M180KI Mouse Enamel: Can something so hard be made simple?

Amelogenin proteins dominate and control the enamel mineralization process. Alternative splicing creates many amelogenins of different lengths and potential functions. Although many tooth formation genes and proteins have been identified, their relations to mechanical function are still largely unknown. Objective: We use a genetic knock-in of the M180 amelogenin to the amelogenin locus, resulting in enamel being fabricated with only one amelogenin protein isoform, allowing us to measure the consequence of this design on bulk enamel hardness and toughness. Methods: Genetic knock-in mice were created, verified and sacrificed. Five freshly extracted lower left 9-week old wild-type and 5 freshly extracted lower left knock-in M180 KI incisors were embedded in epoxy resin. The teeth were ground and polished longitudinally in the sagittal plane for mechanical testing. A Vickers microhardness testing machine was used to make indentions at the center of the enamel thickness in the sagittal plane, as well as in dentin. To evaluate the effect of genetically simplifying enamel formation through the exclusive use of M180 amelogenin isoform on the biological organization and mineralization of enamel, we compared hardness and fracture toughness of the enamel of the wild-type mice and the transgenic mice. Ten repetitions per tooth were averaged to describe each of the 5 teeth in each group; group means and standard deviations were then calculated. Data was analyzed by T tests (p<0.05). Results: Reducing amelogenin isoform complexity 1 order of magnitude to only the M180 amelogenin protein produced enamel that was as hard as, but significantly (p<0.05), 19%, less tough, than wild type enamel. Dentin hardness did not differ between the animal types, suggesting that M180 is sufficient for any epithelial-mesenchymal signaling that is required for tooth formation. Conclusion: A functional, but slightly less tough, enamel can be made using M180 amelogenin exclusively.

NIH/NIDCR DE014189, DE006988, DE013045