Pseudo-3D Cytoskeleton Dynamics and Signal Activation in Osteocytes under Flow

流动下骨细胞的伪 3D 细胞骨架动力学和信号激活

基本信息

批准号：
7978377
负责人：
X. Edward GUO
金额：
$ 18.17万
依托单位：
COLUMBIA UNIV NEW YORK MORNINGSIDE
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-01 至 2012-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7978377
关键词：
3-Dimensional Actins Address Adhesions Apical Area Basic Science Biochemical Biology Biosensor Bone Tissue Cell Adhesion Cell model Cell physiology Cells Cellular Mechanotransduction Cellular biology Complex Computer Simulation Cytoskeleton Dendrites Etiology Event Extracellular Matrix Fluorescence Resonance Energy Transfer Focal Adhesion Kinase 1 Frequencies Grant Health Image Imaging Techniques Individual Liquid substance Mature Bone Measures Mechanical Stimulation Mechanics Microfilaments Microscopy Microtubules Modeling Morphology Nature Osteoblasts Osteoclasts Osteocytes Osteoporosis Pathway interactions Phosphotransferases Physiological Play Printing Process Property Research Resolution Role Scanning Shapes Signal Transduction Solid Stimulus Stress Surface Techniques Technology Testing Time base bone bone cell cell motility cell type fluid flow high risk in vivo migration monolayer neuronal cell body novel public health relevance response shear stress src-Family Kinases time use

项目摘要

DESCRIPTION (provided by applicant): Osteocytes are three-dimensional (3D) ellipsoidal shaped mature bone cells encased in mineralized extracellular matrix. Abundant evidence has shown that the osteocytes are the key mechanonsensor cells that directly regulate bone-forming osteoblast and bone-removing osteoclast activities. Thus, osteocyte functions are critical to the etiology and treatments of osteoporosis, one of the most challenging health issues in the U.S. The proposed project will develop a pseudo-3 dimensional (3D) real-time microscopy technology to simultaneously visualize cell deformation, actin and microtubule cytoskeleton dynamics, and mechano- signaling activation in real-time using biosensors of osteocyte cells under dynamic fluid flow. The proposed technology also incorporates advanced computational fluid dynamics (CFD)-solid modeling of a cell under fluid flow for the estimation of "real-time" cell viscoelastic properties. Given the dynamic nature of fluid flow, cell deformation, and the timescale of mechanically-induced Src/FAK activation (<0.3 seconds), any imaging technique must have a temporal resolution capable of capturing cell deformation at these timescales. Here, we propose using a real-time "pseudo-3D" technique to image two orthogonal views of a cell simultaneously to greater capture the spatial dynamics of actin and microtubule network deformation, Src or FAK activation, and whole cell mechanical properties of micropatterned, ellipsoidally shaped osteocytes under oscillatory flow. The specific aims of the project are: (1) Simultaneously track and analyze individual cell deformation, actin filament networks, and microtubule networks using pseudo-3D imaging of osteocytes under oscillatory fluid flow and correlate the predicted whole cell mechanical properties with the intracellular actin filament or microtubule network strains and (2) Simultaneously track intracellular actin or microtubule network deformation and Src kinase or FAK activation by fluorescence resonance energy transfer (FRET) biosensors using pseudo-3D imaging of osteocytes under oscillatory fluid flow and correlate the localized biosensor activation with the network strains. This study will have significant impact in the basic science research in the field of bone biology, cellular mechanotransduction, cell adhesion, and the mechanobiology of many adherent cell types. PUBLIC HEALTH RELEVANCE: This R21 application will develop a real-time "pseudo-3D" microscopy technique to image two orthogonal views of a cell under dynamic fluid flow simultaneously to greater capture the spatial dynamics of cytoskeletal deformation, Src or FAK activation, and whole cell mechanical properties of osteocytes. This enabling technology will have great impact in the general fields of bone biology and cellular mechanics for an increased understanding of the role that physical forces play in bone cell mechanotransduction.

描述（由申请人提供）：骨细胞是三维（3D）椭圆形成熟的成熟骨细胞，这些骨细胞包裹在矿化细胞外基质中。大量证据表明，骨细胞是直接调节骨形成成骨细胞和骨骼骨化骨细胞活性的关键机械传感器。因此，骨细胞的功能对骨质疏松症的病因和治疗至关重要，骨质疏松症是美国最具挑战性的健康问题之一，拟议项目将开发一种伪-3维（3D）实时显微镜技术，可同时可视化细胞变形，肌动蛋白和微蛋白细胞骨骼的动态学，并在现象中可视化细胞变形。动态流体流动。提出的技术还结合了流体流动下细胞的高级计算流体动力学（CFD） - 固体建模，以估计“实时”细胞粘弹性性能。鉴于流体流动的动态性质，细胞变形和机械诱导的SRC/FAK激活的时间尺度（<0.3秒），任何成像技术都必须具有能够在这些时间尺度捕获细胞变形的时间分辨率。在这里，我们建议使用实时的“伪3D”技术同时对单元的两种正交视图进行映像，以更好地捕获肌动蛋白和微管网络变形，SRC或FAK激活的空间动力学，以及Micropaterned的全细胞机械性能，椭圆形的，椭圆形的骨化体流动。该项目的具体目的是：（1）同时跟踪和分析单个细胞的变形，肌动蛋白细丝网络和微管网络，使用振荡流体流动下骨细胞的伪成像成像，并将预测的全细胞机械特性与近的胞内纤维纤维纤维纤维细胞或2）相关的整个细胞机械性能（2）相关（2）通过荧光共振能量转移（FRET）生物传感器的变形和SRC激酶或FAK激活，使用振荡流体流动下骨细胞的伪3D成像，并将局部生物传感器激活与网络菌株相关。这项研究将对骨生物学，细胞机械转导，细胞粘附和许多粘附细胞类型的机械生物学领域的基础科学研究产生重大影响。公共卫生相关性：此R21应用程序将开发一种实时的“伪-3D”显微镜技术，以同时同时在动态流体流下图像一个细胞的两个正交视图，以更好地捕获细胞骨骼变形，SRC或FAK激活，以及整个细胞机械特性的空间动力学。这种启示技术将对骨骼生物学和细胞力学的一般领域产生巨大影响，以增加对物理力在骨细胞机械转导中的作用的了解。