies. It also opens multiple avenues based on targeting the cell-ECM micromechanical interactions for therapeutic management of disease.