IN VITRO INITIATION OF BIOLOGICAL CALCIFICATION

生物钙化的体外引发

• 批准号: 3158275
• 负责人: ADELE L BOSKEY
• 金额: $ 13.43万
• 依托单位: HOSPITAL FOR SPECIAL SURGERY
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-08-01 至 1992-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3158275
• 关键词: X ray crystallography; ascorbate; calcification; cartilage development; cell differentiation; electron microscopy; immunochemistry; infrared spectrometry; proteoglycan; retinoids; tissue /cell culture; trace elements

Cartilage calcification is an essential step in the processes of fracture healing and bone growth (endochondral ossification). The goal of the proposed studies is to determine the ways in which cartilage calcification is regulated. Since mammals and birds demonstrate different patterns of cartilage calcification they provide excellent models for examining these regulatory process. Mammalian and avian cartilage calcification can be differentially modeled in vitro using a well developed model of chondrogenesis. Chondrogenesis (cartilage formation) occurs in vitro when mesenchymal cells (isolated from embryonic limb buds) are cultured at high density in media containing 1 mM calcium: the matrix will calcify in the presence of 3-4 mM inorganic phosphate. The long range goal of the proposed study is to use chick and mouse mesenchymal cell systems to characterize events in initial calcification. Chemical analyses, light and electron microscopy, x-ray diffraction and infra-red spectroscopy, and immunochemistry will be used to evaluate the mineralized matrix formed in vitro. Experiments are designed initially to optimize the culture conditions to provide maximal "physiologic" matrix mineralization by varying a) ascorbic acid, b) phosphate, c) beta-glycerol phosphate, d) calcium, e) fetal calf serum, and f) atmospheric conditions. The calcified tissue formed in the avian and murine cell culture systems will be compared to each other and to mineralized cartilage produced in vivo, with respect to: a) type of matrix undergoing mineralization, b) mineral characteristics, and c) cells regulating the mineralization process. The cultured cells will than be chemically or physically modified in order to test the hypothesis that viable, as opposed to necrotic, cells are responsible for mineralization. The effect of alteration of matrix properties on mineralization will be examined by using agents that alter: a) cartilage differentiation and development (BrDU and vitamin A), b) proteoglycan synthesis, composition and/or properties (vitamin A, xylosides, hyaluronidase, polylysine), and c) synthesis of other extracellular matrix proteins (cAMP). The culture systems will be examined to determine how clinically relevant trace elements (F, A1, Fe, Ga, Ti, V, Ni, Cr and Co) control the extent of cell mediated mineralization. These cell culture studies are unique due to their emphases on the mechanism of cell-mediated calcification. Their clinical significance comes from the basic information on calcification mechanisms they will provide as well as from the specific data they will generate on the effect of certain therapies on these mechanisms.

软骨钙化是一个重要的步骤， 骨折愈合和骨生长（软骨内骨化）。 拟议研究的目标是确定 软骨钙化是受调节的。 由于哺乳动物和 鸟类表现出不同的软骨钙化模式 他们提供了很好的模型来检查这些监管 过程 哺乳动物和鸟类的软骨钙化可以 在体外使用良好开发的模型进行差异建模， 软骨形成 软骨发生（软骨形成）发生在 当间充质细胞（从胚胎肢体分离）时， 芽）在含有1 mM 钙：基质在3-4 mM钙存在下钙化 无机磷酸盐 拟议研究的长期目标 是用鸡和老鼠的间充质细胞系统 表征初始钙化中的事件。 化学分析， 光学和电子显微镜、x射线衍射和红外线 光谱学和免疫化学将用于评估 体外形成矿化基质。 实验设计 最初优化培养条件以提供最大的 通过改变a）抗坏血酸， B）磷酸盐，c）β-甘油磷酸盐，d）钙，e）胎牛 血清和f）大气条件。 钙化组织 在禽类和鼠类细胞培养系统中形成的细胞将 与其他组织相比， 体内，关于：a）经历的基质类型 矿化，B）矿物质特征，以及c）细胞调节 矿化过程。 培养的细胞将比 化学或物理改性，以检验假设 与坏死细胞相反， 成矿 基质性质改变对 矿化将通过使用改变以下的试剂来检查：a） 软骨分化和发育（BrDU和维生素A）， B）蛋白聚糖的合成、组成和/或性质（维生素 A，木糖苷，透明质酸酶，聚赖氨酸），和c）其他的合成 细胞外基质蛋白（cAMP）。 文化体系将 以确定临床相关的微量元素 （F、Al、Fe、Ga、Ti、V、Ni、Cr和Co）控制晶胞的大小 介导的矿化。 这些细胞培养研究是独一无二的 由于他们强调细胞介导的机制， 钙化 它们的临床意义来自于 他们将提供钙化机制的信息， 以及从他们将产生的影响的具体数据， 这些机制上的某些疗法。