Osteocyte differentiation & MCP-3 chemoattractant in mechanical stress responses

成骨细胞是骨细胞的前体。一个特定的遗传程序和基因表达谱是这一分化为成熟骨细胞的过程的基础，但该程序的详细组成部分尚不清楚。我们已经开始定义一个基因表达模式的子集，代表成骨细胞向被活跃矿化的类骨细胞包围的成骨细胞的状态。利用成骨细胞和成骨细胞模型，我们定义了在成骨细胞中低表达但在骨细胞模型中高表达的基因子集。此外，骨细胞中对液体流动作出反应的基因的子集已经被确定。此外，在骨细胞中对液体流动做出反应的基因的子集已经被确定。其中一个编码单核细胞趋化蛋白MCP-3的基因在骨细胞中比成骨细胞高出13到19倍，成为骨细胞，并识别对骨细胞功能重要的基因。短期目标是确定MCP-3在骨细胞功能中的作用。我们的假设是，MCP-3是成骨细胞分化为骨细胞的标志，也是破骨细胞前体向骨募集的关键分子。在这项建议中，我们将讨论1)单核细胞趋化蛋白-3在骨细胞功能中的作用和2)在体外和体内机械应变下单核细胞趋化蛋白-3基因表达的调节。为了实现这些目标，将使用MPO-Y4细胞系、原代成骨细胞、原代成骨细胞以及在培养中形成含有骨样细胞的矿化结节的2T3细胞系。增加液体流动的幅度和频率将在这些模型中测试对MCP-3产量、基因表达模式和功能的影响。两种啮齿动物体内机械应变模型，牙齿移动模型和尺骨疲劳负荷模型将被用于获取和验证在体外模型中发现的基因表达变化。我们还将使用MCP受体零等位基因的转基因小鼠，CCR2小鼠在破骨细胞前体募集方面会有缺陷。这些研究的结果将极大地扩展我们关于骨细胞功能的重要基因的数据库，并将有助于深入了解破骨细胞重新聚集到骨微骨折的机制。

Osteoblasts are the precursors to osteocytes. A specific genetic program and gene expression profile underlies this differentiation process to a mature osteocyte, but the detailed components of that program are unknown. We have begun to define a subset of gene expression patterns that represent the state of the osteoblast as it progresses towards an osteocyte surrounded by osteoid that is actively mineralizing. Using osteoblast and osteocyte models composed of two cell lines, 2T3, an osteoblast-like cell, and MLO-Y4, an osteocyte-like cell, we have defined a subset of genes that are low in osteoblasts and become highly expressed in the osteocytes model. Additionally, a subset of genes in the osteocyte that respond to fluid flow have been identified Additionally, a subset of genes in the osteocyte that respond to fluid flow have been identified. One genes that encodes for Monocyte Chemoattractant Protein, MCP-3, is 13 TO 19 fold higher in osteocytes compared to osteoblast becomes an osteocyte and identify genes important for osteocyte function. The short- term goal is to determine the role of MCP-3 in osteocyte function. Our hypothesis is that MCP-3 is a marker for differentiation of osteoblasts into osteocytes and a critical molecule in recruitment of osteoclast precursors to bone. In this proposal we will address 1) the role of MCP-3 in osteocyte function and 2) the regulation of MCP-3 gene expression under mechanical strain both in vitro and in vivo. To accomplish these goals, the MPO-Y4 cell line, primary osteocytes, primary osteoblasts and the 2T3 cell line that forms mineralized nodules containing osteocyte-like cells in culture will be used. Increasing magnitude and frequency of fluid flow will be tested for effect on MCP-3 production, gene expression patterns and function in these models. Two rodent in vivo models of mechanical strain, the tooth movement model and ulnae-fatigue loading model will be used to access and validate gene expression changes found in the in vitro models. We will also use transgenic mice with a null allele of the MCP receptor, CCR2 mice will have defects in osteoclast precursor recruitment. Results from these studies will dramatically expand our database on genes important for osteocyte function and should also lead to insights into the mechanism of osteoclast recruitment to sties of bone microfracture.