Theoretical and Experimental Studies of Cell Reorganization on Deformable Materials

该奖项是根据2009年美国复苏和再投资法案（公法111-5）0854129资助的。大多数细胞产生细胞内力，这些力传递给细胞外基质中的力，并被细胞外基质中的力抵消。这种机械力的平衡对于维持细胞的机械和生化平衡，即体内平衡是必要的。当这种平衡被破坏时，细胞骨架就会重组，试图重建体内平衡。这种平衡重建的一个相关例子是细胞及其肌动蛋白应力纤维垂直于循环矩阵拉伸方向的排列。在大多数动脉树中，动脉内皮细胞被拉长并与血管轴排列，但在动脉粥样硬化易发区域缺乏这种排列。首席研究员之前已经证明，内皮细胞的循环拉伸诱导JNK的激活，JNK是一种参与调节促动脉粥样硬化基因表达的信号蛋白，但当细胞及其应力纤维垂直于拉伸时，JNK的激活会减弱。其他体外和体内研究支持细胞排列和抗动脉粥样硬化细胞表型之间的关系；然而，其机制仍然不清楚。该项目的目标是：1)建立一个包含肌动蛋白周转和肌动蛋白-肌球蛋白相互作用的力学模型，以描述基质变形与肌动蛋白细胞骨架的相关反应性重组之间的动态关系；2)使用牵引显微镜、飞秒激光消融和表达荧光标记肌动蛋白的活细胞显微镜对模型进行测试和完善。在这个项目中建立的模型将为理解机械拉伸和细胞骨架重塑在细胞力学、信号转导和细胞功能中的作用提供一个新颖而全面的框架。这一努力将导致一个前所未有的能力来模拟肌动蛋白细胞骨架的动态变化，这些变化发生在不同的空间和时间的拉伸模式下。此外，定量模型将提高重新解释现有数据的能力，并产生新的实验来阐明拉伸诱导的细胞骨架重组的机制。重要的是，该项目将为信号转导模型提供基础，其中输入是机械刺激，而不是生化配体。所提出的模型提供了一个工具，了解贴壁细胞的力学性能如何随时间变化，通过细胞骨架重塑。这些知识将为在组织工程、手术决策和心血管疾病预后中使用机械刺激来调节细胞功能提供指导。通过向公众提供模型软件，并将本项目中发展的概念纳入本科和研究生课程，该模型将得到广泛传播。此外，拟议的项目将为本科生和研究生提供更多的机会，包括来自代表性不足群体的学生，在首席研究员的实验室进行研究。

This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5)0854129KaunasMost cells generate intracellular forces that are transmitted to, and countered by, forces in the extracellular matrix. This mechanical force balance is necessary for maintaining both mechanical and biochemical cell equilibrium, i.e. homeostasis. When this balance is disturbed, the cell cytoskeleton reorganizes in an attempt to reestablish homeostasis. A relevant example of this reestablishment of equilibrium is the alignment of cells and their actin stress fibers perpendicular to the direction of cyclic matrix stretch. Arterial endothelial cells, which are elongated and aligned with the vessel axis in most of the arterial tree, lack such alignment at regions prone to atherosclerosis. The Principal Investigator has previously shown that cyclic stretching of endothelial cells induces activation of JNK - a signaling protein involved in regulating pro-atherogenic gene expression - but that JNK activation subsides as cells and their stress fibers align perpendicular to stretch. Other studies, both in vitro and in vivo, support a relationship between cell alignment and an anti atherogenic cell phenotype; however, the mechanism remains obscure. The goals of this project are to 1) develop a mechanical model that incorporates actin turnover and actin-myosin interactions to describe the dynamic relationships between deformations in the matrix and associated reactive reorganization of the actin cytoskeleton; and 2) test and refine the model using traction microscopy, femtasecond laser ablation, and microscopy of live cells expressing fluorescently-labeled actin. The model developed during this project will provide a novel and comprehensive framework for understanding the roles of mechanical stretch and cytoskeletal remodeling on cell mechanics, signal transduction, and cell function. This effort will result in an unprecedented capability to model the dynamic changes in the actin cytoskeleton that occur in response to diverse spatial and temporal patterns of stretch. Further, a quantitative model will result in an improved ability to reinterpret existing data, as well as generate new experiments to elucidate the mechanisms of stretch-induced cytoskeletal reorganization. Importantly, this project will provide the foundation for models of signal transduction where the inputs are mechanical stimuli, rather than biochemical ligands. The proposed model provides a tool to understand how the mechanical properties of adherent cells change with time through cytoskeletal remodeling. Such knowledge will provide guidance toward the use of mechanical stimuli to regulate cell function in tissue engineering, surgical decision-making, and prognosis of cardiovascular disease. The model will be broadly disseminated by providing public access to the model software and incorporating the concepts developed in this project into undergraduate and graduate courses. Further, the proposed project will provide additional opportunities for undergraduate and graduate research, including students from underrepresented groups, in the laboratory of the Principal Investigator.