Quercetin reduces erosive dentin wear: Evidence from laboratory and clinical studies

ObjectiveThe aim of the present study was to evaluate the effect of quercetin on the acid resistance of human dentin through both laboratory and clinical studies.MethodsTwo hundred and twelve dentin blocks (2 mm × 2 mm × 2 mm) were prepared and used. For the laboratory study, dentin specimens were randomly divided into 8 groups (n = 12): deionized water, ethanol, 1.23 × 10⁴ μg/ml sodium fluoride (NaF), 120 μg/ml chlorhexidine, 183.2 μg/ml epigallocatechin gallate (EGCG), and 75 μg/ml, 150 μg/ml, and 300 μg/ml quercetin (Q75, Q150, and Q300). The specimens were treated with the respective solutions for 2 min and then subjected to in vitro erosion (4 cycles/d for 7 d). The surface microhardness loss (%SMH_l), erosive dentin wear, and surface morphology were evaluated and compared. For the impact on MMP inhibition, the release of crosslinked carboxyterminal telopeptide of type I collagen (ICTP) and the thickness of the demineralized organic matrix (DOM) were measured using additional dentin specimens. For the clinical study, the specimens were treated with NaF or Q300 for 2 min and then subjected to in vivo erosion (4 cycles/d for 7 d). The %SMH_l and erosive dentin wear of the specimens were measured to determine whether quercetin similarly inhibits erosion in situ.ResultsThe quercetin-treated group had a significantly lower %SMH_l and erosive dentin wear than any other group, and the effect was concentration-dependent in vitro (P < 0.05). Dentin treated with quercetin produced significantly less ICTP and had a thicker DOM than the control dentin (P < 0.05). After in vivo erosion, the %SMH_l and erosive dentin wear of the Q300 group were significantly lower than those of the control group (P < 0.05).SignificanceThe application of quercetin was shown, for the first time, to increase the acid resistance of human dentin, possibly through MMP inhibition and DOM preservation.     

The effect of active components from citrus fruits on dentin MMPs

ObjectivesThis study was aimed to evaluate the anti-matrix metalloproteinases (MMPs) ability of active components from citrus fruits (hesperetin: Hst, hesperidin: Hsd and naringenin: Nge).MethodsInactivation effects of citrus flavonoids (Hst, Hsd, Nge) at different concentrations on soluble collagenase were measured using a fluorometric assay. Matrix-bound endogenous MMPs activity was evaluated via dry mass loss and hydroxyproline (HYP) release of demineralized human dentin. Demineralized dentin beams were pretreated with 500 μg/mL citrus flavonoids for 10 min. Chlorhexidine (CHX) was used as inhibitor control. Beams pretreated with distilled water served as blank control. Dentin slabs were used for in situ zymography and evaluated under confocal microscopy. Ultrastructure of demineralized collagen fibers was exhibited by Transmission Electron Microscopy (TEM).ResultsCitrus flavonoids exhibited inactivation function on soluble MMPs and the extent of inactivation increased in a dose-dependent manner. The inactivation percent of citrus flavonoids reached above 90% at the concentration of 500 μg/mL. Compared with control group, citrus flavonoids pretreated demineralized dentin beams exhibited less dry mass loss, lower hydroxyproline release and more intact collagen architecture after 15 days storage. Dentin samples pretreated with citrus flavonoids showed lower enzymes activities in in situ zymography.ConclusionsHst, Hsd or Nge have anti-MMPs ability and can preserve dentin collagen from degradation.Clinical Significance: Hst, Hsd and Nge may have the potential to be used in dentin bonding systems and improve the resin-dentin bonding durability.     

Biomimetic remineralization of artificial caries dentin lesion using Ca/P-PILP

ObjectiveTo assess the ultrastructural change of demineralized dentin collagen during calcium phosphate polymer-induced liquid precursor (Ca/P-PILP) mediated remineralization process and to evaluate the biomimetic remineralization potential of high concentration Ca/P-PILP at demineralized artificial caries dentin lesion, additionally to investigate the bond interfacial integrity as well as the bonding strength of the biomimetic remineralized artificial caries dentin lesion.MethodsDemineralized dentin collagen of 5 μm thick was biomimetically remineralized with low, medium concentration Ca/P-PILP for 10 days and high concentration Ca/P-PILP for 10, 15, 20 days. Artificial caries dentin lesion at a thickness of 150 ± 50 μm was biomimetically remineralized with high concentration Ca/P-PILP for 20 days. The biomimetic remineralization of demineralized dentin collagen was observed by scanning electron microscopy (SEM). The biomimetic remineralization intensity and depth of artificial caries dentin lesion was assessed by Electron Probe Micro Analyzer (EPMA). The bonding interfacial integrity between remineralized artificial caries dentin and composite resin was observed by Swept-source optical coherence tomography (SS-OCT) and the bonding strength of remineralized artificial caries dentin was evaluated by micro-tensile bond strength analysis (μTBS).ResultsSolely PAA-PASP solution and solely saturated Ca/P solution can’t achieve dentin collagen remineralization. Increased concentration of Ca/P-PILP and prolonged remineralization time can enhance the biomimetic remineralization intensity of demineralized dentin collagen. After treating with high concentration Ca/P-PILP, a 150 ± 50 μm thick layer of demineralized artificial caries dentin lesion was not fully remineralized, and the biomimetic remineralization intensity reached up to 88.0%. Furthermore, a better bonding interfacial integrity with less microgap and increased bond strength at both baseline level and aging level were observed when artificial caries dentin lesion was biomimetically remineralized with high concentration Ca/P-PILP.SignificanceBiomimetic remineralization of demineralized caries dentin lesion promotes its clinical properties for resin composited adhesive restoration.     

Distinct effects of polyphenols and solvents on dentin collagen crosslinking interactions and biostability

ObjectiveTo evaluate the effects of different polyphenols and solvents on dentin collagen’s crosslinking interactions and biostabilization against MMPs and collagenase degradation.MethodsTwo polyphenols [proanthocyanidin (PA) and quercetin (QC)] with different water solubility were prepared as treatment solutions using ethanol (EtOH) or dimethyl sulfoxide (DMSO) as solvents. 6-um-thick dentin films were microtomed from dentin slabs of third molars. Following demineralization, films or slabs were subject to 60-s treatment (PA or QC) or no treatment (control) with subsequent extended-rinse with original solvent (EtOH or DMSO) or distilled water (DW). Collagen crosslinking interactions were assessed by FTIR. Biostability was assessed through endogenous MMPs activity via confocal laser scanning microscopy, and exogenous collagenase degradation via weight loss, hydroxyproline release and SEM. Finally, direct collagenase inactivation was also evaluated. Data were analyzed by three-way ANOVA and post-hoc tests (α = 0.05%).ResultsDistinct effects of two polyphenols and solvents on collagen crosslinking and biostabilization were observed. Higher crosslinking and biostability efficacy occurred with PA than QC (p < 0.001) that demonstrated negligible collagen interactions. With DMSO solvent, efficacy results were significantly reduced with both polyphenols (p < 0.05). DMSO-rinse further weakened interactions of PA with collagen, diminishing biostability (p < 0.05). Low biostability was detected with QC and DW-rinse, suggesting direct enzymatic inhibition due to physical presence in collagen.SignificanceCollagen crosslinking interactions and biostability depend on polyphenol chemical characteristics. Treatment-solution solvents may affect interactions between polyphenols and collagen, specifically, DMSO showed detrimental effects on collagen crosslinking and biostability and should be used with caution.     

Potential of Treated Dentin Matrix Xenograft for Dentin-Pulp Tissue Engineering

IntroductionThis study aims to develop and characterize the regenerative potential of an atelopeptidized treated dentin matrix xenograft using in vitro and in vivo models.MethodsFreshly extracted bovine dentin was pulverized into 250- to 500-μm particles and demineralized with 17% EDTA for 1, 7, and 13 days. The samples were atelopeptidized with pepsin. The degree of demineralization and the effect of atelopeptidization were assessed using field emission scanning electron microscopy combined with energy-dispersive X-ray spectroscopy and Fourier transform infrared spectroscopy, respectively. The expression of dentin matrix acidic phosphoprotein 1, dentin sialophosphoprotein, and osteopontin was evaluated in dental pulp stem cells using quantitative real-time polymerase chain reaction. The samples were then implanted intramuscularly in rats for 30 days, and the inflammatory cells were quantified histologically.ResultsField emission scanning electron microscopy combined with energy-dispersive X-ray spectroscopy revealed an exposed tubular structure of dentin after 1 and 7 days of demineralization. Fourier transform infrared spectroscopy confirmed the absence of amide peaks at 1260 to 1640/cm after atelopeptidization. The dental pulp stem cell expression of dentin matrix acidic phosphoprotein 1 and dentin sialophosphoprotein increased in all compared with the untreated control group (P < .05). The maximum expression rates were observed for the 1-day demineralized and atelopeptidized group. The 1-day demineralized group elicited the highest inflammatory response compared with the 7- or 13-day demineralized groups (P < .001). Atelopeptidization significantly decreased the inflammatory response only in the 1-day demineralized dentin group (P < .05).ConclusionsAtelopeptidization of 1-day demineralized dentin xenograft preserved the collagen structure, minimized the immune reaction, and provided sufficient regenerative potential.     

Proanthocyanidin-functionalized hydroxyapatite nanoparticles as dentin biomodifier

ObjectiveThis study evaluated the potential combined effects of nanohydroxyapatite and proanthocyanidin on the remineralization and collagen stabilization of demineralized dentin.MethodsSeventy-five coronal dentin beams (6 × 1 × 1 mm³) were randomly allocated into five experimental groups (n = 15): Sound (no treatment), Control (pH-cycling), nHAp (nanohydroxyapatite), nHAp_PA (Proanthocyanidin-functionalized nanohydroxyapatite), and PA (proanthocyanidin) treatments. The sound group (negative control) were immersed in distilled water over the experimental period. The remaining groups were submitted to a pH-cycling process for 14 days. Following the de-re mineralization process, specimens corresponding to the control group (positive control) were immersed in distilled water whereas the test groups were immersed in 1 mL of respective solution treatment (nHAp, nHAp_PA, or PA) for 1 min. The dentin samples were analyzed to determine their chemical composition (ATR-FTIR and Thermogravimetric) and mineralogical (XRD) characteristics as well as their mechanical response, obtained by three-point bending test.ResultsHigher phosphate content (v₄ PO₄: ATR-FTIR) and amount of mineral (XRD) was observed in the nHAp_PA group. Furthermore, a larger induction of collagen cross-links (ATR-FTIR) and %Organic Matter (TGA) would indicate the PA incorporation and the achievement of dentin matrix stability. These effects on dentin properties were related to increasing flexural strength (MPa), demonstrating that 15% w/v nHAp_PA treatment improved the mechanical properties of the samples.SignificancenHAp_PA shows significant potential for promoting remineralization while improving collagen stability into demineralized dentin in a clinically feasible period of 1 min.     

Efficacy of red propolis hydro-alcoholic extract in controlling Streptococcus mutans biofilm build-up and dental enamel demineralization

ObjectiveThe efficacy of a red propolis hydro-alcoholic extract (RP) in controlling Streptococcus mutans biofilm colonization was evaluated. The effect of RP on dental demineralization was also investigated.MethodsChemical composition was determined by High Performance Liquid Chromatography (HPLC). Minimum Inhibitory and Bactericidal Concentration (MIC and MBC, respectively) were investigated against Streptococcus mutans (ATCC 25175). The cytotoxic potential of 3% RP in oral fibroblasts was observed after 1 and 3 min. Bovine dental enamel blocks (N = 24) were used for S. mutans biofilm formation (48 h), simulating ‘feast or famine’ episodes. Blocks/biofilms were exposed 2×/day, for 3 days, to a cariogenic challenge with sucrose 10% (5 min) and treated (1 min) with: 0.85% saline solution (negative control), 0.12% Chlorhexidine (CHX, positive control for biofilm colonization), 0.05% Sodium Fluoride (NaF, positive control to avoid demineralization) and 3% RP. Biofilms were assessed for viability (CFU/mL), and to observe the concentration of soluble and insoluble extracellular polysaccharides (SEPS and IEPS). Dental demineralization was assessed by the percentage of surface hardness loss (%SHL) and through polarized light microscopy (PLM).ResultsThe RP presented 4.0 pH and ºBrix = 4.8. The p-coumaric acid (17.2 μg/mL) and luteolin (15.23 μg/mL) were the largest contents of phenolic acids and flavonoids, respectively. MIC and MBC of RP were 293 μg/mL and 1172 μg/mL, respectively. The 3% RP showed 43% of viably cells after 1 min. Lower number (p < 0.05) of viable bacteria (CFU/mL) was observed after CHX (1.8 × 10⁵) followed by RP (1.8 × 10⁷) treatments. The lowest concentration (μg/CFU) of SEPS (12.6) and IEPS (25.9) was observed in CHX (p < 0.05) followed by RP (17.1 and 54.3), and both differed from the negative control (34.4 and 63.9) (p < 0.05). Considering the %SHL, all groups differed statistically (p < 0.05) from the negative control (46.6%); but NaF (13.9%), CHX (20.1%) and RP (20.7%) did not differ among them (p > 0.05). After all treatments, suggestive areas of caries lesions were observed by PLM, which were lower for CHX and NaF.ConclusionThe 3% RP reduced S. mutans colonization, decreased concentration of extracellular polysaccharides and reduced dental enamel demineralization.     

Inhibition of endogenous human dentin MMPs by Gluma

ObjectiveThe objective of this study was to determine if Gluma dentin desensitizer (5.0% glutaraldehyde and 35% HEMA in water) can inhibit the endogenous MMPs of dentin matrices in 60 s and to evaluate its effect on dentin matrix stiffness and dry mass weight.MethodsDentin beams of 2 mm × 1 mm × 6 mm were obtained from extracted human third molars coronal dentin. To measure the influence of Gluma treatment time on total MMP activity of dentin, beams were dipped in 37% phosphoric acid (PA) for 15 s and rinsed in water. The acid-etched beams were then dipped in Gluma for 5, 15, 30 or 60 s, rinsed in water and incubated into SensoLyte generic MMP substrate (AnaSpec, Inc.) for 60 min. Controls were dipped in water for 60 s. Additional beams of 1 mm × 1 mm × 6 mm were completely demineralized in 37% PA for 18 h, rinsed and used to evaluate changes on the dry weight and modulus of elasticity (E) after 60 s of Gluma treatment followed by incubation in simulated body fluid buffer for 0, 1 or 4 weeks. E was measured by 3-pt flexure.ResultsGluma treatment inhibited total MMP activity of acid-etched dentin by 44, 50, 84, 86% after 5, 15, 30 or 60 s of exposure, respectively. All completely demineralized dentin beams lost stiffness after 1 and 4 weeks, with no significant differences between the control and Gluma-treated dentin. Gluma treatment for 60 s yielded significantly less dry mass loss than the control after 4 weeks.SignificanceThe use of Gluma may contribute to the preservation of adhesive interfaces by its cross-linking and inhibitory properties of endogenous dentin MMPs.     

Chlorhexidine-loaded poly (amido amine) dendrimer and a dental adhesive containing amorphous calcium phosphate nanofillers for enhancing bonding durability

ObjectiveA novel method of combining chlorhexidine (CHX) loaded poly (amido amine) (PAMAM) dendrimers with a dental adhesive containing amorphous calcium phosphate (ACP) nanofillers are proposed for etch-and-rinse bonding system to enhance resin-dentin bonding durability.MethodsThe CHX-loaded PAMAM and ACP nanofillers were synthesized and characterized. Their effects on the cytotoxicity were tested by MTT assay. Micro-tensile bond strength (μTBS) before and after thermomechanical challenges were used to evaluate the bonding durability. Anti-matrix metalloproteinase (MMPs) property was examined using in-situ zymography. A double-fluorescence technique was used to examine interfacial permeability after bonding. Dentin remineralization in Ca/P lacking solution was observed under scanning electron microscopy.ResultsCompared with a 0.2 wt% CHX solution, the PAMAM loaded CHX had less cytotoxicity, while the in situ zymography showed it could still inhibit MMPs activity within the hybrid layer after released from PAMAM. The application of the novel method maintained the μTBS better than the control group after thermomechanical challenges, and it did not negatively affect water permeability of the bonding interfaces. CHX-loaded PAMAM regulated the calcium (Ca) and phosphate (P) ions provided by the ACP-containing adhesives to remineralize the demineralized dentin surfaces without initial Ca/P in the environment.SignificanceThe novel method can reduce the cytotoxicity of CHX, inhibit MMPs activities, maintain μTBS, and induce dentin remineralization, which are crucial factors for enhancing bonding durability.     

Neurotrophic Proteins in Dentin and Their Effect on Trigeminal Sensory Neurons

IntroductionA plethora of bioactive molecules present during tooth formation become sequestered in the mineralized dentin matrix and can be released into the pulp tissue after demineralization from carious lesions. However, neurotrophic factors are differentially expressed and secreted during various stages of odontogenesis. Thus, the aims of this study were(1) to investigate their presence and relative abundance in crown and root dentin and(2) to evaluate the bioactivity of dentin-derived proteins on neuronal cells.MethodsDentin matrix proteins (DMPs) were isolated from matched roots and crowns of extracted healthy human third molars. The total protein amount as well as the concentration of growth factors and neurotrophic proteins were quantified. The impact on neuritogenesis was determined with mouse trigeminal neurons in vitro and by a hydrogel implant model in vivo. Transient receptor potential cation channel subfamily V member 1 (TRPV1) sensitization of DMP-conditioned neurons was evaluated by single-cell calcium imaging.ResultsThe relative concentration of neurotrophic molecules revealed that nerve growth factor is the most abundant neurotrophin with 3-fold increased expression in radicular dentin. Similarly, brain-derived neurotrophic factor and neurotrophin 3 are more abundant in radicular than coronal dentin. Conversely, glial cell line–derived neurotrophic factor is more abundant in coronal dentin, whereas neurotrophin 4 is equally distributed. Dentin matrix proteins promoted neurite outgrowth in vitro and axonal targeting in vivo, with a greater effect observed by radicular dentin extracts. Furthermore, DMPs sensitized TRPV1 responses in mouse trigeminal neurons with greater activity seen with extracts from root dentin.ConclusionsNeurotrophic factors are differentially distributed between coronal and radicular dentin with different effects of dentin-derived proteins on axonal growth and targeting as well as the sensitization of TRPV1. Thus, extracellular proteins from the dentin matrix are likely involved in neurogenic responses to caries and could be exploited in clinical regenerative endodontics to promote reinnervation and enhance tissue regeneration.     

Effect of Odanacatib on the release of NTX (Amino Terminal Telopeptide) from LPS contaminated type I dentin collagen

ObjectiveTo evaluated the Odanacatib inhibitor treatment on lipopolysaccharide (LPS) contamination effect on cathepsin-K mediated dentin degradation by analysis of type I collagen C- and N-termini telopeptides.MethodsPulverized and disks of human dentin were demineralized and LPS contaminated, or stored in deionized water (DW) for 12 h. Samples were challenged with lactic acid (LA). Aliquots of dentin powder were treated with 1 mL Odanacatib or stored in DW for 30 min. Dentin collagen degradation was determined by sub-product release of C-terminal (ICTP and CTX) and N-terminal (NTX) telopeptides, normalized to total protein (tp) concentration (n = 3). Dentin matrix was evaluated for gravimetric (n = 8) and ultrastructural changes. Data were analyzed by Student t-test, one-way ANOVA and Tukey’s test (α = 5 %).ResultsLA incubation significantly increased telopeptide release compared with DW (p < 0.05). In untreated groups, significantly higher CTXtp, NTXtp telopeptide rates were observed for LA+LPS samples compared with DW (p < 0.01). Odanacatib significantly reduced ICTPtp, CTXtp, and NTXtp telopeptide release for LPS, LA, and LA+LPS conditions. In untreated groups, LPS and LA+LPS challenge significantly increased dentin weight loss (p = 0.02). Within each storage condition, Odanacatib treatment did not affect weight change (p > 0.05) of dentin disks.SignificanceThis study showed that LPS contamination resulted in significantly higher rates of NTX than CTX from dentin matrix. Odanacatib significantly reduced telopeptide release rates of LPS contaminated dentin matrix.     

Effect of biomimetic mineralization on enamel and dentin: A Raman and EDX analysis

ObjectiveTo investigate the effect of an experimental biomimetic mineralization kit (BIMIN) on the chemical composition and crystallinity of caries-free enamel and dentin samples in vitro.MethodsEnamel and dentin samples from 20 human teeth (10 for enamel; 10 for dentin) were divided into a control group without treatment and test samples with BIMIN treatment. Quantitative analysis of tissue penetration of fluoride, phosphate, and calcium was performed using energy-dispersive X-ray spectroscopy (EDX). Mineralization depth was measured by Raman spectroscopy probing the symmetric valence vibration near 960 cm⁻¹ as a marker for crystallinity. EDX data was statistically analyzed using a paired t-test and Raman data was analyzed using the Student’s t-test.ResultsEDX analysis demonstrated a penetration depth of fluoride of 4.10 ± 3.32 μm in enamel and 4.31 ± 2.67 μm in dentin. Calcium infiltrated into enamel 2.65 ± 0.64 μm and into dentin 5.58 ± 1.63 μm, while the penetration depths for phosphate were 4.83 ± 2.81 μm for enamel and 6.75 ± 3.25 μm for dentin. Further, up to 25 μm of a newly mineralized enamel-like layer was observed on the surface of the samples. Raman concentration curves demonstrated an increased degree of mineralization up to 5–10 μm into the dentin and enamel samples.SignificanceBiomimetic mineralization of enamel and dentin samples resulted in an increase of mineralization and a penetration of fluoride into enamel and dentin.     

Effect of Modified Nonequilibrium Plasma with Chlorhexidine Digluconate against Endodontic Biofilms In Vitro

IntroductionNonequilibrium plasma has been reported to effectively kill Enterococcus faecalis in endodontic biofilm compared with chlorhexidine digluconate (CHX). The purpose of this study was to evaluate the antimicrobial in vitro activity of modified nonequilibrium plasma with CHX against E. faecalis and multispecies biofilms on bovine dentin discs.MethodsSterile bovine dentin discs were incubated with E. faecalis or a mixture of bacteria from human dental root canal infections to form 1- and 3-week-old biofilms. The specimens were subjected to nonequilibrium plasma, modified nonequilibrium plasma with CHX, and 2% CHX for 2- and 5-minute exposure. After treatment, the biofilms were stained with viability dyes and examined by confocal laser scanning microscopy and 3-dimensional reconstruction analysis. The proportions of bacterial cells killed by the treatments were calculated.ResultsThe 3-dimensional reconstruction images showed that 1- and 3-week-old biofilms adhered to bovine dentin discs. The proportions of dead cells increased significantly with the longer exposure in each treatment group (P < .05). Modified nonequilibrium plasma was the most effective in killing bacteria in E. faecalis and multispecies biofilms at both 2 and 5 minutes (P < .05). No significant difference was detected between nonequilibrium plasma and CHX groups (P > .05). Significantly more cells were killed in 1-week-old biofilms than in 3-week-old biofilms in all groups (P < .05).ConclusionsThe modified nonequilibrium plasma killed more bacteria than conventional nonequilibrium plasma and 2% CHX in E. faecalis and multispecies endodontic biofilms in vitro and thus shows promise as an additional tool in infection control during endodontic treatment.     

Role of proteoglycans on the biochemical and biomechanical properties of dentin organic matrix

ObjectiveProteoglycans (PGs) are multifunctional biomacromolecules of the extracellular matrix of collagen-based tissues. In teeth, besides a pivotal regulatory role on dentin biomineralization, PGs provide mechanical support to the mineralized tissue and compressive strength to the biosystem. This study assessed enzymatic protocols for selective PGs removal from demineralized dentin to determine the roles of these biomacromolecules in the bulk mechanical properties and biostability of type I collagen.MethodsSelective removal of glycosaminoglycans chains (GAGs) and PGs from demineralized dentin was carried out by enzymatic digestion protocols using chondroitinase ABC (c-ABC) and trypsin (Try). A comprehensive study design included assessment of dentin matrix mass loss, biodegradability of the PGs/GAGs-depleted dentin matrix, ultimate tensile strength (UTS) and energy to fracture tests. Quantitative data was statistically analyzed by two-way and one-way ANOVA followed by the appropriate post hoc tests (α = 0.05).ResultsTransmission electron microscopy images show effective GAGs removal by c-ABC and Try and both enzymatic methods released statistically similar amounts of GAGs from the demineralized dentin. Try digestion resulted in about 25% dentin matrix mass loss and increased susceptibility to collagenolytic digestion when compared to c-ABC (p = 0.0224) and control (p = 0.0901). Moreover, PGs digestion by Try decreased the tensile strengths of dentin. Statistically lower energy to fracture was observed in c-ABC-treated dentin matrix.ConclusionsGAGs plays a pivotal role on tissue mechanics and anisotropy, while the core protein of PGs have a protective role on matrix biostability.     

Evaluation of pH and Calcium Ion Release of Calcium Hydroxide Pastes Containing Different Substances

IntroductionThe objective of this study was to evaluate the pH and calcium ion release of calcium hydroxide pastes associated with different substances.MethodsForty acrylic teeth with simulated root canals were divided into 4 groups according to the substance associated to the calcium hydroxide paste: chlorhexidine (CHX) in 2 formulations (1% solution and 2% gel), Casearia sylvestris Sw extract, and propylene glycol (control). The teeth with pastes and sealed coronal accesses were immersed in 10 mL deionized water. After 10 minutes, 24 hours, 48 hours, and 7, 15, and 30 days, the teeth were removed to another container, and the liquid was analyzed. Calcium ion release was measured by atomic absorption spectrophotometry, and pH readings were made with a pH meter. Data were analyzed statistically by analysis of variance and Tukey test (α = 0.05).ResultsCalcium analysis revealed significant differences (P < .05) for 1% CHX solution and 2% CHX gel at 10 minutes. After 24 hours, 2% CHX gel × Control and 2% CHX gel × 1% CHX solution differed significantly (P < .05). After 48 hours, there were significant differences (P < .05) for 2% CHX gel × Control and Extract × Control. No differences (P > .05) were observed among groups in the other periods. Regarding the pH, there were significant differences (P < .05) for 2% CHX gel × Control and 2% CHX gel × 1% CHX solution after 48 hours and for 2% CHX gel × Control after 15 days. In the other periods, no differences (P > .05) were observed among groups.ConclusionsAll pastes behaved similarly in terms of pH and calcium ion release in the studied periods.     

An in vitro evaluation of microleakage of resin based composites bonded to chlorhexidine-pretreated dentin by different protocols of a universal adhesive system

PurposeThis study compared microleakage of different resin based composite (RBC) materials bonded to dentin, after chlorhexidine (CHX) application, by different adhesion protocols of a universal adhesive system.MethodsClass V cavities were prepared on the buccal and lingual surfaces of 40 premolar teeth. The “etch-and-rinse” technique of a universal bond adhesive system (Single Bond Universal Adhesive) was used on buccal preparations, while the “self-etch” protocol was used on the lingual surfaces. Two RBCs, one bulk fill (Filtek Bulk Fill [FBF]) and one conventional (Filtek Z350 XT [Z350XT]), were used. Teeth were divided into two groups of 20 teeth each, 10 per each RBC (n = 10): (1) control; and (2) pretreatment with 2% CHX. For FBF groups, teeth were restored with a single increment; however, for Z350XT, a layering technique was used. Teeth were aged by thermo-cycling and prepared for microleakage testing. Dye penetration was evaluated and scored from 0 to 4. Data were analyzed at a significance level of P < 0.05.ResultsThe highest microleakage mean scores were found in the control group of the etched margins for both RBCs (2.80 ± 1.033 FBF and 2.10 ± 1.370 Z350XT). The CHX-pretreated group showed significantly lower microleakage than the control for FBF only (P = 0.008). No significant difference was found between groups for the “self-etch” protocol (χ² = 0.884, P = 0.08). No significant differences were found between FBF and Z350XT in all study groups (P > 0.2).ConclusionsWhen the “self-etch” protocol of the universal adhesive system was used, dentin microleakage was not affected by CHX-pretreatment when teeth were restored with bulk fill or conventional RBCs. In the “etch-and-rinse” protocol, CHX application improved the marginal seal before restoration with bulk fill material. However, in the absence of CHX, the “etch-and-rinse” protocol negatively affected marginal integrity.     

Novel bioactive adhesive containing dimethylaminohexadecyl methacrylate and calcium phosphate nanoparticles to inhibit metalloproteinases and nanoleakage with three months of aging in artificial saliva

ObjectivesThe objectives of this study were to: (1) develop a multifunctional adhesive via dimethylaminohexadecyl methacrylate (DMAHDM) and nanoparticles of amorphous calcium phosphate (NACP); and (2) investigate its ability to provide metalloproteinases (MMPs) deactivation and remineralization for long-term dentin bonding durability.MethodsDMAHDM and NACP were incorporated into Adper? Single Bond 2 Adhesive (SB2) at mass fractions of 5% and 20%, respectively. Degree of conversion and contact angle were measured. Endogenous MMP activity of the demineralized dentin beams, Masson’s trichrome staining, nano-indentation, microtensile bond strength and interfacial nanoleakage analyses were investigated after 24 h and 3 months of storage aging in artificial saliva.ResultsAdding DMAHDM and NACP did not compromise the degree of conversion and contact angle of SB2 (p > 0.05). DMAHDM and NACP incorporation reduced the endogenous MMP activity by 53 %, facilitated remineralization, and increased the Young’s modulus of hybrid layer by 49 % after 3 months of aging in artificial saliva, compared to control. For SB2 Control, the dentin bond strength decreased by 38 %, with greater nanoleakage expression, after 3 months of aging (p < 0.05). However, DMAHDM+NACP group showed no loss in bond strength, with much less nanoleakage, after 3 months of aging (p > 0.05).SignificanceDMAHDM+NACP adhesive greatly reduced MMP-degradation activity in demineralized dentin, induced remineralization at adhesive-dentin interface, and maintained the dentin bond strength after aging, without adversely affecting polymerization and dentin wettability. This new adhesive has great potential to help eliminate secondary caries, prevent hybrid layer degradation, and increase the resin-dentin bond longevity.     

Influence of Voxel Size and Filter Application in Detecting Second Mesiobuccal Canals in Cone-beam Computed Tomographic Images

IntroductionThis study assessed the influence of voxel size and filter application in detecting second mesiobuccal (MB2) canals in cone-beam computed tomographic (CBCT) images.MethodsUsing the OP300 CBCT system (Instrumentarium, Tuusula, Finland) and 3 voxel size protocols (80 μm, 125 μm, and 200 μm), we scanned 40 first molars: 20 with an MB2 canal and 20 without. All molars received silver palladium pins on the palatal root, whereas the non-MB2 molars were also filled with gutta-percha. Five oral radiologists assessed the presence of an MB2 canal under 3 filter application conditions: without filter, with sharpen 1 × filter, and with sharpen 2 × filter. Intra- and interobserver reproducibility was evaluated using the weighted kappa index. We compared the area under the receiver operating characteristic curves with SPSS Statistics v.20.0 (IBM Corp, Armonk, NY) using 2-way analysis of variance and the Tukey post hoc test with 5% significance level.ResultsOur analysis found median intra- and interobserver agreement values of 0.70 and 0.56, respectively. The 80-μm voxel with sharpen 1 × filter image group had the highest sensitivity, accuracy, and negative predictive values. As for specificity and positive predictive, the 80-μm voxel group without filter application presented the highest values. The areas under the receiver operating characteristic curve were higher in the 80-μm groups than in the 125-μm and 200-μm voxel size groups (P < .05). We found no differences among the filters used (P = .22) or for the filter–voxel size interactions (P = .88).ConclusionsA smaller voxel size increased the accuracy in detecting MB2 canals, whereas the enhancement filters did not.     

Effect of Long-term Exposure to Peptides on Mono- and Multispecies Biofilms in Dentinal Tubules

IntroductionThe aim of this study was to evaluate the antibiofilm effectiveness of 2% chlorhexidine (CHX) and peptides 1018 and DJK-5 used either alone or in a mixture (peptide and 2% CHX) against Enterococcus faecalis and multispecies biofilms in dentin canals after short-term and long-term exposure.MethodsOne hundred eighty dentin blocks were prepared and filled with E. faecalis or multispecies bacteria by centrifugation. Three-week-old biofilms in dentin were subjected to 2% CHX, DJK-5 (10 μg/mL), 1018 (10 μg/mL), DJK-5 + 2% CHX, or 1018 + 2% CHX for short-term (1 or 3 minutes), short-term exposure after 24 hours, and long-term exposure (24 hours of exposure). The antibacterial efficacy was determined by live/dead bacterial viability staining and confocal laser scanning microscopy.ResultsPeptide DJK-5 with or without CHX was the most effective agent against all the biofilms (P < .05), killing 77% of biofilm bacteria in 1 minute. No significant difference in bacterial killing was detected between the first 3 minutes of exposure (>81%) and after 24 hours of exposure (83%) to DJK-5 or DJK-5 + CHX. Chlorhexidine and peptide 1018 had a weaker antibiofilm effect than DJK-5, and their effect was time dependent (P < .05) with a maximum killing of 60% after 24 hours of exposure.ConclusionsPeptide DJK-5 alone and together with CHX had a rapid antibacterial effect against dentin infection. An additional antibacterial effect by CHX and peptide 1018 was achieved after a 24-hour long-term exposure.     

Relationship between microtensile bond strength and nanoleakage at the composite–dentin interface

ObjectivesThis study evaluated the relationship between microtensile bond strength (μTBS) and occurrence of nanoleakage at the resin–dentin interface using the same specimens.MethodsResin–dentin-bonded micro-specimens (sticks with a size of 300 μm × 300 μm × 8 mm) were prepared using one of two material combinations (group I: Syntac classic/Tetric Ceram Cavifil: n = 57; group II: Prime & Bond NT/Spectrum TPH: n = 52). After immersion of the micro-specimens in 0.1% rhodamine-B solution for 1 h, nanoleakage was imaged nondestructively using a confocal laser scanning microscope (CLSM). Then the specimens were subjected to a μTBS test.ResultsFor the influence of nanoleakage on μTBS with the Syntac classic/Tetric Ceram Cavifil group, the nonparametric Spearman correlation was 0.033 at p = 0.805. For the Prime & Bond NT/Spectrum TPH group, the nonparametric Spearman correlation was 0.077 at p = 0.584.SignificanceThe degree of nanoleakage had no influence on microtensile bond strength for the Syntac classic/Tetric Ceram Cavifil or for the Prime & Bond NT/Spectrum TPH group.