INTEGRIN/ECM INTERACTIONS IN BONE FORMATION AND REMODELING

细胞与细胞外基质和邻近细胞（固态环境）的相互作用对于发育期间的组织形态发生以及整个生命过程中的组织稳态和重塑至关重要。这些实验室先前使用培养的成骨细胞的研究表明，整合素与纤连蛋白的相互作用在成骨细胞分化和存活中起关键作用。拟议研究的总体目标是使用转基因小鼠模型测试这些观察结果的生理相关性。在一个这样的模型中，截短形式的整合素β 1亚基已在转基因小鼠的成熟成骨细胞中表达。表型的初步表征揭示了出生后年轻动物中成骨细胞的骨基质沉积缺陷和成骨细胞的过度骨重塑。在目标1中，将通过与项目6合作，使用组织形态计量学和先进的成像技术分析组织结构和基质以及矿物质质量，在骨组织水平进一步表征这种表型。还将通过测定转基因表达对从野生型和转基因小鼠分离的成骨细胞的粘附、迁移和存活的影响，在细胞水平上表征表型。最后，将与项目2合作研究这种转基因对生物力学特性和修复能力的影响。目的2的研究将集中在纤维连接蛋白在成骨细胞分化和存活中的作用。N-末端70 kDa基质组装体和胶原结合结构域将在转基因小鼠的成骨细胞中表达。该结构域以前被证明可以抑制培养的未成熟的原代成骨细胞的分化，并引发成熟培养的成骨细胞的凋亡。将使用目标1所示的方法和相互作用表征表型。此外，将与项目2合作，在牵张成骨模型中测试完整纤连蛋白或纤连蛋白片段加速或干扰骨骼修复的能力。由于目前还没有动物模型，测试纤维连接蛋白及其整合素受体在成骨细胞功能和骨重建的作用，这些研究应该提供新的见解与骨骼修复和骨骼退行性疾病的相关性。

Cell interactions with extracellular matrix and with neighboring cells (the solid state environment) are critical for tissue morphogenesis during development, and for tissue homeostasis and remodeling throughout life. Previous studies from these laboratories using osteoblastic cells in cultured, have shown that integrin interactions with fibronectin play critical roles in both osteoblast differentiation and survival. The overall goal of the proposed studies is to test the physiologic relevance of these observations using transgenic mouse models. In one such model a truncated form of the integrin beta 1 subunit has been expressed in mature osteoblasts of transgenic mice. Preliminary characterization on the phenotype has revealed defective bone matrix deposition by osteoblasts and excessive bone remodeling by osteoblasts in post-natal young animals. In Aim 1, this phenotype will be characterized further at the bone tissue level by analysis of tissue architecture and matrix and mineral quality, using histomorphometry, and advanced imaging technologies in collaboration with project 6. The phenotype will also be characterized at the cellular level by determining the effects of transgene expression on the adhesion migration and survival of osteoblasts isolated from wildtype and transgenic mice. Finally, effects of this transgene on the biomechanical properties and repair capabilities will be studied in collaboration with Project 2. Studies in Aim 2 will focus on the role of fibronectin in osteoblast differentiation and survival in vivo. The N-terminal 70 kDa matrix assembly and collagen binding domain will be expressed in osteoblasts of transgenic mice. This domain was shown previously to suppress the differentiation of cultured immature primary osteoblasts and to trigger apoptosis of mature cultured osteoblasts. The phenotype will be characterized using the approaches and interactions indicated for Aim 1. In addition, the ability of intact fibronectin or fragments of fibronectin to accelerate or interfere with skeletal repair will be tested in a distraction osteogenesis model in collaboration with Project 2. Since there are currently no animal models that test the roles of fibronectin and its integrin receptors in osteoblast function and bone remodeling, these studies should provide novel insights with relevance to skeletal repair and skeletal degenerative disorders.