Precise timings and spatial progression of human enamel biomineralisation are still largely unknown due to scarcity of developing human enamel specimens available for investigation. This information is crucial for optimising emerging biomimetic regenerative and reparative dentistry routes. Five developing permanent incisors were obtained from an archaeological source and used alongside mature contemporary teeth for comparison. X-ray microtomography (XMT), synchrotron X-ray diffraction (S-XRD) and quantitative back-scattered electrons (qBSE) imaging were used to investigate the mineral density distribution, the crystallites texture magnitude and orientation and the nanostructure of dental enamel at various developmental stages, respectively. XMT revealed that there was a bi-directional mineralisation "front" that starts at the cusp tip and at the enamel-dentine junction (EDJ) travelling cervically and peripherally until the relative mineral density is uniform in the fully mature tooth (2.75 g/cm3 ± 0.01 g/cm3). S-XRD revealed that within one probed region, two populations of crystallite orientations exist simultaneously with an angular separation of 20-50°, with one population being more dominant than the other by a factor of approximately 3:7. Furthermore, one population displayed a higher degree of crystallite texture than the other. These phenomena were observed in all stages of tooth development. The crystallites in both populations were oriented approximately perpendicular to the EDJ regardless of development stage, indicating initial preferred directions of crystallites persist from early through to full maturation. The direction and magnitude of organisation within two distinct populations of crystallites within the developing and mature enamel has not been quantified previously. qBSE analyses suggested that the two observed populations are most likely due to prism decussation and revealed that mineralisation of prism cores precedes that of prism boundaries. These results provide new insights towards building a quantitative spatio-temporal model of human enamel biomineralisation in order to inform emerging biomimetic reparative/regenerative dental technologies.

Yes / For dental human enamel, what is the precise mineralization progression spatially and the precise timings of mineralization? This is an important question in the fundamental understanding of matrix-mediated biomineralization events, but in particular because we can use our understanding of this natural tissue growth in humans to develop biomimetic approaches to repair and replace lost enamel tissue. It is important to understand human tissues in particular since different species have quite distinct spatial and temporal progression of mineralization. In this study, five human central incisors at different stages of enamel maturation/mineralization were spatially mapped using synchrotron X-ray diffraction and X-ray microtomography techniques. From the earliest developmental stage, two crystallite-orientation populations coexist with angular separations between the crystallite populations averaging approximately 40o and varying as a function of position with the tooth crown. In general, population one had significantly lower texture magnitude and contributed a higher percentage to the overall crystalline structure, compared to population two which only contributed 20-30% but had significantly higher texture magnitude. This quantitative analysis allows us to understand the complex and co-operative structure-function relationship between two populations of crystallites within human enamel. There was an increase in the mineral concentration from the enamel-dentin junction peripherally and from the incisal tip cervically as a function of maturation time. Quantitative backscattered-electron analyses revealed that mineralization of prism cores precedes that of prism boundaries. These results provide new insights into the precise understanding of the natural growth of human enamel. / Partly funded by NERC grant ”Timelines in Teeth” NE/F018096/2.

Considerando a eficácia limitada do flúor na proteção dos tecidos duros dentais contra a erosão, o objetivo do presente estudo foi investigar o potencial do laser de gás carbônico (CO2) (10,6 m), associado ou não a produtos fluoretados, na inibição da erosão de esmalte dental. Amostras de esmalte dental humano foram obtidas e divididas aleatoriamente em 8 grupos (n = 11): G1: controle (sem tratamento), G2: gel de Flúor Fosfato Acidulado (FFA), G3: gel de fluoreto de sódio (AmF/NaF), G4: solução de fluoreto estanhoso (AmF/SnF2), G5: laser de CO2 (0,3 J/cm2, 15 s, 226 Hz), G6: laser de CO2 + gel de FFA, G7: laser de CO2 + gel de AmF/NaF e G8: laser de CO2 + solução de AmF/SnF2. Após os tratamentos de superfície, as amostras foram imersas em ácido cítrico a 1% (pH 4,0, 3 minutos). Antes dos tratamentos de superfície e após o desafio ácido, a microdureza de superfície foi mensurada (0,49 N, 20 segundos) e os dados foram analisados estatisticamente através do teste ANOVA com subseqüentes comparações entre pares (p < 0,05). Os resultados mostraram que o grupo G3 (403,5 ± 25,0) apresentou a maior média de microdureza Knoop após o desafio ácido, seguido pelos grupos G2 (396,6 ± 45,0) e G6 (392,8 ± 24,9), sem diferença estatística entre eles. Dentre os grupos tratados com produtos fluoretados, o grupo G4 (305,5 ± 17,7) resultou em microdureza significativamente mais baixa que o G3 (403,5 ± 25,0) e o G2 (396,6 ± 45,0), os quais foram estatisticamente semelhantes entre si. Dentre os grupos tratados com laser, os grupos G5 (341,1 ± 23,8), G7 (374,9 ± 42,1) e G8 (328,6 ± 26,9) apresentaram valores de dureza estatisticamente semelhantes entre si. Dentro dos limites do presente estudo in vitro, pôde-se concluir que o tratamento com gel de AmF/NaF e gel de FFA (com ou sem irradiação prévia com o laser de CO2) mostrou potencial para controlar a progressão da erosão no esmalte dental humano. / Considering the limited effectiveness of fluoride in protecting dental hard tissues against erosion, the aim of the present study was to investigate the potential of the CO2 laser (10.6 m) associated to fluoride agents in inhibiting human enamel erosion. Human enamel samples were obtained and randomly divided into 8 groups (n = 11): G1: control (no treatment), G2: APF gel, G3: AmF/NaF gel, G4: AmF/SnF2 solution, G5: CO2 laser (0.3 J/cm2, 15 s, 226 Hz), G6: CO2 laser + APF gel, G7: CO2 laser + AmF/NaF gel and G8: CO2 laser + AmF/SnF2 solution. After surface treatment, samples were immersed in 1% citric acid (pH 4.0, 3 min). Knoop Surface microhardness (SMH) (0.49 N, 20 s) was measured before surface treatments and after surface softening. The data were statistically analyzed by ANOVA model with subsequent pairwise comparisons (p < 0,05). Group G3 (403.5 ± 25.0) presented the highest SMH means after softening, followed by Groups G2 (396.6 ± 45.0) and G6 (392.8 ± 24.9) with no statistically significant difference between them. Between the fluoride-treated groups, G4 (305.5 ± 17.7) resulted in significantly low SMH mean than G3 (403.5 ± 25.0) and G2 (396.6 ± 45.0), which were statistically similar to each other. Between the laser-treated groups, G5 (341.1 ± 23.8), G7 (374.9 ± 42.1) and G8 (328.6 ± 26.9) presented SMH means statistically similar to each other. Within the limits of the present in vitro study, AmF/NaF and APF application (with or without previous CO2 laser irradiation) have shown a potential to control erosion progression in human dental enamel.

Considerando a eficácia limitada do flúor na proteção dos tecidos duros dentais contra a erosão, o objetivo do presente estudo foi investigar o potencial do laser de gás carbônico (CO2) (10,6 m), associado ou não a produtos fluoretados, na inibição da erosão de esmalte dental. Amostras de esmalte dental humano foram obtidas e divididas aleatoriamente em 8 grupos (n = 11): G1: controle (sem tratamento), G2: gel de Flúor Fosfato Acidulado (FFA), G3: gel de fluoreto de sódio (AmF/NaF), G4: solução de fluoreto estanhoso (AmF/SnF2), G5: laser de CO2 (0,3 J/cm2, 15 s, 226 Hz), G6: laser de CO2 + gel de FFA, G7: laser de CO2 + gel de AmF/NaF e G8: laser de CO2 + solução de AmF/SnF2. Após os tratamentos de superfície, as amostras foram imersas em ácido cítrico a 1% (pH 4,0, 3 minutos). Antes dos tratamentos de superfície e após o desafio ácido, a microdureza de superfície foi mensurada (0,49 N, 20 segundos) e os dados foram analisados estatisticamente através do teste ANOVA com subseqüentes comparações entre pares (p < 0,05). Os resultados mostraram que o grupo G3 (403,5 ± 25,0) apresentou a maior média de microdureza Knoop após o desafio ácido, seguido pelos grupos G2 (396,6 ± 45,0) e G6 (392,8 ± 24,9), sem diferença estatística entre eles. Dentre os grupos tratados com produtos fluoretados, o grupo G4 (305,5 ± 17,7) resultou em microdureza significativamente mais baixa que o G3 (403,5 ± 25,0) e o G2 (396,6 ± 45,0), os quais foram estatisticamente semelhantes entre si. Dentre os grupos tratados com laser, os grupos G5 (341,1 ± 23,8), G7 (374,9 ± 42,1) e G8 (328,6 ± 26,9) apresentaram valores de dureza estatisticamente semelhantes entre si. Dentro dos limites do presente estudo in vitro, pôde-se concluir que o tratamento com gel de AmF/NaF e gel de FFA (com ou sem irradiação prévia com o laser de CO2) mostrou potencial para controlar a progressão da erosão no esmalte dental humano. / Considering the limited effectiveness of fluoride in protecting dental hard tissues against erosion, the aim of the present study was to investigate the potential of the CO2 laser (10.6 m) associated to fluoride agents in inhibiting human enamel erosion. Human enamel samples were obtained and randomly divided into 8 groups (n = 11): G1: control (no treatment), G2: APF gel, G3: AmF/NaF gel, G4: AmF/SnF2 solution, G5: CO2 laser (0.3 J/cm2, 15 s, 226 Hz), G6: CO2 laser + APF gel, G7: CO2 laser + AmF/NaF gel and G8: CO2 laser + AmF/SnF2 solution. After surface treatment, samples were immersed in 1% citric acid (pH 4.0, 3 min). Knoop Surface microhardness (SMH) (0.49 N, 20 s) was measured before surface treatments and after surface softening. The data were statistically analyzed by ANOVA model with subsequent pairwise comparisons (p < 0,05). Group G3 (403.5 ± 25.0) presented the highest SMH means after softening, followed by Groups G2 (396.6 ± 45.0) and G6 (392.8 ± 24.9) with no statistically significant difference between them. Between the fluoride-treated groups, G4 (305.5 ± 17.7) resulted in significantly low SMH mean than G3 (403.5 ± 25.0) and G2 (396.6 ± 45.0), which were statistically similar to each other. Between the laser-treated groups, G5 (341.1 ± 23.8), G7 (374.9 ± 42.1) and G8 (328.6 ± 26.9) presented SMH means statistically similar to each other. Within the limits of the present in vitro study, AmF/NaF and APF application (with or without previous CO2 laser irradiation) have shown a potential to control erosion progression in human dental enamel.