Purification and identification of factors involved in osteoclastic maturation from bovine bone matrix

牛骨基质中破骨细胞成熟相关因子的纯化及鉴定

基本信息

批准号：
05671513
负责人：
AMANO Shigeru
金额：
$ 1.15万
依托单位：
Meikai University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05671513/
关键词：
Osteoclast Differentiation Bone matrix

项目摘要

Several studies have shown that formation of mature osteoclasts significantly increases when osteoclast progenitors are incubated on bone matrix compared with the formation on a plastic substratum. These observations have suggested that some components of bone matrix play a functional role in formation of mature osteoclasts. However, detailed characterization of these components derived from bone matrix has not yet been made.In the present grant-aid for scientific research, we purified the osteoclastic maturation-inducing factors from bovine bone, and obtained the following results.(1) Although tartrate-resistant acid phosphatase (TRAP) -positive cells formed from calvarial cells in the presence of 1alpha, 25- (OH) _2D_3 on plastic culture dishes could not resorb dentine slices, 1alpha, 25- (OH) _2D_3-induced TRAP-positive cells formed in combination with EDTA-extracts from bovine bone could resorb it.(2) The OMIFs were extracted from bovine bone powder during demineralization with EDTA,and fractionated and purified by gel filtration over Superdex 75 prep grade. The peak of OMIFs activity was eluted as an approximately 12KD species from the gel filtration column.(2) The peak fraction having OMIFs activity eluted as a major peak at about 60mM sodium phosphate (pH6.8) by the hydroxyapatite column.(3) The OMIFs eluted as a major peak at about 250mM NaCl by Mono Q equilibrated with 0.02M Bis-Tris propane (pH7.0).(4)The OMIFs were separated into several peaks by reversed-phase HPLC.the hydroxyapatite column. The molecular weights of OMIFs estimated from high-resolution 16.5% polyacrylamide gel electrophoresis were 5.1KD,6.8KD and 8.4KD respectively.In further studies, we will analyze the amino-acid sequence and define physicochemical characteristics of the purified OMIFs.

几项研究表明，与塑料底层上的形成相比，将破骨细胞祖细胞在骨基质上孵育时，成熟破骨细胞的形成显着增加。这些观察结果表明，骨基质的某些成分在成熟破骨细胞的形成中起功能。然而，尚未做出从骨基质中得出的这些成分的详细表征。在目前的科学研究赠款中，我们纯化了牛骨的骨质碎屑成熟因子诱导的整骨成熟因子，并获得了以下结果。 _2D_3在塑料培养皿上无法吸收牙本质切片，1alpha，25-（OH）_2d_3诱导的陷阱阳性细胞与牛骨的EDTA-提取物结合形成，可以将其吸收。（2）用Edta和Edtratied pucareated在Edta和demertation中提取OMIF，并在Edta和demerative中提取omifs，并在Edta和demertife中提取了GEL，并在Edta和demertation putife中提取了GEL，并固定了GEL，并固定了GEL，并固定了GEL，并固定了GEL，并固定了GEL，并将其固定在Edta和demertation demertation puartered中。年级。 OMIFS活性的峰值从凝胶过滤柱中洗脱为大约12kD物种。（2）在约60mm磷酸钠（ph6.8）的主要峰值上，羟基磷灰石柱的峰值分数通过羟基磷灰石柱洗脱了。（PH7.0）。（4）通过反相HPLC将OMIF分为几个峰。羟基磷灰石柱。根据高分辨率16.5％聚丙烯酰胺凝胶电泳估计的OMIF的分子量分别为5.1kd，6.8kd和8.4kd。在进一步的研究中，我们将分析氨基酸序列并定义纯化OMIF的物理化学特性。