Assessment of Corneal Fibroblast Biomechanical Behavior

角膜成纤维细胞生物力学行为的评估

• 批准号: 7498655
• 负责人: W MATTHEW PETROLL
• 金额: $ 6.94万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-02-01 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7498655
• 关键词: 3-Dimensional; ATP phosphohydrolase; Actinin; Adhesives; Behavior; Biology; Biomechanics; Biomedical Engineering; Biophysical Process; Cell Density; Cell physiology; Cells; Collagen; Collagen Fibril; Confocal Microscopy; Cornea; Cytoskeletal Modeling; Data; Development; Developmental Biology; Elements; Equilibrium; Exertion; Experimental Models; Extracellular Matrix; Fibrillar Collagen; Fibroblasts; Focal Adhesions; Funding; Generations; Goals; Growth Factor; Image; Image Analysis; In Vitro; Lasers; Lead; Measures; Mechanical Stress; Mechanics; Mediating; Modeling; Molecular; Monitor; Morphogenesis; Myosin Light Chain Kinase; Myosin Light Chains; Myosin Type II; Pathway interactions; Pattern; Phosphorylation; Platelet-Derived Growth Factor; Play; Process; Proteins; Regulation; Relaxation; Research; Rho-associated kinase; Role; Stress; Stretching; System; Testing; Time; Tissue Engineering; Tissues; Wound Healing; ZYX gene; base; cell behavior; digital imaging; driving force; in vivo; innovation; insight; lysophosphatidic acid; migration; myosin phosphatase; response; rho; time use

Cell-matrix mechanical interactions drive fundamental processes such as developmental morphogenesis, wound healing, and the organization of bioengineered tissues. The overall goal of this research is to determine the underlying cellular and molecular mechanisms that regulate these critical biophysical processes in corneal fibroblasts, which should ultimately lead to more effective approaches to directing their biomechanical behavior in vivo and in vitro. In the first funding period, we developed a new experimental model for directly investigating cell-matrix mechanical interactions inside 3-D fibrillar collagen matrices. Data obtained using this innovative approach has provided new insights into potential mechanisms regulating sub-cellular force generation, matrix reorganization, as well as the modulation of cell behavior by mechanical stress which together lead to the following Hypotheses: 1) The balance between Rac and RhoA activity plays a central role in regulating the mechanical behavior of corneal fibroblasts inside 3-D matrices at the sub- cellular level. These effects are mediated by dynamic changes in cytoskeletal and focal adhesion organization, as well as differential regulation of myosin light chain (MLC) phosphorylation, 2) Corneal fibroblasts actively respond to increases or decreases in local matrix stress (including that produced by neighboring cells); these responses are mediated by compensatory and reciprocal changes in Rho and Rac activation, and 3) The pattern and amount of collagen matrix reorganization can be modulated by altering the balance between Rho and Rac activity, and is enhanced by cell-cell mechanical interactions. To test these hypotheses, we propose the following Specific Aims: 1) Determine the role of Rho and Rac in regulating cytoskeletal organization, mechanical behavior, and sub-cellular force generation by corneal fibroblasts inside 3-D matrices using our time-lapse imaging system, 2) Measure the dynamic mechanical response of corneal fibroblasts to changes in ECM stress; and, 3) Directly assess the process of cell-induced 3-D collagen matrix reorganization (alignment of collagen fibrils), and the roles of Rho and Rac in regulating this process. These studies will be the first to assess the roles of Rho and Rac on the subcellular pattern of force generation and ECM reorganization within 3-D collagen matrices. Overall, this research should provide unique insights into the mechanisms controlling corneal fibroblast migration, contraction, and matrix reorganization," critical ] processes in the.fields qfdevelopmental biology, wound healing and tissue engineering.

细胞-基质机械相互作用驱动基本过程，如发育形态发生， 伤口愈合和生物工程组织的组织。本研究的总体目标是 确定潜在的细胞和分子机制，调节这些关键的生物物理 角膜成纤维细胞的过程，这最终应该导致更有效的方法来指导他们的 在体内和体外的生物力学行为。在第一个资助期，我们开发了一种新的实验性的 直接研究三维纤维胶原基质内细胞-基质机械相互作用的模型。 使用这种创新方法获得的数据为潜在的调节机制提供了新的见解。 亚细胞力的产生，基质重组，以及通过机械调节细胞行为 1）Rac和RhoA活性之间的平衡起着重要的作用， 在调节角膜成纤维细胞的机械行为中的核心作用， 细胞水平。这些作用是由细胞骨架和粘着斑组织的动态变化介导的， 以及肌球蛋白轻链（MLC）磷酸化的差异调节，2）角膜成纤维细胞 积极响应局部基质应力的增加或减少（包括由相邻基质产生的应力）， 细胞）;这些反应由Rho和Rac激活的补偿性和相互变化介导，并且 3)胶原基质重组的模式和数量可以通过改变平衡来调节， Rho和Rac活性之间的相互作用，并通过细胞-细胞机械相互作用增强。为了验证这些假设， 我们提出以下具体目的：1）确定Rho和Rac在调节细胞骨架中的作用 组织、机械行为和三维内角膜成纤维细胞产生的亚细胞力 2）测量角膜的动态力学响应， 3）直接评估细胞诱导的3-D胶原基质的过程 重组（胶原纤维的排列），以及Rho和Rac在调节这一过程中的作用。这些 研究将是第一个评估Rho和Rac对力产生的亚细胞模式的作用， 三维胶原基质中的ECM重组。总的来说，这项研究应该提供独特的见解， 控制角膜成纤维细胞迁移、收缩和基质重组的机制”至关重要， 在发育生物学、伤口愈合和组织工程领域的方法。