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Objective: Experimental procedures have been used to monitor cellular responses at the dentin/pulp interface. Aiming to divert from in vivo studies and oversimplified two-dimensional assays, three-dimensional (3D) models have been developed. This review provides an overview of existing literature, regarding 3D in vitro dentin/pulp reconstruction. Material & Methods: PubMed, Scopus, Cochrane Library and Web of Science- were systematically searched for attributes between 1998 and 2020. The search focused on articles on the development of three-dimensional tools for the reconstruction of a dentin/pulp complex under in vitro conditions, which were then screened and qualitatively assessed. Article grouping according to mode of implementation, resulted in five categories: the customised cell perfusion chamber (CPC) (n = 8), the tooth bud model (TBM) (n = 3), the 3D dentin/pulp complex manufactured by tissue engineering (DPC) (n = 6), the entire tooth culture (ETC) (n = 4) and the tooth slice culture model (TSC) (n = 5). Results: A total of 26 publications, applying nine and eight substances for pulp and dentin representation respectively, were included. Natural materials and dentin components were the most widely utilized. The most diverse category was the DPC, while the CPC group was the test with the highest longevity. The most consistent categories were the ETC and TSC models, while the TBM presented as the most complete de novo approach. Conclusions: All studies presented with experimental protocols with potential upgrades. Solving the limitations of each category will provide a complete in vitro testing and monitoring tool of dental responses to exogenous inputs. Clinical Relevance: The 3D dentin/pulp complexes are valid supplementary tools for in vivo studies and clinical testing. [ABSTRACT FROM AUTHOR]

The potential therapeutic role of the Dental Pulp Stem Cells Secretome (SECR) in a rat model of experimentally induced Temporomandibular Joint (TMJ) Osteoarthritis (OA) was evaluated. Proteomic profiling of the human SECR under specific oxygen tension (5% O2) and stimulation with Tumor Necrosis Factor-alpha (TNF-α) was performed. SECR and respective cell lysates (CL) samples were collected and subjected to SDS-PAGE, followed by LC-MS/MS analysis. The identified proteins were analyzed with Bioinformatic tools. The anti-inflammatory properties of SECR were assessed via an in vitro murine macrophages model, and were further validated in vivo, in a rat model of chemically-induced TMJ-OA by weekly recording of the head withdrawal threshold, the food intake, and the weight change, and radiographically and histologically at 4- and 8-weeks post-treatment. SECR analysis revealed the presence of 50 proteins that were enriched and/or statistically significantly upregulated compared to CL, while many of those proteins were involved in pathways related to "extracellular matrix organization" and "immune system". SECR application in vitro led to a significant downregulation on the expression of pro-inflammatory genes (MMP-13, MMP-9, MMP-3 and MCP-1), while maintaining an increased expression of IL-10 and IL-6. SECR application in vivo had a significant positive effect on all the clinical parameters, resulting in improved food intake, weight, and pain suppression. Radiographically, SECR application had a significant positive effect on trabecular bone thickness and bone density compared to the saline-treated group. Histological analysis indicated that SECR administration reduced inflammation, enhanced ECM and subchondral bone repair and regeneration, thus alleviating TMJ degeneration. [ABSTRACT FROM AUTHOR]

The aim of this systematic review was to evaluate the effectiveness of conservative different therapeutic modalities for temporomandibular disorders (TMD) pain. An electronic systematic search was conducted in the MEDLINE (PubMed) database to identify the randomized clinical trials (RCTs) published between 2001 and 2021. The following, simple or multiple conjunctions, search keywords were selected: TMD pain, TMD management or conservative treatment or treatment strategies and TMD pain, therapeutic modalities or interventions and TMD. Studies included must have patients older than 18 years, with painful TMD, which diagnosis was performed by Research Diagnostic Criteria for TMD or Diagnostic Criteria for TMD. Outcome variables were pain relief and post treatment pain intensity reduction. Data were analysed with non-parametric tests and the level of significance was set at p<.05. Out of 1599 articles obtained, 28 RCTs fulfilled all selection criteria and were included. The results of this study show that there was a significant decrease in short-term post-treatment TMD pain with the use of occlusal splint alone or in combination with other therapeutic modalities when compared with the control group. Statistically significant differences were also detected between laser and photobiomodulation group and the control, in short-term treatment TMD-related pain. The primary findings of the present systematic review showed that the occlusal splint alone or combined with other therapeutic intervention presented positive effect on short-term TMD pain reduction. Secondary outcome suggests that laser and photobiomodulation therapy had, also, a significant role in short term pain relief. [ABSTRACT FROM AUTHOR]

The paper presents computer-aided methods that allocate a dental implant and suggest its size, during the pre-operative planning stage, in conformance with introduced optimization criteria and established clinical requirements. Based on computed tomography data of the jaw and prosthesis anatomy, single tooth cases are planned for the best-suited implant insertion at a user-defined region. An optimum implantation axis line is produced and cylindrical implants of various candidate sizes are then automatically positioned, while their occlusal end is leveled to bone ridge, and evaluated. Radial safety margins are used for the assessment of the implant safety distance from neighboring anatomical structures and bone quantity and quality are estimated and taken into consideration. A case study demonstrates the concept and allows for its discussion. [Copyright &y& Elsevier]

• The dentin/pulp tissue analogue may be used for cytotoxicity assessment of resinous cements. • The composition of the lower compartment (pulp component) affects its stiffness and biological properties. • The resinous cements investigated using the dentin/pulp tissue analogue showed variable but mild cytotoxicity. • Angiogenesis was not severely affected by the resinous cements under investigation. A three-dimensional (3D) dentin/pulp tissue analogue, resembling the human natural tissue has been engineered in an in vitro setup, aiming to assess the cytocompatibility of resin-based dental restorative cements. Stem Cells from Apical Papilla (SCAP) and Human Umbilical Vein Endothelial Cells (HUVEC) were embedded in Collagen-I/Fibrin hydrogels at 1:3 ratio within 24-well plates. Hanging culture inserts were placed over the hydrogels, housing an odontoblast-like cell layer and a human treated-dentin barrier. Shear modulus of the hydrogels at 3.5 and 5 mg/ml was evaluated by dynamic mechanical analysis. Eluates of two resin-based cements, a dual-cure- (Breeze™, Pentron: Cement-1/C1), and a self-adhesive cement (SpeedCEMplus™, Ivoclar-Vivadent: Cement-2/C2) were applied into the dentin/pulp tissue analogue after pre-stimulation with LPS. Cytocompatibility was assessed by MTT assay, live/dead staining and real-time PCR analysis. Both hydrogel concentrations showed similar shear moduli to the natural pulp until day (D) 7, while the 5 mg/ml-hydrogel substantially increased stiffness by D14. Both cements caused no significant toxicity to the dentin/pulp tissue analogue. C1 induced stimulation (p < 0.01) of cell viability (158 ± 3%, 72 h), while pre-stimulation with LPS attenuated this effect. C2 (±LPS) caused minor reduction of viability (15–20%, 24 h) that recovered at 72 h for the LPS+ group. Both cements caused upregulation of VEGF , ANGP-1 , and downregulation of the respective receptors VEGFR-2 and Tie-1. Both resin-based cements showed good cytocompatibility and triggered angiogenic response within the dentin/pulp tissue analogue, indicating initiation of pulp repair responses to the released xenobiotics. [ABSTRACT FROM AUTHOR]

Hybrid chitosan/gelatin/nanohydroxyapatite (CS/Gel/nHA) scaffolds have attracted considerable interest in tissue engineering (TE) of mineralized tissues. The present study aimed to investigate the potential of CS/Gel/nHA scaffolds loaded with dental pulp stem cells (DPSCs) to induce odontogenic differentiation and in vitro biomineralization. CS/Gel/nHA scaffolds were synthesized by freeze-drying, seeded with DPSCs, and characterized with flow cytometry. Scanning Electron Microscopy (SEM), live/dead staining, and MTT assays were used to evaluate cell morphology and viability; real-time PCR for odontogenesis-related gene expression analysis; SEM-EDS (Energy Dispersive X-ray spectroscopy), and X-ray Diffraction analysis (XRD) for structural and chemical characterization of the mineralized constructs, respectively. CS/Gel/nHA scaffolds supported viability and proliferation of DPSCs over 14 days in culture. Gene expression patterns indicated pronounced odontogenic shift of DPSCs, evidenced by upregulation of DSPP, BMP-2, ALP, and the transcription factors RunX2 and Osterix. SEM-EDS showed the production of a nanocrystalline mineralized matrix inside the cell-based and - to a lesser extent - the cell-free constructs, with a time-dependent production of net-like nanocrystals (appr. 25−30 nm in diameter). XRD analysis gave the crystallite size (D = 50 nm) but could not distinguish between the initially incorporated and the biologically produced nHA. This is the first study validating the potential of CS/Gel/nHA scaffolds to support viability and proliferation of DPSCs, and to provide a biomimetic microenvironment favoring odontogenic differentiation and in vitro biomineralization without the addition of any inductive factors, including dexamethasone and/or growth/morphogenetic factors. These results reveal a promising strategy towards TE of mineralized dental tissues. [ABSTRACT FROM AUTHOR]

Introduction: The aim of the present study was to investigate how different aging protocols can affect the flexural strength and phase transformations of yttrium-stabilized zirconia ceramics (Y-TZP) for monolithic restorations. Materials and methods: Bar-shaped specimens from two zirconia ceramics bars were divided into three groups: a. no treatment (c), b. aging in an autoclave (a), and c. thermal cycling (t). The flexural strength was determined by the 3-point bending test and statistical analysis was performed to determine significant differences (p < 0.05). Weibull statistics was used to analyze the dispersion of strength values while surface microstructural analysis was performed through X-ray diffraction analysis (XRD), Fourier transform infrared spectroscopy (FTIR), and scanning electron microscopy (SEM). Results: Aging did not significantly affect the flexural strength but differences were recorded between the two groups, with group A presenting higher strength values and m-phase percentages. Conclusions: The observed differences between the two ceramics could be attributed to variations in composition and processing. [ABSTRACT FROM AUTHOR]

Purpose: Mesenchymal stem cells (MSCs) have appeared as a promising regenerative cell-based therapeutic, for degenerative conditions, such as OA, while the beneficial results from the application of MSCs have been attributed to the MSCs-derived secretome, which is the sum of cytoprotective factors produced by the MSCs. Aim of this study was to systematically review the literature in order to assess whether stem cell secretome (conditioned medium-CM, exosome-Exos or microvesicles-MV)(CM/Exos/MVs) treatment reduces inflammation and enhances cartilage regeneration in preclinical studies of experimental arthritis. Materials and Methods: An extensive electronic search was conducted by 2 independent reviewers by using the PubMed, Cochrane Library, Web of Science, and Scopus database, as well as Google Scholar, in order to identify the studies that met our inclusion criteria until August 2019. Included studies were assessed for quality and Risk of Bias (RoB) using the ARRIVE (Animal Research: Reporting In Vivo Experiments) guidelines and a modification of Systematic Review Centre for Laboratory animal Experimentation (SYRCLE) RoB tool for animal studies, respectively. Results: The initial search provided 525 records, with 28 fulfilling the inclusion criteria. The included studies presented great heterogeneity regarding the stem cells used, the preparation of therapeutic agent as well as the animal models used for testing. In addition, most studies presented with an unclear or high risk bias. Conclusion: In summary, the positive results of CM/Exos/MVs application in preclinical models of experimentally induced OA in terms of resolution of inflammation and cartilage regeneration are highlighted in this review, presenting a promising therapeutic solution for OA. [ABSTRACT FROM AUTHOR]

• The AgNP-doped Ti 6 Al 4 V surfaces preserved high rates of osteoblast cell viability. • The AgNP-doped Ti 6 Al 4 V surfaces preserved high rates of osteoblast cell proliferation. • The AgNP-doped Ti 6 Al 4 V surfaces showed antibacterial effects against P. gingivalis. • The AgNP-doped Ti 6 Al 4 V surfaces showed antibacterial effects against P. intermedia. • The AgNP-doped Ti 6 Al 4 V surfaces could be proposed for dental implant applications. This study aimed to develop silver nanoparticle (AgNP)-doped Ti 6 Al 4 V alloy surfaces and investigate their antibacterial properties against representative periopathogens and potential cytotoxicity on osteoblastic cells. AgNPs of different size distributions (5 and 30 nm) were incorporated onto the Ti 6 Al 4 V surfaces by electrochemical deposition, using colloid silver dispersions with increasing AgNP concentrations (100, 200 and 300 ppm). The time-course silver release from the specimen surfaces to cell culture media was assessed by Atomic Absorption Spectroscopy (AAS). Cell attachment, viability and proliferation were investigated by SEM, live/dead staining MTT and BrdU assays. The antibacterial effects were assessed against P. gingivalis and P. intermedia by serial dilution spotting assays. A time- and concentration-dependent silver release from the experimental surfaces was observed. Overall, cell viability and attachment on the AgNP-doped surfaces, suggested adequate cytocompatibility at all concentrations. A transient cytotoxic effect was detected at 24 h for the 5 nm-sized groups that fully recovered at later time-points, while no cytotoxicity was observed for the 30 nm-sized groups. A statistically significant, concentration-dependent decrease in cell proliferation rates was induced at 48 h in all AgNP groups, followed by recovery at 72 h in the groups coated with 5 nm-sized AgNPs. A statistically significant, concentration-dependent antibacterial effect up to 30% was confirmed against both periopathogens. This study sheds light to the optimal size-related concentrations of AgNP-doped Ti 6 Al 4 V surfaces to achieve antibacterial effects, without subsequent cytotoxicity. These results significantly contribute to the development of antibacterial surfaces for application in oral implantology. [ABSTRACT FROM AUTHOR]

• Aging of Y-TZP influences gingival cell attachment, viability and metabolic activity. • No differences were observed on gingival cell viability among abutment materials. • Lithium disilicate eluates triggered cell metabolic activity. • Surface micro-roughness alterations did not explain differences in biological behavior. This study aimed to investigate the biological effects of yttria-stabilized zirconia (Y-TZP) compared to other dental implant abutment materials, i.e. lithium disilicate (LS2) and titanium alloy (Ti), as well as the effects of aging of Y-TZP on viability/proliferation and attachment properties of Human Gingival Fibroblasts (HGFs). Cylindrical specimens of each material were prepared as per manufacturer's instructions. Y-TZP specimens were divided into three groups: 1. no aging (Zr0), 2. aging for 5 h, 134 °C, 2 bars, 100% humidity (Zr5), 3. aging for 10 h under the same conditions (Zr10). Surface roughness was evaluated by optical profilometry; cell metabolic activity/viability by MTT assay, morphological changes by Scanning Electron Microscopy (SEM) and ratio of live/dead cells by confocal microscopy. Results showed statistically significant reduction of HGF metabolic activity/viability in contact with Y-TZP after aging. Nevertheless, non-aged zirconia showed no significant differences compared with LS2, Ti and control cultures. In contrast, significant stimulation of cell metabolic activity/viability was observed in HGFs exposed to LS2 eluates. Differential morphological patterns were observed for HGF in contact with different materials/treatments, with obviously increased number of dead cells and sparser distribution of HGFs cultured on Zr10 specimens. These effects were not correlated with surface topography, since Y-TZP aging did not alter surface micro-roughness. These findings indicate that Y-TZP shows comparable biological properties to Ti and LS2 as implant abutment material. Nevertheless, Y-TZP aging might influence gingival cell attachment and proliferation properties, providing an alert to a potentially negative effect on the long-term maintenance of gingival architecture. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Chitosan/gelatin scaffolds affect odontogenic differentiation and hydroxyapatite-rich mineralized matrix formation by DPSCs. • Scaffolds with 0.1% GTA promoted abundant matrix (osteoid/ fully mineralized bone) formation in immunocompromised mice. • The in vivo results were enhanced by single pre-exposure of the cell/scaffold constructs to rhBMP-2 before implantation. • Combining DPSCs with chitosan/gelatin scaffolds represents a promising strategy for orofacial bone tissue engineering. Abstract Objective Biomimetic chitosan/gelatin (CS/Gel) scaffolds have attracted great interest in tissue engineering of several tissues. However, limited information exists regarding the potential of combining CS/Gel scaffolds with oral cells, such as dental pulp stem cells (DPSCs), to produce customized constructs targeting alveolar/orofacial bone reconstruction, which has been the aim of the present study. Methods Two scaffold types, designated as CS/Gel-0.1 and CS/Gel-1, were fabricated using 0.1 and 1% (v/v) respectively of the crosslinker glutaraldehyde (GTA). Scaffolds (n = 240) were seeded with DPSCs with/without pre-exposure to recombinant human BMP-2. In vitro assessment included DPSCs characterization (flow cytometry), evaluation of viability/proliferation (live/dead staining, metabolic-based tests), osteo/odontogenic gene expression analysis (qRT-PCR) and structural/chemical characterization (scanning electron microscopy, SEM; energy dispersive X-ray spectroscopy, EDX; X-ray powder diffraction, XRD; thermogravimetry, TG). In vivo assessment included implantation of DPSC-seeded scaffolds in immunocompromised mice, followed by histology and SEM-EDX. Statistical analysis employed one/two-way ANOVA and Tukey's post-hoc tests (significance for p < 0.05). Results Both scaffolds supported cell viability/proliferation over 14 days in culture, showing extensive formation of a hydroxyapatite-rich nanocrystalline calcium phosphate phase. Differential expression patterns indicated GTA concentration to significantly affect the expression of osteo/odontogenic genes, with CS/Gel-0.1 scaffolds being more effective in upregulating DSPP , IBSP and Osterix. In vivo analysis demonstrated time-dependent production of a nanocrystalline, mineralized matrix at 6, 8 and 10 weeks, being more prominent in constructs bearing rhBMP-2 pre-treated cells. The latter showed higher amounts of osteoid and fully mineralized bone, as well as empty space reduction. Significance These results reveal a promising strategy for orofacial bone tissue engineering. [ABSTRACT FROM AUTHOR]

Background: Development of clinical-grade cell preparations is central to meeting the regulatory requirements for cellular therapies under good manufacturing practice-compliant (cGMP) conditions. Since addition of animal serum in culture media may compromise safe and efficient expansion of mesenchymal stem cells (MSCs) for clinical use, this study aimed to investigate the potential of two serum/xeno-free, cGMP culture systems to maintain long-term "stemness" of oral MSCs (dental pulp stem cells (DPSCs) and alveolar bone marrow MSCs (aBMMSCs)), compared to conventional serum-based expansion. Methods: DPSC and aBMMSC cultures (n = 6/cell type) were established from pulp and alveolar osseous biopsies respectively. Three culture systems were used: StemPro_MSC/SFM_XenoFree (Life Technologies); StemMacs_MSC/XF (Miltenyi Biotek); and a-MEM (Life Technologies) with 15% fetal bovine serum. Growth (population doublings (PDs)), immunophenotypic (flow cytometric analysis of MSC markers) and senescence (ß-galactosidase (SA-ß-gal) activity; telomere length) characteristics were determined during prolonged expansion. Gene expression patterns of osteogenic (ALP, BMP-2), adipogenic (LPL, PPAR-β) and chondrogenic (ACAN, SOX-9) markers and maintenance of multilineage differentiation potential were determined by real-time PCR. Results: Similar isolation efficiency and stable growth dynamics up to passage 10 were observed for DPSCs under all expansion conditions. aBMMSCs showed lower cumulative PDs compared to DPSCs, and when StemMacs was used substantial delays in cell proliferation were noted after passages 6-7. Serum/xeno-free expansion produced cultures with homogeneous spindle-shaped phenotypes, while serum-based expansion preserved differential heterogeneous characteristics of each MSC population. Prolonged expansion of both MSC types but in particular the serum/xeno-free-expanded aBMMSCs was associated with downregulation of CD146, CD105, Stro-1, SSEA-1 and SSEA-4, but not CD90, CD73 and CD49f, in parallel with an increase of SA-gal-positive cells, cell size and granularity and a decrease in telomere length. Expansion under both serum-free systems resulted in "osteogenic pre-disposition", evidenced by upregulation of osteogenic markers and elimination of chondrogenic and adipogenic markers, while serum-based expansion produced only minor changes. DPSCs retained a diminishing (CCM, StemPro) or increasing (StemMacs) mineralization potential with passaging, while aBMMSCs lost this potential after passages 6-7 under all expansion conditions. Conclusions: These findings indicate there is still a vacant role for development of qualified protocols for clinical-grade expansion of oral MSCs; a key milestone achievement for translation of research from the bench to clinics. [ABSTRACT FROM AUTHOR]

This study aimed to investigate the potential of low-level laser irradiation (LLLI) to promote odontogenic differentiation and biomineralization by dental pulp stem cells (DPSCs) seeded inside bioceramic scaffolds. Mg-based, Zn-doped bioceramic scaffolds, synthesized by the sol-gel technique, were spotted with DPSCs and exposed to LLLI at 660 nm with maximum output power of 140 mw at fluencies (a) 2 and 4 J/cm2 to evaluate cell viability/proliferation by the MTT assay and (b) 4 J/cm2 to evaluate cell differentiation, using real-time PCR (expression of odontogenic markers) and a p-nitrophenylphosphate (pNPP)-based assay for alkaline phosphatase (ALP) activity measurement. Scanning electron microscopy (SEM) and X-ray diffraction (XRD) analysis were used for structural/chemical characterization of the regenerated tissues. Exposure of the DPSCs/scaffold complexes to the proposed LLLI scheme was associated with statistically significant increase of odontogenesis-related markers (bone morphogenetic protein 2 (BMP-2): 22.4-fold, dentin sialophosphoprotein (DSPP): 28.4-fold, Osterix: 18.5-fold, and Runt-related transcription factor 2 (Runx2): 3.4-fold). ALP activity was significantly increased at 3 and 7 days inside the irradiated compared to that in the non-irradiated SC/DPSC complexes, but gradually decreased until 14 days. Newly formed Ca-P tissue was formed on the SC/DPSC complexes after 28 days of culture that attained the characteristics of bioapatite. Overall, LLLI treatment proved to be beneficial for odontogenic differentiation and biomineralization of DPSCs inside the bioceramic scaffolds, making this therapeutic modality promising for targeted dentin engineering. [ABSTRACT FROM AUTHOR]

Glass-ceramic scaffolds containing Mg have shown recently the potential to enhance the proliferation, differentiation, and biomineralization of stem cells in vitro, property that makes them promising candidates for dental tissue regeneration. An additional property of a scaffold aimed at dental tissue regeneration is to protect the regeneration process against oral bacteria penetration. In this respect, novel bioactive scaffolds containing Mg2+ and Cu2+ or Zn2+, ions known for their antimicrobial properties, were synthesized by the foam replica technique and tested regarding their bioactive response in SBF, mechanical properties, degradation, and porosity. Finally their ability to support the attachment and long-term proliferation of Dental Pulp Stem Cells (DPSCs) was also evaluated. The results showed that conversely to their bioactive response in SBF solution, Zn-doped scaffolds proved to respond adequately regarding their mechanical strength and to be efficient regarding their biological response, in comparison to Cu-doped scaffolds, which makes them promising candidates for targeted dental stem cell odontogenic differentiation and calcified dental tissue engineering. [ABSTRACT FROM AUTHOR]

Stem cells from the apical papilla (SCAP) of human adult teeth are considered an accessible source of cells with angiogenic properties. The aims of this study were to investigate the endothelial transdifferentiation of SCAP, the secretion of pro- and antiangiogenic factors from SCAP, and the paracrine effects of SCAP when exposed to environmental stress to stimulate tissue damage. SCAP were exposed to serum deprivation (SD), glucose deprivation (GD), and oxygen deprivation/hypoxia (OD) conditions, individually or in combination. Endothelial transdifferentiation was evaluated by in vitro capillary-like formation assays, real-time polymerase chain reaction, western blot, and flow cytometric analyses of angiogenesis-related markers; secretome by antibody arrays and enzyme-linked immunosorbent assays (ELISA); and paracrine impact on human umbilical vein endothelial cells (HUVECs) by in vitro transwell migration and capillary-like formation assays. The short-term exposure of SCAP to glucose/oxygen deprivation (GOD) in the presence, but mainly in deprivation, of serum (SGOD) elicited a proangiogenesis effect indicated by expression of angiogenesis-related genes involved in vascular endothelial growth factor (VEGF)/VEGFR and angiopoietins/Tie pathways. This effect was unachievable under SD in normoxia, suggesting that the critical microenvironmental condition inducing rapid endothelial shift of SCAP is the combination of SGOD. Interestingly, SCAP showed high adaptability to these adverse conditions, retaining cell viability and acquiring a capillary-forming phenotype. SCAP secreted higher numbers and amounts of pro- (angiogenin, IGFBP-3, VEGF) and lower amounts of antiangiogenic factors (serpin-E1, TIMP-1, TSP-1) under SGOD compared with SOD or SD alone. Finally, secretome obtained under SGOD was most effective in inducing migration and capillary-like formation by HUVECs. These data provide new evidence on the microenvironmental factors favoring endothelial transdifferentiation of SCAP, uncovering the molecular mechanisms regulating their fate. They also validate the angiogenic properties of their secretome giving insights into preconditioning strategies enhancing their therapeutic potential. [ABSTRACT FROM AUTHOR]

Objectives Aim of this study was to investigate whether Dental Pulp Stem Cells-DPSCs responses to pulp injury caused by resinous monomers is be mediated through activation of Wnt/β-catenin signaling. Methods DPSCs cultures were established from third molars of healthy donors and characterized for stem cell markers with flow cytometry. Cells were exposed to TEGDMA (T: 0.5–2 mM) with or without presence of the Wnt-1 ligand (W:25–100 ng/ml) or the GSK3β inhibitor Lithium (L:1–10 mM), used both as activators of Wnt/β-catenin signaling. Cell viability was evaluated by MTT assay, cell cycle profiles by flow cytometry and expression of key molecules of Wnt/β-catenin signaling by Real-time PCR and Western Blot. Results DPSC exposure to TEGDMA caused a concentration-dependent cytotoxicity, accompanied by G1 arrest at lower and G2/M arrest at higher concentrations or after prolonged exposure. Lithium caused a dual effect, by stimulating/inhibiting cell proliferation at lower/higher concentrations respectively and causing a G2/M arrest in a concentration-dependent manner. Wnt signaling could be activated in DPSCs after Lithium or Wnt-1 treatment, as shown by accumulation of β-catenin, its translocation into the nucleus and enhanced expression of key pathway players, like LEF1 and Cyclin D1. Importantly, exposure to TEGDMA caused a more pronounced activation of the pathway, whereas cumulative effects were observed after T/L or T/W co-treatment, indicating a very strong activation of Wnt signaling after treatment of already “activated” (by Lithium or Wnt-1) cells with TEGDMA. Significance These findings highlight the important role of Wnt canonical signaling in pulp repair responses to common injuries. [ABSTRACT FROM AUTHOR]

Abstract: This study aimed at the chemical, microstructural and textural characterization of the sol–gel derived dental porcelain modified by bioactive glass and the evaluation of its bioactivity. Sol–gel derived specimens of the composite material were constructed and subjected to firing cycle. Specimens of bioactive glass and dental porcelain served as control. All the specimens were characterized using X-ray diffraction (XRD), Raman spectroscopy, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) coupled with energy dispersive spectroscopy (EDS) and N2 porosimetry. The assessment of in vitro bioactivity was carried out by incubating the composite specimens in DMEM solution. Apatite formation was evaluated by SEM/EDS, FTIR and XRD analysis. Microstructural analysis by SEM revealed irregularly shaped particles with broad size distribution, while complex porei network with large pore volume and non-uniform pore size distribution was evident. N2-adsorption isotherms were representative of non-nano-/meso-porous materials. A mixture of a- and b-wollastonite, apatite and leucite phases were detected by FTIR and Raman spectroscopy. The XRD analysis confirmed the previous results, though traces of cristobalite were identified too. The in vitro tests evidenced the bioactivity of the specimens in a 3-day-period. In conclusion, the physicochemical properties of the sol–gel derived composite result in a bioactive material, though further modifications need to be considered in order to fulfill the requirements of application in the clinical reality. [Copyright &y& Elsevier]

Abstract: Objectives: The aim of this study was to evaluate the fracture mode of endodontically treated canines (ETC) restored with glass-fibre posts (GFP) and either metal-ceramic (MC) or all-ceramic (AC) crowns, in the presence or absence of 2mm ferrule height. Methods: Fifty recently extracted human maxillary canines were endodontically treated and randomly divided into 5 groups of 10 specimens each. Four groups (with or without ferrule) were restored with GFP, composite cores and either MC or AC crowns. The fifth group with intact ETC served as control. Teeth were loaded to fracture at a 135° angle to their long axis, at a cross-head speed 0.5mm/min. The fracture modes were divided through optical microscopy to repairable and irreparable. Statistical analysis was performed by means of the Fisher''s Exact test. Results: All groups presented brittle fracture following bending deformation. Irreparable root fractures were the primary fracture mode independently of ferrule. The percentages (%) of repairable fractures were not statistically significant different amongst the MC and AC restored teeth independently of ferrule. However the 2mm ferrule was associated to a higher percentage of repairable fractures, especially in teeth restored with AC crowns. Conclusions: The percentage (%) of repairable fractures was higher in all-ceramic compared to metal-ceramic ETC and in restorations with ferrule, although the differences were not statistically significant. Clinical significance: This in vitro study supports evidence that neither the type of crown restoration nor the presence of ferrule affects statistically significantly the mode of fracture of ETC restored with GFP. [Copyright &y& Elsevier]

Abstract: Objective: The aim of this study was to investigate the effects of resinous monomers on the odontogenic differentiation and mineralization potential of apical papilla stem cells (SCAP). Methods: Cultures were established from developing third molars of healthy donors aged 14–18 years-old and were extensively characterized for proliferation rate, colony forming unit efficiency and expression of stem cell markers (STRO-1, CD146, CD34, CD45, CD105, CD117-c-Kit, CD24, CD90, Nanog, Oct3/4), in order to select those with enhanced stem cell and odontogenic differentiation properties. SCAP enriched cultures were then induced for odontogenic differentiation in the continuous presence of low concentrations (0.05–0.5mM) of the monomers 2-hydroxy-ethyl-methacrylate-HEMA and triethylene-glycol-dimethacrylate-TEGDMA for 3 weeks (long-term exposure). Additionally, the effects of a single exposure (72h) to higher concentrations of HEMA (2mM) and TEGDMA (1mM) were evaluated. Results: The results showed that both types of monomer-exposure significantly delayed the odontogenic differentiation and mineralization processes of SCAP cells. A down-regulation followed by recovery in the expression of differentiation markers, including dentin sialophosphoprotein-DSPP, bone sialoprotein-BSP, osteocalcin-OCN and alkaline phosphatase-ALP was recorded. This was accompanied by reduction of the mineralized matrix produced by monomer-treated-compared to non-treated contol cultures. Furthermore, a concentration-dependence was observed for both monomers during long-term exposure, whereas the effects of HEMA were evident at much lower concentrations compared to TEGDMA. Significance: These findings suggest that resinous monomers can delay the odontogenic differentiation of SCAP cells, potentially disturbing the physiological repair and/or developmental processes of human permanent teeth. [Copyright &y& Elsevier]

Abstract: Objectives: The aim of this study was to investigate the effects of HEMA and TEGDMA on the odontogenic differentiation potential of dental pulp stem/progenitor cells. Methods: Dental stem/progenitor cell cultures were established from pulp biopsies of human deciduous teeth of 1–3 year-old children (Deciduous Teeth Stem Cells-DTSCs). Cultures were characterized for stem cell markers, including STRO-1, CD146, CD34, CD45 using flow cytometry. Cytotoxicity was evaluated with the MTT assay. DTSCs were then induced for osteo/odontogenic differentiation by media containing dexamethasone, KH2PO4,β-glycerophosphate and l-ascorbic acid phosphate in the presence of nontoxic concentrations of HEMA (0.05–0.5mM) and TEGDMA (0.05–0.25mM) for 3weeks. Additionally, the effects of a single exposure (72h) to higher concentrations of HEMA (2mM) and TEGDMA (1mM) were also evaluated. Results: DTSCs cultures were positive for STRO-1 (7.53±2.5%), CD146 (91.79±5.41%), CD34 (11.87±3.02%) and negative for CD45. In the absence of monomers cell migration, differentiation and production of mineralized dentin-like structures could be observed. Cells also progressively expressed differentiation markers, including dentin sialophosphoprotein-DSPP, bone sialoprotein-BSP, osteocalcin-OCN and alkaline phosphatase-ALP. On the contrary, long-term exposure to nontoxic concentrations of HEMA and TEGDMA significantly delayed the differentiation and mineralization processes of DTSCs, whereas, one time exposure to higher concentrations of these monomers almost completed inhibited mineral nodule formation. BSP, OCN, ALP and DSPP expression were also significantly down-regulated. Significance: These findings suggest that HEMA and TEGDMA can severely disturb the odontogenic differentiation potential of pulp stem/progenitor cells, which might have significant consequences for pulp tissue homeostasis and repair. [Copyright &y& Elsevier]

Human deciduous teeth have been proposed as a promising source of mesenchymal stem cells for application in bone and dental tissue engineering. We established cultures of mesenchymal stem cells from the pulp of human deciduous teeth (deciduous teeth stem cells, DTSCs) and analyzed their morphologic, growth, immunophenotypic, and osteo/odontogenic differentiation characteristics using different isolation methods and culturing environments. We compared the biologic behavior of DTSCs isolated either by enzymatic dissociation (DTSCs-ED) or by direct outgrowth from pulp tissue explants (DTSCs-OG). We found that different isolation methods give rise to different populations/lineages of cells with respect to their phenotypic and differentiation characteristics. DTSCs-ED cultures comprised heterogeneous cell populations, whereas DTSCs-OG comprised more homogenous spindle-shaped cells. We have characterized DTSCs as STRO-1/CD146/CD34/CD45 cells. However, the percentage of STRO-1 and CD34 cells was higher in DTSCs-ED (STRO-1, 17.01 ± 5.04%; CD34, 19.79 ± 4.66%) compared to DTSCs-OG cultures (STRO-1, 5.18 ± 2.39%; CD34, 9.94 ± 3.41%), probably as a result of a higher release of stem/progenitor cells from the perivascular niche during enzymatic dissociation. DTSCs isolated using either method displayed an active potential for cellular migration and biomineralization, giving rise to 3D mineralized structures when challenged with dexamethasone, monopotassium phosphate, and β-glycerophosphate. These cellular aggregates progressively expressed differentiation markers of functional odontoblasts, including dentin sialophosphoprotein, bone sialoprotein, osteocalcin, and alkaline phosphatase, having the characteristics of osteodentin. However, in DTSCs-ED, the mineralization rate and the amount of mineralized matrix produced was higher compared to DTSCs-OG cultures. Therefore, DTSCs-ED cells display enhanced biomineralization potential, which might be of advantage for application in clinical therapy. [ABSTRACT FROM AUTHOR]

Abstract: Bioactive glass–alumina composite (BA) pellets were fabricated in the range 95/5–60/40wt.% respectively and were heat-treated under a specific thermal treatment up to 950°C. Control (unheated) and heat-treated pellets were immersed in Simulated Body Fluid (SBF) for bioactivity testing. All pellets before and after immersion in SBF were studied by Fourier Transform Infrared spectroscopy (FTIR), Scanning Electron Microscopy (SEM-EDS) and X-ray Diffraction (XRD) analysis. All composite pellets presented bioactive response. On the surface of the heat-treated pellets the development of a rich biological hydroxyapatite (HAp) layer was delayed for one day, compared to the respective control pellets. Independent of the proportion of the two components, all composites of each group (control and heat-treated) presented the same bioactive response as a function of immersion time in SBF. It was found that by the applied methodology, Al2O3 can be successfully applied in bioactive glass composites without obstructing their bioactive response. [ABSTRACT FROM AUTHOR]

Abstract: Objectives: A wide variety of dental ceramics compositions have been introduced in dental clinical practice in order to combine desired aesthetics with superior mechanical performance. The aim of the present study was to investigate the microstructural changes in three dental ceramics after their sintering according to manufacturers’ instructions and to comparatively evaluate some of their physical, mechanical and biological properties. Methods: The analysis of the phases present in each material before and after sintering was performed with scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction analysis (XRD). The thermal properties of ceramic specimens were evaluated with differential thermal and thermogravimetric analysis (TG-DTA). The mechanical properties evaluated were fracture toughness, Young''s modulus and microhardness with the Vickers indentation method. MTT assay was used for cell proliferation assessment. One-way analysis of variance (ANOVA) with Bonferroni multiple comparisons tests was used to determine statistically significant differences (significance level of p <0.05). Results: Results showed a remarkable variation among the three ceramic compositions of leucite content in the starting unheated ceramic powders ranging between 14 and 32wt.% and in the respective sintered powders ranging between 15 and 41wt.% The low fusing glass–ceramic and the high fusing leucite-based ceramic presented significantly higher fracture toughness (p <0.001) and microhardness and lower modulus of elasticity (p <0.05) compared to the low fusing feldspathic ceramic. The three ceramics were almost equivalent concerning their in vitro biological behavior. Significance: Variations in crystal structure, distribution and composition are related to differences concerning mechanical properties of dental ceramics. [Copyright &y& Elsevier]

Lasers have been well integrated in clinical dentistry for the last two decades, providing clinical alternatives in the management of both soft and hard tissues with an expanding use in the field of dental materials. One of their main advantages is that they can deliver very low to very high concentrated power at an exact point on any substrate by all possible means. The aim of this review is to thoroughly analyze the use of lasers in the processing of dental materials and to enlighten the new trends in laser technology focused on dental material management. New approaches for the elaboration of dental materials that require high energy levels and delicate processing, such as metals, ceramics, and resins are provided, while time consuming laboratory procedures, such as cutting restorative materials, welding, and sintering are facilitated. In addition, surface characteristics of titanium alloys and high strength ceramics can be altered. Finally, the potential of lasers to increase the adhesion of zirconia ceramics to different substrates has been tested for all laser devices, including a new ultrafast generation of lasers. [ABSTRACT FROM AUTHOR]

Objectives Aims of this study were to investigate the optimal conditions of laser irradiation of a novel Bioactive Glass/Dental Ceramic-BP67 composite for acceleration of hydroxyapatite-HA formation and to assess cellular responses on the precipitated HA region. Methods BP67 (Bioactive Glass: 33.3%, Dental Ceramic: 66.7%) was fabricated by the sol–gel method. A laser assisted biomimetic-LAB process was applied to BP67 sintered specimens immersed in 1.5-times concentrated simulated body fluid-1.5×-SBF. The effect of various energy densities of pulsed nanosecond Nd-YAG (1064 nm) laser and irradiation exposure times (30 min, 1 and 3 h) were evaluated for HA precipitation. The HA film was characterized by FTIR, XRD, SEM and micro Raman techniques. ICP-AES was used for revealing changes in chemical composition of the 1.5×-SBF during irradiation. Cell viability and morphological characteristics of periodontal ligament fibroblasts-PDLFs, human gingival fibroblasts-HGFs and SAOS-2 osteoblasts on the HA surface were evaluated by MTT assays and SEM. Results At optimal energy fluence of 1.52 J/cm 2 and irradiation time for 3 h followed by immersion in 1.5×-SBF at 60 °C, a dense HA layer was formed on laser-irradiated BP67 within 7 days. The resulting HA film was tightly bonded to the underlying substrate and had mineral composition similar to cementum. MTT assay showed a consistent reduction of cell proliferation on the HA layer in comparison to conventional control ceramic and BP67 for all 3 cell lines studied. Significance These findings suggest LAB is an effective method for acceleration of HA formation on materials with low bioactivity, while cellular responses need further investigation. [ABSTRACT FROM AUTHOR]

Objective This study aimed to investigate the potential of Mg-based bioceramic scaffolds combined with human treated-dentin matrices (hTDMs) and dentinogenesis-related morphogens to promote odontogenic differentiation and dentin-like tissue formation by Dental Pulp Stem Cells-DPSCs. Methods DPSC cultures were established and characterized by flow cytometry. Experimental cavities were prepared inside crowns of extracted teeth and demineralized by EDTA (hTDMs). Zn-doped, Mg-based bioceramic scaffolds, synthesized by the sol–gel technique, were hosted inside the hTDMs. DPSCs were spotted inside the hTDMs/scaffold constructs with/without additional exposure to DMP-1 or BMP-2 (100 ng/ml, 24 h). Scanning Electron Microscopy-SEM, live/dead fluorescence staining and MTT assay were used to evaluate cell attachment and viability; Real time PCR for expression of osteo/odontogenic markers; Inductively Coupled Plasma-Atomic Emission Spectrometry-ICP/AES for scaffold elemental release analysis; ELISA for hTDM growth factor release analysis; SEM and X-ray Diffraction-XRD for structural/chemical characterization of the regenerated tissues. Results Scaffolds constantly released low concentrations of Mg 2+ , Ca 2+ , Zn 2+ and Si 4+ , while hTDMs growth factors, like DMP-1, BMP-2 and TGFβ-1. hTDMs/scaffold constructs supported DPSC viability, inducing their rapid odontogenic shift, indicated by upregulation of DSPP, BMP-2, osteocalcin and osterix expression. Newly-formed Ca-P tissue overspread the scaffolds partially transforming into bioapatite. Exposure to DMP-1 or BMP-2 pronouncedly enhanced odontogenic differentiation phenomena. Significance This is the first study to validate that combining the bioactivity and ion releasing properties of bioceramic materials with growth factor release by treated natural dentin further supported by exogenous addition of key dentinogenesis-related morphogens (DMP-1, BMP-2) can be a promising strategy for targeted dentin regeneration. [ABSTRACT FROM AUTHOR]

Objectives The temperature variations during the veneering firing cycles of a zirconia dental ceramic can negatively affect its mechanical properties. A possible synergistic effect of both heat-treatment and aging while exposed to the oral environment could result to catastrophic failure. The aim of the present study was to investigate the effect of heat treatment followed during veneering and in vitro aging on the mechanical and microstructural properties of zirconia dental ceramics. Methods Three specimens from each of two zirconia blocks (Ivoclar IPS e.max ZirCAD (IV) and Wieland ZENO Zr (WI)) were cut by CAD/CAM technology, fully sintered and polished. Each one was cut in four equal parts. One part was used as control (C), one was heat-treated (H), one was aged (A) (134 °C, 2 bar, 10 h) and one was heat-treated and subsequently aged (HA). The mechanical properties (nano-hardness ( H ) and elastic modulus ( E* )) were investigated by nano-indentation tests while the surface characterization was carried out with XRD, FTIR and SEM. Results Different treatments on IV and WI samples resulted in a reduction of both H and E* values, however the differences were not statistically significant ( p > 0.05). The combination of treatments imposes an overall effect ( p < 0.001), enhancing the influence on both H and E* values. This reduction in mechanical properties was followed by an increase of monoclinic content. Greater variations in both H and E* values were recorded for WI samples. Significance The clinical performance of zirconia dental ceramics may be affected during firing and aging resulting in increased probability of failure. [ABSTRACT FROM AUTHOR]