Functionalization of PCL-Based Fiber Scaffolds with Different Sources of Calcium and Phosphate and Odontogenic Potential on Human Dental Pulp Cells.

This study investigated the incorporation of sources of calcium, phosphate, or both into electrospun scaffolds and evaluated their bioactivity on human dental pulp cells (HDPCs). Additionally, scaffolds incorporated with calcium hydroxide (CH) were characterized for degradation, calcium release, and odontogenic differentiation by HDPCs. Polycaprolactone (PCL) was electrospun with or without 0.5% w/v of calcium hydroxide (PCL + CH), nano-hydroxyapatite (PCL + nHA), or ß-glycerophosphate (PCL + ßGP). SEM/EDS analysis confirmed fibrillar morphology and particle incorporation. HDPCs were cultured on the scaffolds to assess cell viability, adhesion, spreading, and mineralized matrix formation. PCL + CH was also evaluated for gene expression of odontogenic markers (RT-qPCR). Data were submitted to ANOVA and Student's t-test (α = 5%). Added CH increased fiber diameter and interfibrillar spacing, whereas ßGP decreased both. PCL + CH and PCL + nHA improved HDPC viability, adhesion, and proliferation. Mineralization was increased eightfold with PCL + CH. Scaffolds containing CH gradually degraded over six months, with calcium release within the first 140 days. CH incorporation upregulated DSPP and DMP1 expression after 7 and 14 days. In conclusion, CH- and nHA-laden PCL fiber scaffolds were cytocompatible and promoted HDPC adhesion, proliferation, and mineralized matrix deposition. PCL + CH scaffolds exhibit a slow degradation profile, providing sustained calcium release and stimulating HDPCs to upregulate odontogenesis marker genes.

Strontium-Doped Bioglass-Laden Gelatin Methacryloyl Hydrogels for Vital Pulp Therapy.

This study aimed to develop gelatin methacryloyl (GelMA)-injectable hydrogels incorporated with 58S bioactive glass/BG-doped with strontium for vital pulp therapy applications. GelMA hydrogels containing 0% (control), 5%, 10%, and 20% BG (w/v) were prepared. Their morphological and chemical properties were evaluated by scanning electron microscopy/SEM, energy dispersive spectroscopy/EDS, and Fourier transform infrared spectroscopy/FTIR (n = 3). Their swelling capacity and degradation ratio were also measured (n = 4). Cell viability (n = 8), mineralized matrix formation, cell adhesion, and spreading (n = 6) on DPSCs were evaluated. Data were analyzed using ANOVA/post hoc tests (α = 5%). SEM and EDS characterization confirmed the incorporation of BG particles into the hydrogel matrix, showing GelMA's (C, O) and BG's (Si, Cl, Na, Sr) chemical elements. FTIR revealed the main chemical groups of GelMA and BG, as ~1000 cm-1 corresponds to Si-O and ~1440 cm-1 to C-H. All the formulations were degraded by day 12, with a lower degradation ratio observed for GelMA+BG20%. Increasing the concentration of BG resulted in a lower mass swelling ratio. Biologically, all the groups were compatible with cells (p > 0.6196), and cell adhesion increased over time, irrespective of BG concentration, indicating great biocompatibility. GelMA+BG5% demonstrated a higher deposition of mineral nodules over 21 days (p < 0.0001), evidencing the osteogenic potential of hydrogels. GelMA hydrogels incorporated with BG present great cytocompatibility, support cell adhesion, and have a clinically relevant degradation profile and suitable mineralization potential, supporting their therapeutic potential as promising biomaterials for pulp capping.

Catalysis-based approaches with biopolymers and violet LED to improve in-office dental bleaching.

This in vitro experimental investigation aimed to evaluate the impact of the combined application of a nanofiber scaffold (NS), a polymeric catalyst primer (PCP) containing 10 mg/mL of heme peroxidase enzyme, and violet LED (LEDv) on the esthetic efficacy (EE), trans-amelodentinal cytotoxicity (TC), and procedural duration of conventional in-office bleaching therapy. To achieve this, 96 standardized enamel/dentin discs were individually placed in artificial pulp chambers. A 35% hydrogen peroxide (H2O2) bleaching gel was administered for 45, 30, or 15 min to the enamel, either previously coated with NS + PCP or left uncoated, followed by irradiation with LEDv for 15 min or no irradiation. The established groups were as follows: G1, negative control (no treatment); G2, 35% H2O2/45 min; G3, NS + PCP + LEDv; G4, NS + PCP + 35%H2O2/45 min + LEDv; G5, NS + PCP + 35%H2O2/30 min + LEDv; and G6, NS + PCP + 35%H2O2/15 min + LEDv. Extracts (culture medium + gel components diffused through the discs) were collected and applied to odontoblast-like MDPC-23 cells. EE (ΔE00 and ΔWI) and TC were assessed using ANOVA/Tukey analysis (p < 0.05). The EE analysis revealed no statistical differences between G6 and G2 (p > 0.05). Cells in G6 exhibited higher viability and lower oxidative stress compared to other bleached groups (p < 0.05). In conclusion, employing NS + PCP + LEDv to catalyze a 35%H2O2 bleaching gel applied for 15 min to the enamel resulted in successful esthetic improvements and reduced the cytotoxicity commonly linked with traditional in-office bleaching procedures.

Performance of self-etching adhesives on caries-affected primary dentin treated with glutaraldehyde or silver diamine fluoride.

OBJECTIVES: to evaluate the quality and stability of adhesive interfaces established by self-etching adhesives on caries-affected primary dentin (CAD) treated with glutaraldehyde (GA) or silver diamine fluoride (SDF). METHODS: 42 primary molars were exposed to a microbiological caries-inducing protocol and divided into 6 groups according to the adhesive system (Clearfil SE - CL or FL Bond II - FL) and pretreatment (water, GA or SDF) applied on CAD. One tooth from each group was analyzed for surface modification using infrared spectroscopy. Crowns were restored with resin composite (n = 36) and cut into beams and slices. The beams were subjected to microtensile testing, Raman spectroscopy and SEM after 24 h and 6 months of storage. The slices were analyzed using Micro-Raman spectroscopy to determine the diffusion zone thickness (DZ) in each period. Data were analyzed by ANOVA and Tukey or Kruskal-Wallis and Dunn tests (α = 0.05%). RESULTS: SDF reduced the immediate bond strength for both adhesives. The control groups showed a decrease in BS after 6 months in artificial saliva. GA increased immediate DZ for FL, while SDF had the opposite effect on CL. GA decreased the DZ for FL at 6 months. There was a predominance of adhesive failures with areas of cohesive dentin fractures within control groups. SIGNIFICANCE: Modifications caused by dentin surface treatments may directly affect the performance of adhesive systems and the quality and stability of adhesive restorations.

Pulp cell response to the application of silver diamine fluoride and potassium iodide on caries-like demineralized dentin.

OBJECTIVES: To investigate the response of pulp cells to the application of silver diamine fluoride (SDF) and potassium iodide (KI) on demineralized dentin. MATERIALS AND METHODS: The occlusal surfaces of human dentin discs (0.4 mm thick) with similar permeability were subjected to an artificial caries protocol, and then the discs were adapted into artificial pulp chambers. MDPC-23 cells were seeded on the healthy pulp dentin surface, while the demineralized surface was treated with SDF, KI, SDF + KI, or hydrogen peroxide (positive control-PC) (n = 8). The negative control (NC) received ultrapure water. After 24 h, cell viability (alamarBlue) and morphology (SEM) were evaluated. The extracts were then applied to new MDPC-23 cells seeded in culture plates to assess their viability and the formation of mineralized nodules (MN; Alizarin Red) after seven days. The data were analyzed using one-way analysis of variance/Tukey or Games-Howell tests (α = 5%). RESULTS: SDF and PC significantly reduced the viability of cells seeded on discs (45.6% and 71.0%, respectively). Only cells treated with SDF or PC detached from the dentin substrate, while the remaining cells showed altered morphology. Cells in contact with extracts showed less reduction in viability, but it was still more toxic compared to NC. Only PC reduced MN deposition. SDF + KI or KI alone did not affect the cell response. CONCLUSIONS: SDF applied alone showed a mild to moderate transdentinal cytotoxic effect on pulp cells. However, the combination of SDF + KI reduced the cytotoxic effects. Both materials used alone or in combination did not affect the mineralization ability of pulp cells. CLINICAL RELEVANCE: Besides improving esthetic results, associating potassium iodide with silver diamine fluoride may reduce the transdentinal cytotoxic effects of this cariostatic agent on pulp cells.

Calcium Trimetaphosphate-Loaded Electrospun Poly(Ester Urea) Nanofibers for Periodontal Tissue Engineering.

The objective of this research was to create and appraise biodegradable polymer-based nanofibers containing distinct concentrations of calcium trimetaphosphate (Ca-TMP) for periodontal tissue engineering. Poly(ester urea) (PEU) (5% w/v) solutions containing Ca-TMP (15%, 30%, 45% w/w) were electrospun into fibrous scaffolds. The fibers were evaluated using SEM, EDS, TGA, FTIR, XRD, and mechanical tests. Degradation rate, swelling ratio, and calcium release were also evaluated. Cell/Ca-TMP and cell/scaffold interaction were assessed using stem cells from human exfoliated deciduous teeth (SHEDs) for cell viability, adhesion, and alkaline phosphatase (ALP) activity. Analysis of variance (ANOVA) and post-hoc tests were used (α = 0.05). The PEU and PEU/Ca-TMP-based membranes presented fiber diameters at 469 nm and 414-672 nm, respectively. Chemical characterization attested to the Ca-TMP incorporation into the fibers. Adding Ca-TMP led to higher degradation stability and lower dimensional variation than the pure PEU fibers; however, similar mechanical characteristics were observed. Minimal calcium was released after 21 days of incubation in a lipase-enriched solution. Ca-TMP extracts enhanced cell viability and ALP activity, although no differences were found between the scaffold groups. Overall, Ca-TMP was effectively incorporated into the PEU fibers without compromising the morphological properties but did not promote significant cell function.

Chronic exposure to lipopolysaccharides as an in vitro model to simulate the impaired odontogenic potential of dental pulp cells under pulpitis conditions.

BACKGROUND: Simulating a bacterial-induced pulpitis environment in vitro may contribute to exploring mechanisms and bioactive molecules to counteract these adverse effects. OBJECTIVE: To investigate the chronic exposure of human dental pulp cells (HDPCs) to lipopolysaccharides (LPS) aiming to establish a cell culture protocol to simulate the impaired odontogenic potential under pulpitis conditions. METHODOLOGY: HDPCs were isolated from four healthy molars of different donors and seeded in culture plates in a growth medium. After 24 h, the medium was changed to an odontogenic differentiation medium (DM) supplemented or not with E. coli LPS (0 - control, 0.1, 1, or 10 µg/mL) (n=8). The medium was renewed every two days for up to seven days, then replaced with LPS-free DM for up to 21 days. The activation of NF-κB and F-actin expression were assessed (immunofluorescence) after one and seven days. On day 7, cells were evaluated for both the gene expression (RT-qPCR) of odontogenic markers (COL1A1, ALPL, DSPP, and DMP1) and cytokines (TNF, IL1B, IL8, and IL6) and the production of reactive nitrogen (Griess) and oxygen species (Carboxy-H2DCFDA). Cell viability (alamarBlue) was evaluated weekly, and mineralization was assessed (Alizarin Red) at 14 and 21 days. Data were analyzed with ANOVA and post-hoc tests (α=5%). RESULTS: After one and seven days of exposure to LPS, NF-κB was activated in a dose-dependent fashion. LPS at 1 and 10 µg/mL concentrations down-regulated the gene expression of odontogenic markers and up-regulated cytokines. LPS at 10 µg/mL increased both the production of reactive nitrogen and oxygen species. LPS decreased cell viability seven days after the end of exposure. LPS at 1 and 10 µg/mL decreased hDPCs mineralization in a dose-dependent fashion. CONCLUSION: The exposure to 10 µg/mL LPS for seven days creates an inflammatory environment that is able to impair by more than half the odontogenic potential of HDPCs in vitro, simulating a pulpitis-like condition.

New Technological Approaches for Dental Caries Treatment: From Liquid Crystalline Systems to Nanocarriers.

Dental caries is the most common oral disease, with high prevalence rates in adolescents and low-income and lower-middle-income countries. This disease originates from acid production by bacteria, leading to demineralization of the dental enamel and the formation of cavities. The treatment of caries remains a global challenge and the development of effective drug delivery systems is a potential strategy. In this context, different drug delivery systems have been investigated to remove oral biofilms and remineralize dental enamel. For a successful application of these systems, it is necessary that they remain adhered to the surfaces of the teeth to allow enough time for the removal of biofilms and enamel remineralization, thus, the use of mucoadhesive systems is highly encouraged. Among the systems used for this purpose, liquid crystalline systems, polymer-based nanoparticles, lipid-based nanoparticles, and inorganic nanoparticles have demonstrated great potential for preventing and treating dental caries through their own antimicrobial and remineralization properties or through delivering drugs. Therefore, the present review addresses the main drug delivery systems investigated in the treatment and prevention of dental caries.

Chronic exposure to lipopolysaccharides as an in vitro model to simulate the impaired odontogenic potential of dental pulp cells under pulpitis conditions

Abstract Simulating a bacterial-induced pulpitis environment in vitro may contribute to exploring mechanisms and bioactive molecules to counteract these adverse effects. Objective To investigate the chronic exposure of human dental pulp cells (HDPCs) to lipopolysaccharides (LPS) aiming to establish a cell culture protocol to simulate the impaired odontogenic potential under pulpitis conditions. Methodology HDPCs were isolated from four healthy molars of different donors and seeded in culture plates in a growth medium. After 24 h, the medium was changed to an odontogenic differentiation medium (DM) supplemented or not with E. coli LPS (0 - control, 0.1, 1, or 10 µg/mL) (n=8). The medium was renewed every two days for up to seven days, then replaced with LPS-free DM for up to 21 days. The activation of NF-κB and F-actin expression were assessed (immunofluorescence) after one and seven days. On day 7, cells were evaluated for both the gene expression (RT-qPCR) of odontogenic markers (COL1A1, ALPL, DSPP, and DMP1) and cytokines (TNF, IL1B, IL8, and IL6) and the production of reactive nitrogen (Griess) and oxygen species (Carboxy-H2DCFDA). Cell viability (alamarBlue) was evaluated weekly, and mineralization was assessed (Alizarin Red) at 14 and 21 days. Data were analyzed with ANOVA and post-hoc tests (α=5%). Results After one and seven days of exposure to LPS, NF-κB was activated in a dose-dependent fashion. LPS at 1 and 10 µg/mL concentrations down-regulated the gene expression of odontogenic markers and up-regulated cytokines. LPS at 10 µg/mL increased both the production of reactive nitrogen and oxygen species. LPS decreased cell viability seven days after the end of exposure. LPS at 1 and 10 µg/mL decreased hDPCs mineralization in a dose-dependent fashion. Conclusion The exposure to 10 µg/mL LPS for seven days creates an inflammatory environment that is able to impair by more than half the odontogenic potential of HDPCs in vitro, simulating a pulpitis-like condition.

Calcium silicate-coated porous chitosan scaffold as a cell-free tissue engineering system for direct pulp capping.

OBJECTIVES: This study aimed to develop and characterize different formulations of porous chitosan scaffolds (SCH) associated with calcium silicate (CaSi) and evaluate their chemotactic and bioactive potential on human dental pulp cells (hDPCs). METHODS: Different concentrations of CaSi suspensions (0.5%, 1.0%, and 2.0%, w/v) were incorporated (1:5; v/v) /or not, into 2% chitosan solution, giving rise to the following groups: SCH (control); SCH+ 0.5CaSi; SCH+ 1.0CaSi; SCH+ 2.0 CaSi. The resulting solutions were submitted to thermally induced phase separation followed by freeze-drying procedures to obtain porous scaffolds. The topography, pH, and calcium release kinetics of the scaffolds were assessed. Next, the study evaluated the influence of these scaffolds on cell migration (MG), viability (VB), proliferation (PL), adhesion and spreading (A&S), and on total protein synthesis (TP), alkaline phosphatase (ALP) activity, mineralized matrix deposition (MMD), and gene expression (GE) of odontogenic differentiation markers (ALP, DSPP, and DMP-1). The data were analyzed with ANOVA complemented with the Tukey post-hoc test (α = 5%). RESULTS: Incorporation of the CaSi suspension into the chitosan scaffold formulation increased pore diameter when compared with control. Increased amounts of CaSi in the CH scaffold resulted in higher pH values and Ca release. In Groups SCH+ 1.0CaSi and SCH+ 2.0CaSi, increased VB, PF, A&S, GE of DSPP/DMP-1 and MMD values were shown. However, Group SCH+ 2.0CaSi was the only formulation capable of enhancing MG and showed the highest increase in TP, MMD, and GE of DMP-1 and DSPP values. SIGNIFICANCE: SCH+ 2.0CaSi formulation had the highest chemotactic and bioactive potential on hDPCs and may be considered a potential biomaterial for pulp-dentin complex regeneration.

Manganese oxide increases bleaching efficacy and reduces the cytotoxicity of a 10% hydrogen peroxide bleaching gel.

OBJECTIVE: The study aims to assess the effects of a 10% H2O2 bleaching gel with different MnO2 concentrations on the bleaching efficacy (BE), degradation kinetics (DK) of H2O2, and trans-amelodentinal cytotoxicity (TC). MATERIALS AND METHODS: Standardized bovine enamel/dentin disks (n = 96) were placed in artificial pulp chambers, and the bleaching gels were applied for 45 min. Thus, the following groups were established: (G1) no treatment (negative control/NC); (G2) 35% H2O2 (positive control/PC); (G3) 10% H2O2; (G4) 10% H2O2 + 2 mg/mL MnO2; (G5) 10% H2O2 + 6 mg/mL MnO2; and (G6) 10% H2O2 + 10 mg/mL MnO2. After analyzing bleaching efficacy (ΔE00 and ΔWI), the degradation kinetics of H2O2 and trans-amelodentinal cytotoxicity were determined (n = 8, ANOVA/Tukey; p < 0.05). RESULTS: G6 presented BE (ΔE00 and ΔWI) statistically similar to G2, which represented conventional in-office bleaching (p = 0.6795; p > 0.9999). A significant reduction in the diffusion of H2O2 occurred in G3, G4, G5, and G6 compared to G2 (p < 0.0001). The highest DK of H2O2 occurred in G6 (p < 0.0001), which had the lowest TC in comparison with all other bleached groups (p ≤ 0.0186). CONCLUSION: The addition of 10 mg/mL of MnO2 in a 10% H2O2 bleaching gel potentiates the degradation of this reactive molecule, which increases the BE of the product and decreases TC. CLINICAL SIGNIFICANCE: Replacing a 35% H2O2 gel commonly used for conventional in-office dental bleaching by a 10% H2O2 gel containing 10 mg/mL of MnO2 reduces the cytotoxicity of this professional therapy, maintaining its excellent esthetic efficacy.

The influence of violet LED application time on the esthetic efficacy and cytotoxicity of a 35% H2O2 bleaching gel.

OBJECTIVE: To assess the potential influence of violet LED (V-LED) application time on the esthetic efficacy and cytotoxicity of a 35% H2O2 bleaching gel. METHODOLOGY: Stained and standardized enamel/dentin discs were subjected to one in-office tooth bleaching session (45 min), and the gel was either irradiated or not with V-LED. Thus, the following groups were established (n = 8): G1: No treatment (negative control, NC); G2: 35% H2O2 (positive control, PC); G3: 35%H2O2 + V-LED/15 min; G4: 35%H2O2 + V-LED/30 min; G5: 35%H2O2 + V-LED/45 min. First, esthetic efficacy was assessed (ΔE00 and ΔWI). Discs assembled in artificial pulp chambers were subjected to the same bleaching treatments. Then, the extracts (culture medium + diffused bleaching gel components) were collected and applied to MDPC-23 pulp cells, which were analyzed for viability (Live/Dead, MTT) and oxidative stress (OxS). The amount of H2O2 in the extracts was also determined (leuco crystal-violet/peroxidase). The data were subjected to ANOVA/Tukey at a 5% significance level. RESULTS: Although esthetic efficacy did not differ among the irradiated groups (G3, G4, and G5) (p > 0.05), their results were higher than in G2 (PC; p < 0.05). In the irradiated groups, the cell viability and OxS as well as the amount of H2O2 in the extracts were statistically similar to G2 (PC), regardless of irradiation time (p > 0.05). CONCLUSION: Although V-LED improves the esthetic outcome of in-office tooth bleaching, increasing irradiation time does not effect the color changes and cytotoxicity of this professional therapy.

Inhibitory activity of S-PRG filler on collagen-bound MMPs and dentin matrix degradation.

OBJECTIVES: To evaluate the inhibitory activity of an ion-releasing filler (S-PRG) eluate on dentin collagen-bound metalloproteinases (MMPs) and dentin matrix degradation. METHODS: Dentin beams (5 × 2 × 0.5 mm) from human molars were completely demineralized to produce dentin matrix specimens. The dry mass was measured, and a colorimetric assay (Sensolyte) determined the initial total MMP activity to allocate the beams into four treatment groups (n = 10/group): 1) water for 1 min (negative control); 2) 2% chlorhexidine digluconate (CHX - inhibitor control) for 1 min; 3) S-PRG eluate for 1 min; 4) S-PRG eluate for 30 min. After the treatments, the total MMP activity was reassessed. The specimens were stored in simulated body fluid (SBF) at 37 °C for up to 21 days. The dry mass was reassessed weekly. On day 7, the dentin matrix degradation was analyzed for the presence of collagen fragments (CF; Sirius Red) and hydroxyproline (Hyp) in the SBF. Statistical analyses were performed with ANOVA/Tukey, paired t-tests, and RM-ANOVA/Sidak (α = 5%). RESULTS: S-PRG eluate exposure for 1 and 30 min reduced (p < 0.0001) MMP activity. S-PRG exposure for 30 min presented MMP activity inhibition equivalent to CHX (p = 0.061). S-PRG and CHX decreased CF (p ≤ 0.007) and Hyp (p < 0.046) release. After 21 days of storage, S-PRG-treated beams, regardless of exposure time, presented a reduced (p ≤ 0.017) mass loss, intermediate between CHX and control. CONCLUSION: Treating demineralized dentin with S-PRG eluate for 1 or 30 min reduced matrix-bound MMP activity and dentin matrix degradation for up to 21 days. CLINICAL SIGNIFICANCE: S-PRG filler may hinder the progression of dentin carious/erosive lesions and enhance the stabilization of dentin bonding interfaces.

Influence of ceramic veneer on the transdentinal cytotoxicity, degree of conversion and bond strength of light-cured resin cements to dentin.

OBJECTIVES: To investigate the transdentinal cytotoxicity (TC), degree of conversion (DC), and micro shear bond strength (µSBS) to dentin of light-cured resin cements (LCRCs) photoactivated directly or through a ceramic veneer( ± CV). MATERIALS AND METHODS: The TC was assessed using human dentin discs adapted into artificial pulp chambers. Odontoblast-like cells were seeded on the pulp surface of the discs, then three LCRCs( ± CV) were applied on their etched and hybridized occlusal surface (n = 8/group). The adhesive systems of each LCRCs and sterile phosphate-buffered saline were used as positive and negative controls, respectively. After 24 h, the viability and morphology of cells adhered on discs were assessed. The extracts (culture medium + components of the materials diffused through the discs) were applied on the MDPC-23 to evaluate their viability, adhesion/spreading (A/S), alkaline phosphatase activity (ALP), and mineralized nodule formation (MN). LCRCs( ± CV) specimens were evaluated concerning the DC and µSBS to dentin. Data were analyzed by one-, two-, or three-way ANOVA/Dunnett, Sidak, and Games-Howell tests (α = 5%). RESULTS: All LCRCs( ± CV) reduced cell viability, A/S, ALP, MN, and DC. Except for µSBS, the intensity of reduction was dependent on the LCRC used. LCRCs+CV resulted in lower DC and µSBS but did not increase the TC. SIGNIFICANCE: Besides the presence of CV between the light source and LCRCs reduces the degree of conversion and bond strength to dentin, these materials cause variable level of transdentinal toxicity to pulp cells. Thus, the composition and curing protocols of LCRCs should be revisited and reinforced to prevent mechanical and biological drawbacks.

Nano-hydroxyapatite-incorporated polycaprolactone nanofibrous scaffold as a dentin tissue engineering-based strategy for vital pulp therapy.

OBJECTIVES: Targeting a tissue engineering-based vital pulp therapy (VPT), this study investigated the incorporation of nano-hydroxyapatite (nHA) into polycaprolactone (PCL) nanofibers, and the metabolism of human dental pulp cells (HDPCs) seeded on the scaffolds. METHODS: PCL-based solutions (10% w/v) containing nHA (0 - control; 0.5; 1.0; or 2.0% w/v) were electrospun into nanofibrous scaffolds. The scaffolds were characterized for morphology and composition (MEV/EDS), solubility, the release of calcium/phosphate (C/P), and modulation of medium pH. Then, HDPCs were seeded on the scaffolds and evaluated for cell viability (alamarBlue and live/dead), adhesion and spreading (F-actin), total protein (TP; Lowry), alkaline phosphatase activity (ALP; thymolphthalein assay), expression of odontogenic genes (RT-qPCR), and formation of a mineralized matrix (Alizarin Red). Data were analyzed with ANOVA and post-hocs (α = 5%). RESULTS: Higher nHA concentrations roughened fiber surfaces, whereas PCL+ 2%nHA increased the interfibrillar spaces. PCL+ 1%nHA or PCL+ 2%nHA significantly released more C/P but the medium pH was maintained below 8.0. HDPCs viability was not affected by nHA, while cell adhesion/spreading was favored, especially for PCL+ 2%nHA. Higher protein content and ALP activity were seen for scaffolds incorporated with nHA, after 21 days. PCL+ 1%nHA and PCL+ 2%nHA upregulated the expression of DSPP and DMP1 in 14 days, and COL1A1, ALPL, and DMP1 in 21 days. The formation of a mineralized matrix was nHA concentration-dependent, and it was about 9 × higher for PCL+ 2%nHA. SIGNIFICANCE: nHA-incorporated PCL nanofibrous scaffolds are cytocompatible and can stimulate the adhesion and odontogenic potential of HDPCs. PCL+ 2%nHA formulation is a bioactive tissue engineering-based cell-homing strategy for VPT.

Cytocompatibility and bioactivity of calcium hydroxide-containing nanofiber scaffolds loaded with fibronectin for dentin tissue engineering.

OBJECTIVES: The aim of this study was to characterize polycaprolactone-based nanofiber scaffolds (PCL) incorporated with calcium hydroxide (CH) and evaluate their bioactivity on human dental pulp cells (HDPCs) when loaded with fibronectin (FN). MATERIALS AND METHODS: CH (0.1%; 0.2%; 0.4% w/v; or 0%) was incorporated into PCL (10% w/v) scaffolds prepared by electrospinning. Morphology and composition were characterized using SEM/EDS. HDPCs were seeded on the scaffolds and evaluated for viability (alamarBlue; Live/Dead), and adhesion/spreading (F-actin). Next, scaffolds containing 0.4% CH were loaded with FN (20 µg/mL). HDPCs were evaluated for viability, adhesion/spreading, migration (Trans-well), gene expression (RT-qPCR), alkaline phosphatase activity (ALP), and mineralization nodules (Alizarin Red). Data were submitted to ANOVA and post-hoc tests (α = 5%). RESULTS: Nanofibers with larger diameter were seen as CH concentration increased, while there was no effect on interfibrillar spaces. An increase in cell viability was seen for 0.4% CH, in all periods. Incorporation of CH and FN into the scaffolds increased cellular migration, spread, and viability, all intensified when CH and FN were combined. ALPL and DSPP expression, and ALP activity were not affected by CH and FN. COL1A1 was downregulated in all groups, while DMP1 was upregulated in the presence of CH, with no differences for the groups loaded with FN. CH increased the formation of mineralized matrix, which was not influenced by FN. CONCLUSIONS: In conclusion, the incorporation of CH enhanced the odontogenic potential of HDPCs, irrespective of the presence of FN. The PCL + 0.4% CH formulation may be a useful strategy for use in dentin tissue engineering. CLINICAL RELEVANCE: A change in the form of presentation of calcium hydroxide-based materials used for direct pulp capping can increase biocompatibility and prolong the vitality of dental pulp.

Fibronectin-loaded Collagen/Gelatin Hydrogel Is a Potent Signaling Biomaterial for Dental Pulp Regeneration.

INTRODUCTION: Guided tissue regeneration has been considered a promising biological strategy to replace conventional endodontic therapies of teeth with incomplete root formation. Therefore, in the present study, a collagen/gelatin hydrogel either containing dosages of fibronectin (FN), or not, was developed and assessed concerning their bioactive and chemotactic potential on human apical papilla cells (hAPCs). METHODS: Hydrogels were prepared by varying the ratio of collagen and gelatin (Col/Gel; v/v), and used to establish the following groups: Collagen (positive control); Col/Gel 4:6; Col/Gel 6:4; Col/Gel 8:2. The viability, adhesion, and spreading of cells seeded on the hydrogels were evaluated. Different concentrations of FN (0, 5, or 10 µg/mL) were incorporated into the best formulation of the collagen/gelatin hydrogel selected. Then, the hAPCs seeded on the biomaterials were assessed concerning the cell migration, viability, adhesion and spreading, and gene expression of ITGA5, ITGAV, COL1A1, and COL3A1. RESULTS: The Col/Gel 8:2 group exhibited better cell viability, adhesion and spreading in comparison with Control. Higher values of hAPC migration, viability, adhesion, spreading and gene expression of pulp regeneration markers were found, the higher the concentration was of FN incorporated into the collagen/gelatin hydrogel. CONCLUSION: Collagen/gelatin hydrogel with 10 µg/mL of FN had potent bioactive and chemotactic effects on cultured hAPCs.

Response of pulp cells to resin infiltration of enamel white spot-like lesions.

OBJECTIVES: To investigate the trans-enamel and trans-dentinal biological effects of treating enamel white spot-like lesions (EWSLs) with resin infiltration components (RICs) on odontoblast-like cells (MDPC-23) and human dental pulp cells (HDPCs). METHODS: EWSLs were induced in 60 enamel/dentin discs (4.0 ± 0.2 mm thick) using S. mutans. The discs were adapted into artificial pulp chambers and MDPC-23 were seeded on the dentin surface. The components of a resin infiltration system (Icon) were applied individually or in combination on the enamel surface as following (n = 10/treatment): Etch, Infiltrant, Etch+Infiltrant, or Etch+Dry+Infiltrant. The application of water or hydrogen peroxide served as negative and positive controls, respectively. After 72 h, MDPC-23 viability was evaluated. The extracts were exposed for 72 h to pre-cultured MDPC-23 and HDPCs in 96-well plates to evaluate cell viability, alkaline phosphatase activity (ALP), mineralized nodule formation (MN), and the expression of inflammatory cytokines (ICs) and mineralization-related genes (MRs). Data were analyzed by ANOVA complemented with Tukey or Games-Howell post-hocs (α = 5%). RESULTS: Cell viability, ALP activity, and MN formation were significantly reduced in response to the RICs, presenting intermediate values compared to positive and negative controls. Likewise, ICs were upregulated, whereas MRs were downregulated. Among the RICs, the Etch component caused the most notorious detrimental effects. SIGNIFICANCE: Resin infiltration of EWSLs negatively affected the metabolism of pulp cells in vitro. Therefore, even though resin infiltration is a micro-invasive therapy for non-cavitated caries in enamel, it should be closely followed up seen that components may diffuse and unbalance pulp homeostasis.