Ameloblastin Expression Levels Influence Smoothness of the Enamel Surface

Ameloblastin is expressed and secreted from ameloblasts and contributes to the formation of an enamel layer. After processing by MMP20, the N-terminal Ambn fragment localizes to the sheath space, while the C-terminal Ambn fragment localizes to the surface of the forming enamel layer and is removed by ameloblasts. Objectives: The goal was to determine the surface quality of enamel of mandibular incisors in ameloblastin (Ambn) mutant mice reconstituted with transgenically expressed Ambn. Methods: Ambn mutant mice (lacking exons 5 and 6; Ambn^mutant) were reconstituted with full-length Ambn by mating with mice expressing Ambn from a transgene using the amelogenin promoter. Three transgenic mouse lines (Ambn^mutant/Tg) were identified differing in expression levels in Ambn as determined by Western Blot analysis using molars of PN5. Genotyping was performed by PCR. Hemimandibles were dissected at 7 weeks of age and lyophilized. The bone and enamel organ cell covering mandibular incisors were removed to expose the developing enamel. The enamel surfaces corresponding to secretory stage, maturation stage and erupted portion were analyzed by micro-computed tomography (microCT), light microscopy (LM) and scanning electron microscopy (SEM). Results: The transgene expression ranged from below, similar to, to greater than wildtype controls. MicroCT, LM and SEM revealed that the deposition of enamel started in all three lines of Ambn^mutant/Tg animals during secretory stage. During the maturation stage, the enamel surface changed its texture. In incisors with transgenic Ambn at a level lower than control, the surface of erupted enamel stayed smooth. When transgenic Ambn was similar to control, the surface was grainy. In Ambn^mutant/Tg animals expressing Ambn at highest level, the enamel surface was covered with prominent nodules. Conclusion: Full-length Ambn affects the enamel surface in a dose-dependent manner when expressed in Ambn^mutant mice. This study was funded by University Research Council, UTHSCSA, and US Department of Education.