Mechanical Loading of Cells Through Single Matrix Fibers

描述（由申请人提供） 机械负荷在组织重塑中起着重要作用， 细胞行为的方式依赖于模态，频率和振幅 的装载。然而，细胞能够分化的机制 体内的动态信号仍然是未知的。通过改变 粘连灶内细胞膜-受体-基质相互作用 机械刺激时，细胞可以以一种方式感知和响应动态负荷， 依赖于幅度和频率以及特异性底物-整合素对 涉案为了检验这一假设，本提案的目标是：（1） 开发一种结合细胞外基质的单纤维测力装置 蛋白质在体外向细胞表面上的受体施加动态载荷，（2） 使用以下方法量化纤维表面的粘着斑组装 GFP-桩蛋白和测量发育中细胞骨架增强 在生理频率范围内对加载协议的响应，以及（3） 确定不同配体对粘着斑形成的影响， 纤维表面增强的发展。了解 细胞反应，在局灶性粘连的数量和分布方面， 由细胞骨架强化产生的力，动态载荷 通过各种细胞外基质纤维的应用将阐明 细胞能够区分机械负荷的机制 不同的频率和振幅。

DESCRIPTION (provided by applicant) Mechanical loading plays a fundamental role in tissue remodeling by regulating cell behaviors in a manner dependent on the modality, frequency, and amplitude of loading. However, the mechanisms by which cells are able to differentiate dynamic signals in vivo remain unknown. By altering cytoskeletal-receptor-matrix interactions within focal adhesions in response to mechanical stimuli, cells may sense and respond to dynamic loading in a manner dependent on magnitude and frequency and specific substrate-integrin pair involved. To test this hypothesis, the objectives of this proposal are to: (1) develop a single fiber force-measuring device with bound extracellular matrix proteins to apply dynamic loads to receptors on a cell surface in vitro, (2) quantify the assembly of focal adhesions at the surface of fibers using GFP-paxillin and measure the development cytoskeletal reinforcements in response to loading protocols over a range of physiologic frequencies, and (3) determine the effects of different ligands on focal adhesion formation and the development of reinforcements at the surface of fibers. An understanding of cellular response, in terms of number and distribution of focal adhesions and force generation resulting from cytoskeletal reinforcement, to dynamic loads applied through various extracellular matrix fibers will elucidate the mechanisms by which cells are able to differentiate mechanical loading of various frequencies and amplitudes in vivo.