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

项目摘要/摘要 这项提议的目标是开发和验证激光散斑场微观流变学(SLIM)技术 组织细胞外基质的微机械标测，横向分辨率为10μm，轴向分辨率为60μm， 侵彻深度：5毫米的侵彻深度。ECM僵硬，正如细胞所感知的那样，正在成为一个突出的 在发病之前的微机械线索，并通过协调几乎所有方面来指导其进展 细胞行为的特征。过度和不规则的细胞外基质微机械重构牵涉到广泛的 各种病理疾病，包括心血管疾病、纤维性疾病和癌症，这些疾病加在一起 造成全球50%以上的死亡。尽管如此，我们对潜在机制的理解是严肃的 有限，因为目前没有成像工具可用于ECM的详细微机械标测 与单元格相关的比例。 SLIM测量了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，在以下情况下形成 相干激光光束从组织中向后散射。散射体内粒子的布朗位移 ECM对散斑波动进行动态调制。这些波动反过来又与 成像组织的粘弹性特性。在顺应地区，不受限制的布朗位移引发了 快速波动的斑点，而在刚性区域，受限的运动引起有限的强度变化 有斑点的颗粒。强度波动的像素相关分析提供了2D深度集成地图 组织内的机械特性。然而，该贴图的分辨率仅限于散斑颗粒大小， 由光学的数值孔径(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。AS 因此，评估的地图缺乏深度信息。因此，这项提案的第一个目标是解决这些问题 通过引入创新的超薄平台来解决问题，该平台能够高分辨率、深度分辨率、大视场、 ECM的微机械标测，无需对样品进行物理扫描和重新聚焦。我们的第二个 然后，目标是确定ECM的微观机械特性和已知的疾病特征之间的联系 进展，通过将乳腺癌作为模型来关注。用于微机械的超薄的独特能力 细胞外基质的断层扫描可以确定致病的关键生物力学介质。它还可以打开多个 以细胞-细胞外基质微机械相互作用为基础的治疗疾病管理的途径。

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.