The mechanisms of continuous tooth replacement in the Nile crocodile (Crocodylus niloticus)

Abstract

It is recognised that tooth loss as a consequence of oral diseases affects quality of life in humans. This has directed studies towards biological tooth replacement in vivo. In humans and other mammals, tooth replacement occurs only once (diphyodonty) as opposed to non-mammalian vertebrates where tooth replacement continues throughout life (polyphyodonty). Detailed knowledge of tooth initiation, development and morphology amongst vertebrates and especially amniotes, is necessary to understand the tooth replacement process. Crocodilians provide an interesting model for tooth replacement studies as they also exhibit thecodonty. Regulation of polyphyodonty has not been genetically defined, and it is uncertain whether the molecular mechanisms of continuous tooth replacement are similar to those involved in the primary dentition. The aim of this study was therefore to analyse crocodilian odontogenesis in detail, with the aid of light microscopy and CT scans, in order to provide a structural framework for molecular processes regulating polyphyodonty. Crocodile probes to bmp4 and pitx2 were designed, generated and labelled for use in in situ hybridisation. The expression patterns of pitx2 and bmp4 in embryos and hatchlings of the polyphyodont Nile crocodile (Crocodylus niloticus) were examined at different stages of tooth development. Histologically crocodilian tooth development appears similar to mammals. Interesting variations include the initiation of odontogenesis in the ectomesenchyme, the presence of dental placodes, the ‘null generation teeth’, the two different bell-stage tooth germs and the tooth-family organisation. A direct 1:1 relationship between the status of the erupted tooth and the developmental phase of the replacement tooth was not seen. However in more mature teeth, the replacement tooth germs were at a more advanced developmental stage than those associated with less mature teeth. Molecular data revealed that pitx2 was expressed in the oral epithelium and the dental placode. Bmp4 expression was not evident in the dental placode, but was localised in the odontoblasts of early bell stage tooth germs. Pitx2 and bmp4 were expressed in both the odontoblast and ameloblast layers in late bell stage tooth germs. Expression of pitx2 and bmp4 is conserved across vertebrates and pitx2 may play a role in initiation of primary and successional teeth.