A 4D Traction Force Microscope for the mapping of cellular mechanical stresses

描述（由申请人提供）：细胞产生的机械应力在许多生理过程中至关重要，包括胚胎发生、细胞迁移、细胞增殖和组织形成。然而，目前还没有方法可以测量在3-D支架中粘附和迁移期间施加的细胞牵引力。这些信息对于理解细胞-细胞和细胞-细胞外基质（ECM）相互作用在伤口愈合、癌症转移、心血管疾病和慢性炎症等过程中的作用至关重要。为了部分解决这一需求，几年前开发了一种称为牵引力显微镜的技术，以量化和绘制细胞在其基底上产生的牵引场（1，2）。这项技术提供了有关细胞迁移和粘附机制的有价值的信息，并证明了正常和疾病状态力学的关键差异。然而，这种目前使用的技术是有限的-国家的最先进的是一个2-D计算的细胞产生的力在一个平面基板上。已经确定的是，2D基底缺乏3D ECM在体内提供细胞的生理学现实环境。因此，为了更准确地概括细胞微环境，这些测量需要在3-D中进行，其中细胞的所有侧面都能够与细胞外基质结合，使基质变形并使用细胞产生的牵引应力进行迁移。该提案旨在开发和构建一种仪器（硬件）和一种计算算法（软件），用于计算嵌入3D矩阵中的细胞所施加的应力，以研究ECM中细胞粘附和迁移的动态过程，这些过程模拟了天然的体内环境。我们预计，正如2D牵引力显微镜已被广泛应用一样，我们的工具将引起广大科学家的广泛兴趣，他们对许多不同生理系统和疾病条件下的细胞迁移和粘附机制感兴趣。 公共卫生相关性（由申请人提供）：在该提案中，我们计划开发一种4D牵引力显微镜（硬件和软件），该显微镜可以测量细胞在3D空间和时间中粘附和迁移期间对其基底施加的力。这样的装置对于理解在生理过程中驱动细胞迁移的分子机制是必不可少的，所述生理过程例如伤口愈合、组织形成以及在包括癌症转移和动脉粥样硬化的疾病进展期间。

DESCRIPTION (provided by applicant): Cell-generated mechanical stresses are critical during a number of physiological processes, including embryogenesis, cell migration, cell proliferation and tissue formation. However, there is no method currently available to measure cellular traction forces exerted during adhesion and migration in 3-D scaffolds. This information is critical for understanding the contribution of cell-cell and cell-extracellular matrix (ECM) interactions in processes such as wound healing, cancer metastasis, cardiovascular disease, and chronic inflammation. To partially address this need, a technique called Traction Force Microscopy was developed several years ago to quantify and map the traction field created by a cell on its substrate (1, 2). This technique has provided valuable information about the mechanisms of cell migration and adhesion and has demonstrated key differences in the mechanics of normal and diseased states. However, this currently used technique is limited--- the state-of-the-art is a 2-D calculation of the cell-generated forces on a planar substrate. It is well-established that 2D substrates lack the physiologically realistic environment that a 3D ECM provides cells in vivo. Therefore, to more accurately recapitulate the cellular microenvironment, these measurements need to be done in 3-D, where all sides of the cell are capable of binding to the extracellular matrix, deforming the substrate and using cell-generated traction stresses to migrate. The proposal seeks to develop and build an instrument (hardware) and a computational algorithm (software) that calculates the stresses exerted by cells embedded in 3D matrices for studies of the dynamic processes of cell adhesion and migration in ECMs which mimic the native in vivo environment. We expect that just as 2D Traction Force Microscopy has been widely implemented; our tool will be of wide-spread interest to the large community of scientists interested in mechanism of cell migration and adhesion for a number of different physiological systems and disease conditions. PUBLIC HEALTH RELEVANCE (provided by applicant): In this proposal, we plan to develop a 4D traction force microscope (hardware and software) that can measure the forces that cells exert against their substrate during adhesion and migration in 3D space and time. Such a device is essential to understand the molecular machinery that drives cell migration in physiological processes such as wound healing, tissue formation, and during the progression of diseases including cancer metastasis and atherosclerosis.