Abstract Fluoride induces endoplasmic reticulum (ER) stress in ameloblasts, which is responsible for enamel mineralization disorder. Fluoride induces autophagy in ameloblasts, but the molecular mechanisms through which ameloblasts respond to fluoride‐induced cellular stress and autophagy remain unclear. This study investigated ER stress‐induced autophagy and the regulatory role of the ER molecular chaperone GRP78 in fluoride‐induced autophagy in ameloblast LS8 cells. To explore the relationship between fluoride‐induced ER stress and autophagy, we assessed changes in fluoride‐induced autophagy in LS8 cells following overexpression and/or silencing of the ER stress molecular chaperone GRP78. We found that autophagy induced by fluoride was further increased after GRP78 overexpression in LS8 cells. Fluoride‐induced autophagy was reduced in GRP78‐silenced LS8 cells. Furthermore, we found that ER stress can regulate autophagy in fluoride‐treated ameloblasts (LS8 cells) and that the GRP78/IRE1/TRAF2/JNK pathway is involved in the underlying regulation. Our study suggests that ER stress plays a role in fluoride‐induced damage by inducing ameloblast autophagy.
Enamel fluorosis is characterized by hypomineralization, and forkhead box O1 ( Foxo1 ) is essential for mouse enamel biomineralization. This study investigated the effect of fluoride on Foxo1 expression and its implications for enamel fluorosis. Mandibular incisors were extracted from Sprague Dawley rats treated for 3 months with water containing 0, 50, or 100 p.p.m. F − . Immunohistochemistry was used to localize and quantify FOXO1 expression in dental epithelial layer cells of the incisors. The effect of fluoride on expression of Foxo1 , kallikrein‐4 ( Klk4 ), and amelotin ( Amtn ) mRNA s was analyzed by real‐time RT ‐ PCR , and western blotting was used to measure total and nuclear FOXO 1 protein levels in mature dental epithelial cells. The results revealed that nuclear FOXO 1 was mainly localized in the transition and the mature ameloblasts and exhibited weaker expression in the rats exposed to fluoride. In addition to the reduced levels of Foxo1 , Klk4 , and Amtn mRNA s, the protein levels of total and nuclear FOXO 1 were decreased in the mature dental epithelial cells exposed to fluoride. Thus, excessive fluoride may have an effect on the expression levels of Foxo1 in dental epithelial cells and thereby affect hypomineralization of the enamel during fluorosis.
Objective: To determine whether amoxicillin had an effect on the enamel mineralization of SD rats. Methods: Eighteen SD rats were randomly divided into three groups. The rats in the control group were given distilled water. The rats in two experimental groups were administered 50 or 100 mg/kg amoxicillin by intragastric administration from day 3 to day 17 after birth. The general condition, the structure of liver and kidney, the enamel surface changes of mandibular first molars and incisors were observed. The changes of Ca/P ratio on enamel surface were analyzed by X-ray energy dispersive spectrometer (EDS). The surface morphology after phosphoric acid treatment was examined by scanning electron microscopy (SEM). Histological changes in the ameloblasts of mandibular incisors were analyzed by hematoxylin and eosin (HE) staining. Results: Compared with the control group, the general conditions as well as liver and kidney structures of SD rats in 50 and 100 mg amoxicillin groups had no significant differences. There was no obvious chalky changes on the first mandibular molars of SD rats in each group. All the incisors in 50 and 100 mg groups showed different degrees of chalkiness in the labial incisal 1/3 enamel. X-ray EDS analysis showed that the Ca/P ratios of occlusal and incisal 1/3 enamel in 50 and 100 mg groups (occlusal 1/3 of mandibular first molars: 1.51±0.03 and 1.52±0.02, incisal 1/3 of mandibular incisors: 1.46±0.01 and 1.43±0.01) was significantly lower than that in the control group (occlusal 1/3 of mandibular first molars: 1.67±0.41, incisal1/3 of mandibular incisors: 1.73±0.07) (P 0.05). SEM observation showed that the enamel rods of the first molars and incisors in the 50 and 100 mg group varied in size and arranged disorderly. The spaces between the enamel rods were larger than that in the control group and some areas even appeared large pits. HE staining showed that the gaps between ameloblasts in 50 and 100 mg groups were significantly wider than that in the control group. Conclusions: Intake of amoxicillin during the period of enamel development of SD rats might affect enamel mineralization.
Purpose: To investigate the amelogenesis-inductive effects of surface structures at the nanoscale. For this purpose, variable nanostructured titanium dioxide (TiO 2 ) surfaces were used as a framework to regulate the amelogenic behaviors of ameloblasts with the administration of retinoic acid (RA)/dexamethasone (DEX). Materials and methods: TiO 2 nanotubular (NT) surfaces were fabricated via anodization. Mouse ameloblast-like LS-8 cells were seeded and cultured on NT surfaces in the presence or absence of RA/DEX for 48 h. The amelogenic behaviors and extracellular matrix (ECM) mineralization of LS-8 cells on nanostructured Ti surfaces were characterized using field emission scanning electron microscope, laser scanning confocal microscope, quantitative polymerase chain reaction, MTT assay, and flow cytometry. Results: TiO 2 NT surfaces (tube size ~30 and ~80 nm) were constructed via anodization at 5 or 20 V and denoted as NT5 and NT20, respectively. LS-8 cells exhibited significantly increased spread and proliferation, and lower rates of apoptosis and necrosis on NT surfaces. The amelogenic gene expression and ECM mineralization differed significantly on the NT20 and the NT5 and polished Ti sample surfaces in standard medium. The amelogenic behaviors of LS-8 cells were further changed by RA/DEX pretreatment, which directly drove maturation of LS-8 cells. Conclusion: Controlling the amelogenic behaviors of ameloblast-like LS-8 cells by manipulating the nanostructure of biomaterials surfaces represents an effective tool for the establishment of a systemic framework for supporting enamel regeneration. The administration of RA/DEX is an effective approach for driving the amelogenesis and maturation of ameloblasts. Keywords: TiO 2 nanotube, LS-8, surface nanostructure, amelogenesis, retinoic acid
OBJECTIVE To assess changes in prevalence and degree of dental fluorosis in individuals born before and after the introduction of water with 1.2 mg/L fluoride instead of water with 2.0-10.0 mg/L fluoride previously used in Da Li County in China. METHODS The students (n = 291) were divided into 2 groups. The dental fluorosis was scored according to Dean's classification. The statistical analysis was performed by t-test and chi(2) tests. RESULTS The prevalence of dental fluorosis was significantly lower in the group of the students drinking water from the new well (group 1) as compared to the group of the students drinking the old water (group 2), i.e. 48.8% versus 87.2% (P < 0.01). The percentage of moderate to very severe fluorosis was 13.9% and 0 in group 1 as compared to 32.0% and 8.8% in group 2. The fluorosis community index (FCI), defined by Dean, in group 1 and 2 was medium (1.01) and marked (2.12) respectively. CONCLUSIONS The results showed that: (1) The prevalence of dental fluorosis was significantly lowered by the new source of drinking water. (2) Drinking water, even with 1.2 mg/L fluoride, may cause dental fluorosis during the period of tooth mineralization.
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