miR-23b mediates TNF-α-Inhibited Osteogenic Differentiation of Human Periodontal Ligament Stem Cells by Targeting Runx2.

Periodontitis is the most prevalent oral infection disease, which causes the destruction of periodontal supporting tissues and eventual tooth loss. This study aimed to investigate the molecular mechanism of miRNA-23b (miR-23b) in regulating the osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) in an inflammatory environment. Results revealed that tumor necrosis factor-α (TNF-α), a notoriously inflammatory cytokine, remarkably attenuated the osteogenic differentiation of hPDLSCs, which were partially rescued by SKL2001 (Wnt/ß-catenin agonist). We further explored the underlying roles of miRNAs involved in TNF-α-inhibited osteogenesis of hPDLSCs. The miR-23b significantly increased with TNF-α stimulation, which was abolished by SKL2001. Similar to the effect of TNF-α, miR-23b agonist (agomir-23b) dramatically reduced the expression of runt-related transcription factor 2 (Runx2) and suppressed the osteogenic differentiation of hPDLSCs. The inhibition of miR-23b significantly increased Runx2, which is the major transcription factor during osteogenesis, thereby indicating that miR-23b was an endogenous regulator of Runx2 in hPDLSCs. Bioinformatic analysis and dual luciferase reporter assays confirmed that Runx2 was a target gene of miR-23b. Furthermore, the gain function assay of Runx2 revealed that the Runx2 overexpression efficiently reversed the suppression of the osteogenic differentiation of hPDLSCs with miR-23b agonist, suggesting that the suppressing effect of miR-23b on osteogenesis was mediated by Runx2 inhibition. Our study clarified that miR-23b mediated the TNF-α-inhibited osteogenic differentiation of hPDLSCs by targeting Runx2. Therefore, the expanded function of miR-23b in the osteogenesis of hPDLSCs under inflammatory conditions. This study might provide new insights and a novel therapeutic target for periodontitis.

Status of dental caries and associated factors in Tibetan adults: findings from the fourth China National Oral Health Survey.

BACKGROUND: Tibet, a region where average elevation is above 3500 m and socio-economic development is relatively lower, was not included in National Oral Health Survey over decades. The cross-sectional study aimed to investigate the status of dental caries and associated factors in Tibetan adults. METHODS: Participants aged 35-44, 55-64 and 65-74 years were selected. Decayed, missing, and filled tooth (DMFT), decayed and filled root (DF-Root) and root canal index (RCI) were used to evaluate dental caries. Questionnaire survey on demographic information, socioeconomic status, dietary habits, and oral health knowledge and behavior was conducted. Mann-Whitney U test, logistic regression were used for the statistical analyses. RESULTS: A total of 446 participants were enrolled in the survey. Of these: 222 (49.8%) were females, 224 (50.2%) were males; 149 (33.4%), 151 (33.9%), 146 (32.7%) were aged 35-44, 55-64 and 65-74 years respectively. The mean DMFT (SD) was 7.62 (4.84), 12.46 (8.16), and 21.38 (8.93). The filling rate was very low in all age groups (1.77%, 0.98%, 0.45%). The mean DF-Root (SD) was 0.50 (1.04), 1.04 (2.02), 1.32 (2.14), respectively. Root caries index was 42.27, 44.78 and 57.60%. Older age (65-74 age group) was positively associated with crown caries (odds ratio = 31.20, 95% confidence interval: 10.70-90.96). College degree and above and brushing teeth at least once a day were negatively associated with crown caries (odds ratio = 0.28, 95% confidence interval: 0.09-0.89; odds ratio = 0.39, 95% confidence interval: 0.21-0.72, respectively). Rural area, high income level and brushing teeth at least once a day were negatively and tooth with attachment loss was positively associated with root caries. CONCLUSIONS: The status of dental caries in the adults in Tibet is severe and the treatment rate is very low. The study suggests a correlation between crown caries and the variables age, level of education and frequency of tooth brushing; correlation between root caries and residence, income level, frequency of tooth brushing and exposed root surfaces. These findings could be as reference to develop community based interventions to reduce the prevalence of caries in Tibet.

Application of fluoride disturbs plaque microecology and promotes remineralization of enamel initial caries.

Background: The caries-preventive effect of topical fluoride application has been corroborated by a number of clinical studies. However, the effect of fluoride on oral microecology remains unclear. Objective: To monitor the effect of fluoride on dental plaque microecology and demineralization/remineralization balance of enamel initial caries. Methods: Three-year-old children were enrolled and treated with fluoride at baseline and 6 months. International Caries Detection and Assessment System II indices of 52 subjects were measured at baseline, 3, 6, and 12 months. Supragingival plaque samples of 12 subjects were collected at baseline, 3 and 14 days for 16S rRNA sequencing. Results: Changes in microbial community structure were observed at 3 days after fluoridation. Significant changes in the relative abundance of microorganisms were observed after fluoride application, especially Capnocytophaga, unidentified Prevotellaceae and Rothia. Functional prediction revealed that cell movement, carbohydrate and energy metabolism were affected significantly after fluoride application. Fluoride significantly inhibited enamel demineralization and promoted remineralization of early demineralized caries enamel at 3 months. Conclusion: Fluoride application significantly inhibited the progression of enamel initial caries and reversed the demineralization process, possibly by disturbing dental plaque microecology and modulating the physicochemical action of demineralization/remineralization. This deepened our understanding of caries-preventive effects and mechanisms of fluoride.

Preparation of a highly crosslinked biosafe dental nanocomposite resin with a tetrafunctional methacrylate quaternary ammonium salt monomer.

The design of antimicrobial dental nanocomposite resin to prevent secondary dental caries and minimize biosafety problems is an important endeavor with both fundamental and practical implications. In the present work, a novel tetrafunctional methacrylate-based polymerizable quaternary ammonium monomer (TMQA) was synthesized with the aim of using it as an immobilized antibacterial agent in methacrylate dental composites, and its structure was characterized. The antibacterial action of TMQA and polymerized resin specimens against suspected cariogenic bacteria Streptococcus mutans were evaluated. Furthermore, the double bond conversion, contact angle, water sorption, solubility, heterogeneity, and crosslink density of the experimental resins with different concentrations of TMQA were investigated. CCK-8 and real-time cell analyses were used to evaluate the cytotoxicity of the experimental resins. The results showed that TMQA was successfully synthesized and had strong antibacterial properties against Streptococcus mutans. The experimental resins with different concentrations of TMQA had a similar degree of conversion and contact angle to the neat resin. With the addition of 4% TMQA to the resins, water absorption and solubility were reduced while their heterogeneity and crosslink density increased. The cell viability of each experimental group was similar to that of the neat resin group and was higher than that of the commercial adhesive single bond 2 group. Therefore, TMQA can be used to impart antibacterial properties to resins and increase the crosslink density of dental resin composites.

Antibacterial and thermomechanical properties of experimental dental resins containing quaternary ammonium monomers with two or four methacrylate groups.

Resins with strong antibacterial and thermomechanical properties are critical for application in oral cavities. In this study, we first evaluated the antibacterial effect of an unfilled resin incorporating 1, 4, and 7 mass% of quaternary ammonium salt (QAS) monomers containing two methacrylate groups (MAE-DB) and four methacrylate groups (TMH-DB) against Streptococcus mutans, and tested the cytotoxicity and thermomechanical properties of the 4 mass% MAE-DB and TMH-DB modified resin materials. A neat resin without a QAS monomer served as the control. As the concentration of both QAS monomers increases, the formation of a Streptococcus mutans biofilm on the experimental material is increasingly inhibited. The results of colony forming unit counts and the metabolic activity showed that both the MAE-DB and TMH-DB modified resins have a strong bactericidal effect on the bacteria in a biofilm, but no bactericidal effect on the bacteria in a solution. The viability-staining and morphology results also demonstrate that the bacteria deform, lyse, shrink, and die on the surface of the two QAS-modified resins. Cytotoxicity results show that the addition of TMH-DB can reduce the cytotoxicity of the resin, while the addition of MAE-DB increases the cytotoxicity of the resin. DMA results show that a TMH-DB modified resin has a higher storage modulus than a MAE-DB modified resin owing to its better crosslink density. The two groups of experimental resins showed a similar glass transition temperature. These data indicate that the two QAS monomers can impart similar antibacterial properties upon contact with a dental resin, whereas TMH-DB can endow the resin with a higher crosslink density and storage modulus than MAE-DB because it has more polymerizable groups.

Effect of Er,Cr:YSGG Laser at Different Output Powers on the Micromorphology and the Bond Property of Non-Carious Sclerotic Dentin to Resin Composites.

BACKGROUND: The objective of this study was to investigate the influence of Er,Cr:YSGG laser irradiated at different powers on the micromorphology and the bonding property of non-carious sclerotic dentin to resin composites. METHODS: Two hundred bovine incisors characterized by non-carious sclerotic dentin were selected, and the seventy-two teeth of which for surface morphological analysis were divided into nine groups according to various treatments (A: the control group, B: only treated with the adhesive Adper Easy One, C: diamond bur polishing followed by Adper Easy One, D-I: Er,Cr:YSGG laser irradiating at 1W, 2W, 3W, 4W, 5W, 6W output power, respectively, followed by Adper Easy One). The surface roughness values were measured by the non-contact three-dimensional morphology scanner, then the surface micromorphologies of surfaces in all groups were assessed by scanning electron microscopy (SEM); meanwhile, Image Pro-Plus 6.0 software was used to measure the relative percentage of open tubules on SEM images. The rest, one hundred twenty-eight teeth for bond strength test, were divided into eight groups according to the different treatments (A: only treated with the adhesive Adper Easy One, B: diamond bur polishing followed by the above adhesive, C-H: Er,Cr:YSGG laser irradiating at 1 W, 2 W, 3 W, 4 W, 5 W, 6 W output power, respectively, followed by the above adhesive), and each group was subsequently divided into two subgroups according to whether aging is performed (immediately tested and after thermocycling). Micro-shear bond strength test was used to evaluate the bond strength. RESULTS: The 4W laser group showed the highest roughness value (30.84±1.93µm), which was statistically higher than the control group and the diamond bur groups (p<0.05). The mean percentages ((27.8±1.8)%, (28.0±2.2)%, (30.0±1.9)%) of open tubules area in the 4W, 5W, 6W group were higher than other groups (p<0.05). The 4W laser group showed the highest micro-shear bond strength not only in immediately tested (17.60±2.55 PMa) but after thermocycling (14.35±2.08MPa). CONCLUSION: The Er,Cr:YSGG laser at 4W power can effectively improve the bonding property between non-carious sclerotic dentin and resin composites by increasing the roughness and mean percentage area of open tubules.

N-Acetyl Cysteine (NAC)-Directed Detoxification of Methacryloxylethyl Cetyl Ammonium Chloride (DMAE-CB).

Methacryloxylethyl cetyl ammonium chloride (DMAE-CB) is a polymerizable antibacterial monomer and has been proved as an effective strategy to achieve bioactive bonding with reliable bacterial inhibitory effects. However, the toxicity of DMAE-CB may hamper its wide application in clinical situations. Thus, this study was designed to investigate the toxicity of DMAE-CB and explore the possible protective effects of N-acetyl cysteine (NAC). High performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) analysis showed that chemical binding of NAC and DMAE-CB occurred in a time dependent manner. Pre-incubation of fourty-eight hours is required for adequate reaction between DMAE-CB and NAC. DMAE-CB reduced human dental pulp cells (hDPCs) viability in a dose-dependent manner. The toxic effects of DMAE-CB were accompanied by increased reactive oxygen species (ROS) level and reduced glutathione (GSH) content. NAC alleviated DMAE-CB-induced oxidative stress. Annexin V/ Propidium Iodide (PI) staining and Hoechst 33342 staining indicated that DMAE-CB induced apoptosis. Collapsed mitochondrial membrane potential (MMP) and activation of caspase-3 were also observed after DMAE-CB treatment. NAC rescued hDPCs from DMAE-CB-induced apoptosis, accompanied by lower level of MMP loss and caspase-3 activity. This study assists to elucidate the mechanism underlying the cytotoxic effects of DMAE-CB and provides theoretical supports for the searching of effective strategies to reduce toxicity of quaternary ammonium dental monomers.

Structure-property relationships in hybrid dental nanocomposite resins containing monofunctional and multifunctional polyhedral oligomeric silsesquioxanes.

Organic-inorganic hybrid materials, such as polyhedral oligomeric silsesquioxanes (POSS), have the potential to improve the mechanical properties of the methacrylate-based composites and resins used in dentistry. In this article, nanocomposites of methacryl isobutyl POSS (MI-POSS [bears only one methacrylate functional group]) and methacryl POSS (MA-POSS [bears eight methacrylate functional groups]) were investigated to determine the effect of structures on the properties of dental resin. The structures of the POSS-containing networks were determined by scanning electron microscopy, transmission electron microscopy, X-ray diffraction, and Fourier transform infrared spectroscopy. Monofunctional POSS showed a strong tendency toward aggregation and crystallization, while multifunctional POSS showed higher miscibility with the dimethacrylate monomer. The mechanical properties and wear resistance decreased with increasing amounts of MI-POSS, indicating that the MI-POSS agglomerates act as the mechanical weak point in the dental resins. The addition of small amounts of MA-POSS improved the mechanical and shrinkage properties. However, samples with a higher MA-POSS concentration showed lower flexural strength and flexural modulus, indicating that there is a limited range in which the reinforcement properties of MA-POSS can operate. This concentration dependence is attributed to phase separation at higher concentrations of POSS, which affects the structural integrity, and thus, the mechanical and shrinkage properties of the dental resin. Our results show that resin with 3% MA-POSS is a potential candidate for resin-based dental materials.

In vitro antibacterial activity of a novel resin-based pulp capping material containing the quaternary ammonium salt MAE-DB and Portland cement.

BACKGROUND: Vital pulp preservation in the treatment of deep caries is challenging due to bacterial infection. The objectives of this study were to synthesize a novel, light-cured composite material containing bioactive calcium-silicate (Portland cement, PC) and the antimicrobial quaternary ammonium salt monomer 2-methacryloxylethyl dodecyl methyl ammonium bromide (MAE-DB) and to evaluate its effects on Streptococcus mutans growth in vitro. METHODS: The experimental material was prepared from a 2 : 1 ratio of PC mixed with a resin of 2-hydroxyethylmethacrylate, bisphenol glycerolate dimethacrylate, and triethylene glycol dimethacrylate (4 : 3 : 1) containing 5 wt% MAE-DB. Cured resin containing 5% MAE-DB without PC served as the positive control material, and resin without MAE-DB or PC served as the negative control material. Mineral trioxide aggregate (MTA) and calcium hydroxide (Dycal) served as commercial controls. S. mutans biofilm formation on material surfaces and growth in the culture medium were tested according to colony-forming units (CFUs) and metabolic activity after 24 h incubation over freshly prepared samples or samples aged in water for 6 months. Biofilm formation was also assessed by Live/Dead staining and scanning electron microscopy. RESULTS: S. mutans biofilm formation on the experimental material was significantly inhibited, with CFU counts, metabolic activity, viability staining, and morphology similar to those of biofilms on the positive control material. None of the materials affected bacterial growth in solution. Contact-inhibition of biofilm formation was retained by the aged experimental material. Significant biofilm formation was observed on MTA and Dycal. CONCLUSION: The synthesized material containing HEMA-BisGMA-TEGDMA resin with MAE-DB as the antimicrobial agent and PC to support mineralized tissue formation inhibited S. mutans biofilm formation even after aging in water for 6 months, but had no inhibitory effect on bacteria in solution. Therefore, this material shows promise as a pulp capping material for vital pulp preservation in the treatment of deep caries.