METHODS: Root canal preparation was performed using stainless steel K-files™ and F4 size protaper with irrigation protocols of 6% NaOCl + 2% CHX; 3.5% QIS; 2% QIS and sterile saline. Biofilms were prepared using E. faecalis adjusted and allowed to grow for 3 days, treated with irrigants, and allowed to grow for 7 days. AFM was performed and surface free energy calculated. MC3T3 cells were infected with endo irrigant treated E. faecalis biofilms. Raman spectroscopy of biofilms were performed after bacterial re-growth on root dentine and exposed to different irrigation protocols and collagen fibers analysed collagen fibers using TEM. Antimicrobial potency against E. faecalis biofilms and cytoxicity against 3T3 NIH cells were also. Resin penetration and MitoTracker green were also evaluated for sealer penetration and mitochondrial viability. Data were analysed using One-way ANOVA, principal component analysis and post-hoc Fisher's least-significant difference.
RESULTS: Elastic moduli were maintained amongst control (5.5 ± 0.9) and 3.5% QIS (4.4 ± 1.1) specimens with surface free energy higher in QIS specimens. MC3T3 cells showed reduced viability in 6%NaOCl+2%CHX specimens compared to QIS specimens. DNA/purine were expressed in increased intensities in control and 6% NaOCl + 2% CHX specimens with bands around 480-490 cm-1 reduced in QIS specimens. 3.5% QIS specimens showed intact collagen fibrillar network and predominantly dead bacterial cells in confocal microscopy. 3.5% QIS irrigant formed a thin crust-type surface layer with cytoplasmic extensions of 3T3NIH spread over root dentine. Experiments confirmed MitoTracker accumulation in 3.5% treated cells.
SIGNIFICANCE: Novel QIS root canal irrigant achieved optimum antimicrobial protection inside the root canals facilitating a toxic effect against the Enterococcus faecalis biofilm. Root dentine substrates exhibited optimum mechanical properties and there was viability of fibroblastic mitochondria.
METHODS: VE-TPGS was added to RF-solution, at RF/VE-TPGS (w/w) ratios of 0.125/0.250 and 0.125/0.500. Demineralized dentine beams were used (10wt.% phosphoric acid), rinsed using deionized-water and analysed using ELISA (Human MMP2 ELISA; Human CTSK/Cathepsin-K for MMP2 and Cathepsin K analysis). AFM of dentine collagen-fibrils structure was done before and after dentine specimens' placement in mineralization solution and tested after 14days in artificial saliva/collagenase (AS/Co) solution. The specimens were tested after 24h in mineralization solution for surface/bulk elastic modulus. Nano-indentation was carried out for each specimen on intertubular-dentine with lateral spacing of 400nm. Reduced elastic-modulus and nano-hardness were calculated and collagen content was determined using hydroxyproline-assay. Micro-Raman were performed. TEM was carried out to study structural variations of dentine-collagen in artificial-saliva (collagenase). Data were presented as mean±standard deviation and analyzed by SPSS v.15, by analysis of variance.
RESULTS: Synergetic effect of VE-TPGS was observed with RF through higher structural integrity of dentine collagen-fibrils shown by TEM/AFM. Superior surface/bulk mechanical stability was shown by nano-indentation/mechanical testing. Improvement in collagenase degradation resistance for hydroxyproline release was observed and lower endogenous-protease release of MMP-2/Cathepsin-K. Raman-analysis analysed chemical interactions between RF and collagen confirming structural-integrity of collagen fibrils after crosslinking. After 24h mineralization, AFM showed mineral depositions in close association with dentine-collagen fibrils with RF/VE-TPGS formulations.
SIGNIFICANCE: Potential synergetic effect of RF/VE-TPGS was observed by reflection of higher structural integrity and conformational-stability of dentine-collagen fibrils.
METHODS: The interaction between HIFU waves and dentin-surface in terms of structural, mechanical and chemical variations were investigated by SEM, TEM, AFM, nano-indentation and Raman-analysis. The bonding between HIFU-treated dentin and two-step, etch-and-rinse, adhesive was preliminary explored by characterizing dentin-bound proteases activities, resin-dentin interfacial morphology and bond-durability with HIFU exposure at different time-points of 60, 90 and 120 s compared to conventional acid-etching technique.
RESULTS: With the increase in HIFU exposure-time from 60-to-120 s, HIFU waves were able to remove the smear-layer, expose dentinal-tubules and creating textured/rough dentin surface. In addition, dentin surfaces showed a pattern of interlocking ribbon-like minerals-coated collagen-fibrils protruding from the underlaying amorphous dentin-background with HIFU exposure for 90 s and 120 s. This characteristic pattern of dentin-surface showing inorganic-minerals associated/aligned with collagen-fibrils, with 90-to-120 s HIFU-treatment, was confirmed by the Raman-analysis. HIFU-treated specimens showed higher nano-indentation properties and lower concentrations of active MMP-2 and Cathepsin-K compared to the acid-etched specimens. The resin-dentin bonded interface revealed the partial/complete absence of the characteristic hybrid-layer formed with conventional etch-and-rinse bonding strategy. Additionally, resin-infiltration and resin-tags formation were enhanced with the increase in HIFU exposure-time to 120 s. Although, all groups showed significant decrease in bond-strength after 12 months compared to 24 h storage in artificial saliva, groups exposed to HIFU for 90 s and 120 s showed significantly higher μTBS compared to the control acid-etched group.
SIGNIFICANCE: The implementation of HIFU-technology for dental hard-tissues treatment could be of potential significance in adhesive/restorative dentistry owing to its ability of controlled, selective and localised combined tissue alteration/ablation effects.
METHODS: As a preliminary work, water droplets of 1.5 μL were placed on the surfaces of hydroxyapatite (HA) discs of different densities. The water droplet profile was dynamically recorded every second over a period of 10 s using a contact angle meter to determine the relationship between sorptivity and density. To measure and calculate sorptivity on enamel surfaces, varnish was painted on the labial surface of 96 extracted caries-free human teeth, leaving two 1.4 ± 0.1 mm diameter circular exposed test sites. The specimens were randomly divided into 6 groups (n = 16) and subjected to 0(G0), 7(G7), 14(G14), 21(G21), 28(G28) and 35(G35) days of pH cycling, respectively. A 0.7 μL water droplet was placed on each exposed site and Optical Coherence Tomography was used to measure its height every 10 seconds for 2 min. Sorptivity was computed by considering sorption equations and Washburn's analysis of capillary kinetics and correction for evaporation was also performed. Micro-Computed Tomography scans of the specimens were obtained and delta Z (ΔZ) is the parameter used to measure mineral loss. ΔZ at 10 μm (ΔZ10) and 50 μm (ΔZ50) from the surface were calculated. One-way ANOVA and Post-hoc Tukey tests were used to compare sorptivity between groups and bivariate correlations were used to analyze the association between sorptivity and ΔZ.
RESULTS: Sorptivity was found to be inversely and linearly correlated with HA density with R2 value of 0.95. With enamel, there is a general trend of increase in mean sorptivity from G0 to G35, except for a decrease in G21. The same trends were observed for both ΔZ10 and ΔZ50. The decrease in sorptivity in G21 coincided with the presence of a surface hypermineralized layer in G21 samples. Post-hoc Tukey showed significant differences in mean sorptivity between G0 and G14, G0 and G21 as well as G14 and G21. Post-hoc Dunnett's T3 showed significant differences for ΔZ10 between G0 and G14 as well as G14 and G21. Significant correlation between mean sorptivity and ΔZ10 was detected with Pearson correlation coefficient of 0.461. For ΔZ50, post-hoc Tukey showed significant differences between G0 and G14 but no significant difference was detected between G14 and G21. No correlations were detected between mean sorptivity and ΔZ50.
SIGNIFICANCE: Sorptivity was found to be inversely and linearly correlated with HA density with R2 value of 0.95. With enamel, there is a general trend of increase in mean sorptivity from G0 to G35, except for a decrease in G21. The same trends were observed for both ΔZ10 and ΔZ50. The decrease in sorptivity in G21 coincided with the presence of a surface hypermineralized layer in G21 samples.
METHODS: Dentine surfaces were etched with 37% phosphoric acid, bonded with respective in vitro ethanol and acetone adhesives modified with (m/m, 0, 1%, 2% and 3% ribose), restored with restorative composite-resin, and sectioned into resin-dentine slabs and beams to be stored for 24h or 12 months in artificial saliva. Bond-strength testing was performed with bond failure analysis. Pentosidine assay was performed on demineralized ribose modified dentine specimens with HPLC sensitive fluorescent detection. The structural variations of ribose-modified dentine were analysed using TEM and human dental pulpal cells were used for cell viability. Three-point bending test of ribose-modified dentine beams were performed and depth of penetration of adhesives evaluated with micro-Raman spectroscopy. The MMP-2 and cathepsin K activities in ribose-treated dentine powder were also quantified using ELISA. Bond strength data was expressed using two-way ANOVA followed by Tukey's test. Paired T tests were used to analyse the specimens for pentosidine crosslinks. The modulus of elasticity and dentinal MMP-2 and cathepsin K concentrations was separately analyzed using one-way ANOVA.
RESULTS: The incorporation of RB in the experimental two-step etch-and-rinse adhesive at 1% improved the adhesive bond strength without adversely affecting the degree of polymerisation. The newly developed adhesive increases the resistance of dentine collagen to degradation by inhibiting endogenous matrix metalloproteinases and cysteine cathepsins. The application of RB to acid-etched dentine helps maintain the mechanical properties.
SIGNIFICANCE: The incorporation of 1%RB can be considered as a potential candidate stabilizing resin dentine bond.
METHODS: 15wt% of zirconia (ZrO2) as well as 30, 35, and 40wt% of beta-tricalcium phosphate (β-TCP) were compounded with PA 12, followed by the fabrication of filament feedstocks using a single screw extruder. The fabricated filament feedstocks were used to print the impact specimens. The melt flow rate, tensile properties of fabricated filament feedstocks, and 3D printed impact properties of the specimens were assessed using melt flow indexer, universal testing machine, and Izod pendulum tester, respectively. The microstructure of selected filament feedstocks and broken impact specimens were analysed using a field emission scanning electron microscope and universal testing machine. Human periodontal ligament fibroblast cells (HPdLF) were used to evaluate the cytotoxicity of the materials by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromid) (MTT) assay.
RESULTS: Hybrid ceramics filled PA 12 indicated sufficient flowability for FDM 3D printing. The tensile strength of hybrid ceramics filled PA 12 filament feedstocks slightly reduced as compared to unfilled PA 12. However, the tensile modulus and impact strength of hybrid ceramics filled PA 12 increased by 8%-31% and 98%-181%, respectively. A significant increase was also detected in the cell viability of the developed composites at concentrations of 12.5, 25, 50 and 100mg/ml.
SIGNIFICANCE: The newly developed hybrid ceramics filled PA 12 filament feedstock with improved properties is suitable for an FDM-based 3D printer, which enables the creation of patient-specific craniofacial implant at a lower cost to serve low-income patients.
METHODS: NIH 3T3 mouse fibroblasts were cultured in Dulbecco's Modified Eagle's Medium and incubated for 3 days. The cells (3×104) were seeded on the pulpal side of dentine discs and the occlusal side of the discs were treated with different cavity disinfectants: Group 1: de-ionized water (control); Group 2: 2% chlorhexidine (CHX); Group 3: 2% QAS; Group 4: 5% QAS, and Group 5: 10% QAS. Cell morphology of NIH 3T3 cells was examined using scanning electron microscopy (SEM) and cell viability was assessed using Trypan blue assay. The eluates were collected and applied on cells seeded in 24-well plates. The total protein production, alkaline phosphatase activity and deposition of mineralized nodules were evaluated after 7 and 14 days. Immunofluorescence staining was performed on the samples with primary antibodies of CD68+, CD80+, and CD163+ assessing the macrophage M1/M2 phenotypes. The macrophages were imaged using a confocal scanning light microscope with an excitation wavelength of 488nm.
RESULTS: No significant difference in cell viability (p<0.0001), total protein production (p<0.01) and mineralized nodule production (p<0.05) was found between 2% QAS and the control, which was significantly higher than 2% CHX, 5% and 10% QAS after 14 days. Alkaline phosphatase production of 2% QAS was significantly lower than the control (p<0.001), but higher than 2% CHX at 14 days. The M1/M2 macrophage ratio was also significantly lower in the 2% and 10% QAS groups (p<0.05) compared to the control and 2% CHX groups.
SIGNIFICANCE: The 2% QAS cavity disinfectant does not have cytotoxic effects on 3T3 NIH mouse fibroblast cells and the predominance of the anti-inflammatory phenotype after its application may stimulate healing and tissue repair.
MATERIALS AND METHODS: An experimental adhesive system based on bis-GMA, HEMA and hydrophobic monomer was doped with RF0.125 (RF - Riboflavin) or RF/VE-TPGS (0.25/0.50) and submitted to μTBS evaluation. Resin dentine slabs were prepared and examined using SEM and TEM. Adhesion force was analysed on ends of AFM cantilevers deflection. Quenched peptide assays were performed using fluorescence scanner and wavelengths set to 320nm and 405nm. Cytotoxicity was assessed using human peripheral blood mononuclear cell line. Molecular docking studies were carried out using Schrödinger small-molecule drug discovery suite 2018-2. Data from viable cell results was analyzed using one-way ANOVA. Bond strength values were analysed by two-way ANOVA. Nonparametric results were analyzed using a Kruskal-Wallis test at a 0.05 significance level.
RESULTS: RF/VE-TPGS0.25 groups showed highest bond strength results after 24-h storage in artificial saliva (p<0.05). RF/VE-TPGS0.50 groups showed increased bond strength after 12-months of ageing. RF/VE-TPGS modified adhesives showed appreciable presence of a hybrid layer. Packing fraction indicated solid angle profiles describing well sized density and topology relations for the RF/VE-TPGS adhesives, in particular with the RF/VE-TPGS0.50 specimens. Qualitative analysis of the phenotype of macrophages was prominently CD163+ in the RF/VE-TPGS0.50. Both the compounds showed favourable negative binding energies as expressed in terms of 'XP GScore'.
CONCLUSION: New formulations based on the incorporation of RF/VE-TPGS in universal adhesives may be of significant potential in facilitating penetration, distribution and uptake of riboflavin within the dentine surface.
METHODS: TPAu nanoparticles were fabricated from 0.31-g tetrachloroauric acid and 0.38-g of N-(2-mercaptopropionyl) glycine (2.4-mmol). Then co-dissolved using 35-mL of 6:1 methanol/acetic acid and mixed using NaBH4. EDC (0.3-M) was conjugated to TPAu nanoparticles at TPAU/EDC-0.25:1, and TPAU/EDC-0.5:1 treatment formulations ratios. Dentin specimens treated with 0.3-M EDC solution alone or left untreated were used as control. Nanoparticles formulations were characterized in term of particles morphology and size, Zeta potential, thermogravimetric analysis and small-angle X-ray scattering. Dentin substrates were characterized in term of TEM investigation, dentin proteases characterization, hydroxyproline liberation, elastic modulus measurement, Raman analysis and confocal microscopy viewing.
RESULTS: TEM evaluation of tiopronin protected gold nanoparticles dispersion revealed nano-clusters formations in both groups. However, based on our TEM measurements, the particle-size was ranging from ˜20 to 50 nm with spherical core-shape which were almost similar for both TPAu/EDC ratios (0.5:1 and 0.25:1). Zeta potential measurements indicate negative nanoparticles surface charge. SAXS profiles for both formulations, suggest a typical profile for uni-lamellar nanoparticles. Superior dentin collagen cross-linking effect was found with the TPAu/EDC nanoparticles formulations compared to the control and EDC treated groups.
SIGNIFICANCE: Cross-linking of dentin collagen using TPAu coupled with EDC through TPAu/EDC nanoparticles formulations is of potential significance in improving the biodegradation resistance, proteases inhibition, mechanical and structural stability of demineralized dentin substrates. In addition, the cross-linking effect is dependent on TPAu/EDC ratio, whereas higher cross-linking effect was found at TPAu/EDC ratio of 0.5:1.
METHODS: Root canal was prepared using stainless steel K-files™ and ProTaper™ and subjected to manual and ultrasonic irrigation using 6% NaOCl+2% CHX, 6% NaOCl+2% QAS and saline as control. For confocal-microscopy, Raman spectroscopy and SEM analysis before and after treatment, Enterococcus faecalis cultured for 7 days. Raman spectroscopy analysis was done across cut section of gutta percha/sealer-dentine to detect resin infiltration. Indentation of mechanical properties was evaluated using a Berkovich indenter. The contact angle of irrigants and surface free energy were evaluated. Mineralization nodules were detected through Alazarin red after 14 days.
RESULTS: Control biofilms showed dense green colonies. Majority of E. faecalis bacteria were present in biofilm fluoresced red in NaOCl+2% QAS group. There was reduction of 484cm-1 Raman band and its intensity reached lowest with NaOCl+2% QAS. There was an increase in 1350-1420cm-1 intensity in the NaOCl+2% CHX groups. Gradual decrease in 1639cm-1 and 1609cm-1 Raman signal ratios were seen in the resin-depth region of 17μm>, 14.1μm> and 13.2μm for NaOCl+2% QAS, NaOCl+2% CHX and control groups respectively. All obturated groups showed an intact sealer/dentine interface with a few notable differences. 0.771 and 83.5% creep indentation distance for NaOCl+2% QAS ultrasonic groups were observed. Highest proportion of polar component was significantly found in the NaOCl+2% QAS groups which was significantly higher as compared to other groups. Mineralized nodules were increased in NaOCl+2% QAS.
SIGNIFICANCE: Favorable antimicrobial and endodontic profile of the NaOCl+2% QAS solution might suggest clinical use for it for more predictable reduction of intracanal bacteria.
METHODS: HA having nanorods structure were synthetized using ultrasonication with precipitation method. HA nanorods were characterized by TEM for average-size/shape. Following phosphoric acid demineralization, dentine specimens were treated with HA-nanorods with/without subsequent HIFU exposure for 5 s, 10 s and 20 s then stored in artificial saliva for 1-month. Dentine specimens were characterized using different SEM and Raman spectroscopic techniques. In addition, the biochemical stability and HA-nanorods were examined using ATR-FTIR to observe attachment of nanoparticles. Also, surface nanoindentation properties were evaluated using AFM in tapping-mode.
RESULTS: HA-nanorods displayed well-defined, homogenous plate-like nanostructure. TEM revealed intact collagen-fibrils network structure with high density due to obliteration of interfibrillar spaces with clear evidence of remineralization in combined HA/HIFU treatment. With HA-nanorods treatment collagen-network structure was visible, consisting of fibrils interlaced into a compact pattern with evidence of minerals deposition. AFM investigation revealed clear mineral formation with the increase of HIFU exposure time. Bands associated with inorganic phase dominate well in HIFU exposed specimens with PO stretching within dentine mineral identified at 960 cm-1. Characteristic dentine structure for control and HIFU 20 s specimens is reflected as oscillatory mean Amide-I intensity with measurement giving a precise sinusoidal response of polarization angle β within dentinal tissue. Nanoindentation testing showed a gradual significant increase in elastic-modulus with the increase in HIFU exposure time after 1-month storage. FTIR spectrum of the HIFU exposed dentine displayed bands at 1650 cm-1, 1580 cm-1 and 1510 cm-1 that can be attributed to Amide-I, II and III.
SIGNIFICANCE: The synergetic effect of HIFU exposure on remineralization potential of demineralized dentine-matrix following nano-hydroxyapatite treatment was revealed. This synergetic effect is dependent on HIFU exposure time.
METHODS: Root canals were instrumented and randomly divided into the following groups: irrigation with saline, 6% NaOCl (sodium hypochlorite), 6% NaOCl+2% CHX (Chlorhexidine), 2% CHX, 0.5% k21/E (k21 - quaternary ammonium silane) and 1% k21/E. E. faecalis were grown (3-days) (1×107CFU mL-1), treated, and further cultured for 11-days. Specimens were subjected to SEM, confocal and Raman analysis and macrophage vesicles characterized along with effect of lipopolysaccharide treatment. 3T3 mouse-fibroblasts were cultured for alizarin-red with Sortase-A active sites and Schrödinger docking was performed. TEM analysis of root dentin substrate with matrix metalloproteinases profilometry was also included. A cytotoxic test analysis for cell viability was measured by absorbance of human dental pulp cells after exposure to different irrigant solutions for 24h. The test percentages have been highlighted in Table 1.
RESULTS: Among experimental groups, irrigation with 0.5% k21/E showed phase separation revealing significant bacterial reduction and lower phenylalanine 1003cm-1 and Amide III 1245cm-1 intensities. Damage was observed on bacterial cell membrane after use of k21/E. No difference in exosomes distribution between control and 0.5%k21/E was observed with less TNFα (*p<0.05) and preferential binding of SrtA. TEM images demonstrated integrated collagen fibers in control and 0.5%k21/E specimens and inner bacterial membrane damage after k21/E treatment. The k21 groups appeared to be biocompatible to the dental pulpal cells grown for 24h.
SIGNIFICANCE: Current investigations highlight potential advantages of 0.5% k21/E as irrigation solution for root canal disinfection.
METHODS: N-TiO2 powder was manufactured by calcining commercial TiO2 with urea. Free radical release from the N-TiO2 powder under visible light irradiation was analysed using UV-Vis spectrophotometry. The N-TiO2 powder was incorporated into a dental resin and the photocatalytic activity assessed using a dye under both visible light and dark conditions. Using XTT assay to measure the cellular metabolic activity, the antibacterial properties of the N-TiO2 /resin composite discs were tested using Streptococcus mutans.
RESULTS: Doping nitrogen of TiO2 resulted in a band gap shift towards the visible light spectrum, which enabled the powder to release reactive oxygen species when exposed to visible light. When incorporated into a dental resin, the N-TiO2/resin composite still demonstrated sustained release of reactive oxygen species, maintaining its photocatalytic activity and showing an antibacterial effect towards Streptococcus mutans under visible light conditions.
SIGNIFICANCE: N-TiO2 filled resin composite shows great promise as a potential aesthetic resin based adhesive for orthodontic bonding.
METHODS: A total of 40 Tetric EvoCeram™ resin composite specimens against either a Lava™ Plus zirconia antagonist (n=20) or IPS e.max Press lithium disilicate antagonist (n=20) were prepared for the study. The surface roughness profiles of each resin composite before and after an in-vitro simulated chewing test were analysed using a 3D profilometer and Talymap software. After the simulated chewing, the surface profiles of representative Tetric EvoCeram specimens from each group were analysed using scanning electron microscopy. Independent t-test and paired t-test were used for statistical analysis.
RESULTS: For both lithium disilicate and zirconia groups, all surface roughness parameters (Ra, Rt, Sa, Sq,) of Tetric EvoCeram were significantly higher post-chewing compared to pre-chewing (p<0.05); the post-chewing surface roughness parameters of Tetric EvoCeram for the lithium disilicate group were significantly higher (p<0.05) than in the zirconia group.
SIGNIFICANCE: This chewing simulation test showed that Tetric EvoCeram composites exhibited a rougher surface when opposing lithium disilicate ceramic compared to opposing zirconia ceramic.
METHODS: PMMA disks containing GO (0.01, 0.05, 0.1, or 0.5 wt%) were subjected to a biaxial flexural test to determine the Weibull parameters (m: modulus of Weibull; σ0: characteristic strength; n = 30 at 1 MPa/s) and slow crack growth (SCG) parameters (n: subcritical crack growth susceptibility coefficient, σf0: scaling parameter; n = 10 at 10-2, 10-1, 101, 100 and 102 MPa/s). Strength-probability-time (SPT) diagrams were plotted by merging SCG and Weibull parameters.
RESULTS: There was no significant difference in the m value of all materials. However, 0.5 GO presented the lowest σ0, whereas all other groups were similar. The lowest n value obtained for all GO-modified PMMA groups (27.4 for 0.05 GO) was higher than the Control (15.6). The strength degradation predicted after 15 years for Control was 12%, followed by 0.01 GO (7%), 0.05 GO (9%), 0.1 GO (5%), and 0.5 GO (1%).
SIGNIFICANCE: The hypothesis was partially accepted as GO increased PMMA's fatigue resistance and lifetime but did not significantly improve its Weibull parameters. GO added to PMMA did not significantly affect the initial strength and reliability but significantly increased PMMA's predicted lifetime. All the GO-containing groups presented higher resistance to fracture at all times analyzed compared with the Control, with the best overall results observed for 0.1 GO.