The mechanism of bone formation with reference t the phosphoprotein in the bone matrix

应用超微结构、免疫组化和生化方法研究了酸性磷蛋白与骨髓基质钙化的关系。超微结构分析显示，类骨质深层分布有直径约0.5 μm的小球状物，其分布方式与钙化球状物相似。全染色显示的蓝色区域的分布模式也与基质中钙化区域的分布模式相似。通过染色-全染色将基质成分染成蓝色的生化分析显示与鸡骨桥蛋白相似。此外，随着钙化的进展，鹌鹑骨桥蛋白mRNA在髓骨中的表达增强。分别针对骨髓矿相蛋白组分和基于鹌鹑骨桥蛋白部分氨基酸序列的合成肽制备了多克隆抗体。免疫组化反应是阳性的基质和骨细胞与使用的抗体从蛋白质，但阳性反应仅限于破骨细胞和成骨细胞，使用抗体对肽。这些结果表明骨桥蛋白大量存在于骨髓基质中，并随着基质钙化的进展而增加，提示骨桥蛋白与骨髓钙化有关。

The relationship between acidic phosphoprotein and the calcification of medullary bone matrix was examined employing ultrastructural, immunohistochemical and biochemical methods. Ultrastructural analysis showed that small globules (〜0.5 μm) were distributed at the deeper parts of osteoid, and the distribution pattern of the globules was similar to that of calcifying globules. The distribution pattern of the areas stained blue by stains-all staining was also similar to that of calcified areas in the matrix. Biochemical analyses of the matrix component stained blue by stains-all staining showed the similarity to chicken osteopontin. Furthermore, according as calcification progressed, the expression of quail osteopontin mRNA was enhanced in medullary bone. Polyclonal antibodies were raised against the protein fraction obtained from mineral phase of medullary bone and the synthetic peptide based the partial sequence of amino acid of quail osteopontin, respectively. Immunohistochemical reaction was positive in the matrix and bone cells with use of the antibody raised from the protein, but the positive reaction was confined to osteoclasts and osteoblasts, using the antibody raised against the peptide. These results indicate that osteopontin abundantly exists in medullary bone matrix and increases according as the matrix calcification advances, and suggest that osteopontin relates to medullary bone calcification.