ROLE OF MATRIX VESICLES IN CALCIFICATION

细胞外基质囊泡被认为在细胞凋亡中起关键作用。 软骨、骨和牙本质中钙化的起始，但它们的 作用机制尚未完全了解。 建议的目标 研究旨在阐明软骨内钙化的机制， 特别强调基质囊泡的作用。 注意力将集中 1）基质形成和释放的决定因素 囊泡的细胞，2）继续表征其酶， 蛋白质、矿物质形态和膜成分，以及3）进一步研究 诱导过程中基质囊泡对Ca和Pi的代谢 矿物形成 研究的目的是：a）阐明 局部营养因子对能量代谢的影响及其对细胞 Ca代谢，B）测定细胞Ca代谢对细胞内Ca代谢的影响， 可钙化基质囊泡的形成，c）进一步阐明 碱性磷酸酶及其相关蛋白在矿物质诱导中的作用 由基质囊泡形成。 我们将继续使用骨骺生长 以快速生长的鸡长骨的板软骨为来源， 我们的组织和细胞。 这个来源提供了丰富的，积极的 用于分离细胞和基质囊泡的钙化材料，以及用于 研究他们的新陈代谢 使用的技术将是组织培养、细胞培养、 分馏、45 Ca-和32 Pi-代谢、化学和物理方法： 色谱法（柱、薄层、HPLC、二维等） 电泳（PAGE、SDS-PAGE、等电聚焦）、酶学 （纯化，动力学分析），光谱学（UV，IR，NMR，MAS-NMR）， 电子显微镜（透射、扫描、冷冻断裂）、X射线 衍射等。

Extracellular matrix vesicles are believed to play a key role in the initiation of calcification in cartilage, bone and dentin, but their mechanism of action is not fully understood. The objective of the proposed research is to elucidate the mechanism of endochondral calcification with special emphasis on the role of matrix vesicles. Attention will be focused on 1) determining factors involved in the formation and release of matrix vesicles by cells, 2) continued characterization of their enzymes, proteins, mineral forms and membrane constituents, and 3) further study of the metabolism of Ca and Pi by matrix vesicles during the induction of mineral formation. Studies will be aimed at a) elucidating the role of local nutritional factors on energy metabolism and its effect on cellular Ca metabolism, b) determining the effect of cellular Ca metabolism on the formation of calcifiable matrix vesicles, c) further elucidating the role of alkaline phosphatase and associated proteins in the induction of mineral formation by matrix vesicles. We will continue to use epiphyseal growth plate cartilage from long bones of rapidly growing chickens as the source of our tissue and cells. This source provides abundant, actively calcifying material for isolation of cells and matrix vesicles, and for studying their metabolism. Techniques used will be tissue culture, cell fractionation, 45Ca- and 32Pi-metabolism, chemical and physical methods: chromatography (column, thin-layer, HPLC, 2-dimensional, etc.) electrophoresis (PAGE, SDS-PAGE, isoelectric focusing), enzymology (purification, kinetic analyses), spectroscopy (UV, IR, NMR, MAS-NMR), electron microscopy (transmission, scanning, freeze-fracture), x-ray diffraction, etc.