Mechanical Signaling through Osteoblast Focal Adhesions

描述（由申请人提供）：本提案的总体目标是了解介导骨对机械负荷的合成代谢反应的细胞机制。骨的机械负荷引起骨内空间内的间质液的运动。流体剪切应力（FSS）引起的成骨细胞和骨细胞的机械刺激被假设为在骨对机械载荷的响应中起作用。支持这一假设的观察来自于这样的观察，即与在静态培养条件下维持的细胞相比，在体外由FSS刺激的成骨细胞和骨细胞表现出增加的代谢活性。本申请的中心焦点是确定介导骨细胞对其局部环境的机械敏感性的细胞机制。具体而言，我们建议检查的作用，粘附网站（通常称为焦点接触或焦点粘连）之间的成骨细胞和骨细胞的细胞外基质，介导的整合素细胞粘附分子在调节骨细胞的反应，FSS。这项研究的结果应该提供一个更好的理解的基本细胞和分子机制，介导的骨细胞对机械负荷的反应，通过测试的假设，即局灶性粘连的功能作为mechanoreceptors通过协调与整合素相互作用的细胞质信号分子的活性。骨细胞对单向和振荡流体流的生理相关反应包括增殖活性改变和向合成代谢或成骨细胞表型的分化增强、基因表达改变、前列腺素产生增加、促分裂原活化蛋白激酶（MAPK）和粘着斑激酶（FAK）的活化以及胰岛素样生长因子受体（IGF-1 R）对IGF-1刺激的敏感性。提出了三个具体目标。在目的1中，我们将确定破坏局部粘连的结构完整性对成骨细胞对FSS的生化反应的影响。在目的2中，我们将确定粘着斑激酶（FAK）介导的信号转导通路激活的FSS在机械转导的作用。在目标3中，我们将确定转录因子NMP 4/CIZ在介导的信号转导通路中的作用，该信号转导通路在流体剪切应力作用下通过局部粘连激活。总之，这些研究应该提供新的信息，介导骨机械转导的细胞机制。

DESCRIPTION (provided by applicant): The overall goal of this proposal is understand the cellular mechanisms that mediate the anabolic response of bone to mechanical loading. Mechanical loading of bone induces the movement of interstitial fluid within the spaces inside bone. The resulting mechanical stimulation of osteoblasts and osteocytes caused by fluid shear stress (FSS) is hypothesized to play a role in the response of bone to mechanical loading. Support for this hypothesis comes from the observation that osteoblasts and osteocytes that are stimulated by FSS in vitro exhibit increased metabolic activity compared to cells that are maintained under static culture conditions. The central focus of this application is to determine the cellular mechanisms that mediate the mechanical sensitivity of bone cells to their local environment. Specifically, we propose to examine the role of adhesion sites (commonly referred to as focal contacts or focal adhesions) between osteoblasts and osteocytes to the extracellular matrix that are mediated by integrin cell adhesion molecules in regulating the response of bone cells to FSS. The results of this study should provide an improved understanding of the fundamental cellular and molecular mechanisms that mediate the response of bone cells to mechanical loading by testing the hypothesis that focal adhesions function as mechanoreceptors by coordinating the activity of cytoplasmic signaling molecules that interact with integrins. Physiologically relevant responses by bone cells to unidirectional and oscillatory fluid flow include altered proliferative activity and enhanced differentiation toward an anabolic or osteoblastic phenotype, altered gene expression, increased prostaglandin production, activation of mitogen activated protein kinases (MAPK) and focal adhesion kinase (FAK) and sensitization of the insulin-like growth factor receptor (IGF-1R) to stimulation by IGF-1. Three specific aims are proposed. In Aim 1 we will determine the effect of disrupting the structural integrity of focal adhesions on biochemical response of osteoblasts to FSS. In Aim 2 we will determine the role of focal adhesion kinase (FAK) mediated signal transduction pathways activated by FSS in mechanotransduction. In Aim 3 we will determine the role of the transcription factor NMP4/CIZ in mediating signal transduction pathways activated through focal adhesions in response to fluid shear stress. Together these studies should provide novel information on the cellular mechanisms that mediate mechanotransduction in bone.