Effect of Dimethyloxalylglycine on Stem Cells Osteogenic Differentiation and Bone Tissue Regeneration-A Systematic Review.

Dimethyloxalylglycine (DMOG) has been found to stimulate osteogenesis and angiogenesis of stem cells, promoting neo-angiogenesis in bone tissue regeneration. In this review, we conducted a comprehensive search of the literature to investigate the effects of DMOG on osteogenesis and bone regeneration. We screened the studies based on specific inclusion criteria and extracted relevant information from both in vitro and in vivo experiments. The risk of bias in animal studies was evaluated using the SYRCLE tool. Out of the 174 studies retrieved, 34 studies met the inclusion criteria (34 studies were analyzed in vitro and 20 studies were analyzed in vivo). The findings of the included studies revealed that DMOG stimulated stem cells' differentiation toward osteogenic, angiogenic, and chondrogenic lineages, leading to vascularized bone and cartilage regeneration. Addtionally, DMOG demonstrated therapeutic effects on bone loss caused by bone-related diseases. However, the culture environment in vitro is notably distinct from that in vivo, and the animal models used in vivo experiments differ significantly from humans. In summary, DMOG has the ability to enhance the osteogenic and angiogenic differentiation potential of stem cells, thereby improving bone regeneration in cases of bone defects. This highlights DMOG as a potential focus for research in the field of bone tissue regeneration engineering.

Small Molecules Promote the Rapid Generation of Dental Epithelial Cells from Human-Induced Pluripotent Stem Cells.

Human-induced pluripotent stem cells (hiPSCs) offer a promising source for generating dental epithelial (DE) cells. Whereas the existing differentiation protocols were time-consuming and relied heavily on growth factors, herein, we developed a three-step protocol to convert hiPSCs into DE cells in 8 days. In the first phase, hiPSCs were differentiated into non-neural ectoderm using SU5402 (an FGF signaling inhibitor). The second phase involved differentiating non-neural ectoderm into pan-placodal ectoderm and simultaneously inducing the formation of oral ectoderm (OE) using LDN193189 (a BMP signaling inhibitor) and purmorphamine (a SHH signaling activator). In the final phase, OE cells were differentiated into DE through the application of Purmorphamine, XAV939 (a WNT signaling inhibitor), and BMP4. qRT-PCR and immunostaining were performed to examine the expression of lineage-specific markers. ARS staining was performed to evaluate the formation of the mineralization nodule. The expression of PITX2, SP6, and AMBN, the emergence of mineralization nodules, and the enhanced expression of AMBN and AMELX in spheroid culture implied the generation of DE cells. This study delineates the developmental signaling pathways and uses small molecules to streamline the induction of hiPSCs into DE cells. Our findings present a simplified and quicker method for generating DE cells, contributing valuable insights for dental regeneration and dental disease research.

Endoplasmic reticulum chaperone GRP78 participates in fluoride-induced autophagy in LS8 cells by regulating the IRE1-TRAF2-JNK pathway.

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.

MAP kinase phosphatase MKP-1 regulates p-ERK1/2 signaling pathway with fluoride treatment.

Dental fluorosis is characterized by hypomineralization of tooth enamel caused by ingestion of excessive fluoride during enamel formation. Excess fluoride could have effects on the ERK signaling, which is essential for the ameloblasts differentiation and tooth development. MAP kinase phosphatase-1 (MKP-1) plays a critical role in regulating ERK related kinases. However, the role of MKP-1 in ameloblast and the mechanisms of MKP-1/ERK signaling in the pathogenesis of dental fluorosis are incompletely understood. Here, we adopted an in vitro fluorosis cell model using murine ameloblasts-like LS8 cells by employing sodium fluoride (NaF) as inducer. Using this system, we demonstrated that fluoride exposure led to an inhibition of p-MEK and p-ERK1/2 with a subsequent increase in MKP-1 expression in a dose-dependent manner. We further identified, under high dose fluoride, MKP-1 acted as a negative regulator of the fluoride-induced p-ERK1/2 signaling, leading to downregulation of CREB, c-myc, and Elk-1. Our results identify a novel MKP-1/ERK signaling mechanism that regulates dental fluorosis and provide a framework for studying the molecular mechanisms of intervention and fluorosis remodeling under normal and pathological conditions. MKP-1 inhibitors may prove to be a benefit therapeutic strategy for dental fluorosis treatment.

Prevalence of dentin hypersensitivity among the residents of Xi'an city, China.

OBJECTIVE: To investigate the prevalence and intra-oral distribution of dentine hypersensitivity (DH) and to evaluate the related risk factors. MATERIAL AND METHODS: A total of 1320 subjects, aged between 20 and 69 years old, were selected from six communities in the urban areas of Xi'an, China. The data were collected by conducting individual interviews using a standard questionnaire; then, the clinical examination was performed for patients who reported about the discomforts they felt in their teeth when subjected to chemical, mechanical and thermal stimuli. Dentine hypersensitivity (DH) was diagnosed by a subject short, sharp pain in response to a blast of cold air from a triple syringe. RESULTS: While replying to the questionnaire, 445 subjects reported about signs of discomfort in the teeth. DH was diagnosed in 336 persons by clinic examination. Thus, the overall prevalence of DH was 33.7% in the questionnaire and 25.5% in the intraoral test. The prevalence of DH was higher in females (33.8%) than in males (22.2%). Furthermore, we found that the prevalence of DH was highest in the age group of 50-59 years (39.3%). The most common initiation factors were acid (37.7%) followed by cold stimuli (35.8%). In general, most subjects with sensitive teeth had a higher educational background. CONCLUSIONS: The prevalence of DH was 25.5% in the population of Xi'an City in China. More emphasis should give to middle-aged and old females while planning oral health intervention campaigns. In addition, premolars and cervical surfaces should be examined for the prevention of DH.

LS8 cell apoptosis induced by NaF through p-ERK and p-JNK - a mechanism study of dental fluorosis.

OBJECTIVE: To investigate the possible biological mechanism of dental fluorosis at a molecular level. MATERIAL AND METHODS: Cultured LS8 were incubated with serum-free medium containing selected concentrations of NaF (0 ∼ 2 mM) for either 24 or 48 h. Subcellular microanatomy was characterized using TEM; meanwhile, selected biomolecules were analysed using various biochemistry techniques. Transient transfection was used to modulate a molecular pathway for apoptosis. RESULTS: Apoptosis of LS8 was induced by NaF treatment that showed both time and concentration dependency. The activity of caspase-3, -8, -9 was found to be increased with NaF in a dose-dependent manner. Western blot revealed that the protein expression of p-ERK and p-JNK were decreased, while the expression of p-P38 was increased. Inhibition of the p-ERK and p-JNK pathways resulted in a similar decrease for caspase-3. CONCLUSION: During NaF-induced apoptosis of LS8, p-ERK and p-JNK were closely associated with induction of apoptosis, which might be a mechanism of dental fluorosis.

Excessive fluoride reduces Foxo1 expression in dental epithelial cells of the rat incisor.

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) mRNAs was analyzed by real-time RT-PCR, and western blotting was used to measure total and nuclear FOXO1 protein levels in mature dental epithelial cells. The results revealed that nuclear FOXO1 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 AmtnmRNAs, the protein levels of total and nuclearFOXO1 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.

Oral health status and oral health knowledge, attitudes and behavior among rural children in Shaanxi, western China: a cross-sectional survey.

BACKGROUND: The current oral health status and possible dental risk factors among children in rural Shaanxi Province, western China are unreported. This study aimed to describe the oral health status and to analyze the possible risk factors for the oral health status in this population. METHODS: A multi-stage cluster sampling method was used to survey 12- to 15-year-olds and 4- to 6-year-olds in villages in Shaanxi Province. The structured questionnaires were provided to the 12- to 15-year-olds and to the caregivers of the 4- to 6-year-olds to collect information on the subjects' oral health knowledge, attitudes and behavior. A clinical examination was performed to assess dental caries and gingival bleeding (only 12- to 15-year-olds). SPSS 17.0 statistical software was used to analyze the data. RESULTS: The decayed, missing, filled teeth (DMFT) index scores of 12- to 15-year-olds and 4-to 6-year-olds averaged 0.45 and 3.05, respectively. The caries prevalence was 23.9% in 12- to 15-year-olds and 67% in 4-to 6-year-olds. Additionally, 45.2% of the 12- to 15-year-olds had gingival bleeding and 62.8% had calculus. The oral health knowledge of the subjects was generally poor, whereas they held very positive attitudes toward oral health. A low number of participants reported that they brushed their teeth at least twice daily. Moreover, a statistically significant relationship was found between oral health knowledge scores, tooth brushing frequency and DMFT scores as well as gingival bleeding in the 12- to 15-year-olds. Frequency of sweets consumption was strongly related to dmft scores in the 4- to 6-year-olds. CONCLUSION: The oral health status, oral health knowledge and behaviors among village children in Shaanxi Province are poor. Oral health education to improve oral health knowledge and to increase the frequency of tooth brushing should be undertaken in the rural schools in western China.

Epigallocatechin-3-gallate attenuates fluoride induced apoptosis via PI3K/FoxO1 pathway in ameloblast-like cells.

Fluoride is a double-edged sword. It was widely used for early caries prevention while excessive intake caused a toxicology effect, affected enamel development, and resulted in dental fluorosis. The study aimed to evaluate the protective effect and mechanism of Epigallocatechin-3-gallate (EGCG) on the apoptosis induced by fluoride in ameloblast-like cells. We observed that NaF triggered apoptotic alterations in cell morphology, excessive NaF arrested cell cycle at the G1, and induced apoptosis by up-regulating Bax and down-regulating Bcl-2. NaF activated the insulin-like growth factor receptor (IGFR), and phosphatidylinositol-3-hydroxylase (p-PI3K), while dose-dependently down-regulating the expression of Forkhead box O1 (FoxO1). EGCG supplements reversed the changes in LS8 morphology, the cell cycle, and apoptosis induced by fluoride. These results indicated that EGCG possesses a protective effect against fluoride toxicity. Furthermore, EGCG suppressed the activation of p-PI3K and the down-regulation of FoxO1 caused by fluoride. Collectively, our findings suggested that EGCG attenuated fluoride-induced apoptosis by inhibiting the PI3K/FoxO1 signaling pathway. EGCG may serve as a new alternative method for dental fluorosis prevention, control, and treatment.

Novel pit and fissure sealant with nano-CaF2 and antibacterial monomer: Fluoride recharge, microleakage, sealing ability and cytotoxicity.

Conventional resin-based sealants release minimal fluoride ions (F) and lack antibacterial activity. The objectives of this study were to: (1) develop a novel bioactive sealant containing calcium fluoride nanoparticles (nCaF2) and antibacterial dimethylaminohexadecyl methacrylate (DMAHDM), and (2) investigate mechanical performance, F recharge and re-release, microleakage, sealing ability and cytotoxicity. Helioseal F served as commercial control. The initial F release from sealant containing 20% nCaF2 was 25-fold that of Helioseal F. After ion exhaustion and recharge, the F re-release from bioactive sealant did not decrease with increasing number of recharge and re-release cycles. Elastic modulus of new bioactive sealant was 44% higher than Helioseal F. The new sealant had excellent sealing, minimal microleakage, and good cytocompatibility. Hence, the nanostructured sealant had substantial and sustained F release and antibacterial activity, good sealing ability and biocompatibility. The novel bioactive nCaF2 sealant is promising to provide long-term F ions for caries prevention.

Intravoxel incoherent motion diffusion-weighted imaging in quantitative evaluation of Ileal Crohn's disease - A comparison with dynamic contrast-enhanced magnetic resonance imaging and ileocolonoscopy.

OBJECTIVES: To investigate the prospective role of intravoxel incoherent motion diffusion-weighted imaging (IVIM-DWI) in evaluating terminal ileal Crohn's disease (CD) inflammation quantitatively, compared with quantitative dynamic contrastenhanced magnetic resonance imaging (DCE-MRI) and ileocolonoscopic segmental score. METHODS: Fifty CD patients underwent magnetic resonance enterography (MRE) including IVIM-DWI and quantitative DCE-MRI from Jan. 2017 to Nov. 2019. ADC, D, D* and f value of IVIM-DWI and Ktrans, Kep, and Ve value of DCE-MRI in normal (n = 50) and inflamed bowel segments (n = 50), defined during the clinical MRI analysis, were calculated and compared using Wilcoxon signed-rank tests respectively. Receiver operating characteristic (ROC) analysis was performed. Correlations between IVIM-DWI and DCE-MRI parameters in comparison with ileocolonoscopic segmental score were assessed using Spearman's rank correlation analysis. RESULTS: For IVIM-DWI, ADC, D, D* and f value showed significant differences respectively between normal and inflamed bowel segments (p < 0.05). ADC value presented the highest diagnostic accuracy (AUC = 0.813) and sensitivity (92%), and D value presented the highest specificity (84%) for the evaluation of inflamed bowel segments. For DCE-MRI, Ktrans value presented the highest diagnostic accuracy (AUC = 0.835), the highest sensitivity for Kep value (88%) and the highest specificity for Ve value (96%). ADC, f and Ktrans value had high correlations with ileocolonoscopic score respectively (r = -0.739-0.876, p < 0.01). The logarithm of normalized signal intensity/b-values for IVIM-DWI could also indicate directly the evident difference between the normal and inflamed bowel segments of terminal ileal CD. CONCLUSION: IVIM-DWI will be another promising noninvasive tool to provide precise quantitative-indicators in evaluating inflamed bowel segments of terminal ileal CD with little contrast-agent damage worries.

NaF reduces KLK4 expression by decreasing Foxo1/Runx2 expression in LS8 cells.

OBJECTIVE: This study aimed to investigate the effect of high fluoride on runt-related transcription factor 2 (Runx2) expression and to explore the possible relationship among Runx2, forkhead box o1 (Foxo1) and kallikrein 4 (KLK4) in high fluoride-treated ameloblasts. DESIGN: Ameloblast-like cells (LS8 cells) were exposed to various concentrations of sodium fluoride (NaF) for up to 48 h. Runx2 expression was downregulated by gene silencing, and Foxo1 expression was up- and downregulated by gene overexpression and silencing, respectively. The mRNA and protein levels of Runx2, Foxo1, KLK4 and matrix metalloproteinase 20 (MMP20) were detected by qRT-PCR and western blotting. RESULTS: Runx2 expression was decreased in a dose- and time-dependent manner in NaF-treated LS8 cells. The knockdown of Runx2 markedly decreased KLK4 expression in LS8 cells under NaF conditions. However, the variation trend of MMP20 was unclear. In addition, forced Foxo1 expression led to significant upregulation of Runx2 in LS8 cells under NaF conditions. In contrast, the knockdown of Foxo1 markedly decreased the Runx2 protein levels under NaF conditions. Moreover, Foxo1 downregulation markedly decreased runx2 mRNA levels, and this inhibition in LS8 cells was intensified when combined with NaF treatment. CONCLUSION: The results indicated that NaF reduces Runx2 expression in LS8 cells and that decreased Foxo1/Runx2 expression induced by high fluoride is a cause of low KLK4 expression.

Enhanced healing process of tooth sockets using strontium-doped TiO2.

Prevention of residual ridge resorption is important for tooth socket healing in clinical treatment. As a well known biomaterial, titanium dioxide (TiO2) has been reported to show desirable bone regeneration capability. On the other hand, strontium plays a role in maintaining normal function in organisms and balancing bone remodeling. Hence, we synthesized strontium-doped titanium dioxide mesoporous nanospheres functionalized with amino-group using diphenyl diisocyanate. After incorporation with segmented polyurethane, the obtained injectable SPU/Sr-TiO2/MDI nanocomposite adhesive showed satisfactory antibacterial activity and cell nontoxicity. This nanocomposite was used for tooth socket healing, and greatly promoted the formation of new bone tissue in the tooth extraction socket.

Evaluation of Palatal Furcation Groove and Root Canal Anatomy of Maxillary First Premolar: A CBCT and Micro-CT Study.

OBJECTIVES: This study is aimed at investigating the root and root canal morphology by cone beam computed tomography (CBCT) and palatal furcation groove of the buccal root by microcomputed tomography (micro-CT) of maxillary first premolars in a Chinese subpopulation. METHODS: This study assessed CBCT images of 440 patients aged 14-80 years. Based on Vertucci's classification, the number of roots and the canal configuration were determined. Forty-eight maxillary first premolars with furcation grooves were analyzed by micro-CT in patients aged 18-25 years. RESULTS: Based on the CBCT assay, 70.22% and 29.32% of maxillary first premolars were 1 root and 2 roots, respectively. The configuration indicated statistical difference (P < 0.05) between male and female patients. The most common canal type was type IV and was found in 44.32% of cases, followed by type I in 27.84%, and then type II in 20.57%. Root bifurcations had 40.13% prevalence which was distributed more in the middle third than in the cervical and the apical third. For the micro-CT study, 95.83% of the furcation groove configuration was found in the bifurcated maxillary first premolars. The length varied from 1.02 to 7.63 mm. The mean depth of this groove was 0.57 mm in the root coronal, 0.47 mm in the root middle, and 0.22 mm in the root apical level. Palatal dentin width was smaller than 1 mm. CONCLUSION: The anatomy of the root and root canal system and the irregular wall width of maxillary first premolars with furcation grooves may help dentists to understand the anatomical morphology and improve the outcomes of endodontic treatment.

Calcium mitigates fluoride-induced kallikrein 4 inhibition via PERK/eIF2α/ATF4/CHOP endoplasmic reticulum stress pathway in ameloblast-lineage cells.

OBJECTIVES: The present study aimed to investigated the effect and mechanism of Ca2+ treatment on fluoride in ameloblast-lineage cells (ALCs). MATERIALS AND METHODS: The effects of fluoride and different Ca2+ levels treatment on the proliferative activity, cell apoptosis, cell cycle, intracellular free Ca2+, were firstly determined. Kallikrein 4 (KLK4), glucose-responsive protein 78 (GRP78), Protein kinase R -like endoplasmic reticulum kinase (PERK), the α subunit of eukaryotic initiation factor 2 (eIF2α), activating transcription factor 4 (ATF4), CCAAT enhancer-binding protein homologous protein (CHOP), were investigated in ALCs. RESULTS: The proliferative activity was obviously inhibited under concentrations of single fluoride high than 1 mM, and indicated highest proliferation at single 2.5 mM Ca2+ concentration in ALC cells. In addition, we found that single fluoride markedly induced intracellular free Ca2+ increasing, G2/M phase arrest, apoptosis. GRP78 and endoplasmic reticulum stress pathway of PERK/eIF2α/ATF4/CHOP were significantly increased, while the proliferation and KLK4 were markedly reduced in ALCs. Ca2+ additional treatment can obviously reverse the effect of fluoride-induced apoptosis and inhibition of KLK4. The effect of GRP78 and endoplasmic reticulum stress pathway of PERK/eIF2α/ATF4/CHOP were also alleviated under Ca2+ additional treatment in ALCs. More important, the results of 2.5 mmol/L Ca2+ treatment on the proliferation, cell cycle and apoptosis suggest this concentration is relatively better to mediate the intracellular Ca2+ homeostasis in ALCs. CONCLUSIONS: In sum, Ca2+-supplementation exerts antagonistic the toxic effects on fluoride and this inhibitory effect suggests the potential implications for Ca2+-supplementation on fluorosis.

Effect of co-precipitation plus spray-drying of nano-CaF2 on mechanical and fluoride properties of nanocomposite.

OBJECTIVE: Fluoride (F)-releasing restoratives typically are either weak mechanically or release only low levels of F ions. The objectives of this study were to: (1) develop a novel photo-cured nanocomposite with strong mechanical properties and high levels of sustained F ion release via a two-step "co-precipitation + spray-drying" technique to synthesize CaF2 nanoparticles (nCaF2); and (2) investigate the effect of spray-drying treatment after co-precipitation of nCaF2 on mechanical properties and F ion release of composite. METHODS: Two types of CaF2 particles were synthesized: A co-precipitation method yielded CaF2cp; "co-precipitation + spray-drying" yielded nCaF2cpsd. Composites were fabricated with fillers of: (1) 0% CaF2 + 70% glass; (2) 10% CaF2cp + 60% glass; (3) 15% CaF2cp + 55% glass; (4) 20% CaF2cp + 50% glass; (5) 10% nCaF2cpsd + 60% glass; (6) 15% nCaF2cpsd + 55% glass; and (7) 20% nCaF2cpsd + 50% glass. A commercial F-releasing nanocomposite served as control. RESULTS: The nCaF2cpsd had much smaller particle size (median = 32 nm) and narrower distribution (22-57 nm) than CaF2cp (median = 5.25 µm, 162 nm-67 µm). The composite containing nCaF2cpsd had greater flowability, flexural strength, elastic modulus and hardness than CaF2cp composite and commercial control composite. At 84-day immersion in water, the nanocomposites containing 20% nCaF2cpsd had 65 times higher cumulative F release, and 77 times greater long-term F-release rate, than commercial control. CONCLUSIONS: A novel two-step "co-precipitation + spray-drying" technique of synthesizing nCaF2 was developed. The photo-cured nanocomposite containing 20% nCaF2cpsd possessed strong mechanical properties and excellent long-term F-release ability, and hence is promising for dental restoration applications to inhibit secondary caries.

A multi-centre and cross-sectional study of dentine hypersensitivity in China.

AIM: The objective of this study was to establish the prevalence of dentine hypersensitivity (DH) in Chinese urban adults and the possible effects of smoking on DH. MATERIALS AND METHODS: A total of 2640 subjects were distributed equally in 12 communities in Chengdu and Xian City, respectively, and of all age groups (10 years for an age group) including the same number of male and female subjects in each community. Each subject completed a structured interview and the subjects who reported hypersensitivity symptoms were examined with cold air from a dental triple syringe in order to confirm the diagnosis of DH. Attachment loss and gingival recession of sensitive teeth were measured by a Williams periodontal probe. RESULTS: The diagnosis of DH established following a clinical assessment yielded an overall prevalence of 25.5%. The 50-59-year-old age group had the greatest number of subjects with DH (p<0.05). 78.6% and 31.4% of sensitive teeth were associated with attachment loss and gingival loss, respectively. Subjects who smoked did not have more sensitive teeth on average than subjects who did not smoke (p>0.05). CONCLUSIONS: The prevalence of DH in a selected community population was 25.5%.

Effects of applying amoxicillin in juvenile mice on enamel mineralization and the expression of kallikrein­related peptidase 4 and tight junction proteins in ameloblasts.

Amoxicillin is a common pediatric drug. However, to the best of our knowledge, the role of amoxicillin in enamel hypomineralization has not yet been fully elucidated. The aim of the present study was to assess the effects of amoxicillin on enamel mineralization, the morphology of ameloblasts, as well as the expression of kallikrein­related peptidase 4 (KLK4), and the tight junction proteins, claudin 1 (CLDN1), claudin 4 (CLDN4) and occludin (OCLN), in ameloblasts of juvenile mice. A total of 36 3­day­old Kunming mice were randomly divided into three groups. The mice were administered 0, 50 or 100 mg/kg amoxicillin by intragastric administration for 19 days. The surface morphology and calcium (Ca), phosphorous (P) and carbon contents of mandibular incisors and first molars were examined by scanning electron microscopy and energy dispersive X­ray spectroscopy. Histological changes in the ameloblasts of mandibular incisors were analyzed by hematoxylin and eosin staining. The KLK4, CLDN1, CLDN4 and OCLN expression levels of ameloblasts were observed by immunohistochemical staining. The incidence of white patches in the incisor was 100% in the 100 mg/kg amoxicillin­treated groups. A greater number of enamel defects were observed in the incisal/occlusal half of mandibular incisors/molars compared with in the cervical half in the amoxicillin­treated groups. Following phosphoric­acid treatment, the enamel rod and interrod were aligned in a disorderly manner in the amoxicillin­treated groups. Amoxicillin decreased the Ca/P ratio in the enamel of mandibular incisors and molars. More intercellular spaces among maturation ameloblasts were observed in the amoxicillin­treated groups. Amoxicillin decreased KLK4 and CLDN1, CLDN4 and OCLN expression in mature ameloblasts. The administration of amoxicillin in juvenile mice induced enamel hypomineralization, and the effects of amoxicillin on enamel hypomineralization may be mediated via multiple pathways.

Calcium promotes differentiation in ameloblast-like LS8 cells by downregulation of phosphatidylinositol 3 kinase /protein kinase B pathway.

OBJECTIVES: To investigate the effect and mechanism of calcium on LS8 cell differentiation, especially on phosphatidylinositol 3 kinase (PI3K) /protein kinase B(AKT) pathway. MATERIALS AND METHODS: Ameloblast-like LS8 cell line was used and additional 0-3.5 mmol/L calcium chloride was treated for 24 h, 48 h. Cell viability and morphological changes, cell cycle and associated regulatory proteins were analyzed. RESULTS: No significant effects on morphological changes were observed. Decreased cell viability and increased S phase cells were accompanied by the significant decrease of cyclin A and cyclin B proteins, and significant increase of cyclin D protein in LS8 cells. Additionally, kallikrein-4 and amelotin expressions were significantly increased. Finally, the levels of PI3K, AKT, p-AKT and forkhead box O3 (FOXO3) significantly downregulated after calcium treatment in LS8 cells. CONCLUSIONS: Calcium inhibit proliferation and promotes differentiation in LS8 cells, this is closely related to the downregulation of PI3K/AKT signal in LS8 cells.

Novel pit and fissure sealant containing nano-CaF2 and dimethylaminohexadecyl methacrylate with double benefits of fluoride release and antibacterial function.

OBJECTIVE: Pit and fissure sealants with antibacterial and remineralization properties have broad application prospects in caries prevention. The objectives of this study were to: (1) develop a novel pit and fissure sealant containing CaF2 nanoparticles (nCaF2) and dimethylaminohexadecyl methacrylate (DMAHDM); and (2) investigate the effects of nCaF2 and DMAHDM on biofilm response and fluoride (F) ion release for the first time. METHODS: Helioseal F was used as a control. Bioactive sealants were formulated with DMAHDM and nCaF2. Flow properties, enamel shear bond strength, hardness and F ion releases were measured. Streptococcus mutans (S. mutans) biofilms were grown on sealants. Biofilm metabolic activity, lactic acid production, colony-forming units (CFU), and pH of biofilm culture medium were measured. RESULTS: Adding 5% DMAHDM and 20% nCaF2 did not reduce the paste flow and enamel bond strength, compared to control (p < 0.05). Hardness of sealants with 20% nCaF2 and DMAHDM was higher than control (p < 0.05). The F ion release from 20% nCaF2 was much higher than that of commercial control (p < 0.05). The sealant with DMAHDM reduced the S. mutans biofilm CFU by 4 logs. The pH in biofilm medium of the new bioactive sealant was much higher (pH 6.8) than that of commercial sealant (pH 4.66) (p < 0.05). SIGNIFICANCE: The new bioactive pit and fissure sealant with nCaF2 and DMAHDM achieved high fluoride release and strong antibacterial performance. This novel fluoride-releasing and antibacterial sealant is promising to inhibit caries and promote the remineralizaton of enamel and dentin.