Comparative immunochemical analyses of the developmental expression and distribution of ameloblastin and amelogenin in rat incisors
Comparative immunochemical analyses of the developmental expression and distribution of ameloblastin and amelogenin in rat incisors
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DOI：
10.1177/002215549804600806
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发表时间：
1998-08-01
影响因子：
3.2
通讯作者：
Smith, CE
中科院分区：
文献类型：
--
作者：
Nanci, A;Zalzal, S;Smith, CE
矿化组织在使用蛋白质吸引和组织钙和磷酸盐离子进入结构化矿物质相方面是独特的。因此，精确了解基质蛋白的表达和细胞外分布对于理解其功能非常重要。本研究的目的是获得比较信息的表达，细胞内和细胞外的分布，和动态的蛋白质代表的两个主要类别的釉基质蛋白。使用针对啮齿类动物中主要的选择性剪接同种型制备的抗体和mRNA探针来可视化釉原蛋白，并且通过对一种釉质蛋白特异性的抗体和mRNA探针来可视化非釉原蛋白，所述釉质蛋白由三个名称来指代：成釉蛋白、釉蛋白和鞘蛋白。定性和定量的免疫细胞化学，结合免疫印迹和原位杂交，表明在早期前分泌期和中晚期成熟期，检测到mRNA信号，但似乎没有蛋白质被分泌，在mRNA信号和蛋白质分泌位点之间存在相关性。细胞外釉原蛋白的免疫反应性一般弱分泌表面附近，增加超过约1.25 μ m的距离，达到一个水平略高于预期的量，如果蛋白质被均匀地沉积在整个釉质层。成釉蛋白的免疫标记显示出相反的模式，分泌表面附近的金颗粒相对较多，而更深处的釉质层则少得多。布雷菲德菌素A和放线菌酮停止蛋白分泌的管理显示，釉原蛋白的免疫印迹模式是相对稳定的，而成釉蛋白迅速分解成较低分子量的片段。釉原蛋白免疫标记稳定的细胞表面的距离通常对应于成釉蛋白开始显示净减少的点。这些数据表明，釉原蛋白和成釉蛋白的分布之间的相关性，完整的成釉蛋白具有促进/稳定晶体伸长的短暂作用。
Mineralized tissues are unique in using proteins to attract and organize calcium and phosphate ions into a structured mineral phase. A precise knowledge of the expression and extracellular distribution of matrix proteins is therefore very important in understanding their function. The purpose of This investigation was to obtain comparative information on the expression, intracellular and extracellular distribution, and dynamics of proteins representative of the two main classes of enamel matrix proteins. Amelogenins were visualized using an antibody and an mRNA probe prepared against the major alternatively spliced isoform in rodents, and nonamelogenins by antibodies and mRNA probes specific to one enamel protein referred to by three names: ameloblastin, amelin, and sheathlin. qualitative and quantitative immunocytochemistry, in combination with immunoblotting and in situ hybridization, indicated a correlation between mRNA signal and sites of protein secretion for amelogenin, but not for ameloblastin, during the early presecretory and mid- to late maturation stages, during which mRNA signals were detected but no proteins appeared to be secreted. Extracellular amelogenin immunoreactivity was generally weak near secretory surfaces, increasing over a distance of about 1.25 mu m to reach a level slightly above an amount expected if the protein were being deposited evenly across the enamel layer. Immunolabeling for ameloblastin showed an inverse pattern, with relatively more gold particles near secretory surfaces and much fewer deeper into the enamel layer. Administration of brefeldin A and cycloheximide to stop protein secretion revealed that the immunoblotting pattern of amelogenin was relatively stable, whereas ameloblastin broke down rapidly into lower molecular weight fragments. The distance from the cell surface at which immunolabeling for amelogenin stabilized generally corresponded to the point at which that for ameloblastin started to show a net reduction. These data suggest a correlation between the distribution of amelogenin and ameloblastin and that intact ameloblastin has a transient role in promoting/stabilizing crystal elongation.