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Functional analysis of osteocyte-specific acidic phosphoprotein by gene transfection

基因转染对骨细胞特异性酸性磷蛋白的功能分析

基本信息

批准号：
13671898
负责人：
TOYOSAWA Satoru
金额：
$ 2.37万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13671898/
关键词：
Dentin matrix protein 1 Transgenic mice Function Mineralization Osteocytes Osteoblasts

项目摘要

Dentin matrix protein 1 (DMP1) is one of the acidic phosphoproteins originally identified from a rat incisor cDNA library. DMP1 has abundant acidic domains and a large number of phosphorylation consensus sites for casein kinase I and II, which are negatively charged at physiological pH. Therefore, DMP1 likely binds with calcium and may regulate matrix mineralization.We demonstrated that DMP1 mRNA was predominantly expressed in osteocytes but not in osteoblasts and that DMP1 protein was in the pericellular bone matrix of osteocytes, while other bone matrix proteins including osteohalcin, osteopontin and bone sialoprotein were expressed in osteoblasts. We determined the precise expression pattern of DMP1 mRNA and localization of its protein in dentin and cementum. The localization of DMP1 mRNA and DMP1 protein in bone and tooth was closely related to their mineralization, suggesting that DMP1 plays an important role in mineraliztion.To elucidate the function of DMPI in vivo, we generated transenic mice that overexpressed DMP1 in osteoblasts and odontoblasts using type I collagen promoter. In DMP1 transgenic mice, trabecular bone the metaphysis was dramatically decrease. However, the number of osteoclasts was not increased, although the functional level of osteoclasts was not studied. The bone density of cortical bone near the metaphysis was increased. Northern blot analysis demonstrated no change of the production of the bone matrix. These findings suggested that DMP1 promotes the rate of bone mineralization process, although the significance of DMP1 expression specific for osteocytes was not yet known.

牙本质基质蛋白1 （DMP1）是最初从大鼠门牙cDNA文库中鉴定出的酸性磷酸化蛋白之一。DMP1具有丰富的酸性结构域和大量酪蛋白激酶I和II的磷酸化一致位点，这些位点在生理ph下带负电荷。因此，DMP1可能与钙结合并调节基质矿化。我们证明DMP1 mRNA主要在骨细胞中表达，而在成骨细胞中不表达，DMP1蛋白存在于骨细胞的细胞周骨基质中，而其他骨基质蛋白包括骨卤素、骨桥蛋白和骨涎蛋白在成骨细胞中表达。我们确定了DMP1 mRNA的精确表达模式及其蛋白在牙本质和牙骨质中的定位。DMP1 mRNA和DMP1蛋白在骨骼和牙齿中的定位与其矿化密切相关，提示DMP1在矿化中起重要作用。为了阐明DMPI在体内的功能，我们使用I型胶原启动子培养了在成骨细胞和成牙细胞中过表达DMP1的转基因小鼠。在DMP1转基因小鼠中，骨小梁干骺端明显减少。然而，破骨细胞的数量没有增加，尽管没有研究破骨细胞的功能水平。干骺端附近皮质骨的骨密度增加。Northern blot分析显示骨基质的生成没有变化。这些发现表明，DMP1促进骨矿化过程的速率，尽管DMP1表达对骨细胞特异性的意义尚不清楚。