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In the developing tooth, 1-hydroxyethylidene-1,1-bisphosphonate (HEBP) causes hypoplasia and hypomineralization of enamel and dentine. The present study was undertaken to clarify the effects of HEBP on the formation of dental tissues of tooth germs in an organ culture system. Mandibular first molars from 17.5-day-old mouse embryos were cultured with or without 250 microM HEBP in culture medium. Cultured tooth germs were analyzed by histological examination and by immunohistochemical localization using anti-amelogenin antibody. In cultured tooth germs treated with HEBP before the commencement of calcification in dentine, calcification of dentine matrix was inhibited completely and enamel formation was not observed. Ameloblasts were directly adjacent to dentine matrix. However, immunohistochemical data indicated that these ameloblasts secreted amelogenin. In the experiments of adding HEBP to cultured tooth germs on culture day 13, calcified dentine and enamel had formed before the administration of HEBP, but the dentine matrix newly formed after the administration of HEBP had not calcified. It was confirmed by immunohistochemical observations that enamel matrix-like material had penetrated into uncalcified dentine matrix and accumulated in dental papilla of tooth germs. However, no enamel matrix-like material was observed in calcified dentine and predentine underneath the calcified dentine by immunohistochemical staining. From these results, it might be concluded that ameloblasts secreted enamel matrix in the presence of HEBP and diffused through uncalcified dentine matrix into dental papilla. These findings suggests the calcification of dentine might be essential for the physical barrier to accumulate the enamel matrix and form a distinct layer of enamel as enamel.

Effects of 1-hydroxyethylidene-1,1-bisphosphonate (HEBP) on the Formation of Dental Hard Tissue of Mouse Molar Tooth Germs in Organ Culture System.