Immunohistochemical and molecular biological study of enamel proteins with special reference to molecular isoforms.

釉质蛋白，如釉原蛋白和鞘蛋白，有几种异构体，由mRNAs的选择性剪接产生。为了阐明鞘蛋白分子异构体的作用，我们开发了能够识别特殊分子形式的区域特异性抗体。用免疫组织化学和免疫组织化学方法对大鼠切牙和猪牙胚的釉质器官和未成熟的釉质基质进行了分析。研究结果总结如下：1.研究成果。对大鼠切牙基质形成期和早期成熟期的釉质器官的免疫化学分析表明，成釉蛋白和Amelin是大鼠鞘蛋白的两种亚型，分别合成56 kDa和54 kDa的核心蛋白。在基质形成阶段，釉质器官中釉质蛋白和成釉蛋白的含量差异不大，而在早期成熟期，前者的含量显著低于后者。大鼠切牙的免疫组织化学研究表明，成釉蛋白和Amelin的免疫定位基本相同。然而，由于釉质基质中深层的Amelin免疫反应较弱，Amelin的降解速度似乎比成釉蛋白快。对猪未成熟牙釉质的免疫组织化学分析表明，识别短形鞘蛋白(猪鞘蛋白)的抗体在整个牙釉质的棱柱鞘中明显染色。但经识别短型鞘蛋白的抗体免疫染色后，表层的棱柱鞘并不清晰。在向成熟早期过渡的后期，表层短型鞘蛋白的免疫反应明显弱于长型鞘蛋白，说明短型鞘蛋白的降解速度快于长型鞘蛋白。

Enamel proteins, such as amelogenins and sheath proteins, have several isoforms produced by alternative splicing of mRNAs. In order to clarify the role of molecular isoforms of sheath proteins, we have developed region specific antibodies that can recognize special molecular forms. Enamel organ and immature enamel matrix from the rat incisor and the porcine tooth germ were analyzed by means of immunochemistry and immunohistochemistry. The results are summarized as follows.1. Immunochemical analysis of the enamel organ from the matrix formation stage and the early maturation stage in the rat incisor showed that ameloblastin and amelin, two isoforms of rat sheath proteins, were synthesized as 56 kDa and 54 kDa core proteins, respectively. The amount of amelin and ameloblastin contained in the enamel organ was not so different at the matrix formation stage, while the former was significantly lower than the latter at the early maturation stage.2. Immunohistochemistry of rat incisor has showed that immunolocalization of ameloblastin and amelin was essentially identical. However, the degradation of amelin seemed to be faster than that of ameloblastin because of weak immunoreaction of amelin at the middle to deep layer of enamel matrix.3. Immunohistochemical analysis of porcine immature enamel has revealed that antibody that recognize short-form of sheathlin (porcine sheath protein) clearly stained prism sheath at the entire layer. However, after the immunostaining with antibody that recognize short-form of sheathlin, prism sheath at the surface layer was not so clear.4. At the late transition to early maturation stages, immunoreactivity of the short-form of sheathlin at the surface layer was significantly weaker than that of the long-form of sheathlin, suggesting the degradation of short-form of sheathlin occurs faster than the long-form of sheathlin.