Osteoblastic differentiation and changes in the redox state in pulp stem cells by laser treatment.

The aim of this study was to determine the effect of low-level laser therapy (LLLT) on cell proliferation, mitochondrial membrane potential changes (∆Ψm), reactive oxygen species (ROS), and osteoblast differentiation of human dental pulp stem cells (hDPSCs). These cells were irradiated with 660- and 940-nm lasers for 5 s, 50 s, and 180 s. Cell proliferation was assessed using the resazurin assay, cell differentiation by RUNX2 and BMP2 expression, and the presence of calcification nodules using alizarin-red S staining. ROS was determined by the dichlorofluorescein-diacetate technique and changes in ∆Ψm by the tetramethylrhodamine-ester assay. Data were analyzed by a Student's t-test and Mann-Whitney U test. The 940-nm wavelength for 5 and 50 s increased proliferation at 4 days postirradiation. After 8 days, a significant decrease in proliferation was observed in all groups. Calcification nodules were evident in all groups, with a greater staining intensity in cells treated with a 940-nm laser for 50 s, an effect that correlated with increased RUNX2 and BMP2 expression. ROS production and Δψm increased independently of irradiation time. In conclusion, photobiomodulation (PBM) with LLLT induced morphological changes and reduced cell proliferation rate, which was associated with osteoblastic differentiation and increased ROS and Δψm, independent of wavelength and time.

Effect of flavonoids from grape seed and cranberry extracts on the microbiological activity of Streptococcus mutans: a systematic review of in vitro studies.

OBJECTIVE: To provide an overview of the available scientific evidence from in vitro studies regarding the effect induced by the flavonoids contained in grape seed extracts (GSE) and cranberry on the microbiological activity of Streptococcus mutans (S. mutans). METHODS: This systematic review was performed following the parameters of the PRISMA statement (Preferred Reporting Items for Systematic Reviews and Meta-Analysis). Electronic and manual searches were conducted using PubMed, ScienceDirect, Web of Science, EBSCO, and Cochrane databases. Reference lists of selected articles were reviewed to identify relevant studies. The search was not limited by year and was conducted solely in English. Eligible studies comprised publications describing in vitro studies that evaluated the effect of flavonoids derived from GSE and cranberry extracts on the microbiological activity of S. mutans. Common variables were identified to consolidate the data. Authors of this review independently screened search results, extracted data, and assessed the risk of bias. RESULTS: Of the 420 studies identified from the different databases, 22 publications were finally selected for review. The risk of bias was low in 13 articles and moderate in 9. The studies analyzed in this review revealed that cranberry extract has an inhibitory effect on the bacterial growth of S. mutans in ranges from 0.5 mg/mL to 25 mg/mL, and GSE exerts a similar effect from 0.5 mg/mL to 250 mg/mL. Additionally, the extracts or their fractions showed reduced biofilm formation capacity, decreased polymicrobial biofilm biomass, deregulation of glycosyltransferases (Gtf) B and C expression, and buffering of pH drop. In addition to adequate antioxidant activity related to polyphenol content. CONCLUSIONS: The overall results showed that the extracts of cranberry and grape seed were effective in reducing the virulence factors of the oral pathogen. According to the data, proanthocyanidins are the active components in cranberry and grape seed that effectively resist S. mutans. They can inhibit the formation of insoluble polysaccharides in the extracellular matrix and prevent glycan-mediated adhesion, cohesion, and aggregation of the proteins in S. mutans. This suggests that these natural extracts could play an important role in the prevention of cariogenic bacterial colonization, as well as induce a decrease in their microbiological activity.

Response of osteoblastic cells to low-level laser treatment: a systematic review.

Low-level laser therapy (LLLT)-induced photobiomodulation (PBM) stimulates bone tissue regeneration by inducing osteoblast differentiation and mitochondrial activation. However, the role of reactive oxygen species (ROS) in this process remains controversial. The aim of this systematic review was to collect and analyze the available literature on the cellular and molecular effects of LLLT on osteoblasts and the role of ROS in this process. A search was conducted in PubMed, ScienceDirect, Scopus, and Web of Science. Studies published in English over the past 15 years were selected. Fourteen articles were included with moderate (n = 9) and low risk of bias (n = 5). Thirteen studies reported the use of diode lasers with wavelengths (λ) between 635 and 980 nm. One study used an Nd:YAG laser (λ1064 nm). The most commonly used λ values were 808 and 635 nm. The energy densities ranged from 0.378 to 78.75 J/cm2, and irradiation times from 1.5 to 300 s. Most studies found increases in proliferation, ATP synthesis, mitochondrial activity, and osteoblastic differentiation related to moderate and dose-dependent increases in intracellular ROS levels. Only two studies reported no significant changes. The data presented heterogeneity owing to the variety of LLLT protocols. Although several studies have shown a positive role of ROS in the induction of proliferation, migration, and differentiation of different cell types, further research is required to determine the specific role of ROS in the osteoblastic cell response and the molecular mechanisms involved in triggering previously reported cellular events.

Hydrothermal Transformation of Eggshell Calcium Carbonate into Apatite Micro-Nanoparticles: Cytocompatibility and Osteoinductive Properties.

The eggshell is a biomineral consisting of CaCO3 in the form of calcite phase and a pervading organic matrix (1-3.5 wt.%). Transforming eggshell calcite particles into calcium phosphate (apatite) micro-nanoparticles opens the door to repurposing the eggshell waste as materials with potential biomedical applications, fulfilling the principles of the circular economy. Previous methods to obtain these particles consisted mainly of two steps, the first one involving the calcination of the eggshell. In this research, direct transformation by a one-pot hydrothermal method ranging from 100-200 °C was studied, using suspensions with a stoichiometric P/CaCO3 ratio, K2HPO4 as P reagent, and eggshells particles (Ø < 50 µm) both untreated and treated with NaClO to remove surface organic matter. In the untreated group, the complete conversion was achieved at 160 °C, and most particles displayed a hexagonal plate morphology, eventually with a central hole. In the treated group, this replacement occurred at 180 °C, yielding granular (spherulitic) apatite nanoparticles. The eggshell particles and apatite micro-nanoparticles were cytocompatible when incubated with MG-63 human osteosarcoma cells and m17.ASC murine mesenchymal stem cells and promoted the osteogenic differentiation of m17.ASC cells. The study results are useful for designing and fabricating biocompatible microstructured materials with osteoinductive properties for applications in bone tissue engineering and dentistry.

Eggshell Membrane as a Biomaterial for Bone Regeneration.

The physicochemical features of the avian eggshell membrane play an essential role in the process of calcium carbonate deposition during shell mineralization, giving rise to a porous mineralized tissue with remarkable mechanical properties and biological functions. The membrane could be useful by itself or as a bi-dimensional scaffold to build future bone-regenerative materials. This review focuses on the biological, physical, and mechanical properties of the eggshell membrane that could be useful for that purpose. Due to its low cost and wide availability as a waste byproduct of the egg processing industry, repurposing the eggshell membrane for bone bio-material manufacturing fulfills the principles of a circular economy. In addition, eggshell membrane particles have has the potential to be used as bio-ink for 3D printing of tailored implantable scaffolds. Herein, a literature review was conducted to ascertain the degree to which the properties of the eggshell membrane satisfy the requirements for the development of bone scaffolds. In principle, it is biocompatible and non-cytotoxic, and induces proliferation and differentiation of different cell types. Moreover, when implanted in animal models, it elicits a mild inflammatory response and displays characteristics of stability and biodegradability. Furthermore, the eggshell membrane possesses a mechanical viscoelastic behavior comparable to other collagen-based systems. Overall, the biological, physical, and mechanical features of the eggshell membrane, which can be further tuned and improved, make this natural polymer suitable as a basic component for developing new bone graft materials.

Apatite-coated outer layer eggshell membrane: A novel osteoinductive biohybrid composite for guided bone/tissue regeneration.

Hybrid biomimetic materials aim to replicate the organic-inorganic constructs of mineralized tissues. During eggshell formation, the outer surface of the eggshell membrane (ESM) promotes calcium carbonate nucleation, while the inner one prevents mineralization toward the egg white and yolk. In the current study, the outer surface of the ESM acted as a heteronucleant in calcium phosphate precipitation by the vapor diffusion sitting drop method, while the inner one remained unmineralized. The aim was to fabricate a 2D biomaterial with dual functions, osteoinductive on one side and protective against cell invasion on the other side. The microstructural, physicochemical, morphological, and mechanical properties of the mineralized ESM were characterized by XRD, TGA, XPS, FTIR/Raman, HR-SEM, and mechanical testing techniques. The cytocompatibility and osteoinductive ability were assessed by biological assays of cell viability, proliferation, and osteogenic differentiation on human mesenchymal stromal cells (hMSCs). Results indicate that the outer surface of the ESM induces the heterogeneous precipitation of carbonate-apatite phase depicting biomimetic features. In addition, the apatite/ESM shows a much higher cytocompatibility than the pristine ESM and promotes the osteogenic differentiation of hMSCs more efficiently. Overall, the apatite/ESM composite exhibits compositional, crystalline, mechanical, and biological properties that resemble those of mineralized tissues, rendering it an approachable and novel material especially useful in guided tissue/bone regeneration.

Efficacy of CAD/CAM Glass Fiber Posts for the Restoration of Endodontically Treated Teeth.

OBJECTIVE: The aim of this systematic review was to provide an overview of available scientific evidence regarding the comparative efficacy of computer-aided design (CAD) and computer-aided manufacturing (CAM) glass fiber posts with prefabricated and metal cast posts for the restoration of endodontically treated teeth (ETT). METHODS: This systematic review was conducted following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Electronic and manual searches were performed using the PubMed, SciELO, Cochrane, ScienceDirect, Web of Science, and EBSCO databases. The reference lists of the selected papers were reviewed to identify relevant papers. There were no year restrictions, and eligible studies were those in English publications and describing in vitro studies evaluating intraradicular retainers (IRs) for (i) fracture resistance, (ii) bond strength, (iii) adaptation, and (iv) cement layer thickness. Literature reviews, systematic reviews, meta-analyses, case reports, in vitro studies with <8 specimens, and noncomparative trials involving prefabricated or metal cast posts were excluded. The authors of this review independently screened the search results, extracted data, and assessed the risk of bias. RESULTS: No significant differences were found in fracture resistance between prefabricated and CAD/CAM glass fiber posts or between CAD/CAM glass fiber and metal cast posts, although the latter demonstrated higher fracture resistance than the prefabricated glass fiber posts. Restoration with a full crown was not necessary to increase the fracture resistance in the presence of the ferrule effect. CAD/CAM glass fiber and metal cast posts had higher bond strength, lower nanoleakage, and better adaptation to the root canal. CONCLUSIONS: Despite the heterogeneity of methodologies and results reported, the results of these studies indicated that the CAD/CAM glass fiber and metal cast posts showed greater efficacy in terms of fracture resistance, retention, and adaptation, compared to prefabricated glass fiber posts.

Dataset on the effect of flavonoids on the stabilization of the resin-dentin interface.

Data in this article are associated with our research article "Effect of Myricetin on Odontoblast-like Cells and its Potential to Preserve Resin-Dentin Bonds." Both a poor infiltration of resin monomers into the demineralized dentin matrix and hydrolytic degradation of the adhesive could lead to the instability of the resin-dentin interface. The degradation of collagen is caused by matrix metalloproteinases (MMP) and cysteine cathepsins. These collagenolytic enzymes are contained in their latent form as pro-MMPs in the dentinal structure, and undergo activation during the adhesive process. Given that the integrity of the collagen matrix is essential for the preservation of the dentin bond strength in both the medium and long term, the inhibition of these proteases is necessary to improve the durability of adhesive restorations. Among the different strategies suggested to improve both the behavior of the substrate against enzymatic degradation and the biomechanical behavior of the adhesive interface, the use of protease inhibitors and collagen crosslinking agents has been recommended, such as polyphenols. Research has focused on flavonoids such as proanthocyanidins (PAC), a class of phenolic compounds found in a variety of plants such as blueberry and grape whose chemical structure favors their action as cross-linking agents. However, the focus has recently shifted towards myricetin (MYR) due to its chemical structure: a greater amount of hydroxyl groups at the substitution positions, which form bonds with the carbonyl groups of the side chains of collagen amino acids and generate interfiber bonds. Our previous study has shown the efficacy of MYR both as a cross-linking agent and as a MMP inhibitor without any immediate effects on microtensile bond strength (µTBS) and preserving it for six months after storage, and maintaining the odontoblastic phenotype without affecting cell viability. The objective of this article is to present a dataset on the effect of flavonoids PAC and MYR on the resin-dentin interface. Given that durability of the resin-dentin bond holds great importance for the clinical longevity of adhesive restorations, our data aims to show the effects of these flavonoids on resin-dentin µTBS after 18-month storage. Test groups for the µTBS assay were set as follows: G1 (negative control), conventional adhesion technique; G2 (vehicle control), 100% ethanol (EtOH) for 120 s; G3, 0.2% chlorhexidine (CHX) for 60 s; G4, 1% glutaraldehyde (GA) for 60 s; and G5, 600 µM myricetin (MYR) for 120 s. Datasets were exported to SPSS software, version 21.0 (SPSS, Chicago, IL, USA) for analysis using the Shapiro-Wilk, a two-way analysis of variance including factor interactions (treatment and storage time). Data are presented as mean ± standard deviation (SD). Differences with p-values < 0.05 were considered significant. Our data can be used as a basis for comparison among other natural and synthetic substances that could work as MMP inhibitors and crosslinking agents. These findings could be useful for designing an effective strategy towards the stabilization of the hybrid layer in a relevant clinical protocol.

Viability determination data for odontoblast-like cells exposed to resin monomers.

Data in this article are associated with our research article "Dental Resin Monomers Induce Early and Potent Oxidative Damage on Human Odontoblast-like Cells." Dental adhesives are polymeric compounds consisting of several chemical substances, including resin monomers, such as 2-hydroxyethyl methacrylate (HEMA) and triethylene glycol dimethacrylate (TEGDMA), together with other comonomers, making up the organic matrix of the adhesive and whose composition is based on the methyl methacrylate chemistry. The release of residual monomers, susceptible to biodegradation, acts as a source of bioactive compounds, which can interact with tissues and induce a cytotoxic cellular response. The most used techniques to evaluate cytotoxicity, proliferation, or metabolic activity of cells exposed to different substances, are MTT and resazurin. Each chemistry evaluates cell viability differently, so the data obtained could vary depending on the technique sensitivity to detect changes in cell metabolism. The objective of this article was to present viability data as a function of the metabolic activity in human odontoblast-like cells (hOLCs), exposed to 3, 6, 9, and 12 mM HEMA, or 0.75, 1.5, 3, and 6 mM TEGDMA evaluated by the MTT, and resazurin techniques in the first 24 hours of exposure, at different time points. The absorbance data for the MTT test and the fluorescence intensity for the resazurin test were obtained by spectrometry. SIMSTAT software 2.6.5 for Windows was used to confirm the normal data distribution (Levene's test). Subsequently, an analysis of variance (one-way ANOVA) was performed to compare the control with each HEMA and TEGDMA concentration. Where a p < 0.05 indicated a high F value, a Fisher's least significant differences post-hoc analysis was performed, using an alpha value < 0.05. Data from the different time points were compared with a Student's t-test for each concentration. These data may be useful to compare the cytotoxic response of hOLCs with other cell types or the cell response to other resin monomers.

Dental resin monomers induce early and potent oxidative damage on human odontoblast-like cells.

Resin-based dental materials consist of filler particles and different monomers that are light cured in situ to re-establish dental function and aesthetics. Due to the degree of conversion of adhesive polymers, the monomers triethyleneglycol dimethacrylate (TEGDMA) and 2-hydroxyethyl methacrylate (HEMA) are released in relatively high amounts and are susceptible to degradation, acting as bioactive compounds and affecting cell and tissues. This study aimed to assess the effect of HEMA and TEGDMA exposure on metabolic activity, membrane integrity, and cell survival of human odontoblast-like cell (hOLCs). Exposure to resin monomers for 24 h induced major changes in cell membrane integrity, metabolic activity, and survival, which were measured by the calcein method and lactate dehydrogenase release. Increased and early reactive oxygen species (ROS) production was observed leading to degradative oxidation of membrane lipids identified as malondialdehyde production. Severe alteration in mitochondria occurred due to transmembrane mitochondrial potential collapse, possibly inducing activation of apoptotic cell death. hOLCs exposure to resin monomers modified the cell redox potential, with consequences on membrane permeability and integrity, including mitochondrial function. Lipid peroxidation appears to be a key phenomenon for the membrane structures oxidation after HEMA and TEGDMA exposure, leading to cell death and cytotoxicity. hOLCs respond early by differential induction of adaptive mechanisms to maintain cell homeostasis. Modulation of oxidative stress-induced response involves the regulation of genes that encode for antioxidant proteins such as catalase and heme oxygenase-1; regulation that functions as a critical protection mechanism against oxidative cell damage induced by HEMA and TEGDMA. Ascorbic acid as an antioxidant substance mitigates the oxidative damage associated with exposure to monomers.

Infection Risk Prediction Model for COVID-19 Based on an Analysis of the Settlement of Particles Generated during Dental Procedures in Dental Clinics.

BACKGROUND: The health emergency declaration owing to severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has drawn attention toward nosocomial transmission. The transmission of the disease varies depending on the environmental conditions. Saliva is a recognized SARS-CoV-2 reservoir in infected individuals. Therefore, exposure to fluids during dental procedures leads to a high risk of contagion. OBJECTIVE: This study aimed to develop an infection risk prediction model for COVID-19 based on an analysis of the settlement of the aerosolized particles generated during dental procedures. MATERIALS AND METHODS: The settlement of aerosolized particles during dental aerosol-generating procedures (AGPs) performed on phantoms was evaluated using colored saliva. The gravity-deposited particles were registered using a filter paper within the perimeter of the phantom head, and the settled particles were recorded in standardized photographs. Digital images were processed to analyze the stained area. A logistic regression model was built with the variables ventilation, distance from the mouth, instrument used, area of the mouth treated, and location within the perimeter area. RESULTS: The largest percentage of the areas stained by settled particles ranged from 1 to 5 µm. The maximum settlement range from the mouth of the phantom head was 320 cm, with a high-risk cutoff distance of 78 cm. Ventilation, distance, instrument used, area of the mouth being treated, and location within the perimeter showed association with the amount of settled particles. These variables were used for constructing a scale to determine the risk of exposure to settled particles in dentistry within an infection risk prediction model. CONCLUSION: The greatest risk of particle settlement occurs at a distance up to 78 cm from the phantom mouth, with inadequate ventilation, and when working with a high-speed handpiece. The majority of the settled particles generated during the AGPs presented stained areas ranging from 1 to 5 µm. This model was useful for predicting the risk of exposure to COVID-19 in dental practice.

Effect of myricetin on odontoblast-like cells and its potential to preserve resin-dentin Bonds.

Stabilization of the resin-dentin interface to increase the durability of adhesive dental restorations is a challenging task. The use of naturally occurring collagen crosslinking agents has been proposed to prevent degradation of the hybrid layer. Myricetin (MYR) is a flavonoid with a wide variety of beneficial effects and it has been used for the treatment of different systemic pathologies. The chemical structure of MYR makes it a powerful antioxidant, an inhibitor of matrix metalloproteinase (MMP) activity, and a collagen cross-linker. This study presents MYR as a novel treatment in operative dentistry to stabilize the resin-dentin interface by inhibiting MMPs and crosslinking the collagen. Viability tests carried out using a resazurin assay showed that MYR had no cytotoxic effects on human odontoblast-like cells and the phenotype was preserved. Fluorometric MMP activity assay and fluorescence microscopy revealed that the MMPs in the demineralized dentin were effectively inhibited by the application of MYR (600 µM for 120 s). A microtensile bond strength test was performed immediately and after six months of storage. The bond strength to dentin was not affected by MYR and was preserved over time. Demineralized dentin beams were evaluated to determine the dentin biomodification using microtensile strength and elastic modulus assays. MYR improved the biomechanical behavior of the demineralized dentin similarly to glutaraldehyde, a recognized crosslinking agent. These findings indicated that MYR acts as an MMP inhibitor, collagen cross-linker, and preserver of the bond strength. Furthermore, MYR is an ethanol-soluble molecule with a lower molecular weight than the other polyphenols; hence, it can be applied for a short time and diffuses deeply through the dentin without any associated cytotoxicity. This molecule has beneficial effects on the biological and mechanical behavior of the resin-dentin interface and may be used to effectively stabilize the hybrid layer in a clinical setting.

Polymodal Activation and Desensitization of TRPV1 Receptor in Human Odontoblasts-Like Cells with Eugenol.

Dentinal hypersensitivity is a frequent reason for dental consultation, and its pathophysiology has not been fully clarified. Previous findings have made it possible to establish a relationship between the cellular sensory capacity and the activation of the polymodal transient receptor potential vanilloid 1 (TRPV1), which is responsible for the nociceptive response and whose desensitization could cause analgesia. Thus, the objective of this study was to determine the expression, localization, and functional activity of TRPV1 in human odontoblasts-like-cells (hOLCs) and the effect of eugenol (EUG) on its activation and desensitization. Human dental pulp stem cells (hDPSCs) were obtained from third molars and were characterized using flow cytometry, and their differentiation potential toward the osteoblastic, chondrogenic, and adipogenic lineages was investigated. Subsequently, the hDPSCs underwent odontogenic differentiation for 7, 14, and 21 days, and their phenotype (odontogenic markers dentin matrix protein-1 (DMP-1) and dentin sialoprotein (DSP)) was evaluated using immunofluorescence. The TRPV1 gene expression in hOLCs was estimated using RT-qPCR, and its localization was analyzed using immunofluorescence. Half-maximal effective concentration (EC50) from both eugenol (EUG) and capsaicin (CAP) was determined; in addition, receptor activation was evaluated against chemical, thermal, and pH stimuli. For the statistical analysis, a one-way ANOVA with a Tukey post hoc test (p < 0.05) was used. After establishing the in vitro model of hOLCs and the membrane location of TRPV1, its chemical activation with EUG and CAP was demonstrated, as well as its thermal activation at ≥ 43°C and with an acidic (<6) or basic pH (between 9 and 12). Receptor desensitization was achieved after 20 min of exposure to two concentrations of EUG (603.5 and 1000 µM). These findings represent a stepping-stone for the construction of a pulp pain study model oriented toward a therapeutic alternative for the treatment of dentinal hypersensitivity.

Odontoblast-Like Cells Differentiated from Dental Pulp Stem Cells Retain Their Phenotype after Subcultivation.

Odontoblasts, the main cell type in teeth pulp tissue, are not cultivable and they are responsible for the first line of response after dental restauration. Studies on dental materials cytotoxicity and odontoblast cells physiology require large quantity of homogenous cells retaining most of the phenotype characteristics. Odontoblast-like cells (OLC) were differentiated from human dental pulp stem cells using differentiation medium (containing TGF-ß1), and OLC expanded after trypsinization (EXP-21) were evaluated and compared. Despite a slower cell growth curve, EXP-21 cells express similarly the odontoblast markers dentinal sialophosphoprotein and dentin matrix protein-1 concomitantly with RUNX2 transcripts and low alkaline phosphatase activity as expected. Both OLC and EXP-21 cells showed similar mineral deposition activity evidenced by alizarin red and von Kossa staining. These results pointed out minor changes in phenotype of subcultured EXP-21 regarding the primarily differentiated OLC, making the subcultivation of these cells a useful strategy to obtain odontoblasts for biocompatibility or cell physiology studies in dentistry.