GAPD and tubulin are suitable internal controls for qPCR analysis of oral squamous cell carcinoma cell lines

The selection of housekeeping genes is critical for gene expression studies. To address this issue, four candidate housekeeping genes, including several commonly used ones, were investigated in oral squamous cell carcinoma cell lines. A simple quantitative RT-PCR approach was employed by comparing relative expression of the four candidate genes within two cancerous cell tines (HN6 and HN31) and one noncancerous cell tine (HaCaT) treated or not with EGF and TGF-beta 1. Data were analyzed using ANOVA followed by the NormFinder software program. On this basis, stability of the candidate housekeeping genes was ranked and non statistical differences were found using ANOVA test. On the other hand, the NormFinder was able to show that GAPD and TUBB presented the less variable results, representing appropriated housekeeping genes for the samples and conditions anatyzed. In conclusion, this study suggests that the GAPD and the TUBB represent adequate normalizers for gene profiting studies in OSCC cell. tines, covering, respectively, high and low expression levels genes. (C) 2008 Elsevier Ltd. All rights reserved.

Long-term stability of dentin matrix following treatment with various natural collagen cross-linkers

Objectives: Collagen disorganization is one of the main degradation patterns found in unsuccessful adhesive restorations. The hypothesis of this study was that pretreatment using natural collagen cross-linking agents rich in proanthocyanidin (PA) would improve mechanical properties and stability over time of the dentin collagen and, thus, confer a more resistant and lasting substrate for adhesive restorations. Methods: PA-based extracts, from grape seed (GSE), cocoa seed (CSE), cranberry (CRE), cinnamon (CNE) and acai berry (ACE) were applied over the demineralized dentin. The apparent elastic modulus (E) of the treated dentin collagen was analyzed over a 12 month period. Specimens were immersed in the respective solution and E values were obtained by a micro-flexural test at baseline, 10, 30, 60, 120 and 240 min. Samples were stored in artificial saliva and re-tested after 3, 6 and 12 months. Data was analyzed using ANOVA and Tukey test. Results: GSE and CSE extracts showed a time-dependent effect and were able to improve [240 min (MPa): GSE = 108.96 +/- 56.08: CSE = 59.21 +/- 24.87] and stabilize the E of the organic matrix [12 months (MPa): GSE = 40.91 +/- 19.69; CSE = 42.11 +/- 13.46]. CRE and CNE extracts were able to maintain the E of collagen matrices constant over 12 months [CRE = 11.17 +/- 7.22; CNE = 9.96 +/- 6.11; MPa]. ACE (2.64 +/- 1.22 MPa) and control groups immersed in neat distilled water (1.37 +/- 0.69 MPa) and ethanol-water (0.95 +/- 0.33 MPa) showed no effect over dentin organic matrix and enable their degradation and reduction of mechanical properties. Significance: Some PA-based extracts were capable of improving and stabilizing collagen matrices through exogenous cross-links induction. (C) 2011 Elsevier Ltd. All rights reserved.

EFFECT OF METAL PRIMERS ON MICROTENSILE BOND STRENGTH BETWEEN ZIRCONIA AND RESIN CEMENTS

Statement of problem. There are no established clinical procedures for bonding zirconia to tooth structure using resin cements. Purpose. The purpose of this study was to evaluate the influence of metal primers, resin cements, and aging on bonding to zirconia. Material and methods. Zirconia was treated with commercial primers developed for bonding to metal alloys (Metaltite, Metal Primer II, Alloy Primer or Totalbond). Non-primed specimens were considered as controls. One-hundred disk-shaped specimens (19 x 4 mm) were cemented to composite resin substrates using Panavia or RelyX Unicem (n=5). Microtensile bond strength specimens were tested after 48 hours and 5 months (150 days), and failure modes were classified as type 1 (between ceramic/cement), 2 (between composite resin/cement) or 3 (mixed). Data were analyzed by 3-way ANOVA and Multiple Comparison Tukey test (alpha=.05). Results. The interactions primer/luting system (P=.016) and luting system/storage time (P=.004) were statistically significant. The use of Alloy Primer significantly improved the bond strength of RelyX Unicem (P<.001), while for Panavia, none of the primers increased the bond strength compared to the control group. At 48 hours, Panavia had statistically higher bond strength (P=.004) than Unicem (13.9 +/- 4.4MPa and 10.2 +/- 6.6MPa, respectively). However, both luting systems presented decreasing, statistically similar; values after aging (Panavia: 3.6 +/- 2.2MPa; Unicem: 6.1 +/- 5.3MPa). At 48 hours, Alloy Primer/Unicem had the lowest incidence of type 1 failure (8%). After aging, all the groups showed a predominance of type 1 failures. Conclusions. The use of Alloy Primer improved bond strength between RelyX Unicem and zirconia. Though the initial values obtained with Panavia were significantly higher than RelyX Unicem, after aging, both luting agents presented statistically similar performances. (J Prosthet Dent 2011;105:296-303)

Effects of adhesive temperature on the early and 6-month dentin bonding

Objectives: The aim of this study was to test the effect of adhesive temperature on the bond strength to dentin (mu TBS) and silver nitrate uptake (SNU) of an ethanol/water (Adper Single Bond 2 [SB]) and an acetone-based (Prime&Bond 2.1 [PB]) etch-and-rinse adhesive system. Methods: The bottles of each adhesive were kept in various temperatures (5 degrees C, 20 degrees C, 37 degrees C and 50 degrees C) for 1 h previously to its application in the occlusal demineralized dentin of 40 molars. Bonded sticks (0.8 mm(2)) were tested in tension (0.5 mm/min) immediately (IM) or after 6 months (6 M) of water storage. Two bonded sticks from each hemi-tooth were immersed in silver nitrate and analyzed by SEM. Data were analyzed by two-way repeated measures ANOVA and Tukey`s test (alpha = 0.05). Results: No significant difference in mu TBS was detected for both adhesives at 5 degrees C and 20 degrees C. The highest bond strength for PB was observed in the 37 degrees C group while for SB it was in the 50 degrees C. Significant reductions of bond strengths were observed for PB at 37 degrees C and SB at 50 degrees C after 6 M of water storage. Silver nitrate deposition was seen in all hybrid layers, irrespective of the group. Lower silver nitrate deposition (water trees) in the adhesive layer was seen for PB and SB at higher temperatures. Conclusions: The heating or refrigeration of the adhesives did not improve their resin-dentin bond resistance to water degradation over time. (C) 2009 Elsevier Ltd. All rights reserved.

Can the durability of one-step self-etch adhesives be improved by double application or by an extra layer of hydrophobic resin?

Objectives: This study evaluated the immediate and 6-month resin-dentin mu-bond strength (mu TBS) of one-step self-etch systems (Adper Prompt L-Pop [AD] 3M ESPE; Xeno III [XE] Dentsply De Trey; iBond [iB] Heraeus Kulzer) under different application modes. Materials and methods: Dentin oclusal surfaces were exposed by grinding with 600-grit SiC paper. The adhesives were applied according to the manufacturer`s directions [MD], or with double application of the adhesive layer [DA] or following the manufacturer`s directions plus a hydrophobic resin layer coating [HL]. After applying the adhesive resins, composite crowns were built up incrementally. After 24-h water storage, the specimens were serially sectioned in ""x"" and ""y"" directions to obtain bonded sticks of about 0.8 mm 2 to be tested immediately [IM] or after 6 months of water storage [6M] at a crosshead speed of 0.5 mm/min. The data from each adhesive was analyzed by a two-way repeated measures ANOVA (mode of application vs. storage time) and Tukey`s test (alpha = 0.05). Results: The adhesives performed differently according to the application mode. The DA and HL either improved the immediate performance of the adhesive or did not differ from the MD. The resin-dentin bond strength values observed after 6 months were higher when a hydrophobic resin coat was used than compared to those values observed under the manufacturer`s directions. Conclusions: The double application of one-step self-etch system can be safety performed however the application of an additional hydrophobic resin layer can improve the immediate resin-dentin bonds and reduce the degradation of resin bonds over time. (c) 2008 Elsevier Ltd. All rights reserved.

Effect of prolonged application times on the durability of resin-dentin bonds

Objectives. To examine the effect of prolonged application time on the early and 3-year resin-dentin microtensile bond strength. Methods. Water/ethanol (Single Bond [SB]) and acetone-based systems (One Step [OS]) were employed. A flat superficial dentin surface was exposed in third human molars by wet abrasion. The adhesives were applied to a delimited area of 52 mm(2) on wet surfaces, for 40, 90, 150 and 300s. Four teeth were assigned for each experimental condition. Composite build-ups were constructed incrementally After water storage at 37 degrees C for 24 h, teeth were sectioned to obtain sticks with cross-sectional areas of 0.8 mm(2) to be tested in tension (0.5 mm/min) either immediately (IM) or after 3 years (3Y) of water storage. The microtensile bond strength (mu TBS) values were analyzed by two way repeated measures ANOVA and Tukey`s test (alpha = 0.05). Results. The 90- and 150-s groups achieved the highest IM mu TBS for OS (p < 0.01). For SB, the highest IM mu TBS values were observed after 300-s application (p < 0.01). Significant decreases in mu TBS were observed for OS in the 40- and 90-s groups after 3Y, except for the 150-s group. With regard to SB, after 3Y significant drops in mu TBS values were observed for the 40- and 150-s groups, except for the 300-s group. Significance. Prolonged application times can increase the immediate LTBS of two-step etch-and-rinse adhesive systems and make the adhesive layer more stable over time. (c) 2007 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Do Low-shrink Composites Reduce Polymerization Shrinkage Effects?

Progress in polymer science has led to continuous reduction of polymerization shrinkage, exemplified by a new generation of ""low-shrink composites"". The common inference that shrinkage stress effects will be reduced in teeth restored with such restoratives with lower shrinkage was tested in extracted human premolars. Mesio-occluso-distal slot-shaped cavities were cut and restored with a conventional (SupremePlus) or low-shrink (RefleXions, Premise, Kalore, and LS) composite (N = 5). We digitized the coronal surfaces before and 10 min after restoration to determine cuspal deflection from the buccal and lingual volume change/area. We also determined the main properties involved (total shrinkage, post-gel shrinkage, degree of conversion, and elastic modulus), as well as microleakage, to verify adequate bonding. It was shown that, due to shrinkage stresses, buccal and lingual surfaces pulled inward after restoration (9-14 microns). Only Kalore and LS resulted in significantly lower tooth deformation (ANOVA/Student-Newman-Keuls post hoc, p = 0.05). The other two low-shrink composites, despite having the lowest and highest total shrinkage values, did not cause significant differences in cuspal deflection. Deflection seemed most related to the combination of post-gel shrinkage and elastic modulus. Therefore, even for significantly lower total shrinkage values, shrinkage stress is not necessarily reduced.

Micro-Shear Bond Strength of Different Adhesives to Human Dental Enamel

The aim of this study was to evaluate the micro-shear bond strength of 5 adhesive systems to enamel, one single-bottle acid-etch adhesive (O), two self-etching primers (P) and two all-in-one self-etching adhesives (S). Method: Sixty premolar enamel surfaces (buccal or lingual) were ground flat with 400- and 600-grit SiC papers and randomly divided into 5 groups (n=12), according to the adhesive system.. SB2 - Single Bond 2 (O); CSE - Clearfil SE Bond (P); ADS - AdheSE (P); PLP - Adper Prompt L-Pop (S); XE3 - Xeno III (S). Tygon tubing (inner diameter of 0.8mm) restricted the bonding area to obtain the resin composite (Z250) cylinders. After storage in distilled water at 37 degrees C for 24h and thermocycling, micro-shear testing was performed (crosshead speed of 0.5mm/min). Data were submitted to one-way ANOVA and Tukey test (a=5%). Samples were also subjected to stereomicroscopic and SEM evaluations after micro-shear testing. Mean bond strength values (MPa +/- SD) and the results of Tukey test were: SB2: 36.36(+/- 3.34)a; ADS: 33.03(+/- 7.83)a; XE3: 32.76(+/- 5.61)a; CSE: 30.61(+/- 6.68)a; PLP: 22.17(+/- 6.05)b. Groups with the same letter were not statistically different. It can be concluded that no significant difference was there between SB2, ADS, XE3 and CSE, in spite of different etching patterns of these adhesives. Only PLP presented statistically lower bond strengths compared with others. J Clin Pediatr Dent 35(3): 301-304, 2011

Polymerization stress, shrinkage and elastic modulus of current low-shrinkage restorative composites

Objective. To compare currently available low-shrinkage composites with others regarding polymerization stress, volumetric shrinkage (total and post-gel), shrinkage rate and elastic modulus. Methods. Seven BisGMA-based composites (Durafill/DU, Filtek Z250/FZ, Heliomolar/HM, Aelite LS Posterior/AP, Point 4/P4, Filtek Supreme/SU, ELS/EL), a silorane-based (Filtek LS, LS), a urethane-based (Venus Diamond, VD) and one based on a dimethacrylate-derivative of dimer acid (N`Durance, ND) were tested. Polymerization stress was determined in 1-mm high specimens inserted between two PMMA rods attached to a universal testing machine. Total volumetric shrinkage was measured using a mercury dilatometer. Maximum shrinkage rate was used as a parameter of the reaction speed. Post-gel shrinkage was measured using strain-gages. Elastic modulus was obtained by three-point bending. Data were submitted to one-way ANOVA/Tukey test (p = 0.05), except for elastic modulus (Kruskal-Wallis). Results. Composites ranked differently for total and post-gel shrinkage. Among the materials considered as ""low-shrinkage"" by the respective manufacturers, LS, EL and VD presented low post-gel shrinkage, while AP and ND presented relatively high values. Polymerization stress showed a strong correlation with post-gel shrinkage except for LS, which presented high stress. Elastic modulus and shrinkage rate showed weak relationships with polymerization stress. Significance. Not all low-shrinkage composites demonstrated reduced polymerization shrinkage. Also, in order to effectively reduce polymerization stress, a low post-gel shrinkage must be associated to a relatively low elastic modulus. (C) 2010 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Effects of Ultramorphological Changes on Adhesion to Lased Dentin-Scanning Electron Microscopy and Transmission Electron Microscopy Analysis

Dentin irradiation with erbium lasers has been reported to alter the composite resin bond to this treated surface. There is still a lack of studies reporting the effect of erbium lasers on dentin organic content and elucidating how laser treatment could interfere in the quality of the resin-dentin interface. This study aimed to evaluate the effect of erbium laser irradiation on dentin morphology and microtensile bond strength (lTBS) of an adhesive to dentin. Seventy-two dentin disks were divided into nine groups (n = 8): G1-Control (600-grit SiC paper); Er:YAG groups: G2-250 mJ/4 Hz; G3-200 mJ/4 Hz; G4-180 mJ/10 Hz; G5-160 mJ/10 Hz; Er, Cr:YSGG groups: G6-2 W/20 Hz; G7-2.5 W/20 Hz; G8-3 W/20 Hz; G9-4 W/20 Hz. Specimens were processed for cross-sectional analysis by scanning electron microscopy (SEM) (n = 3), transmission electron microscopy (TEM) (n = 2), and adhesive interface (n = 3). Forty-five dentin samples (n = 5) were restored and submitted to lTBS testing. ANOVA (alpha = 5%) revealed that G1 presented the highest lTBS values and irradiated groups did not differ from each other. TEM micrographs showed a superficial layer of denatured collagen fibrils. For SEM micrographs, it was possible to verify the laser effects extending to dentin subsurface presenting a rough aspect. Cross-sectional dentin micrographs of this hybridized surface revealed a pattern of modified tags with ringlike structures around it. This in vitro study showed that erbium laser irradiation interacts with the dental hard tissue resulting in a specific morphological pattern of dentin and collagen fibrils that negatively affected the bond strength to composite resin. Microsc. Res. Tech. 74:720-726, 2011. (C) 2010 Wiley-Liss, Inc.

Comparison of dentin root canal permeability and morphology after irradiation with Nd:YAG, Er:YAG, and diode lasers

The aim of this study was to compare the effects of Nd:YAG, Er:YAG, and diode lasers on the morphology and permeability of root canal walls. The three laser wavelengths mentioned interact differently with dentin and therefore it is possible that the permeability changes caused will determine different indications during endodontic treatment. Twenty-eight human single-rooted teeth were instrumented up to ISO 40 and divided into four groups: group C, control (GC), non-laser irradiated; group N (GN), irradiated with Nd:YAG laser; group E (GE), with Er:YAG laser and group D (GD) with diode laser. After that, the roots were filled with a 2% methylene blue dye, divided into two halves and then photographed. The images were analyzed using Image J software and the percentage of dye penetration in the cervical, middle, and apical root thirds were calculated. Additional scanning electron microscopy (SEM) analyses were also performed. The analysis of variance (ANOVA) showed significant permeability differences between all groups in the middle and cervical thirds (p < 0.05). The Tukey test showed that in the cervical third, GN presented means of dye penetration statistically significantly lower than all of the other groups. In the middle third, GE and GD showed statistically higher dye penetration means than GC and GN. SEM analysis showed melted surfaces for GN, clean wall surfaces with open dentinal tubules for GE, and mostly obliterated dentinal tubules for GD. Er:YAG (2,094 nm) laser and diode laser (808 nm) root canal irradiation increase dentinal permeability and Nd:YAG (1,064 nm) laser decreases dentin permeability, within the studied parameters.

Long-term effect of carbodiimide on dentin matrix and resin-dentin bonds

Objectives: To characterize the interaction of 1-Ethyl-3-[3-dimethylaminopropyl] carbodiimide Hydrochloride (EDC) with dentin matrix and its effect on the resin-dentin bond. Methods: Changes to the stiffness of demineralized dentin fragments treated with EDC/N-hydroxysuccinimide (NHS) in different solutions were evaluated at different time points. The resistance against enzymatic degradation was indirectly evaluated by ultimate tensile strength (UTS) test of demineralized dentin treated or not with EDC/NHS and subjected to collagenase digestion. Short- and long-term evaluations of the strength of resin-dentin interfaces treated with EDC/NHS for 1 h were performed using microtensile bond strength (mu TBS) test. All data (MPa) were individually analyzed using ANOVA and Tukey HSD tests (alpha = 0.05). Results: The different exposure times significantly increased the stiffness of dentin (p < 0.0001, control-5.15 and EDC/NHS-29.50), while no differences were observed among the different solutions of EDC/NHS (p = 0.063). Collagenase challenge did not affect the UTS values of EDC/NHS group (6.08) (p > 0.05), while complete degradation was observed for the control group (p = 0.0008, control-20.84 and EDC/NHS-43.15). EDC/NHS treatment did not significantly increase resin-dentin mu TBS, but the values remained stable after 12 months water storage (p < 0.05). Conclusions: Biomimetic use of EDC/NHS to induce exogenous collagen cross-links resulted in increased mechanical properties and stability of dentin matrix and dentin-resin interfaces. (C) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 94B: 250-255, 2010.

Influence of matrix composition on polymerization stress development of experimental composites

Objective. Stress development at the tooth/restoration interface is one of the most important reasons for failure of adhesive restorations. The aim of this study was to evaluate the influence of BisGMA/TEGDMA (B/T) and UDMA/TEGDMA (U/T) ratios on polymerization stress (PS) and on the variables related to its development: degree of conversion (DC), polymerization maximum rate (Rp(max)), volumetric shrinkage (VS), elastic modulus (E), stress relaxation (SR) and viscosity of experimental composites. Method. Composites were formulated containing B/T or U/T in mol% ratios of 2: 8, 3: 7, 4: 6, 5: 5, 6: 4, 7: 3 and 8: 2, and 15 wt% of fumed silica. PS was determined with a universal testing machine. VS was measured with a linometer. E and SR were obtained in three-point bending. DC and Rp(max) were determined by real time NIR spectroscopy and viscosity was measured in viscometer. Data were submitted to one-way ANOVA, Tukey test (alpha = 0.05%) and regression analyses. Results. PS, VS, E and DC decreased and viscosity and Rp(max) increased with base monomer content in both series. PS showed strong correlation with VS, DC and viscosity. PS, VS and DC were higher and viscosity was lower for UDMA-based materials. Significance. Reduced viscosity, kinetics parameters and molecular characteristics led UDMA-based composites to elevated conversion and relatively lower PS at lower TEGDMA contents, compared to B/T composites. (C) 2010 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Microshear Bond Strength of Self-etching Systems Associated with a Hydrophobic Resin Layer

Purpose: To evaluate in vitro the microshear bond strength of adhesive systems applied to dentin according to manufacturers` instructions, associated or not with a hydrophobic layer of unfilled resin. Materials and Methods: Six self-etching adhesives (Clearfil SE Bond, Kuraray Medical; AdheSE, lvoclar Vivadent; Xeno III, Dentsply; I Bond, Heraeus-Kulzer; Bond Force, Tokuyama; Futurabond DC, Voco) were tested. The labial dentin of sixty bovine incisors was exposed, and the teeth were divided into two groups according to the application or not of an extra hydrophobic resin layer (Scotchbond Multi Purpose Plus, bottle 3). Six composite cylinders (Filtek Z250, 3M ESPE) were built up on each treated surface. Specimens were stored in distilled water at 37 C for 24 h and then subjected to the microshear bond strength test in a universal testing machine at a crosshead speed of 0.5 mm/min. Microshear bond strength values were analyzed by 2-way ANOVA and Tukey`s post-hoc test. Failure mode was determined using a stereomicroscope under 20X magnification. Results: The application of the hydrophobic resin layer did not affect bond strength, except for AdheSE. However, the bond strengths with the hydrophobic layer were similar among the six tested systems (Clearfil: 17.1 +/- 7.9; AdheSE: 14.5 +/- 7.1; Xeno III: 12.8 +/- 7.7; I Bond: 9.5 +/- 5.8; Bond Force: 17.5 +/- 4.1; Futurabond: 7.7 +/- 2.3). When used as recommended by the manufacturers, Bond Force presented statistically higher bond strength than AdheSE and I Bond (p < 0.05) (Clearfil 10.4 +/- 4.9; AdheSE 1.6 +/- 1.6; Xeno III: 9.0 +/- 3.8; I Bond: 3.0 +/- 1.5; Bond Force: 14 +/- 3.9; Futurabond: 8.8 +/- 3.8). Failure mode was predominantly adhesive. Conclusion: The bond strength of the self-etching systems tested was not significantly affected by the application of a hydrophobic layer, but a significant improvement was observed in AdheSE.

One-year stability of resin-dentin bonds created with a hydrophobic ethanol-wet bonding technique

Dentin bonding performed with hydrophobic resins using ethanol-wet bonding should be less susceptible to degradation but this hypothesis has never been validated. Objectives. This in vitro study evaluated stability of resin-dentin bonds created with an experimental three-step BisGMA/TEGDMA hydrophobic adhesive or a three-step hydrophilic adhesive after one year of accelerated aging in artificial saliva. Methods. Flat surfaces in mid-coronal dentin were obtained from 45 sound human molars and randomly divided into three groups (n = 15): an experimental three-step BisGMA/TEGDMA hydrophobic adhesive applied to ethanol (ethanol-wet bonding-GI) or water-saturated dentin (water-wet bonding-GII) and Adper Scotchbond Multi-Purpose [MP-GIII] applied, according to manufacturer instructions, to water-saturated dentin. Resin composite crowns were incrementally formed and light-cured to approximately 5 mm in height. Bonded specimens were stored in artificial saliva at 37 degrees C for 24h and sectioned into sticks. They were subjected to microtensile bond test and TEM analysis immediately and after one year. Data were analyzed with two-way ANOVA and Tukey tests. Results. MP exhibited significant reduction in microtensile bond strength after aging (24 h: 40.6 +/- 2.5(a); one year: 27.5 +/- 3.3(b); in MPa). Hybrid layer degradation was evident in all specimens examined by TEM. The hydrophobic adhesive with ethanol-wet bonding preserved bond strength (24 h: 43.7 +/- 7.4(a); one year: 39.8 +/- 2.7(a)) and hybrid layer integrity, with the latter demonstrating intact collagen fibrils and wide interfibrillar spaces. Significance. Coaxing hydrophobic resins into acid-etched dentin using ethanol-wet bonding preserves resin-dentin bond integrity without the adjunctive use of MMPs inhibitors and warrants further biocompatibility and patient safety`s studies and clinical testing. (C) 2009 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.