GENE EXPRESSION PATTERNS IN OSTEOCYTES IN RESPONSE TO LOAD

现在已知，骨骼对机械负荷的适应性的微小变化可导致骨骼的大的变化。 骨骼抗断裂性。骨细胞被认为是骨接收的机械感觉细胞 这些生理信号并以调节其局部微环境的方式做出反应， 全面控制骨的选择性区域中的骨形成和骨吸收。牙本质基质蛋白1， DMP1和基质细胞外磷酸糖蛋白MEPE在骨细胞中高度表达， 响应机械负载。这两种蛋白质都高度定位于骨细胞的小管和陷窝中， 而DMP1主要存在于泪小管壁上。我们的目标是利用这两个基因， 代表骨细胞选择性基因对机械应变的响应，以识别分子信号传导 负责改变骨特性的机制。我们的假设是，特定的骨细胞选择性 机械响应增强子区域存在于DMP1和MEPE的启动子中， 由特定的转录家族途径控制。为了验证这一假设， 提出了具体目标：具体目标1。确定DMP1与MEPE基因的关系 表达模式与体内机械负荷后的应变场分析。具体目标2。确定 小鼠尺骨和股骨骨细胞变形与不同应变和基因水平的关系 DMP1和MEPE顺式调节区的激活。具体目标3。确定顺式调节 DMP1和MEPE基因的区域，其控制骨细胞对选择性负荷的反应。这 该项目的独特之处在于，DMP1和MEPE基因表达将与体内肉眼可见的应变相关 并且具有离体的局部细胞变形。这些基因及其相应的顺式调控区与 报告基因将作为不同负荷条件下骨细胞反应性的灵敏读数， 不同的基因背景。该项目将致力于了解顺式调节系统的 DMP1和MEPE基因对骨细胞的选择性和识别转录因子 负责这种对机械负载的选择性和响应性。该项目的目标将是 使用细胞模型来鉴定分子机制，使用动物模型进行体内验证， 结合工程原理，结合分子和系统生物学方法。 提高抗疲劳性是防止断裂的主要手段。定位骨细胞基因， 选择性响应负荷的通路将提供对预防或治疗重要的信息 骨疾病如废用性骨质疏松症、绝经后骨质疏松症和其他病理性骨质疏松症 骨质流失的情况。

It is now known that small changes in bone adaption to mechanical load can lead to large changes in skeletal resistance to fracture. Osteocytes are believed to be the mechanosensory cells of bone receiving these physiological signals and responding in a manner to regulate their local microenvironment and to globally control bone formation and bone resorption in selective regions of bone. Dentin Matrix Protein 1, DMP1, and Matrix Extracellular Phosphoglycoprotein, MEPE, are highly expressed in osteocytes and respond to mechanical load. Both proteins are highly localized in the canaliculi and lacunae of osteocytes, with DMP1 found predominately on the canalicular walls. Our goal is to use these two genes as representative of osteocyte selective genes responsive to mechanical strain to identify molecular signalling mechanisms responsible for changes in bone properties. Our hypothesis is that specific osteocyte selective and mechanically responsive enhancer regions exist in the promoters of DMP1 and MEPE that are controlled by specific transcription family pathways in response to strain. To test this hypothesis three specific aims are proposed: Specific Aim 1. Determine the relationship between DMP1 and MEPE gene expression patterns with strain field analysis upon mechanical loading in vivo. Specific Aim 2. Determine the relationship of osteocyte deformation in the mouse ulna and femur to different levels of strain and gene activation of the DMP1 and MEPE cis-regulatory regions. Specific Aim 3. Determine the cis-regulatory regions of the DMP1 and MEPE genes that control the response to loading selectively in osteocytes. This project is unique in that DMP1 and MEPE gene expression will be correlated with macroscopic strain in vivo and with local cell deformation ex vivo. These genes and their appropriate cis-regulatory regions linked to reporters will serve as sensitive read-outs of osteocyte responsiveness in different loading conditions in different genetic backgrounds. This project will be devoted to understanding the cis-regulatory systems of both the DMP1 and MEPE genes in terms of their osteocyte selectivity and to identifying transcription factors responsible for this selectivity and responsiveness to mechanical loading. The goals of this project will be accomplished using cell models to identify molecular mechanisms, animal models for in vivo validation, together with engineering principles, combined with a molecular and a systems biology approach. Increased fatigue resistance is a major means to prevent fracture. Mapping osteocyte genes and pathways that are selectively responsive to load will provide information important to prevention or treatment of bone disease such as disuse osteoporosis, post menopausal osteoporosis and other pathological conditions of bone loss.