ROLE OF MATRIX VESICLES IN CALCIFICATION

基质囊泡在钙化中的作用

• 批准号: 3155011
• 负责人: ROY E WUTHIER
• 金额: $ 25.05万
• 依托单位: UNIVERSITY OF SOUTH CAROLINA AT COLUMBIA
• 依托单位国家: 美国
• 财政年份: 1979
• 资助国家: 美国
• 起止时间: 1979-01-01 至 1989-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3155011
• 关键词: Raman spectrometry; X ray crystallography; adenosinetriphosphatase; alkaline phosphatase; bone metabolism; calcification; calcium binding protein; calcium metabolism; calcium phosphate; calcium transporting ATPase; cartilage; cartilage metabolism; chickens; chondrocytes; connective tissue metabolism; electron microscopy; epiphysis; extracellular matrix; freeze etching; gas chromatography; gel electrophoresis; infrared spectrometry; laboratory rabbit; lipid biosynthesis; magnesium; membrane structure; normal ossification; nuclear magnetic resonance spectroscopy; paper chromatography; phospholipids; protein biosynthesis; pyrophosphates; radiotracer; thin layer chromatography; transport proteins; tritium; ultraviolet spectrometry

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.

细胞外基质囊泡被认为在 软骨、骨和牙本质钙化的开始，但它们 作用机制尚不完全清楚。 拟议的目标 研究目的是阐明软骨内钙化的机制 特别强调基质囊泡的作用。 注意力将会集中 1) 确定参与基质形成和释放的因素 细胞囊泡，2) 对其酶的持续表征， 蛋白质、矿物质形式和膜成分，以及 3) 的进一步研究 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 局部营养因素对能量代谢及其对细胞的影响 钙代谢，b) 确定细胞钙代谢对 可钙化基质囊泡的形成，c) 进一步阐明其作用 碱性磷酸酶和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 以快速生长的鸡的长骨为来源的板软骨 我们的组织和细胞。 该来源提供了丰富的、积极的 用于分离细胞和基质囊泡的钙化材料，以及 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、45Ca-和32Pi-代谢、化学和物理方法： 色谱（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（UV、IR、NMR、MAS-NMR）、 电子显微镜（透射、扫描、冷冻断裂）、X 射线 衍射等