Meeting report: a hard look at the state of enamel research.

The Encouraging Novel Amelogenesis Models and Ex vivo cell Lines (ENAMEL) Development workshop was held on 23 June 2017 at the Bethesda headquarters of the National Institute of Dental and Craniofacial Research (NIDCR). Discussion topics included model organisms, stem cells/cell lines, and tissues/3D cell culture/organoids. Scientists from a number of disciplines, representing institutions from across the United States, gathered to discuss advances in our understanding of enamel, as well as future directions for the field

The effect of fluoride on enamel and dentin formation in the uremic rat incisor

Renal impairment in children is associated with tooth defects that include enamel pitting and hypoplasia. However, the specific effects of uremia on tooth formation are not known. In this study, we used rat mandibular incisors, which continuously erupt and contain all stages of tooth formation, to characterize the effects of uremia on tooth formation. We also tested the hypothesis that uremia aggravates the fluoride (F)-induced changes in developing teeth. Rats were subjected to a two-stage 5/6 nephrectomy or sham operation and then exposed to 0 (control) or 50 ppm NaF in drinking water for 14 days. The effects of these treatments on food intake, body growth rate, and biochemical serum parameters for renal function and calcium metabolism were monitored. Nephrectomy reduced food intake and weight gain. Intake of F by nephrectomized rats increased plasma F levels twofold and further decreased food intake and body weight gain. Uremia affected formation of dentin and enamel and was more extensive than the effect of F alone. Uremia also significantly increased predentin width and induced deposition of large amounts of osteodentin-like matrix-containing cells in the pulp chamber. In enamel formation, the cells most sensitive to uremia were the transitional-stage ameloblasts. These data demonstrate that intake of F by rats with reduced renal function impairs F clearance from the plasma and aggravates the already negative effects of uremia on incisor tooth development

Oral Sciences PhD Program Enrollment, Graduates, and Placement: 1994 to 2016

For decades, dental schools in the United States have endured a significant faculty shortage. Studies have determined that the top 2 sources of dental faculty are advanced education programs and private practice. Those who have completed both DDS and PhD training are considered prime candidates for dental faculty positions. However, there is no national database to track those trainees and no evidence to indicate that they entered academia upon graduation. The objective of this study was to assess outcomes of dental school–affiliated oral sciences PhD program enrollment, graduates, and placement between 1994 and 2016. Using the American Dental Association annual survey of advanced dental education programs not accredited by the Commission on Dental Accreditation and data obtained from 22 oral sciences PhD programs, we assessed student demographics, enrollment, graduation, and placement. Based on the data provided by program directors, the average new enrollment was 33, and graduation was 26 per year. A total of 605 graduated; 39 did not complete; and 168 were still in training. Among those 605 graduates, 211 were faculty in U.S. academic institutions, and 77 were faculty in foreign institutions. Given that vacant budgeted full-time faculty positions averaged 257 per year during this period, graduates from those oral sciences PhD programs who entered academia in the United States would have filled 9 (3.6%) vacant faculty positions per year. Therefore, PhD programs have consistently generated only a small pipeline of dental school faculty. Better mentoring to retain talent in academia is necessary. Stronger support and creative funding plans are essential to sustain the PhD program. Furthermore, the oral sciences PhD program database should be established and maintained by dental professional organizations to allow assessments of training models, trends of enrollment, graduation, and placement outcomes

The Acid Test of Fluoride: How pH Modulates Toxicity

Background: It is not known why the ameloblasts responsible for dental enamel formation are uniquely sensitive to fluoride ($F^−$). Herein, we present a novel theory with supporting data to show that the low pH environment of maturating stage ameloblasts enhances their sensitivity to a given dose of $F^−$. Enamel formation is initiated in a neutral pH environment (secretory stage); however, the pH can fall to below 6.0 as most of the mineral precipitates (maturation stage). Low pH can facilitate entry of $F^−$ into cells. Here, we asked if $F^−$ was more toxic at low pH, as measured by increased cell stress and decreased cell function. Methodology/Principal Findings: Treatment of ameloblast-derived LS8 cells with $F^−$ at low pH reduced the threshold dose of $F^−$ required to phosphorylate stress-related proteins, PERK, eIF2α, JNK and c-jun. To assess protein secretion, LS8 cells were stably transduced with a secreted reporter, Gaussia luciferase, and secretion was quantified as a function of $F^−$ dose and pH. Luciferase secretion significantly decreased within 2 hr of $F^−$ treatment at low pH versus neutral pH, indicating increased functional toxicity. Rats given 100 ppm $F^−$ in their drinking water exhibited increased stress-mediated phosphorylation of eIF2α in maturation stage ameloblasts (pH<6.0) as compared to secretory stage ameloblasts (pH∼7.2). Intriguingly, $F^−$-treated rats demonstrated a striking decrease in transcripts expressed during the maturation stage of enamel development (Klk4 and Amtn). In contrast, the expression of secretory stage genes, AmelX, Ambn, Enam and Mmp20, was unaffected. Conclusions: The low pH environment of maturation stage ameloblasts facilitates the uptake of $F^−$, causing increased cell stress that compromises ameloblast function, resulting in dental fluorosis

Effect of fluoride toothpastes on enamel demineralization

BACKGROUND: It was the aim of this study to investigate the effect of four different toothpastes with differing fluoride compounds on enamel remineralization. METHODS: A 3 × 3 mm window on the enamel surface of 90 human premolars was demineralized in a hydroxyethylcellulose solution at pH 4.8. The teeth were divided into 6 groups and the lower half of the window was covered with varnish serving as control. The teeth were immersed in a toothpaste slurry containing: placebo tooth paste (group 1); remineralization solution (group 2); Elmex Anticaries (group 3); Elmex Sensitive (group 4); Blend-a-med Complete (group 5) and Colgate GRF (group 6). Ten teeth of each group were used for the determination of the F(- )content in the superficial enamel layer and acid solubility of enamel expressed in soluble phosphorus. Of 6 teeth of each group serial sections were cut and investigated with polarization light microscopy (PLM) and quantitative energy dispersive X-ray analysis (EDX). RESULTS: The PLM results showed an increased remineralization of the lesion body in the Elmex Anticaries, Elmex Sensitive and Colgate GRF group but not in the Blend-a-med group. A statistically significant higher Ca content was found in the Elmex Anticaries group. The fluoride content in the superficial enamel layer was significantly increased in both Elmex groups and the Blend-a-med group. Phosphorus solubility was significantly decreased in both Elmex groups and the Blend-a-med group. CONCLUSION: It can be concluded that amine fluoride compounds in toothpastes result in a clearly marked remineralization of caries like enamel lesions followed by sodium fluoride and sodium monofluorophosphate formulations

The VicGeneration study - a birth cohort to examine the environmental, behavioural and biological predictors of early childhood caries: background, aims and methods

Background Dental caries (decay) during childhood is largely preventable however it remains a significant and costly public health concern, identified as the most prevalent chronic disease of childhood. Caries in children aged less than five years (early childhood caries) is a rapid and progressive disease that can be painful and debilitating, and significantly increases the likelihood of poor child growth, development and social outcomes. Early childhood caries may also result in a substantial social burden on families and significant costs to the public health system. A disproportionate burden of disease is also experienced by disadvantaged populations. Methods/Design This study involves the establishment of a birth cohort in disadvantaged communities in Victoria, Australia. Children will be followed for at least 18 months and the data gathered will explore longitudinal relationships and generate new evidence on the natural history of early childhood caries, the prevalence of the disease and relative contributions of risk and protective biological, environmental and behavioural factors. Specifically, the study aims to: 1. Describe the natural history of early childhood caries (at ages 1, 6, 12 and 18 months), tracking pathways from early bacterial colonisation, through non-cavitated enamel white spot lesions to cavitated lesions extending into dentine. 2. Enumerate oral bacterial species in the saliva of infants and their primary care giver. 3. Identify the strength of concurrent associations between early childhood caries and putative risk and protective factors, including biological (eg microbiota, saliva), environmental (fluoride exposure) and socio-behavioural factors (proximal factors such as: feeding practices and oral hygiene; and distal factors such as parental health behaviours, physical health, coping and broader socio-economic conditions). 4. Quantify the longitudinal relationships between these factors and the development and progression of early childhood caries from age 1-18 months. Discussion There is currently a lack of research describing the natural history of early childhood caries in very young children, or exploring the interactions between risk and protective factors that extend to include contemporary measures of socio-behavioural factors. This study will generate knowledge about pathways, prevalence and preventive opportunities for early childhood caries, the most prevalent child health inequality

Effects of fluoride on the interactions between amelogenin and apatite crystals.

Fluorosed enamel is more porous and less mineralized, possibly related to altered amelogenin-modulated crystal growth. The purpose of this study was to examine the role of fluoride in interactions between amelogenin and apatite crystals. Recombinant human amelogenin (rh174) was bound to carbonated hydroxyapatite containing various amounts of fluoride, and analyzed by protein assay, SDS PAGE, and AFM. Interactions between rh174 and fluoride were assayed by isothermal titration calorimetry (ITC). The initial binding rate of rh174, as well as total amount of rh174 bound to fluoride-containing carbonated hydroxyapatite, was greater than that in the control carbonated hydroxyapatite. Fluoride in solution at physiologic (5.3 micromolar, or 0.1 ppm) concentrations showed no significant effect on binding, but higher fluoride levels significantly decreased protein binding. ITC showed no interactions between fluoride and rh174. These results suggest that fluoride incorporation into the crystal lattice alters the crystal surface to enhance amelogenin binding, with no direct interactions between fluoride and amelogenin

Evolutionary Story of Mammalian-specific Amelogenin Exons 4, ``4b'', 8, and 9

International audienceAmelogenin gene organization varies from 6 exons (1,2,3,5,6,7) in amphibians and sauropsids to 10 in rodents. The additional exons are exons 4, 8, 9, and ``4b'', the latter being as yet unidentified in AMELX transcripts. To learn more about the evolutionary origin of these exons, we used an in silico approach to find them in 39 tetrapod genomes. AMEL organization with 6 exons was the ancestral condition. Exon 4 was created in an ancestral therian (marsupials + placentals), then exon 9 in an ancestral placental, and finally exons ``4b'' and 8 in rodents, after divergence of the squirrel lineage. These exons were either inactivated in some lineages or remained functional: Exon 4 is functional from artiodactyls onward; exon 9 is known, to date, only in rodents, but could be coding in various mammals; and exon ``4b'' was probably coding in some rodents. We performed PCR of cDNA isolated from mouse and human tooth buds to identify the presence of these transcripts. A sequence analogous to exon ``4b'', and to exon 9, could not be amplified from the respective tooth cDNA, indicating that even though sequences similar to these exons are present, they are not transcribed in these species