MINERAL MATRIX RELATIONS IN CALCIFYING TISSUES

描述：（改编自研究者摘要）本提案 希望获得有关脊椎动物骨骼的新信息， 骨、软骨和肌腱钙化。 正常的小鸡，老鼠， 和火鸡;骨和软骨培养;以及异常的骨生成 将使用突变的Alkata小鼠作为模型。 这些研究 示例将检查某些结构和化学相互作用， 它们各自的细胞外基质、细胞内基质，以及 细胞内和细胞外界面。 将系统地开展工作， 定义蛋白质分泌途径，细胞基质粘附关系， 矿物基质组合 超微结构，生化， 免疫化学和生物物理方法将被应用于延长 申请人先前的结果，描述了该车辆的存在、性质、位置， 脊椎动物体内有机基质和矿物质成分的相互作用 钙化 要检验的具体假设是： 脊椎动物钙化组织的基质严重影响 矿物在晶体大小、形状、位置 定位和分布。 具体目标，解决结构问题- 在原子、分子和大分子水平上的功能关系， 层次，是：（1）确定胶原的存在和位置， 和参与矿化的非胶原蛋白，以及 选择整合素和细胞骨架元素在细胞基质 界面，利用常规和高压电子显微镜， 三维（3D）图像重建和视频技术，以及2D 和3D免疫化学;（2）确定矿物沉积的位置， 类似的方法以及电子衍射，X射线探针显微分析， 和新型傅里叶变换红外光谱;（3）表征 细胞外基质蛋白与矿物质相互作用 上述方法和原子力显微镜的相关性， 计算机建模（分子动力学）;（4）评估 骨细胞培养对骨基质-矿物质相互作用的影响 蛋白质转运抑制剂莫能菌素。 建议在 结果数据将有助于了解结构和功能 细胞、细胞-基质界面、细胞外基质和事件 是脊椎动物钙化的基础

DESCRIPTION: (Adapted from investigator's abstract) This proposal intends to obtain new information concerning vertebrate skeletal calcification in bone, cartilage, and tendon. Normal chicks, mice, rats, and turkeys; bone and cartilage cultures; and an abnormal osteogenesis imperfecta mouse mutant will be used as models. Studies in these examples will examine certain structural and chemical interactions in their respective extracellular matrices, intracellular matrices, and intra-and extracellular interfaces. Work will be done systematically, defining protein secretory pathways, cell matrix adhesion relations, and mineral-matrix associations. Ultrastructural, biochemical, immunochemical, and biophysical methods will be applied to extend the applicants' previous result describing the presence, nature, location, and interaction of organic matrix and mineral components in vertebrate calcification. The specific hypothesis to be examined is: the organic matrix of vertebrate calcifying tissues critically influences the deposition of mineral in terms of crystal size, shape, location, orientation, and distribution. Specific Aims, to address structure- function relations at atomic, molecular, and macromolecular levels of hierarchy, are to: (1) identify the presence and location of collagenous and non-collagenous proteins involved in mineralization, as well as selected integrins and cytoskeletal elements at the cell-matrix interface, utilizing conventional and high voltage electron microscopy, three-dimensional (3D) image reconstruction and video techniques, and 2D and 3D immunochemistry; (2) determine the sites of mineral deposition by similar means and also electron diffraction, x-ray probe microanalysis, and novel Fourier transform infrared spectroscopy; (3) characterize interaction between extracellular matrix proteins and mineral through correlation of the methods above and by atomic force microscopy and computer modeling (molecular dynamics); and (4) assess the effects in bone cell culture on matrix-mineral interaction following modulation of protein transport with the inhibitor, monensin. It is suggested that outcome data will contribute to knowledge of structure and function of cells, the cell-matrix interface, and extracellular matrices and events fundamental to vertebrate calcification.