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Journal of Prosthetic Dentistry

Influence of silver nanoparticles on the resin-dentin bond strength and antibacterial activity of a self-etch adhesive system

  • Jia Wang
    Affiliations
    Attending Physician, Department of Endodontics and Operative Dentistry, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China

    College of Stomatology, Shanghai Jiao Tong University, Shanghai, PR China

    National Center for Stomatology, National Clinical Research Center for Oral Disease and Shanghai Key Laboratory of Stomatology, Shanghai, PR China
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  • Wei Jiang
    Affiliations
    Associate Chief Physician, Department of Endodontics and Operative Dentistry, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China

    College of Stomatology, Shanghai Jiao Tong University, Shanghai, PR China

    National Center for Stomatology, National Clinical Research Center for Oral Disease and Shanghai Key Laboratory of Stomatology, Shanghai, PR China
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  • Jingping Liang
    Affiliations
    Professor, Department of Endodontics and Operative Dentistry, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China

    College of Stomatology, Shanghai Jiao Tong University; National Center for Stomatology, National Clinical Research Center for Oral Disease and Shanghai Key Laboratory of Stomatology, Shanghai, PR China
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  • Shujun Ran
    Correspondence
    Corresponding author: Dr Shujun Ran, Department of Endodontics and Operative Dentistry, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, College of Stomatology, Shanghai Jiao Tong University, No. 639, Zhizaoju Rd, Shanghai 200011, PR CHINA
    Affiliations
    Attending Physician, Department of Endodontics and Operative Dentistry, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China

    College of Stomatology, Shanghai Jiao Tong University, Shanghai, PR China

    National Center for Stomatology, National Clinical Research Center for Oral Disease and Shanghai Key Laboratory of Stomatology, Shanghai, PR China
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Open AccessPublished:November 14, 2022DOI:https://doi.org/10.1016/j.prosdent.2022.09.015

      Abstract

      Statement of problem

      The influence of silver nanoparticles on adhesive properties of Single Bond Universal (3M ESPE) and the antibacterial activity of silver nanoparticles-modified adhesives against Streptococcus mutans is unclear.

      Purpose

      The purpose of this in vitro study was to evaluate the effect of silver nanoparticles on the dentin bond strength of modified adhesives and the antibacterial activity against the cariogenic pathogen S. mutans.

      Material and methods

      Single Bond Universal adhesive was used as the control. Silver nanoparticles were added to adhesives at 0.05% and 0.1% (by weight) (experimental groups), and scanning electron microscopy was used to observe the uniformity of the modified adhesives. The Single Bond Universal adhesive and the modified adhesives were then used to prepare dentin-composite resin blocks. The microtensile bond strength and microleakage of the prepared dentin-composite resin blocks were determined with or without thermocycling. The colony-forming units (CFU) of S. mutans cultured with the adhesives were evaluated, and the microtensile bond strength and microleakage of each group were tested after treatment with S. mutans. The differences in the microtensile bond strength or CFU were analyzed by using the 2-way analysis of variance and independent sample t test. The differences in microleakage between the groups were evaluated by using the Mann-Whitney test (α=.05).

      Results

      Silver nanoparticle-modified adhesives exhibited uniform morphologies without agglomeration and exhibited a homogeneous adhesive layer in dentin-composite resin blocks. The microtensile bond strength and microleakage of the modified adhesives were similar to those of the control group, with or without thermocycling (P>.05). However, thermocycling reduced the bond strength significantly (P<.001). Self-etch adhesives incorporated with silver nanoparticles showed significant antibacterial activities after less than 6 months of aging treatment. The modified adhesives did not exhibit a decreased bond strength after S. mutans exposure (P>.05), while the control group exhibited a markedly decreased bond strength after S. mutans exposure (P<.05).

      Conclusions

      Silver nanoparticle-modified adhesives showed excellent antibacterial activities against S. mutans and resisted the destruction of dentin bond strength caused by S. mutans while not compromising the bonding properties of Single Bond Universal self-etch adhesives.
      Clinical Implications
      Silver nanoparticle-modified adhesives can reduce the adhesion of cariogenic bacteria to the bonding interface, which may help reduce secondary caries.
      Dental caries is the most common dental disease with approximately 2.3 billion people globally having caries in permanent teeth in 2017 and 0.5 billion children having caries in primary teeth.
      Global, regional, and national incidence, prevalence, and years lived with disability for 354 diseases and injuries for 195 countries and territories, 1990-2017: a systematic analysis for the global burden of disease study 2017.
      Composite resin is the most frequently used material to restore carious teeth or teeth with defects because of its good esthetics and bonding.
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      Proteins, pathogens, and failure at the composite-tooth interface.
      and composite resin restorations present a higher risk of developing secondary caries than amalgam restorations.
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      may cause interfacial defects and marginal microleakage, which is one of the main causes of restoration failure leading to marginal staining, debonding, and recurrent caries.
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      The World Health Organization (WHO) has specified methylene blue dye as a grading tool for evaluating microleakage.
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      Streptococcus mutans has been reported to be the primary cariogenic pathogen because of its ability to synthesize large quantities of extracellular glucan polymers to form biofilms, acidogenicity, and aciduricity.
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      The acid challenge can significantly damage the adhesive interface of composite resins,
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      Calibration of a lactic-acid model for simulating biofilm-induced degradation of the dentin-composite interface.
      and cariogenic bacteria also degrade composite resins and adhesives.
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      Studies have been conducted on composite resins modified with antibacterial agents, including silver, gold, titanium, and zinc.
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      Silver nanoparticles have been used as antibacterial agents and reported to have broad-spectrum antibacterial and antiviral properties, to have good cell biocompatibility, and not to reduce the mechanical properties of dental composite resins.
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      Dentin pretreatment with silver nanoparticles has been reported to reduce the degradation of resin-dentin surfaces and increase the microshear bond strength (μSBS).
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      • Koohpeima F.
      Influence of silver nanoparticles on resin-dentin bond strength durability in a self-etch and an etch-and-rinse adhesive system.
      Single Bond Universal (3M ESPE) is a commercially available self-etch dental adhesive which is widely used in composite resin bonding and for bonding porcelain restorations.
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      ,
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      The null hypotheses were that the bond strength and microleakage of Single Bond Universal (3M ESPE) would not be affected by silver nanoparticles and that adhesives modified with silver nanoparticles would have no antibacterial activity against S. mutans.

      Material and methods

      A self-etch adhesive (Single Bond Universal, SBU; 3M ESPE) was used as the parent bonding system and control group. Silver nanoparticles (silver 2-ethylhexanoate; Strem Chemicals, Inc) Ø20 nm in size were dissolved in 2-(tertbutylamino) ethyl methacrylate (TBAEMA; Sigma) at a concentration of 0.08 g of silver salt per 1 g of TBAEMA.
      • Melo M.A.
      • Cheng L.
      • Zhang K.
      • Weir M.D.
      • Rodrigues L.K.
      • Xu H.H.
      Novel dental adhesives containing nanoparticles of silver and amorphous calcium phosphate.
      ,
      • Cheng L.
      • Weir M.D.
      • Xu H.H.
      • Antonucci J.M.
      • Kraigsley A.M.
      • Lin N.J.
      • et al.
      Antibacterial amorphous calcium phosphate nanocomposites with a quaternary ammonium dimethacrylate and silver nanoparticles.
      The prepared Ag-TBAEMA solution was then mixed with the Single Bond Universal (SBU) at a silver 2-ethylhexanoate concentration of 0.05% and 0.1% (referred to as 0.05% NAg-SBU and 0.1% NAg-SBU), regarded as the experimental groups.
      • Melo M.A.
      • Cheng L.
      • Zhang K.
      • Weir M.D.
      • Rodrigues L.K.
      • Xu H.H.
      Novel dental adhesives containing nanoparticles of silver and amorphous calcium phosphate.
      To observe the dispersion of silver nanoparticles in adhesives, the adhesives were respectively applied to slides and polymerized with a light-emitting diode (LED) polymerization unit (Elipar TriLight; 3M ESPE). The slide surfaces were then sputter-coated with gold and examined with a scanning electron microscope (SEM) (Quanta 200; FEI Co).
      For the preparation of dentin-composite resin blocks, 78 caries-free extracted human third molars were collected and stored in saline at 4 °C. Approval was obtained from the Ethics Committee of the Shanghai Ninth People’s Hospital affiliated with the Shanghai Jiao Tong University, School of Medicine, Shanghai, China. For the microtensile bond strength (μTBS) test and microleakage experiment, dentin-composite resin blocks were prepared, as shown in Figure 1. Dentin blocks were immersed in 2% sodium hypochlorite for 10 minutes and flushed with running water to clean the dentin surface. The 3 groups of adhesives (SBU, 0.05% NAg-SBU, and 0.1% NAg-SBU) were respectively applied to the dentin with a microbrush for 20 seconds, air-dried for 5 seconds, and light-polymerized for 10 seconds by using an LED polymerizing unit (Elipar TriLight). Composite resin blocks were built with a nanofilled composite resin (Filtek Z350 XT; 3M ESPE) in 2 layers of 2 mm.
      Figure thumbnail gr1
      Figure 1Flow diagram showing preparation of dentin-composite resin disks from intact human third molars.
      S. mutans UA159 was used for antibacterial assays. The S. mutans was cultured in a bacteria culture medium (Brain Heart Infusion Broth, BHI; BD Difco) and on solid BHI plates containing 2% agar under anaerobic conditions (80% N2, 10% H2, 10% CO2 at 37 °C) for 24 hours.
      For the μTBS test, 11 dentin-composite resin blocks in each group were embedded in an autopolymerizing acrylic resin (Refine Bright; KenDao Co). The blocks were then sectioned longitudinally in 2-mm thicknesses in the mesiodistal and buccolingual planes across the bonded interface to obtain resin-dentin sticks (Fig. 1). One resin-dentin stick in each group was polished with 800-grit and 1200-grit silicon carbide abrasive papers for 1 minute under water and then ultrasonically cleaned for 10 minutes in distilled water (Ultrasonic Cleaner; DongHang Co). After air-drying, the sticks were gold sputter-coated and examined under the SEM (Quanta 200). Fifteen sticks in each group were μTBS tested within 24 hours, and 15 sticks were tested after 5000 thermocycles (Cold-thermal Cycler TC-501F; Weier laboratory equipment Co) (5 °C and 55 °C water baths, 30 seconds holding time). A total of 30 resin-dentin sticks in each group were sterilized with epoxy hexane (E51830; Shangai Acmec Biochemical Co), and 15 sticks were cultured with S. mutans for 14 days and replaced with fresh BHI media every 2 days. Another 15 resin-dentin sticks in each group were cultured in saline for 14 days. For μTBS testing, the sticks were measured on a universal testing machine (Vitrodyne V1000; Liveco Inc) with a crosshead speed of 1 mm/min. The cross-sectional bonding area of each stick was measured, and the maximum failure load was divided by the bonding area to calculate the μTBS value in MPa. The failure modes of the resin-dentin bond disks after μTBS testing were also examined under the SEM (Quanta 200) at magnifications of ×50 and ×1000. Failure modes were classified as adhesive failure, cohesive failure in dentin or resin composite, and mixed failure.
      • Niyomsujarit N.
      • Senawongse P.
      • Harnirattisai C.
      Bond strength of self-etching adhesives to dentin surface after smear layer removal with ultrasonic brushing.
      A few specimens were lost during the mechanical test.
      To evaluate microleakage, 15 dentin-composite resin blocks were prepared in each group. Five blocks in each group were allocated to 5000 thermocycles. Five dentin blocks from each group were cultured with 15 mL of the S. mutans culture (OD600 nm=0.8) at 37 °C for 14 days. The dentin and roots of the blocks were coated with nail polish (S013; Noiro Co), leaving 1 mm of the bonding surface uncoated. After air-drying, the blocks were immersed in 1% methylene blue solution (2 ml:20 mg; Jumpcan Pharmaceutical Co) for 72 hours and rinsed thoroughly in distilled water. The blocks were embedded in the autopolymerizing resin (Refine Bright) with cubic molds and longitudinally sectioned at a 2-mm thickness in the mesiodistal direction across the bonded interface to obtain resin-dentin slices (Fig. 1). Three slices from each tooth were evaluated under a stereomicroscope (S9; LEICA). Each slice had 2 margins of bonded interface, so there were 28 to 30 images to be evaluated, excluding the images with enamel that had been incompletely removed.
      For the antibacterial tests, 72 Ø10×3-mm resin disks were prepared from a nanofilled composite resin (Filtek Z250 XT; 3M ESPE). The disks were polished by using 1000-grit silicon carbide papers. A total of 8 μL of SBU, 0.1% Ag-TBAEMA (as positive control group), 0.05% Nag-SBU, and 0.1% Nag-SBU solutions was respectively applied to the resin disks for 20 seconds and then air-dried and polymerized for 10 seconds under an LED light (Elipar TriLight). Eighteen disks were prepared in each group. After complete polymerization, the disks were immersed in sterile distilled water and vibrated for 2 hours to remove unpolymerized components. After drying, the disks were sterilized with epoxy hexane (E51830) and immersed in saline at 37 °C for 0, 7, 30, 60, 90, and 180 days. The saline was refreshed twice each week. Three disks in each group at every time point were transferred to 24-well plates, cultured with 2 mL of BHI and inoculated with 20 μL of the S. mutans suspension (OD600 nm=1.0). Samples were then grown in anaerobic conditions at 37 °C for 24 hours, and the disks were washed gently in PBS 3 times to remove the planktonic cells and shaken aggressively in 1 mL of PBS for 2 minutes. A total of 100 μL of 10-fold serial dilutions was plated in triplicate on BHI agar plates. After 48 hours of growth, the plates were photographed, and the colony-forming units (CFUs) were enumerated.
      The data were analyzed with a statistical software program (IBM SPSS Statistics, v19.0; IBM Corp). The data for CFUs and microtensile bond strength tests were analyzed for their conformance to a normal distribution by using the Kolmogorov-Smirnov test. The differences in μTBS or CFUs were analyzed by using a 2-way analysis of variance with the Tukey post hoc test. The differences in the μTBS of the same adhesives with and without S. mutans exposure, or with and without thermocycling, were analyzed by using the independent sample t test. The adhesive failure types were analyzed with the Fisher exact test, and the differences in microleakage between groups were evaluated with the Mann-Whitney test (α=.05).

      Results

      From the SEM morphological assessments of the Single Bond Universal (SBU), 0.05% NAg-SBU, and 0.1% NAg-SBU adhesives prepared on slides, it was found that the silver nanoparticle was uniformly dispersed in SBU adhesives without agglomeration (Fig. 2). The adhesive layers in the 3 groups were homogeneous on resin-dentin sticks at 8- to 12-μm thickness, and no adhesive defects or gaps were present in the 3 groups (Fig. 3).
      Figure thumbnail gr2
      Figure 2Scanning electron microscope images of polymerized adhesives. A, Original magnification ×5000 and a, original magnification ×20 000 SBU; B, original magnification ×5000 and b, original magnification ×20 000 0.05% NAg-SBU; C, original magnification ×5000 and c, original magnification ×20 000 0.1% NAg-SBU. SBU, Single Bond Universal.
      Figure thumbnail gr3
      Figure 3Dentin-composite disks. A, Original magnification ×300 and a, original magnification ×2000 SBU; B, original magnification ×300 and b, original magnification ×2000 0.05% NAg-SBU; C, original magnification ×300 and c, original magnification ×2000 0.1% NAg-SBU. SBU, Single Bond Universal.
      The μTBS data from the 3 adhesives are shown in Table 1. The type of adhesives showed no significant influence (P=.394), but the test conditions influenced the bond strength significantly (P<.001). The interactions between adhesives and test conditions were not significant (P=.682). The bond strengths of the 3 test groups decreased after thermocycling, with the difference in the 0.1% NAg-SBU group being statistically significant (P=.012).
      Table 1Mean ±standard deviations microtensile bond strengths (μTBS) of dentin-composite resin blocks before and after thermocycling (MPa)
      AdhesiveImmediateAfter ThermocyclingP
      SBU21.75 ±4.6617.81 ±4.85.089
      0.05% Ag-SBU19.94 ±5.6615.12 ±4.67.062
      0.1% Ag-SBU21.45 ±7.4414.39 ±4.95.012
      P<.05.
      Two-way analysis of varianceFa=13.952, Pa<.001Fb=0.946, Pb=.395

      Fc=0.47, Pc=.628
      SBU, Single Bond Universal.
      Fa and Pa, statistical results of analyzing test condition (with or without thermocycling).
      Fb and Pb, statistical results of analyzing type of adhesive.
      Fc and Pc, statistical results of analyzing interactions between type of adhesive and testing conditions.
      SBU, Single Bond Universal.
      P<.05.
      The failure modes of the different groups of dentin-composite resin disks following the μTBS tests are presented in Table 2 and Figure 4. In the SUB group, the failure modes were mainly mixed failure (62.5%) and adhesive failure (25%). For the 0.05% and 0.1% NAg-SBU groups, mixed fracture failures were also the most frequent failure types (62.5% and 100%, respectively). The constituent ratio of failure modes between the groups had no significant difference before (P=.442) or after thermocycling (P=1.0). The adhesive failure was increased in the 3 test groups after thermocycling but with no statistical difference (P>.05).
      Table 2Failure mode analysis before and after thermocycling
      ModeGroupMixed FailureAdhesive FailureCohesive Failure in CompositeCohesive Failure in DentinP
      ImmediateSBU62.5%25%12.5%0Pa=.442
      0.05% Ag-SBU62.5%12.5%12.5%12.5%
      0.1% Ag-SBU100%000
      After thermocyclingSBU44.4%22.2%33.3%0Pb=.620Pa=1.0
      0.05% Ag-SBU60%20%20%0Pb=.783
      0.1% Ag-SBU60%40%00Pb=.128
      SBU, Single Bond Universal.
      Pa, statistical results of analyzing constituent ratio between groups under same conditions.
      Pb, statistical results of analyzing constituent ratio of same group before and after thermocycling.
      Figure thumbnail gr4
      Figure 4Scanning electron images of failure modes of resin-dentin bond disks. A, Original magnification ×50 and a, original magnification ×1000. Mixed failure mode, images presented adhesive, composite, and dentin; B, original magnification ×50 and b, original magnification ×1000. Adhesive failure; C, original magnification ×50 and c, original magnification ×1000. Cohesive failure within composite resin; D, original magnification ×50 and d, original magnification ×1000. Cohesive failure within dentin.
      The marginal leakage of dentin-composite resin blocks was divided into 4 levels by using methylene blue dye, as depicted in Figure 5. The marginal leakage levels in the 0.05% NAg-SBU and 0.1% NAg-SBU groups were not significantly different compared with that in the SBU group (P>.05), and the changes of microleakage in each group were not significant after thermocycling (P>.05) (Table 3).
      Figure thumbnail gr5
      Figure 5Microleakage of dentin-composite disks dyed with 1% methylene blue. A, No leakage present, level 1. B, Microleakage less than 0.5 mm, level 2. C, Microleakage between 0.5 and 1.0 mm, level 3. D, Microleakage greater than 1.0 mm, level 4.
      Table 3Microleakage evaluation before and after thermocycling
      GroupThermocyclingImages for Evaluation0<0.5 mm0.5 to 1.0 mm>1.0 mmP
      Control-SBUBefore29131321Pb=.333
      After30161210
      0.05% Ag-SBUBefore30171210Pa=.279Pb=.921
      After30171120Pa=1.0
      0.1% Ag-SBUBefore30181200Pa=.149Pb=.330
      After3022710Pa=.635
      SBU, Single Bond Universal.
      Pa, compared to control group at same condition.
      Pb, compared to same groups before thermocycling.
      The antibacterial effect of the modified adhesives on cariogenic bacteria is shown in Figure 6. The 0.05% NAg-SBU and 0.1% NAg-SBU adhesives exhibited significant antibacterial activity against S. mutans after 7, 30, 60, and 90 days of aging treatment (P<.05). However, the antibacterial activity ceased after 6 months (P>.05). In each group, the antibacterial activity decreased over time because of material aging (P<.05) (Supplemental Table 1, available online).
      Figure thumbnail gr6
      Figure 6Streptococcus mutans colony-forming units per disk in different groups immediately or after immersion in saline at 37 °C for 7, 30, 60, 90, and 180 days. ∗, P<.05; ∗∗, P<.01; ∗∗∗, P<.001. SBU, Single Bond Universal.
      The μTBS in the SBU group decreased markedly after S. mutans exposure (P<.001) (Table 4). In the 0.05% NAg-SBU and 0.1% NAg-SBU groups, μTBS was not affected by S. mutans (P>.05). Both variables, including the type of adhesives (P=.014) and the culture conditions (P=.002), significantly influenced the bond strength. The bond failure types of the different groups of dentin-composite resin disks after the μTBS tests are presented in Table 5. In all 3 groups, the adhesive failure was statistically similar after S. mutans exposure (P>.05).
      Table 4Mean ±standard deviation microtensile bond strengths (μTBS) of dentin-composite resin blocks with or without S. mutans exposure (MPa)
      AdhesiveImmediateAfter ThermocyclingP
      SBU18.35 ±1.9712.31 ±2.56<.001
      0.05% Ag-SBU19.72 ±4.1519.76 ±5.08
      Significant difference, compared with SBU after thermocycling.
      .986
      0.1% Ag-SBU16.68 ±3.5214.71 ±3.48.281
      Two-way analysis of varianceFa=6.514 Pa=.014Fb=7.388, Pb=.002

      Fc=2.965, Pc=.062
      SBU, Single Bond Universal.
      Fa and Pa, statistical results of analyzing test conditions (with or without S. mutans exposure).
      Fb and Pb, statistical results of analyzing type of adhesive.
      Fc and Pc, statistical results of analyzing interactions between type of adhesives and culture conditions.
      Significant difference, compared with SBU after thermocycling.
      Table 5Failure mode analysis with and without S. mutans exposure
      ModeGroupMixed FailureAdhesive FailureCohesive Failure in CompositeCohesive Failure in DentinP
      No S. mutansSBU66.7%11.1%11.1%11.1%Pa=1.0
      0.05% Ag-SBU75%25%00
      0.1% Ag-SBU66.7%22.2%11.1%0
      S. mutansSBU50%40%10%0Pb=.628Pa=.840
      0.05% Ag-SBU54.5%27.3%18.2%0Pb=.659
      0.1% Ag-SBU50%50%00Pb=.464
      SBU, Single Bond Universal.
      Pa, statistical results of analyzing constituent ratio between groups under same conditions.
      Pb, statistical results of analyzing constituent ratio in each group with and without S. mutans exposure.
      The microleakage levels of all 3 test groups were statistically similar for both with and without S. mutans exposure (P>.05) (Table 6). With S. mutans exposure, the 0.1% NAg-SBU group exhibited less microleakage than the SBU group (P=.038) but showed statistically similar levels to the 0.05% NAg-SBU group (P=.152).
      Table 6Microleakage evaluation with and without S. mutans exposure
      GroupS. mutans ExposureImages for Evaluation0<0.5 mm0.5 to 1.0 mm>1.0 mmP
      Control-SBUNo29131321Pb=.121
      Yes2891090
      0.05% Ag-SBUNo30171210Pa=.279Pb=.157
      Yes30121530Pa=.152
      0.1% Ag-SBUNo30181200Pa=.149Pb=.334
      Yes28151320Pa=.038
      P<.05.
      SBU, Single Bond Universal.
      Pa, compared with control group at same condition.
      Pb, compared with same group without S. mutans exposure.
      P<.05.

      Discussion

      The null hypothesis that silver nanoparticles would not influence the physicochemical properties of the Single Bond Universal adhesive was not rejected. However, because the modified adhesives exhibited marked antimicrobial activity and avoided the decrease in bond strength and increase in microleakage caused by S. mutans, the null hypothesis was rejected.
      Silver nanoparticles have attracted considerable attention by dental manufacturers because of their excellent antibacterial properties and good biocompatibility at low concentrations.
      • Jowkar Z.
      • Shafiei F.
      • Asadmanesh E.
      • Koohpeima F.
      Influence of silver nanoparticles on resin-dentin bond strength durability in a self-etch and an etch-and-rinse adhesive system.
      However, the authors are unaware of a previous study on the influence of silver nanoparticles on the mechanical properties of the SBU adhesive. The results showed that silver nanoparticles at concentrations of 0.05 wt% and 0.1 wt% did not influence the μTBS of SBU. In addition, SEM images showed that the modified SBU presented well-dispersed silver nanoparticles, without agglomeration or defects in the bond interfaces. The good compatibility and mechanical properties were probably because the silver nanoparticles were dissolved in the TBAEMA monomer, which was mixed with the resin and could be photopolymerized.
      • Melo M.A.
      • Cheng L.
      • Zhang K.
      • Weir M.D.
      • Rodrigues L.K.
      • Xu H.H.
      Novel dental adhesives containing nanoparticles of silver and amorphous calcium phosphate.
      However, thermocycling reduced the μTBS in all groups. Failure mode analyses also revealed increases in adhesive failures in the 3 groups after thermocycling, consistent with a previous study that reported that thermocycling significantly reduced the bond strength.
      • Asiry M.A.
      • AlShahrani I.
      • Alaqeel S.M.
      • Durgesh B.H.
      • Ramakrishnaiah R.
      Effect of two-step and one-step surface conditioning of glass ceramic on adhesion strength of orthodontic bracket and effect of thermo-cycling on adhesion strength.
      The SBU adhesive contains silane. The hydrolysis of silane and the luting agent at the adhesive interface was probably responsible for the reduction in bond strength after thermocycling.
      • Asiry M.A.
      • AlShahrani I.
      • Alaqeel S.M.
      • Durgesh B.H.
      • Ramakrishnaiah R.
      Effect of two-step and one-step surface conditioning of glass ceramic on adhesion strength of orthodontic bracket and effect of thermo-cycling on adhesion strength.
      To obtain excellent bond strengths, the adhesive must uniformly penetrate the collagen system of dentin and be effectively polymerized.
      • Zecin-Deren A.
      • Sokolowski J.
      • Szczesio-Wlodarczyk A.
      • Piwonski I.
      • Lukomska-Szymanska M.
      • Lapinska B.
      Multi-layer application of self-etch and universal adhesives and the effect on dentin bond strength.
      Hybrid layers are formed by infiltration of the adhesive into the collagen fibrils and dentin tubules and are important for sealing and micromechanical retention between the resin and dentin.
      • Nakabayashi N.
      • Kojima K.
      • Masuhara E.
      The promotion of adhesion by the infiltration of monomers into tooth substrates.
      The hybrid layer in the self-etch adhesive system was narrow because of the mildly acidic monomers used.
      • Tran X.V.
      • Tran K.Q.
      Microleakage and characteristics of resin-tooth tissues interface of a self-etch and an etch-and-rinse adhesive systems.
      In the present study, SEM images showed that the adhesive layer was approximately 8- to 12-μm thick in the 3 groups, consistent with a previous study.
      • Zecin-Deren A.
      • Sokolowski J.
      • Szczesio-Wlodarczyk A.
      • Piwonski I.
      • Lukomska-Szymanska M.
      • Lapinska B.
      Multi-layer application of self-etch and universal adhesives and the effect on dentin bond strength.
      The adhesive layer was uniform, with no gaps or defects present in the 3 test groups. Thus, the marginal microleakage levels in the 0.05% NAg-SBU and 0.1% NAg-SBU groups were not significantly different from that in the SBU group with or without thermocycling. The good sealing performance may have been because silver nanoparticles of about 20 nm in size could infiltrate into dentinal tubules. SBU includes methacryloyloxydecyl dihydrogen phosphate (MDP) monomers, which contain a phosphoric acid functional group and interact chemically with hydroxyapatite crystals. The chemical interaction between MDP and dentin is thought to be responsible for the good bond durability.
      • Van Meerbeek B.
      • Yoshihara K.
      • Yoshida Y.
      • Mine A.
      • De Munck J.
      • Van Landuyt K.L.
      State of the art of self-etch adhesives.
      A complete seal of the tooth-restoration interface is highly desirable but is difficult to achieve. Microgaps have been reported to be mainly between the adhesive and dentin or between the adhesive resin and the hybrid layer.
      • Loguercio A.D.
      • Reis A.
      • Bortoli G.
      • Patzlaft R.
      • Kenshima S.
      • Rodrigues Filho L.E.
      • et al.
      Influence of adhesive systems on interfacial dentin gap formation in vitro.
      ,
      • Duarte S.J.
      • Lolato A.L.
      • de Freitas C.R.
      • Dinelli W.
      SEM analysis of internal adaptation of adhesive restorations after contamination with saliva.
      Interfacial defects at adhesive interfaces could allow for bacteria adhesion and lead to discoloration, interfacial degradation, and recurrent caries.
      • Zhu L.
      • Li Y.
      • Carrera C.A.
      • Chen Y.C.
      • Li M.
      • Fok A.
      Calibration of a lactic-acid model for simulating biofilm-induced degradation of the dentin-composite interface.
      S. mutans is a well-known oral cariogenic bacterium because of its acidogenicity and aciduricity.
      • Lemos J.A.
      • Palmer S.R.
      • Zeng L.
      • Wen Z.T.
      • Kajfasz J.K.
      • Freires I.A.
      • et al.
      The biology of streptococcus mutans.
      An acid challenge has been reported to degrade the dentin-composite resin interface and reduce bond strength.
      • Zhu L.
      • Li Y.
      • Carrera C.A.
      • Chen Y.C.
      • Li M.
      • Fok A.
      Calibration of a lactic-acid model for simulating biofilm-induced degradation of the dentin-composite interface.
      Therefore, an antibacterial adhesive is desirable to prevent recurrent caries, and this study incorporated silver nanoparticles into the SBU adhesive. Silver ions inhibit bacterial growth by inactivating the vital enzymes of bacteria and blocking deoxyribonucleic acid (DNA) replication, leading to cell death.
      • Rai M.
      • Yadav A.
      • Gade A.
      Silver nanoparticles as a new generation of antimicrobials.
      The silver nanoparticle-modified adhesives showed significant antibacterial activity against S. mutans and resisted the destruction of dentin bond strength caused by S. mutans. However, silver nanoparticle-modified adhesives exhibited antibacterial activities for less than 6 months in this study although other studies have reported that silver-modified composites exhibited long-lasting inhibitory effects against S. mutans.
      • Yoshida K.
      • Tanagawa M.
      • Atsuta M.
      Characterization and inhibitory effect of antibacterial dental resin composites incorporating silver-supported materials.
      ,
      • Tanagawa M.
      • Yoshida K.
      • Matsumoto S.
      • Yamada T.
      • Atsuta M.
      Inhibitory effect of antibacterial resin composite against streptococcus mutans.
      The difference may have been because only 8 μL of the modified adhesives was used in these studies.
      • Yoshida K.
      • Tanagawa M.
      • Atsuta M.
      Characterization and inhibitory effect of antibacterial dental resin composites incorporating silver-supported materials.
      ,
      • Tanagawa M.
      • Yoshida K.
      • Matsumoto S.
      • Yamada T.
      • Atsuta M.
      Inhibitory effect of antibacterial resin composite against streptococcus mutans.
      Limitations of the present study included the in vitro design and that a single organism (S. mutans) was studied. The actual oral environment is much more complex. Determining whether the modified adhesives can reduce recurrent caries in patients requires clinical studies.

      Conclusions

      Based on the findings of this in vitro study, the following conclusions were drawn:
      • 1.
        Silver nanoparticles incorporated into the Single Bond Universal self-etch adhesive exhibited marked antibacterial activities and did not compromise the bond strength or microleakage.
      • 2.
        Silver nanoparticles protected the bond interface and bond strength from the destructive effects of S. mutans.

      CRediT authorship contribution statement

      Jia Wang: Methodology, Software, Project administration. Wei Jiang: Data curation, Writing – original draft. Jingping Liang: Conceptualization, Supervision. Shujun Ran: Conceptualization, Writing – review & editing, Resources.

      Supplementary data

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