Elsevier

Dental Materials

Volume 30, Issue 8, August 2014, Pages 926-935
Dental Materials

Hydrophilicity of dentin bonding systems influences in vitro Streptococcus mutans biofilm formation

https://doi.org/10.1016/j.dental.2014.05.009Get rights and content

Abstract

Objective

To evaluate in vitro Streptococcus mutans (S. mutans) biofilm formation on the surface of five light-curing experimental dental bonding systems (DBS) with increasing hydrophilicity. The null hypothesis tested was that resin chemical composition and hydrophilicity does not affect S. mutans biofilm formation.

Methods

Five light-curing versions of experimental resin blends with increasing hydrophilicity were investigated (R1, R2, R3, R4 and R5). R1 and R2 contained ethoxylated BisGMA/TEGDMA or BisGMA/TEGDMA, respectively, and were very hydrophobic, were representative of pit-and-fissure bonding agents. R3 was representative of a typical two-step etch-and-rinse adhesive, while R4 and R5 were very hydrophilic resins analogous to self-etching adhesives. Twenty-eight disks were prepared for each resin blend. After a 24 h-incubation at 37 °C, a multilayer monospecific biofilm of S. mutans was obtained on the surface of each disk. The adherent biomass was determined using the MTT assay and evaluated morphologically with confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM).

Results

R2 and R3 surfaces showed the highest biofilm formation while R1 and R4 showed a similar intermediate biofilm formation. R5 was more hydrophilic and acidic and was significantly less colonized than all the other resins. A significant quadratic relationship between biofilm formation and hydrophilicity of the resin blends was found. CLSM and SEM evaluation confirmed MTT assay results.

Conclusions

The null hypothesis was rejected since S. mutans biofilm formation was influenced by hydrophilicity, surface acidity and chemical composition of the experimental resins. Further studies using a bioreactor are needed to confirm the results and clarify the role of the single factors.

Introduction

Resin-based composites are increasingly used because of their excellent esthetic properties and improved mechanical characteristics [1], [2]. The evolution of polymer matrices and filler particles composition enhanced the performances of these materials [3], [4], [5] along with more reliable bonds to dental hard tissues. Nevertheless, the main reason for failure of resin composite restorations is still secondary caries occurring at the interface between the composite material/dentin bonding systems (DBS) and enamel and dentin [6], [7], [8], [9], [10], [11].

As the development of such lesions is mainly related to the effect of acids and enzymes produced by bacteria colonizing the interface, the interactions between the surface of resin composites, DBS and the overlying biofilm greatly influence the lifespan of the adhesive restorations. A relevant problem is that resin composite and DBS surfaces are more heavily colonized by oral biofilms than surfaces of other restorative materials (such as amalgam, glass-ionomer cements and ceramics), as well as sound enamel surfaces [12], [13], [14].

Dentin bonding systems are blends of hydrophilic and hydrophobic monomers with the ability to couple the hydrophobic resin composite materials to hydrophilic surfaces such as dentin or enamel. They may be classified as etch-and-rinse or self-etching adhesives. These adhesives may be further classified as multi-step (i.e. three-step etch-and-rinse and two-step self-etching) or simplified by combining the number of steps required for the clinical application (i.e. two-step etch-and rinse and one-step self-etching) [15], [16], [17]. Since simplified formulations involve mixing of nonsolvated adhesives with solvated primers (i.e. two-step etch-and-rinse) or with self-etching primers (i.e. one-step self-etch), DBS simplification strongly increases the hydrophilicity of the mixture and of the bond [18]. It has been previously shown that the chemical composition of DBS influences bacterial colonization. The results suggest that variations in the chemical structure of the monomers, solvents or application techniques can extensively influence biofilm formation even when a DBS does not contain any specific antibacterial formulation [19], [20], [21], [22], [23], [24], [25].

Previous studies investigated the effect of commercially available DBS on cariogenic bacteria colonization. Pinheiro et al. [26] studied Streptococcus mutans biofilm formation on the surface of different DBS. The authors concluded that different materials produce diverse colonization levels in relation to their variable chemical composition, solvent and application technique. Comparison between etch-and-rinse and self-etching adhesives showed lower S. mutans colonization for the latter, when compared to the etch-and-rinse approach [27], [28]. Additionally, discrepancies in bacterial growth induced by different one-step self-etching adhesives and self-etching primers have been reported [29].

Experimental resins that cover a wide range of hydrophilicity and other properties of contemporary DBS have been formulated (R1–R5); these resins rank from very hydrophobic to very hydrophilic, as manifested by their solubility parameters [17]. Previous studies have evaluated their tensile strength, modulus of elasticity, degree of conversion, influence of solvent content, water sorption and solubility [17], [30], [31], [32], [33], [34], [35], [36]. However, no data is available regarding the influence of the hydrophilicity of these experimental resin blends on surface biofilm formation. Since S. mutans is considered to be one of the most important microorganisms responsible for primary and secondary caries [37], [38], [39], [40], the aim of the present study was to investigate the influence of experimental DBS chemical composition and hydrophilicity on S. mutans colonization in vitro. The null hypothesis tested was that hydrophilicity of DBS does not affect S. mutans biofilm formation.

Section snippets

Specimen preparation

All reagents and multi-well plates used in the present study were purchased from Sigma–Aldrich (St. Louis, MO) unless otherwise specified.

Five light-curing versions of neat experimental resin blends with increasing hydrophilicity were investigated (R1, R2, R3, R4 and R5). Their compositions are listed in Table 1. All blends included 0.25% camphorquinone and 1% 2-ethyl-dimethyl-4-aminobenzoate as the photoinitiator and accelerator, respectively. Resin blend R1 and R2 are similar to nonsolvated

Results

The means and standard errors of the MTT assay on the 5 experimental resin blends are reported in Fig. 1. Resin blends R2 and R3 showed the highest S. mutans colonization. Resin blends R1 and R4 showed similar intermediate biofilm formation while R5 surfaces were significantly less colonized than the other resin blends, although its composition differed from R3 only for the presence of an acidic phosphate monomer Bis[2-(methacryloyloxy)ethyl]phosphate (p < 0.001). Similarly, R4 showed a

Discussion

Reduction of biofilm formation on the surface of resin-based materials is considered an important target to reduce secondary caries risk and improve the longevity of restorations. Previous studies investigating biofilm formation [12] were performed using the agar diffusion test. In contrast, the method employed in the present study directly evaluated adherent biomass in experimental conditions that simulated in vivo clinical conditions. The results of the present study warrant rejection of the

Conflict of interest

The authors report no conflict of interest.

Acknowledgements

The authors wish to thank Mr. Aurelio Valmori for photographical assistance. The study was funded by grants from MIUR (Italy): FIRB RBAP1095CR to Breschi L (P.I.), PRIN 2009SAN9K5 to Breschi L (P.I.) and R21 DE 091213 (P.I. FRT).

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