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Concept: Dentin

266

Evidence of prehistoric dentistry has been limited to a few cases, the most ancient dating back to the Neolithic. Here we report a 6500-year-old human mandible from Slovenia whose left canine crown bears the traces of a filling with beeswax. The use of different analytical techniques, including synchrotron radiation computed micro-tomography (micro-CT), Accelerator Mass Spectrometry (AMS) radiocarbon dating, Infrared (IR) Spectroscopy and Scanning Electron Microscopy (SEM), has shown that the exposed area of dentine resulting from occlusal wear and the upper part of a vertical crack affecting enamel and dentin tissues were filled with beeswax shortly before or after the individual’s death. If the filling was done when the person was still alive, the intervention was likely aimed to relieve tooth sensitivity derived from either exposed dentine and/or the pain resulting from chewing on a cracked tooth: this would provide the earliest known direct evidence of therapeutic-palliative dental filling.

Concepts: Electron, Spectroscopy, Mass spectrometry, Teeth, Dental caries, Scanning electron microscope, Tooth enamel, Dentin

250

Dentin-pulp complex regeneration is a promising alternative treatment for the irreversible pulpitis caused by tooth trauma or dental caries. This process mainly relies on the recruitment of endogenous or the transplanted dental pulp stem cells (DPSCs) to guide dentin-pulp tissue formation. Platelet-derived growth factor (PDGF), a well-known potent mitogenic, angiogenic, and chemoattractive agent, has been widely used in tissue regeneration. However, the mechanisms underlying the therapeutic effects of PDGF on dentin-pulp complex regeneration are still unclear. In this study, we tested the effect of PDGF-BB on dentin-pulp tissue regeneration by establishing PDGF-BB gene-modified human dental pulp stem cells (hDPSCs) using a lentivirus. Our results showed that PDGF-BB can significantly enhance hDPSC proliferation and odontoblastic differentiation. Furthermore, PDGF-BB and vascular endothelial growth factor (VEGF) secreted by hDPSCs enhanced angiogenesis. The chemoattractive effect of PDGF-BB on hDPSCs was also confirmed using a Transwell chemotactic migration model. We further determined that PDGF-BB facilitates hDPSCs migration via the activation of the phosphatidylinositol 3 kinase (PI3K)/Akt signaling pathway. In vivo, CM-DiI-labeled hDPSCs were injected subcutaneously into mice, and our results showed that more labeled cells were recruited to the sites implanted with calcium phosphate cement scaffolds containing PDGF-BB gene-modified hDPSCs. Finally, the tissue-engineered complexes were implanted subcutaneously in mice for 12 weeks, the Lenti-PDGF group generated more dentin-like mineralized tissue which showed positive staining for the DSPP protein, similar to tooth dentin tissue, and was surrounded by highly vascularized dental pulp-like connective tissue. Taken together, our data demonstrated that the PDGF-BB possesses a powerful function in prompting stem cell-based dentin-pulp tissue regeneration. Stem Cells Translational Medicine 2017.

Concepts: Angiogenesis, Growth factor, Vascular endothelial growth factor, Cellular differentiation, Teeth, Dental caries, Dentin, Platelet-derived growth factor

186

The requirement for immediate vascularization of engineered dental pulp poses a major hurdle towards successful implementation of pulp regeneration as an effective therapeutic strategy for root canal therapy, especially in adult teeth. Here, we demonstrate a novel strategy to engineer pre-vascularized, cell-laden hydrogel pulp-like tissue constructs in full-length root canals for dental pulp regeneration. We utilized gelatin methacryloyl (GelMA) hydrogels with tunable physical and mechanical properties to determine the microenvironmental conditions (microstructure, degradation, swelling and elastic modulus) that enhanced viability, spreading and proliferation of encapsulated odontoblast-like cells (OD21), and the formation of endothelial monolayers by endothelial colony forming cells (ECFCs). GelMA hydrogels with higher polymer concentration (15% w/v) and stiffness enhanced OD21 cell viability, spreading and proliferation, as well as endothelial cell spreading and monolayer formation. We then fabricated pre-vascularized, full-length, dental pulp-like tissue constructs by dispensing OD21 cell-laden GelMA hydrogel prepolymer in root canals of extracted teeth and fabricating 500 µm channels throughout the root canals. ECFCs seeded into the microchannels successfully formed monolayers and underwent angiogenic sprouting within 7 days in culture. In summary, the proposed approach is a simple and effective strategy for engineering of pre-vascularized dental pulp constructs offering potentially beneficial translational outcomes.

Concepts: Inflammation, Angiogenesis, Engineering, Teeth, Root canal, Endodontic therapy, Young's modulus, Dentin

169

BACKGROUND: Recent studies reported on the very complex morphology of the pulp system in equine cheek teeth. The continuous production of secondary dentine leads to distinct age-related changes of the endodontic cavity. Detailed anatomical knowledge of the dental cavities in all ages is required to explain the aetiopathology of typical equine endodontic diseases. Furthermore, data on mandibular and maxillary pulp systems is in high demand to provide a basis for the development of endodontic therapies. However, until now examination of the pulp cavity has been based on either sectioned teeth or clinical computed tomography. More precise results were expected by using micro-computed tomography with a resolution of about 0.1 mm and three-dimensional reconstructions based on previous greyscale analyses and histological verification. The aim of the present study was to describe the physiological configurations of the pulp system within a wide spectrum of tooth ages. RESULTS: Maxillary teeth: All morphological constituents of the endodontic cavity were present in teeth between 4 and 16 years: Triadan 06s displayed six pulp horns and five root canals, Triadan 07-10s five pulp horns and four root canals and Triadan 11s seven pulp horns and four to six root canals. A common pulp chamber was most frequent in teeth <=5 years, but was found even in a tooth of 9 years. A large variety of pulp configurations was observed within 2.5 and 16 years post eruption, but most commonly a separation into mesial and distal pulp compartments was seen. Maxillary cheek teeth showed up to four separate pulp compartments but the frequency of two, three and four pulp compartments was not related to tooth age (P > 0.05). In Triadan 06s, pulp horn 6 was always connected to pulp horns 1 and 3 and root canal I. In Triadan 11s, pulp horns 7 and 8 were present in variable constitutions. Mandibular teeth: A common pulp chamber was present in teeth up to 15 years, but most commonly seen in teeth <=5 years. A segmented pulp system was found in 72% of the investigated teeth. Segmentation into separate mesial and distal pulp compartments was most commonly present. Pulp horn 4 coalesced either with the mesial pulp horns 1 and 3 or with the distal pulp horns 2 and 5. CONCLUSIONS: Details of the pulpar anatomy of equine cheek teeth are provided, supporting the continuous advancement in endodontic therapy. Numerous individual configurations of the pulp system were obtained in maxillary cheek teeth, but much less variability was seen in mandibular cheek teeth.

Concepts: Teeth, Tooth, Dental caries, Root canal, Endodontic therapy, Endodontics, Dentin, Animal anatomy

168

In order to endow environmental protection features to dentifrice, hydroxyapatite (HA) was added to ordinary dentifrice. The effects on dentinal tubule occlusion and surface mineralization were compared after brushing dentine discs with dentifrice with or without HA. The two types of dentifrice were then added to 100 µg/ml of hexavalent chromium cation (Cr(6+)) solution in order to evaluate their capacities of adsorbing Cr(6+) from water. Our results showed that the dentifrice containing HA was significantly better than the ordinary dentifrice in occluding the dentinal tubules with a plugging rate greater than 90%. Moreover, the effect of the HA dentifrice was persistent and energy-dispersive spectrometer (EDS) revealed that the atomic percentages of calcium and phosphorus on the surface of dentine discs increased significantly. Adding HA to ordinary dentifrice significantly enhanced the ability of dentifrice to adsorb Cr(6+) from water with the removal rate up to 52.36%. In addition, the sorption was stable. Our study suggests that HA can be added to ordinary dentifrice to obtain dentifrice that has both relieving dentin hypersensitivity benefits and also helps to control environmental pollution.

Concepts: Chemistry, Atom, Adsorption, Pollution, Dentin, Hydroxylapatite, Dentine hypersensitivity, Environmentalism

143

Endodontic sealer residues remaining within the pulp chamber dentin after root canal obturation and cleaning with various solvents may compromise the appearance and the durability of dental restorations. Acid etching is routinely performed prior to application of dentine adhesive systems, but is effect on residual sealer material and the optimal time-point for performing etching, are unknown. Here, we evaluated the effect of acid etching on the dentin surface when performed either immediately or 7 days after removal of the endodontic sealer with two solvents, i.e., 95% ethanol or xylol. Forty crowns fragments from bovine incisors were impregnated with sealer and divided into 4 groups (n = 10 each), according to the dentin cleaning protocol and to the acid etching time-point: G1, 95.0% ethanol and immediate acid etching; G2, xylol and immediate acid etching; G3, 95.0% ethanol and acid etching after 7 days; and G4, xylol and acid etching after 7 days. Scanning electron microscopy (SEM) images (2000 ×) were obtained from each specimen and the number of open dentinal tubules counted and compared. Another 40 fragments were similarly prepared, and SEM images were obtained (500 ×) to score and compare the persistence of sealer residues on the dentin. G4 showed the most open dentinal tubules and the least epoxy resin-based sealer residues on the dentin surface (p < 0.05). The least epoxy resin-based sealer residues was obtained when acid etching, using 37% phosphoric acid, was performed after 7 days after cleaning the dentin with xylol.

Concepts: Acid, Ethanol, Phosphoric acid, Teeth, Dental caries, Scanning electron microscope, Dentin, Pulp

28

Caries lesions in dental hard tissues autofluoresce when exposed to light of certain wavelengths, whereas sound tissues do not, and this can be used as an in vitro histological marker for dental caries. Detection of autofluorescence is the basis of KaVo DIAGNOdent™ technology, and provides objective feedback control of laser-stimulated ablation of dental caries for the KaVo Key Laser 3™. This Er:YAG laser operates at 2940nm wavelength, and is effective at removal of infected dental hard tissues. Micro-computed tomography (micro-CT) allows the non-invasive investigation of three-dimensional structures and analysis of mineral density profiles of dentine following laser ablation.

Concepts: Fluorescence, Light, Control theory, Wavelength, Feedback, Dental caries, Dentin, Control system

28

Background: Dentin hypersensitivity (DH) is a painful, exaggerated response to normal stimuli, such as cold, sweetness, and brushing. The aim of the present controlled, randomized, double-masked, non-inferiority clinical trial is to evaluate the effectiveness of cyanoacrylate in the treatment of DH when compared to the application of low-intensity laser. Methods: The study includes 434 sensitive teeth from 62 patients. A total of 216 teeth were treated with laser and 218 with cyanoacrylate. A numeric rating scale was used to record the parameters of pain related to the stimuli at baseline and after the treatment at intervals of 24 hours and 30, 90, and 180 days. Results: Both groups had significant reductions in DH. However, there was no significant difference between the two groups ≤6 months. Intragroup analysis showed that the effect of cyanoacrylate obtained at 24 hours remained for 90 days in response to air-jet test and 30 days for cold-spray test. There was a statistically significant difference between all other intragroup comparisons at the time intervals (P <0.001). Conclusions: It was concluded that cyanoacrylate is as effective as low-intensity laser in reducing DH. In addition, it is a more accessible and low-cost procedure and can be safely used in the treatment of DH.

Concepts: Clinical trial, Statistics, Effect, Statistical significance, Effectiveness, Harshad number, Dentin, Dentine hypersensitivity

28

Dentinal hypersensitivity is one of the oldest recorded complaints of discomfort to mankind and yet there appears to be no permanent treatment for this clinical condition. This study was designed to evaluate the clinical efficacy of potassium binoxalate gel and neodymium:yttrium-aluminum-garnet (Nd:YAG) laser on dentin hypersensitivity for a period of 9 months. Eighty teeth (20 subjects, 25-55 years old, M = F) were evaluated in a split-mouth design to receive potassium binoxalate (group A, 40 teeth) and Nd:YAG (group B, 40 teeth: 1 W, 10 Hz, and 60 s, irradiated twice). The diameter of output beam was about 300 μm with a distance of 2 mm between laser fiber or tip and tooth surface. The clinical efficacy was evaluated by air-blast test and cold-water test using visual analog scale. Electron microscopy photomicrographs were taken to confirm the results. Analysis was done at baseline; immediately post-treatment; and at 3, 6, 9 months post-treatment. Student’s paired and unpaired T tests were used to evaluate the statistical analysis. Both treatment modalities were effective in reducing dentine hypersensitivity. However, Nd:YAG laser was better when intragroup comparison was made at 9 months post-treatment. Nd:YAG lasers is better in long-term treatment (up to 9 months) owing to the melting of dentinal tubules. However, due to depth of penetration of microcrystals, gel was better when ease of the procedure is considered. Nevertheless, both treatment modalities resulted in recurrence. Hence, further studies are needed to discover an agent, which can be considered as a “gold standard”.

Concepts: Clinical trial, Statistics, Effectiveness, Gold, Teeth, Dentin, Dentine hypersensitivity, Potassium nitrate

28

A 3 day randomised clinical study investigating the efficacy of two toothpastes, designed to occlude dentine tubules, for the treatment of dentine hypersensitivity. OBJECTIVES: A product comparison study to compare the short term clinical efficacy of a strontium acetate/silica toothpaste with an arginine/calcium carbonate paste for pain reduction in dentine hypersensitivity. METHODS: The study was examiner blind of two arm parallel design. Eighty healthy adult subjects from general dental practice with ≥ 2 sensitive teeth but otherwise good oral health, were enrolled and randomised to 1 of 2 toothpaste treatments, schedule provided by the sponsor. Almost equal numbers received each treatment. Tooth sensitivity was measured in three ways; evaporative (Schiff score; Visual Analogue Scale) and tactile stimuli (Yeaple probe), prior to and immediately after subjects' self application of a single pea sized dose of toothpaste, and following subsequent twice daily brushing for three days with the paste. RESULTS: All 80 subjects completed the study. Results confirm that for both treatments, pain was reduced immediately and relief was sustained after 3 days use. For all 3 measures, benefit was similar between the two pastes, with no statistical or clinical difference demonstrated, apart from response to evaporative stimulus at 3 days, where Schiff scores were significantly lower in the arginine group, p=0.02. CONCLUSIONS: It can be concluded that both desensitising, occluding toothpastes provided reduction of pain from dentine hypersensitivity on a short term basis: toothpastes appearing to be clinically similarly effective both after a single subject dab on application and post twice daily brushing for three days. National Research Ethics Service register number 09/H020/57:

Concepts: Clinical trial, Effectiveness, Teeth, Dentin, Dentistry, Dentine hypersensitivity, Potassium nitrate, Oral pathology