Volumetric time-lapse imaging of biophysical cell-extracellular matrix interactions for systems mechanobiology research

用于系统力学生物学研究的生物物理细胞-细胞外基质相互作用的体积延时成像

• 批准号: 10389834
• 负责人: Steven Graham Adie
• 金额: $ 8.91万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10389834
• 关键词: 3-Dimensional; Address; Adoption; Atomic Force Microscopy; Behavior; Biological Markers; Biophysics; Cell Culture Techniques; Cell Density; Cells; Collagen; Confocal Microscopy; Diagnostic; Environment; Extracellular Matrix; Fluorescence; Future; Image; Lead; Malignant Neoplasms; Measurement; Mechanics; Methods; Neoplasm Metastasis; Population; Research; Resolution; Role; Stress; Surface; Symptoms; System; Techniques; Testing; Three-Dimensional Imaging; Time; Traction; base; biophysical analysis; cancer cell; carcinogenesis; cell behavior; cell motility; cell type; cellular imaging; design; experimental study; fluorescence imaging; imaging capabilities; imaging modality; imaging platform; imaging study; mechanical behavior; mechanical properties; migration; millimeter; novel; reconstruction; targeted treatment; tumor

Project Summary The understanding of cancer has evolved rapidly over the last decade, particularly with discoveries regarding the role of physical factors, such as extracellular matrix (ECM) stiffness and cellular forces, in carcinogenesis. This research has shown that altered ECM stiffness is not just a symptom of tumors, but is now known to trigger the actual onset of and progression of malignancy. Another key finding is that cellular traction stresses increase with increasing metastatic potential, suggesting that cell traction forces could be a biomarker for the likelihood of metastasis. Additionally, it has been found that (2D) collective behavior of cell populations can be significantly different from that of isolated cancer cells, and that cell migratory behavior in 3D matrices is significantly different migration on 2D surfaces. Although this has motivated the adoption of 3D microenvironments in cancer mechanobiology research, current imaging methods to quantify ECM mechanical properties and local cellular forces only provide 2D imaging, or when they do support 3D imaging, they do not provide long-range volumetric measurements of collective mechanical behavior with cellular resolution. The central objective of this proposal is to develop quantitative reconstruction capabilities for OCT-based techniques recently developed by the PI's group for volumetric imaging of cell traction forces and ECM mechanical properties. These new quantitative capabilities will be integrated with a fluorescence confocal microscopy module, to demonstrate a novel imaging platform with unprecedented capabilities for time-lapse imaging studies of biophysical cell-ECM interactions in 3D environments. Aim 1 will develop the capabilities for quantitative 3D reconstruction of ECM mechanical properties and validate it against rheometry and atomic force microscopy (AFM). Aim 2 will demonstrate our OCT-based imaging of 3D cell traction forces using low-density cell cultures, integrating cellular resolution imaging of ECM mechanical properties over millimeter-scale volumes. The demonstration of these novel, integrated imaging capabilities in low-density cell cultures will be followed by a demonstration in dense tumor spheroid cell cultures, where we will compare traction forces and ECM remodeling at the main spheroid boundary versus surrounding invasion strands. Aim 3 will add a confocal fluorescence imaging module to our OCT system, and we will demonstrate that this imaging platform can perform time-lapse reconstruction of 3D cell traction forces and cell-induced changes in ECM mechanical properties in a multiple-cell population migrating in 3D collagen. This will enable the first direct comparison of the time-varying traction forces of different cell types simultaneously migrating in 3D collagen. Our novel 3D imaging platform for systems mechanobiology research could lead to a deeper understanding of potential biophysical (mechanical) hallmarks of cancer, that can be used in the future to design and test new `mechano-therapies' that target/modulate the mechanical properties of the ECM.

项目摘要 在过去的十年中，对癌症的理解迅速发展，尤其是关于 物理因素的作用，例如细胞外基质（ECM）刚度和细胞力，在致癌作用中。 这项研究表明，ECM刚度的改变不仅是肿瘤的症状，而且现在已知会触发 恶性肿瘤的实际发作和进展。另一个关键发现是细胞牵引力增加 随着转移潜力的增加，这表明细胞牵引力可能是生物标志物 转移。此外，已经发现（2D）细胞种群的集体行为可以显着 与孤立的癌细胞不同，3D矩阵中的细胞迁移行为显着不同 在2D表面上迁移。尽管这促使在癌症中采用3D微环境 机械生物学研究，当前量化ECM机械性能和局部细胞的成像方法 力仅提供2D成像，或者当它们支持3D成像时，它们不提供远程体积 通过细胞分辨率测量集体机械行为。该提议的核心目标 是为PI最近开发的基于OCT的技术开发定量重建功能 细胞牵引力和ECM机械性能的体积成像组。这些新的定量 功能将与荧光共聚焦显微镜模块集成，以演示新型成像 具有前所未有的功能的平台，可用于延时成像研究生物物理细胞-ECM相互作用 3D环境。 AIM 1将开发ECM机械定量3D重建的功能 性质并验证其针对流变计和原子力显微镜（AFM）。 AIM 2将证明我们的 使用低密度细胞培养物对3D细胞牵引力的OCT基于OCT的成像，整合细胞分辨率 ECM机械性能的成像在毫米级体积上。这些小说的演示， 低密度细胞培养物中的综合成像功能将在致密肿瘤中进行示范 球形细胞培养物，我们将比较主要球形边界的牵引力和ECM重塑 与周围的入侵链相对。 AIM 3将在我们的OCT系统中添加共聚焦荧光成像模块， 我们将证明该成像平台可以执行3D细胞牵引力的延时重建 在3D中迁移的多细胞种群中的力和细胞诱导的ECM机械性能变化 胶原。这将使不同细胞类型的时变牵引力进行首次直接比较 同时在3D胶原蛋白中迁移。我们的新颖的3D系统机械生物学研究平台 可能会更深入地了解癌症的潜在生物物理（机械）标志，可以使用 将来设计和测试针对/调节机械性能的新的“机械策略” ECM。