Facultad de Ciencias de la Salud

Objectives: This study aimed at evaluating the cytotoxicity, chemical and structural properties of experimental fluoride-doped calcium-phosphates as potential remineralising materials for dental applications. Methods: Experimental calcium phosphates were formulated using β-tricalcium phosphate, monocalcium phosphate monohydrate, calcium hydroxide, and different concentrations of calcium/sodium fluoride salts [(5 wt%: VSG5F), (10 wt%: VSG10F), (20 wt%: VSG20F)]. A fluoride-free calcium phosphate (VSG) was used as control. Each tested material was immersed in simulated body fluid (SBF), (24 h, 15 and 30 days) to assess their ability to crystallise into apatite-like. Cumulative fluoride release was assayed up to 45 days. Moreover, each powder was placed into a medium containing human dental pulp stem cells (200 mg/mL) and their cytotoxicity was analysed using the 3-(4,5-dimethylthiazol-2-yl)− 2,5-diphenyltetrazolium bromide (MTT) assay (24 h, 48 h and 72 h incubation). These latter results were statistically analysed by ANOVA and Tukey’s test (α = 0.05). Results: All the experimental VSG-F materials produced fluoride-containing apatite-like crystals after SBF immersion. VSG20F presented prolonged release of fluoride ions into the storage media (45d). VSG, VSG10F and VSG20F showed a significant cytotoxicity at dilution of 1:1, while at 1:5, only VSG and VSG20F demonstrated a reduction in cell viability. At lower dilutions (1:10, 1:50 and 1:100) all specimens showed no significant toxicity to hDPSCs, but an increase in cell proliferation. Significance: The experimental fluoride-doped calcium-phosphates are biocompatible and possess a clear ability to evoke fluoride-containing apatite-like crystallisation. Hence, they may be promising remineralising materials for dental applications.

Objective: Remineralising composites with antibacterial properties may seal the cavity and prevent secondary caries. This study aimed at developing experimental flowable composites containing different concentrations of fluoride-doped calcium phosphate fillers and evaluating their remineralising and antibacterial properties. Methods: Experimental resin-based composites containing different concentrations (0–20 %) of fluoride-doped calcium phosphate fillers (VS10/VS20) were formulated. The release of calcium (Ca), phosphate (PO) and fluoride (F) ions was assessed for 30 days. Remineralisation properties were evaluated through ATR-FTIR and SEM/EDX after storage in simulated body fluid (SBF). The metabolic activity and viability of Streptococcus gordonii was also evaluated through ATP, CFU and live/dead confocal microscopy. The evaluation of specific monomer elution from the experimental composites was conducted using high-performance liquid chromatography (HPLC). Results: The composites containing VS10 showed the highest release of Ca, those containing VS20 released more F over time (p < 0.05), while there was no significant difference in terms of PO ions release between the groups (p > 0.05). A quick 7-day mineral precipitation was observed in the tested composites containing VS10 or VS20 at 10 %; these materials also showed the greatest antibacterial activity (p < 0.05). Moreover, the tested composites containing VS10 presented the lowest elution of monomers (p < 0.05). Conclusions: Innovative composites were developed with low monomers elution, evident antibacterial activity against S. gordonii and important remineralisation properties due to specific ions release. Clinical significance: Novel composites containing fluoride-doped calcium phosphates may be promising to modulate bacteria growth, promote remineralisation and reduce the risk of cytotoxicity related to monomers’ elution.

To evaluate the biomodification ability of lignin used as pre-treatment in human dentin before the application of an etch-and-rinse adhesive. Experimental hydroethanolic solutions with different cross-linking agents were used: 6.5% proanthocyanidins (PAC, from grape-seed extract); 2% cardanol (CARD, from cashew-nut shell liquid); lignin (LIG, from eucalyptus) at 1, 2 or 4% concentrations. The negative control (NC) was ethanol 50 v%. Extracted molars were prepared, and dentin microtensile bond strength ( TBS) was evaluated after 24 h water storage or 10,000 thermocycling aging. Further specimens were processed for SEM nanoleakage, micropermeability confocal microscopy evaluation and in situ degree of conversion (DC) through micro-Raman spectroscopy. Demineralized dentin sticks were submitted to a three-point bending test to evaluate the elastic modulus (E) before and after 1 min biomodification using the tested solutions. Moreover, it was also evaluated the mass changes and hydroxyproline (HYP) release after 4-weeks of water storage. Vibrational collagen crosslinking identification was evaluated through micro-Raman spectroscopy. The results were analyzed by analysis of variance (ANOVA) and Tukey’s test ( = 0.05). A significant reduction in TBS was observed in groups NC (p < 0.001) and CARD (p = 0.026). LIG-4% showed no significant reduction in TBS after aging (p = 0.022). Nanoleakage micrographs showed hybrid layer protection with all agents, but reduced micropermeability was attained only with lignin. Polymerization was negatively affected in the presence of all tested cross-linking agents, except LIG-1%. Lignin and cardanol increased the dentin E values, but only lignin reduced the mass loss in dentin specimens. Effective collagen crosslinking (1117 cm􀀀1 and 1235 cm􀀀1) was detected for all agents. HYP release was significantly lower with LIG-1% than NC (p < 0.001). Lignin was able to perform collagen cross-linking and prevent the degradation of unprotected dentin collagen, thereby improving the bonding performance of the composite restorations performed in this study.

The study investigated allogenic pulp transplantation as an innovative method of regenerative endodontic therapy. Three patients were selected for the endodontic treatment of single-root teeth, who also had a son/daughter with deciduous teeth or third molars scheduled for extraction. Receptor teeth were endodontically instrumented and irrigated using a tri-antibiotic solution. During the transplant procedures, the teeth from the son/daughter were extracted, sectioned, and the pulp was carefully removed. The harvested pulp from the donor was inserted into the root canal of the host tooth (father/mother), followed by direct pulp capping and resin composite restoration. The teeth were followed-up with for 2 years and were surveyed with computed tomography, the electric pulp vitality test, and Doppler ultrasound examination. At the 6-month follow-up, positive pulp vitality and the formation of periapical lesions were verified in cases 1 and 2. Case 3 showed remarkable periapical radiolucency before transplantation, but after 1 year, such lesions disappeared and there was positive vitality. All teeth were revascularized as determined by Doppler imaging after 2 years with no signs of endodontic/periodontal radiolucency. In conclusion, although this was a case series with only three patients and four teeth treated, it is possible to suppose that this allogenic pulp transplantation protocol could represent a potential strategy for pulp revitalization in specific endodontic cases.

Objective: To assess the effects of different concentrations of 10-methacryloyloxydecyl dihydrogen phosphate (10-MDP) included in experimental ceramic primers on the degree of conversion (DC) and microshear bond strength (μSBS) of a dual-cure resin cement, and on the acidity neutralization potential of zirconia (ZrO2) in comparison to hydroxyapatite (HAp). Methods: Experimental ceramic primers were formulated using 5 wt%, 10 wt%, 20 wt%, or 40 wt% 10-MDP as an acidic functional monomer and camphorquinone (CQ)/amine or 1-phenyl-1,2- propanedione (PPD) as a photoinitiator system. Clearfil Ceramic Primer (Kuraray Dental, Tokyo, Japan) was used as the commercial control. Micro- Raman spectroscopy was used to assess the DC of uncured and light-cured resin cements applied onto primer-treated ZrO2 surfaces. The μSBS and pH of primers were assayed in a universal testing machine and by a digital pH meter (Tec-3MP; Tecnal, Piracicaba, Brazil), respectively. Statistical analysis was performed by one-way analysis of variance (ANOVA) and Tukey’s test (p<0.05). Results: DC was not affected until a concentration of 10% 10-MDP in CQ primer and 5% 10-MDP in PPD primer was reached, when compared with the positive control (p>0.05). Groups 10-MDP 5% in CQ and PPD primers showed the highest μSBS compared with the positive control (p>0.05); however, higher concentrations of 10-MDP induced significant DC and μSBS reduction (p<0.05). HAp neutralized 10-MDP primers, but ZrO2 provided higher acidity to the primers’ pH. Conclusion: 10-MDP monomer should be used in low concentrations in ZrO2 primers to avoid reduction of the polymerization and bond strength of resin cement.

This study aimed at evaluating the shear bond strength (SBS) of modern self-adhesive resin cements and resin-modified glass ionomer cements applied to di erent prosthetic substrates. Zirconia, lithium-disilicate glass-ceramic and a noble metal alloy were used as bonding substrates. They were all sand-blasted with alumina, while LD was further etched with 9.6% hydrofluoric acid (10 s). A light-curing resin-modified glass ionomer cement (3M-GIC: Ketac Cem Plus) and a self-curing resin-modified glass ionomer cement (GC-GIC: FujiCEM 2) were compared to self-adhesive resin cements (PAN: Panavia SA Universal) and (3M-RES: Rely X Unicem 2). Ten specimens for each substrate were produced and up to five cylinders of each cement were bonded to each substrate. The shear bond strength (SBS) was evaluated after 24 h or after thermocycling (TC) aging (5000 cycles). The data was statistically analysed by two-way ANOVA and Student–Newman–Keuls test ( = 0.05). Failure modes were analysed through stereoscopic microscopy. The greatest SBS was attained with PAN, whilst 3M-GIC showed the lowest SBS and failed prevalently in adhesive mode. No di erence in SBS was observed between GC-GIC and 3M-RES. After TC aging, all cements showed significant drop (p < 0.05) in SBS, but PAN showed the greatest SBS. Reliable bond strength to prosthetic substrates can be achieved with specific universal resin-luting cements and may be an alternative to glass ionomer cements when luting alloy substrates.

The aim of this study was to evaluate the physicochemical properties of experimental adhesives containing tailored fluoridated calcium phosphate fillers. Five experimental resins were formulated, one contained no calcium phosphate filler (Control) and the other four resins contained 40 wt% of calcium phosphate fillers doped with increasing concentrations of fluoride: adhesives CaP (no fluoride), CaP-2F (2 wt% F), CaP-4F (4 wt% F), and CaP-8F (8 wt% F). Resin specimens were prepared for a three-point bending test to evaluate the elastic modulus (E) and flexural strength (FS) at 24 h and after 2 months of water storage. Disk-shaped specimens were prepared to analyze the water sorption (Ws) according to ISO 4049. The degree of conversion (DC) of the tested resins was also assessed through FTIR. Statistical analysis was performed with ANOVA and Tukey’s test (p < 0.05). The addition of F-CaP particles at 0 and 2 wt% fluoride had no effect on the DC (p > 0.05) compared to the filler-free resin. However, these same experimental resins showed the highest E after water storage. All resins had a significant reduction of the FS after water storage (p < 0.001), but at a lower percentage rate for those experimental resins containing fluoride-free CaP or fluoride-doped CaP- 2F. These latter experimental resins achieved similar Ws to control the filler-free resin. Conversely, those resins containing 4 or 8 wt% fluoride-doped CaP had a significant Ws increase (p < 0.05). In conclusion, the incorporation of calcium phosphate particles without fluoride or with 2 wt% fluoride may represent a promising strategy to generate adhesive resins with specific physicochemical properties. Fluoride incorporation in CaP fillers for dental resin adhesive should be performed at low concentrations in order to avoid excessive water sorption and a decrease of polymerization.

Purpose: To compare after five-year simulated pulpal pressure (SPP) the degradation of adhesive-dentin interfaces created using two simplified adhesives applied with different bonding strategies. Materials and Methods: A two-step self-etch (CSE: Clearfil SE Bond) adhesive was used as a control multistep adhesive. The tested experimental materials were two simplified adhesives, a one-step self-etch (CS3: Clearfil S3 Bond) and a self-priming etch-and-rinse adhesive (SB2: Adper Single-Bond 2). Half of the bonded specimens were submitted to microtensile bond strength (μTBS) testing after 24 h. The other half submitted to SPP for five years before μTBS testing. Nonfractured sticks were evaluated using transmission electron microscopy (TEM). Scanning electron microscopy (SEM) was used to evaluate silver-nitrate nanoleakage within the interface. Data were statistically analyzed by two-way ANOVA and Tukey’s test (p < 0.05). Results: Prolonged SPP induced bond-strength reduction for both SB2 and CS3. All bonding approaches showed increased nanoleakage after aging. The two simplified adhesives showed severe degradation at the resin-dentin interface. TEM revealed that the main degradation patterns for the etch-and-rinse adhesive SB2 was collagen breakdown, while polymer hydrolysis along with filler debonding was mainly observed in CS3. Conclusions: Simplified adhesives applied in etch-and-rinse mode are mainly characterized by hydrolysis and collagen degradation. In self-etch mode, simplified adhesives may principally show hydrolysis of the polymeric matrix and/or at the interface of fillers and coupling agent. The use of multistep self-etching adhesives may guarantee greater dentin bond durability compared to simplified adhesives.