LPS-induced autophagy in human dental pulp cells is associated with p38.

Lipopolysaccharides (LPS), which are components of the cell wall of Gram-negative bacteria, are among the important factors that induce inflammation, including pulpitis. Autophagy in human dental pulp cells (hDPCs) acts as a protective mechanism that promotes cell survival under adverse conditions through different signaling pathways. In this study, we examined whether LPS increases autophagy in hDPCs and investigated the role of mitogen-activated protein kinases signaling and nuclear factor κB (NF-κB) in this process. We found that stimulation of hDPCs with 0.1 µg/mL LPS increased the protein and mRNA levels of autophagy markers, beclin1 and microtubule associated protein light chain 3II (LC3II). In addition, acridine orange staining and transmission electron microscopy demonstrated the induction of autophagy upon the treatment of LPS. Furthermore, LPS affected phosphorylation of p38, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK), and the nuclear translocation of NF-κB. While p38 inhibitor suppressed the LPS-induced increase in protein levels of beclin1 and LC3-II. Our results suggest that LPS induced autophagy in hDPCs and affected the phosphorylation of p38, ERK, and JNK, as well as the nuclear translocation of NF-κB. Phosphorylation of p38 may be involved in LPS-induced autophagy in hDPCs.

Characterization of Living Dental Pulp Cells in Direct Contact with Mineral Trioxide Aggregate.

Mineral trioxide aggregate (MTA) was introduced as a material for dental endodontic regenerative therapy. Here, we show the dynamics of living dental pulp cells in direct contact with an MTA disk. A red fluorescence protein (DsRed) was introduced into immortalized porcine dental pulp cells (PPU7) and cloned. DsRed-PPU7 cells were cultured on the MTA disk and cell proliferation, chemotaxis, the effects of growth factors and the gene expression of cells were investigated at the biological, histomorphological and genetic cell levels. Mineralized precipitates formed in the DsRed-PPU7 cells were characterized with crystal structural analysis. DsRed-PPU7 cells proliferated in the central part of the MTA disk until Day 6 and displayed a tendency to move to the outer circumference. Both transforming growth factor beta and bone morphogenetic protein promoted the proliferation and movement of DsRed-PPU7 cells and also enhanced the expression levels of odontoblastic gene differentiation markers. Mineralized precipitates formed in DsRed-PPU7 were composed of calcium and phosphate but its crystals were different in each position. Our investigation showed that DsRed-PPU7 cells in direct contact with the MTA disk could differentiate into odontoblasts by controlling cell-cell and cell-substrate interactions depending on cell adhesion and the surrounding environment of the MTA.

Bond strength and adaptation of pulp capping materials to dentin.

This study evaluated the shear bond strength (SBS) and internal marginal adaptation of pulp-capping materials to dentin. Flat occlusal deep dentin surfaces were produced and randomly assigned to two groups (sound or artificial caries-affected dentin). The specimens in each group were assigned to one of seven subgroups according to the materials used: Biodentine, Theracal LC, Ultra-Blend plus, Calcimol LC, ApaCal ART, EQUIA Forte, and Ionoseal. Buildups (3-mm inner diameter and 2-mm deep) were made over the dentin surfaces. The bonded specimens were tested under shear forces at a crosshead speed of 0.8 mm/min and fracture modes were determined using a stereomicroscope at 25× magnification. The materials were applied to the pulp floor of prepared Class I cavities and then the cavities were restored with composite resin. Restored molar teeth were subjected to 5,000 thermocycles and sectioned in a bucco-lingual direction. Resin replicas were made to determine the adaptation at the pulp floor with scanning electron microscopy. Significant differences were determined among both bond strengths and gap formations of the materials. EQUIA Forte applied to both dentin substrates had a significantly higher SBS than the other materials. The bond strength of each material was not influenced by the dentin condition. Biodentine (3.03%), EQUIA Forte (7.83%), and Theracal LC (13.37%) had lower gap formations compared to other materials but were not significantly different from each other.

Isolamento e diferenciação das células-tronco da polpa dentária canina em células progenitoras neurais / Isolation and differentiation of canine dental pulp stem cells in neural progenitor cells

O objetivo deste estudo foi verificar a capacidade de diferenciação das células-tronco da polpa dentária canina em células progenitoras neurais bem como quantificar obtenção e viabilidade celular, durante três passagens em cultura. As células foram extraídas da polpa dentária de dois cadáveres caninos, com aproximadamente dez meses de idade, que foram a óbito em decorrência de traumatismo automotivo. Após três subculturas, realizou-se avaliação da viabilidade celular por quantificação em câmara de Neubauer. A partir disso, induziu-se diferenciação neural em meio de cultura neurobasal (Gibco™), com células aderidas ao plástico ou suspensas em placas tratadas com agarose. Após sete e 14 dias em cultivo indutor, observou-se morfologia e perfil imunofenotípico utilizando citometria de fluxo e imunocitoquímica fluorescente. Aos 14 dias as células apresentaram alto grau de expressão para marcadores anti-nestina e anti-glial fibrillary acidic protein (anti-GFAP). Anteriormente, obteve-se ao 25º dia, média de 18x106 células viáveis indiferenciadas oriundas do tecido pulpar. Sugere-se que as células-tronco indiferenciadas da polpa dentária canina apresentem índices satisfatórios de diferenciação em células progenitoras neurais, aderidas ou suspensas em cultura. A polpa dentária dos dentes decíduos caninos, fornece células indiferenciadas viáveis em quantidade adequada.(AU)

The objective of this study was to verify the differentiation capacity of canine tooth pulp stem cells in neural progenitor cells as well as to quantify the attainment and viability during three culture passages. The cells were extracted from the dental pulp of two canine cadavers, with approximately ten months of age, which died due to automotive trauma. After three subcultures, cell viability evaluation was performed by Neubauer chamber quantification. Neural differentiation was induced in neurobasal culture medium (Gibco ™), with cells adhered to the plastic or suspended in agarose-treated plates. After seven and 14 days in inducer culture, morphology and immunophenotypic profile were observed using flow cytometry and fluorescent immunocytochemistry. At 14 days the cells had a high degree of expression for anti-nestin and anti-glial fibrillary acidic (anti-GFAP) markers. Previously, an average of 18x106 undifferentiated viable cells from the pulp tissue were obtained on the 25th day. It is suggested that the undifferentiated canine pulp stem cells present satisfactory differentiation indices in neural progenitor cells, adhered or suspended in culture. The dental pulp of deciduous canine teeth provides viable undifferentiated cells in adequate quantity.(AU)

Artificial Caries Resistance in Enamel after Topical Fluoride Treatment and 445 nm Laser Irradiation.

OBJECTIVE: This in vitro study is aimed at investigating the caries preventive effectiveness of 445 nm diode laser in combination with topical fluoridation. MATERIALS AND METHODS: A total of 30 caries-free bovine teeth were used in this study. Eighteen teeth were covered with nail varnish except four windows on the labial surface. The windows were assigned to no treatment/control (C), laser (L) (0.3 W, 60 s, and 90 J/cm2), fluoride (F), and fluoride followed by laser (FL) treatment groups. Artificial caries lesions were created, and the teeth were sectioned and investigated under polarized light microscopy for quantitative measurement of the resulted lesion depth. Ten teeth were used for surface temperature measurement and two teeth for scanning electron microscopy (SEM). Extra twelve human molars were used for the intrapulpal temperature measurement. The absorbance of fluoride at 445 nm was measured. RESULTS: The means of lesion depth for the C, L, F, and FL groups were 123.48 (±21.93), 112.33 (±20.42), 99.58 (±30.68), and 89.03 (±30.38) µm, respectively. The pairwise differences of the L, F, and FL groups compared with the C group were significant (p < 0.05). The differences between groups were tested: FL versus L p=0.02, F versus L p=0.16, and FL versus F p=0.91, and the difference of the F versus FL was not significant (p=0.91). Temperature increment at the enamel surface and pulp roof were ∆T = 16.67 (±4.11) and 2.12 (±0.66)°C, respectively. The topical fluoride absorbance at 445 nm is five orders higher than that at 810 nm. SEM shows that after laser irradiation the enamel surface was intact and without thermal damage. CONCLUSIONS: The 445 nm laser irradiation may be useful for caries prevention, and its effectiveness is lower than those previously achieved using the argon ion laser.

The effect of hyaluronic acid hydrogels on dental pulp stem cells behavior.

Dental caries and trauma, particularly in childhood, are among the most prevalent teeth problems, which result in the creation of cavities and probably tooth loss. Thus, novel regenerative approaches with high efficiency and less toxicity are required. Stem cell therapy along with the implementation of scaffolds has provided excellent opportunities in the regeneration of teeth structure. Hyaluronic acid (HA) hydrogels have enticed great attention in the field of regenerative medicine. The unique chemical and structural properties of HA and its derivatives have enabled their application in tissue engineering. Several factors such as the location and type of the lesion, teeth age, the type of capping materials determine the success rate of pulp therapy. HA hydrogels have been considered as biocompatible and safe scaffold supports in human dental cell therapies.

Protection against HEMA-Induced Mitochondrial Injury In Vitro by Nrf2 Activation.

Dental resin monomers such as 2-hydroxyethyl methacrylate (HEMA) disturb vital cell functions and induce mitochondrial intrinsic apoptosis via generation of oxidative stress. Nuclear factor erythroid 2-related factor 2 (Nrf2) regulates the gene expression of antioxidative enzymes and plays a crucial role in the maintenance of cellular redox equilibrium and mitochondrial homeostasis. The present study investigated the functional significance of Nrf2 in cellular response toward HEMA. It was found that HEMA stimulation promoted nuclear translocation of Nrf2 and increased Nrf2 and heme oxygenase-1 (HO-1) expression, which was further enhanced by Nrf2 activator tert-butylhydroquinone (tBHQ), but suppressed by Nrf2 inhibitor ML385. Pretreatment of primary human dental pulp cells (hDPCs) with tBHQ protected the cells from HEMA-induced oxidative injury (increased reactive oxygen species production and apoptosis) and mitochondrial impairment (morphological alterations, decreased ATP production, suppressed oxidative phosphorylation activity, depolarization of mitochondrial membrane potential, and disrupted electron transport chain). In contrast, pretreatment with ML385 increased cell sensitivity to these injurious processes. This protective effect on mitochondria could be related to peroxisome proliferator-activated receptor γ coactivator 1α (PGC1α)/nuclear respiratory factor 1 (NRF1) pathway. These results contribute to the understanding of the function of Nrf2 and the development of novel therapies to counteract the adverse effects of dental resin monomers.

Dental properties, ultrastructure, and pulp cells associated with a novel DSPP mutation.

OBJECTIVE: To investigate physical characteristics and behaviours of dental pulp cells of teeth isolated from a dentinogenesis imperfecta (DGI) patient with a novel dentin sialophosphoprotein (DSPP) mutation. SUBJECTS AND METHODS: Whole exome and Sanger sequencing were employed to identify mutations. Physical characteristics of the teeth were examined. Pulp cells' behaviours including cell proliferation, colony-forming unit, osteogenic differentiation, pluripotent markers, and mesenchymal stem cell markers were investigated. RESULTS: The proband had opalescent brown primary teeth with extensive loss of enamel. Mutation analysis revealed a novel heterozygous 4-bp deletion, c.1915_1918delAAGT (p.K639QfsX674), in exon 5 of the DSPP associated with DGI. Analysis of the extracted primary incisor demonstrated a decrease in brightness but an increase in yellow and red chroma. The dentin showed reduced mineral density. The dentinal tubules were present in the predentin, but progressively collapsed in the dentin. The pulp cells exhibited markedly reduced CD105 expression, decreased cell proliferation, and smaller colony-forming units. CONCLUSIONS: We identified a novel mutation in the DSPP gene which disturbed dentin characteristics and pulp cells' behaviours. Our study expands the mutation spectrum and understanding of pathologic dentin phenotypes related to the frameshift deletion in the dentin phosphoprotein (DPP) region of the DSPP gene.

Expression and distribution of three transient receptor potential vanilloid(TRPV) channel proteins in human odontoblast-like cells.

Odontoblasts have been suggested to contribute to nociceptive sensation in the tooth via expression of the transient receptor potential (TRP) channels. The TRP channels as a family of nonselective cation permeable channels play an important role in sensory transduction of human. In this study, we examined the expression of transient receptor potential vanilloid-1 (TRPV1), transient receptor potential vanilloid-2 (TRPV2) and transient receptor potential vanilloid-3 (TRPV3) channels in native human odontoblasts (HODs) and long-term cultured human dental pulp cells with odontoblast phenotyoe (LHOPs) obtained from healthy wisdom teeth with the use of immunohistochemistry (IHC), immunofluorescence (IF), quantitative real-time polymerase chain reaction (qRT-PCR),western blotting (WB) and immunoelectron microscopy (IEM) assay. LHOPs samples were made into ultrathin sections, mounted on nickel grids, floated of three TRPV antibodies conjugated with 10 nm colloidal gold particles and observed under IEM at 60,000 magnifications. The relative intracellular distributions of these three channels were analyzed quantitatively on IEM images using a robust sampling, stereological estimation and statistical evaluation method. The results of IHC and IF convinced that TRPV1, TRPV2 and TRPV3 channels were expressed in native HODs and (LHOPs). The result of qRT-PCR and WB confirmed that the gene and protein expression of TRPV1, TRPV2, and TRPV3 channels and TRPV1 mRNA are more abundantly expressed than TRPV2 and TRPV3 in HODs (P < 0.05). Quantitative analysis of IEM images showed that the relative intracellular distributions of these three channels are similar, and TRPV1, TRPV2 and TRPV3 proteins were preferential labeled in human odontoblast processes, mitochondria, and endoplasmic reticulum. Thus, HODs could play an important role in mediating pulp thermo-sensation due to the expression of these three TRPV channels. The difference of relative intracellular distributions of three channels suggests that special structures such as processes may have an important role to sensing of the outer stimuli first.

Cell-derived micro-environment helps dental pulp stem cells promote dental pulp regeneration.

OBJECTIVES: The function of the dental pulp is closely connected to the extracellular matrix (ECM) structure, and ECM has received significant attention due to its biological functions for regulating cells. As such, the interaction between the ECM niche and cells is worth exploring for potential clinical uses. MATERIALS AND METHODS: In this study, dental pulp stem cell (DPSC)-derived ECM (DPM) was prepared through cell culture and decellularization to function as the cell niche, and changes in DPSC behaviour and histological analysis of dental pulp tissue regeneration were evaluated following the DPM culture. DPM promoted the replication of DPSCs and exhibited retention of their mineralization. Then, the DPM-based culture strategy under odontogenic culture medium was further investigated, and the mineralization-related markers showed that DPSCs were regulated towards odontogenic differentiation. Dental pulp-like tissue with well-arranged ECM was harvested after a 2-month subcutaneous implantation in nude mice with DPM application. Additionally, DPSCs cultured on the plastic culture surface showed the up-regulation of mineralization makers in vitro, but there was a disorder in matrix formation and mineralization when the cells were cultured in vivo. RESULTS AND CONCLUSIONS: DPM-based cultivation could serve as a cell niche and modulate DPSC behaviour, and this method also provided an alternative to harvest tissue-specific ECM and provided a strategy for ECM-cell interaction.

Response of Dental Pulp Stem Cells to Synthetic, Allograft, and Xenograft Bone Scaffolds.

Different degrees of clinical success have been reported for synthetic, allograft, and xenograft bone substitutes in human trials. Although these substitutes have been clinically investigated, their in vitro effects on cell differentiation remain unclear. Proliferation, differentiation, and attachment of dental pulp stem cells (DPSCs) to ß-tricalcium phosphate (ß-TCP), freeze-dried bone allograft (FDBA), and deproteinized bovine bone mineral (DBBM) were compared in this study. MTT assay, measurement of total DNA, and reverse transcriptase polymerase chain reaction were performed. ß-TCP had the highest potential for DPSC attachment and proliferation, while FDBA induced osteoblastic differentiation of DPSCs. Further in vivo investigations are necessary to select a clinically appropriate scaffold.

Effect of silver diamine fluoride (SDF) on the dentin-pulp complex: ex vivo histological analysis on human primary teeth and rat molars / Efecto del diamino fluoruro de plata (DFP) sobre complejo dentino-pulpar. Análisis histológico ex vivo en dientes primarios humanosy molares de rata

The aim of this study was to determine the effect of SDF on thedentin­pulp complex using two models: teeth after SDFapplication (ex vivo) and experimental animal molars. Adescriptive study was performed using two models. In the firstmodel, primary teeth (ex vivo) with enamel­dentin caries, withoutpulp involvement and previously treated with 38% SDF, wereevaluated by means of two techniques: (a) Scanning ElectronMicroscopy (SEM) and energy­dispersive X­ray detector (EDS)to determine qualitative and quantitative composition, and (b)brightfield optical microscopy (OM) after decalcification. Thesecond model used laboratory animal molars from 12 maleWistar rats. Standardized enamel­dentin cavities approximately0.5 mm deep were made the distal fossa of the occlusal face ofboth first lower molars, to one of which a 38% SDF solution wasapplied, while the other was used as a control. Histologicalsections were prepared and dental pulp was evaluatedqualitatively in both groups. SEM on ex vivo teeth showed areasof hypermineralization in the intertubular dentin and few blockedtubules, while EDS detected Ag in the center of the lesion(7.34%), its concentration declining at the edges (1.71%), withnone in the areas farthest from the lesion...

El objetivo del trabajo fue determinar del efecto del DFP encomplejo dentino­pulpar aplicando dos modelos: piezasdentarias luego de su aplicación (ex vivo) y en molares deanimales experimentales. Se realizó un estudio descriptivoaplicando dos modelos: en piezas dentarias primarias (ex vivo)con caries amelodentinarias sin compromiso pulpar que hayansido sometidas previamente con DFP 38%, mediante dosevaluaciones: Microscopía electrónica de Barrido (MEB) ydetector de energía dispersiva de rayos X (EDS) a fin dedeterminar su composición cuali y cuantitativa y Microscopíaóptica de campo claro (MCC) mediante la técnica descalcifi ­cación y en molares de animales de laboratorio donde seutilizaron 12 ratas Wistar macho. La técnica fue estandarizadaen la fosa distal de la cara oclusal del primer molar inferior, serealizó una cavidad amelodentinaria aprox. 0.5 mm de profun ­didad, en ambos molares. En un molar se aplicó la soluciónDFP al 38 % y el opuesto como control. Se realizaron corteshistológicos y se evaluó en forma cualitativa la pulpa dentalen ambos grupos. En las piezas ex vivas mediante MEBse observaron áreas de hipermineralización en la dentinaintertubular y escasos conductillos obliterados y por EDS sedetectó Ag en el centro de la lesión (7.34%), disminuyendo suconcentración en los límites (1,71%) y no se detectó en las zonasmás alejadas de la misma. En MCC se observó DFP sellando losconductillos sólo en sitio de colocación y con una penetraciónlimitada, por debajo, los conductillos se observaron de aspectonormal y el tejido pulpar asociado con la caries tratadaha mostrado un infiltrado inflamatorio crónico y formaciónde dentina terciaria, sin observarse precipitado de Ag...

Understanding External Cervical Resorption in Vital Teeth.

INTRODUCTION: The aim of this study was to investigate the 3-dimensional (3D) structure and the cellular and tissue characteristics of external cervical resorption (ECR) in vital teeth and to understand the phenomenon of ECR by combining histomorphological and radiographic findings. METHODS: Twenty-seven cases of vital permanent teeth displaying ECR were investigated. ECR diagnosis was based on clinical and radiographic examination with cone-beam computed tomographic imaging. The extracted teeth were further analyzed by using nanofocus computed tomographic imaging, hard tissue histology, and scanning electron microscopy. RESULTS: All examined teeth showed some common characteristics. Based on the clinical and experimental findings, a 3-stage mechanism of ECR was proposed. At the first stage (ie, the initiation stage), ECR was initiated at the cementum below the gingival epithelial attachment. At the second stage (ie, the resorption stage), the resorption invaded the tooth structure 3-dimensionally toward the pulp space. However, it did not penetrate the pulp space because of the presence of a pericanalar resorption-resistant sheet. This layer was observed to consist of predentin, dentin, and occasionally reparative mineralized (bonelike) tissue, having a fluctuating thickness averaging 210 µm. At the last advanced stage (ie, the repair stage), repair took place by an ingrowth and apposition of bonelike tissue into the resorption cavity. During the reparative stage, repair and remodeling phenomena evolve simultaneously, whereas both resorption and reparative stages progress in parallel at different areas of the tooth. CONCLUSIONS: ECR is a dynamic and complex condition that involves periodontal and endodontic tissues. Using clinical, histologic, radiographic, and scanning microscopic analysis, a better understanding of the evolution of ECR is possible. Based on the experimental findings, a 3-stage mechanism for the initiation and growth of ECR is proposed.

Effects of sildenafil on dental pulp: immunohistochemical and ultrastructural evaluation / Efectos de sildenafil en la pulpa dental: evaluación inmunohistoquímica y ultraestructural

Sildenafil is a strong peripheral vasodilator and is used to treat cardiovascular and neurosurgery. The purpose of this study was to investigate the immunohistochemical and ultrastructural effects of sildenafil on dental pulp of rats. The study was performed with adult female Wistar-Albino rats. Control group (n= 7) were fed on standard laboratory diet until surgery. The study group (n= 7) were administered sildenafil orally with orogastric tube 10 mg·kg-1 once a day for 30 days. Each rat was anesthetized and incisor teeth were removed. This study examined the immunohistochemical and ultrastructural effects of sildenafil on the dental pulp in rats. The relaxation from the vessel, endothelial cell hyperplasia, moderate degeneration of collagen fibers were observed to cause degenerative changes in odontoblast with sildenafil. In the pulp tissue long-term use sildenafil is thought to cause degeneration and new vessel formation.

El sildenafil es un vasodilatador periférico importante y se utiliza para tratar enfermedades cardiovasculares y en neurocirugía. El propósito de este estudio fue investigar los efectos inmunohistoquímicos y ultraestructurales del sildenafil sobre la pulpa dental de ratas. El estudio se realizó con ratas Wistar albinas, hembras adultas. El grupo de control (n= 7) fue alimentado con una dieta estándar de laboratorio hasta que se realizó la cirugía. El grupo de estudio (n= 7) fue tratado con sildenafil por vía oral y sonda orogástrica 10 mg·kg-1 una vez al día durante 30 días. Cada rata fue anestesiada y se extrajeron los dientes incisivos. Se examinaron los efectos inmunohistoquímicos y ultraestructurales del sildenafil sobre la pulpa dentaria. Con la administración de sildenafil se observó la relajación de los vasos, la hiperplasia de las células endoteliales y una degeneración moderada de fibras colágenas causando cambios degenerativos en los odontoblastos. En el tejido pulpar, el uso de sildenafil a largo plazo puede causar la degeneración y neoformación de vasos.

In Vitro Evaluation of Microleakage and Microhardness of Ethanolic Extracts of Propolis in Different Proportions Added to Glass Ionomer Cement.

OBJECTIVE: To evaluate the effect of ethanolic extracts of propolis (EEP) addition in different proportions to glass ionomer cement (GIC) on microleakage and microhardness of GIC. STUDY DESIGN: The cement was divided into four groups: one using the original composition and three with 10%, 25%, and 50% EEP added to the liquid and then manipulated. For microleakage assessment, sixty primary molars were randomly divided into four groups (n=15). Standard Class II cavities were prepared and then filled with EEP in different proportions added to GICs. Microleakage test was performed using a dye penetration method. The data were analyzed using one-way ANOVA and Mann-Whitney U tests (α = 0.05). Disc shaped specimens were prepared from the tested GIC to determine Vickers hardness (VHN). The data were analyzed using one-way ANOVA and post hoc Tukey test (α = 0.05). RESULTS: There were no statistically significant differences between the groups in terms of microleakage (p > 0.05). There were statistically significant differences between the VHN values of groups (p < 0.05). Increasing addition of EEP to GIC statistically significantly increased VHN value of GIC (p < 0.05). CONCLUSIONS: The addition of EEP to GIC increased the microhardness of the GIC and did not adversely affect the microleakage. Thus, it might be used during routine dental practice due to its antibacterial properties.

Morphological aspects and physical properties of enamel and dentine of Sus domesticus: A tooth model in laboratory research.

This study aims to describe and analyze morphological and physical properties of deciduous teeth of Sus domesticus. Ultrastructural analysis, mineral composition and microhardness of enamel and dentine tissues were performed on 10 skulls of S. domesticus. External anatomic characteristics and the internal anatomy of the teeth were also described. Data regarding microhardness and ultrastructural analysis were subjected to statistical tests. For ultrastructural analysis, we used the analysis of variance (ANOVA) with Tukey's post hoc (p≤0.05) test. In the analysis of microhardness, the difference between the enamel and dentine tissues was analyzed by a Student's t test. Values were expressed as mean with standard error. The results of ultrastructural analysis showed the presence of an enamel prism pattern. A dentinal tubule pattern was also observed, with a larger diameter in the pulp chamber and the cervical third, in comparison to middle and apical thirds. We observed an average microhardness of 259.2kgf/mm(2) for enamel and 55.17kgf/mm(2) for dentine. In porcine enamel and dentine, the chemical elements Ca and P showed the highest concentration. The analysis of internal anatomy revealed the presence of a simple root canal system and the occurrence of main canals in the roots. The observed features are compatible with the functional demand of these animals, following a pattern very similar to that seen in other groups of mammals, which can encourage the development of research using dental elements from the pig as a substitute for human teeth in laboratory research.

Three-dimensional Imaging Reveals New Compartments and Structural Adaptations in Odontoblasts.

In organized tissues, the precise geometry and the overall shape are critical for the specialized functions that the cells carry out. Odontoblasts are major matrix-producing cells of the tooth and have also been suggested to participate in sensory transmission. However, refined morphologic data on these important cells are limited, which hampers the analysis and understanding of their cellular functions. We took advantage of fluorescent color-coding genetic tracing to visualize and reconstruct in 3 dimensions single odontoblasts, pulp cells, and their assemblages. Our results show distinct structural features and compartments of odontoblasts at different stages of maturation, with regard to overall cellular shape, formation of the main process, orientation, and matrix deposition. We demonstrate previously unanticipated contacts between the processes of pulp cells and odontoblasts. All reported data are related to mouse incisor tooth. We also show that odontoblasts express TRPM5 and Piezo2 ion channels. Piezo2 is expressed ubiquitously, while TRPM5 is asymmetrically distributed with distinct localization to regions proximal to and within odontoblast processes.

Immediate human pulp response to ethanol-wet bonding technique.

OBJECTIVES: To evaluate the short-term response of human pulps to ethanol-wet bonding technique. METHODS: Deep class V cavities were prepared on 17 sound premolars and divided into three groups. After acid-etching, the cavities from groups 1 (G1) and 2 (G2) were filled with 100% ethanol or distilled water, respectively, for 60 s before the application of Single Bond 2. In group 3 (G3, control), the cavity floor was lined with calcium hydroxide before etching and bonding. All cavities were restored with resin composite. Two teeth were used as intact control. The teeth were extracted 48h after the clinical procedures. From each tooth serial sections were obtained and stained with haematoxylin and eosin (H/E) and Masson's trichrome. Bacteria microleakage was assessed using Brown & Brenn. All sections were blindly evaluated for five histological features. RESULTS: Mean remaining dentine thickness was 463±65µm (G1); 425±184µm (G2); and 348±194µm (G3). Similar pulp reactions followed ethanol- or water-wet bonding techniques. Slight inflammatory responses and disruption of the odontoblast layer related to the cavity floor were seen in all groups. Stained bacteria were not detected in any cavities. Normal pulp tissue was observed in G3 except for one case. CONCLUSIONS: After 48h, ethanol-wet bonding does not increase pulpal damage compared to water-wet bonding technique. CLINICAL SIGNIFICANCE: Ethanol-wet bonding may increase resin-dentine bond durability. This study reported the in vivo response of human pulp tissue when 100% ethanol was applied previously to an etch-and-rinse simplified adhesive system.

Investigation on the physical-mechanical properties of dental resin composites reinforced with novel bimodal silica nanostructures.

The aim of this study was to investigate the influence of bimodal silica nanostructures comprising of SiO2 nanoparticles (SiO2 NPs, ~70 nm) and SiO2 nanoclusters (SiO2 NCs, 0.07-2.70 µm) on physical-mechanical properties of resin-based composites (RBCs). SiO2 NPs and SiO2 NCs were prepared with the Stöber method and the coupling reaction, respectively, then silanized and employed as fillers to construct RBCs using a mixture of bisphenol A glycerolate dimethacrylate (Bis-GMA) and tri(ethylene glycol) dimethacrylate (TEGDMA) as the organic matrix. Results showed that the properties of RBCs were influenced by the filler ratios of bimodal silica nanostructures, and the appropriate amount of SiO2 NPs could effectively increase the activating light efficiency and filler packing density of RBCs. Among all experimental RBCs, RBC 50-20 (SiO2 NPs:SiO2 NCs=50:20, wt/wt) presented the highest degree of conversion (71.6±1.1%), the lowest polymerization shrinkage (2.6±0.1%), and the enhanced flexural strength (104.8±4.4 MPa), flexural modulus (6.2±0.3 GPa), and compressive strength (205.8±14.3 MPa), which were improved by 44%, 19%, 28%, 48%, and 42% in comparison with those of RBC 0-60 (SiO2 NPs:SiO2 NCs=0:60, wt/wt), respectively. Besides, in vitro cytotoxicity evaluation of RBC 50-20 indicated its acceptable cytotoxicity. Although the best performance was achieved by commercial Z350 XT, the introduction of bimodal silica nanostructures might provide the enhanced physical-mechanical properties of RBCs, compared with those of RBC 0-60 reinforced with unimodal SiO2 NCs.

The expression of hyperpolarization-activated cyclic nucleotide-gated channel 1 (HCN1) and HCN2 in the rat trigeminal ganglion, sensory root, and dental pulp.

Hyperpolarization-activated cyclic nucleotide-gated channel 1 (HCN1) and 2 (HCN2) are abundantly expressed in primary sensory neurons and contribute to neuronal excitability and pathological pain. We studied the expression of HCN1 and HCN2 in the rat trigeminal ganglion (TG) neurons and axons in the dental pulp, and the changes in their expression following inflammation, using light- and electron-microscopic immunocytochemistry and quantitative analysis. HCN1 and HCN2 were expressed predominantly in large-sized, neurofilament 200-immunopositive (+) or parvalbumin+ soma in the TG whereas they were expressed mostly in unmyelinated and small myelinated axons in the sensory root. The expression was particularly strong along the plasma membrane in the soma. In the dental pulp, majority of HCN1+ and HCN2+ axons coexpressed calcitonin gene-related peptide. They were expressed mainly in the peripheral pulp and pulp horn where the axons branch extensively in the dental pulp. The expression of HCN1 and HCN2 in TG neurons increased significantly in rats with experimentally induced inflammation of the dental pulp. Our findings support the notion that HCN1 and HCN2 are expressed mainly by both the soma of mechanosensitive neurons in the TG and peripheral axons of nociceptive neurons in the sensory root, and may play a role in the mechanisms of inflammatory pain from the dental pulp.