Introduction

In dentistry, a strong and enduring adhesion to tooth surfaces has a crucial role in the success of a restoration. Restorations used in prosthetic treatment mostly cover dentin and are bonded to the tooth by mechanical and chemical factors. Surface roughness is an important mechanical factor that contributes to the retention of a restoration because it increases the surface area1. Mechanical retention can be formed inside of a restoration or on the tooth surface. In clinical practice, the dentin surface is roughened by acid etching or laser treatments. The main disadvantage of acid etching is demineralization of the tooth structure; therefore, cavity pretreatment with lasers has been proposed as an alternative for dentin etching2.

In dentistry, lasers are used in conservative treatment; in addition, they are also used to roughen cavities or a tooth surface for bonding resins and prosthodontic attachment3. Although many laser devices have been employed for this purpose, erbium laser devices [erbium, chromium: yttrium scandium gallium garnet (Er, Cr: YSGG) and erbium: yttrium aluminum garnet (Er: YAG)] are the most effective and safe laser systems, and frequently used in dental clinics4. Erbium lasers have high absorption properties in water and hydroxyapatite; they can therefore ablate enamel and dentin with minimal adverse effects on the pulp and surrounding tissues5. Studies6,7,8 investigating the roughening of dentin by erbium lasers reported that after laser treatment, rough surfaces were produced, dentin tubules opened, and the mineral content of dental tissues changed.

Some studies9,10,11,12 have demonstrated that dental hard tissues can be precisely ablated with femtosecond lasers as an alternative to erbium lasers. Femtosecond lasers use ultrashort laser pulses. The main advantages of femtosecond lasers are their speed, accuracy, and ability to roughen the material surface with negligible heat loss9,10,11,12.

In prosthetic treatments, all ceramic or adhesive fixed restorations are generally cemented to the tooth surface by a resin cement. Depending on the resin cement type, bonding agents are usually used with these cements. Bonding agents have an important role in the chemical adhesion of resin cements to the tooth surface13,14, and they protect the demineralized collagen structure from oral bacteria and liquids of the oral environment15. Some bonding agents can elute ions from their structure, provide remineralization, and contribute to the longevity of restorations16,17.

Studies reported in literature are limited to the investigation of the morphological content10,11,12,18, mineral content7,8,19, both morphologic and mineral content20, and bond strength of laser-treated dentin21,22,23,24,25,26,27. There is still a lack of knowledge regarding the effect of different lasers (Er: YAG and femtosecond lasers) and phosphate-containing bonding types on mineral content and bond strength of dentin. Therefore, the purposes of this research were (1) to assess the mineral changes in dentin after different surface treatments (Er: YAG or femtosecond laser) and phosphate-containing bonding application and (2) to assess the bond strength of dentin to resin cement. Our hypotheses were as follows: (1) different surface treatments and/or bonding types can affect the mineral content of dentin and (2) different surface treatments and/or bonding types can affect the bond strength of dentin to resin cement.

Materials and methods

A power analysis was performed before this study was conducted. Based on this analysis (with 80% power), each group needed to consist of a minimum of five specimens. Therefore, six specimens were prepared for each subgroup of this study. The Ethical Committee of the University of Selcuk (Konya, Turkey) approved the present study (committee number, 2016/04). Informed consent was obtained from all participants.

Thirty-nine freshly extracted human molar teeth were embedded in self-cure acrylic resin (Meliodent; Heraeus Kulzer, Hanau, Germany) to 2 mm below the enamel-cement intersection line. Using a low-speed sectioning device (Isomet 1000; Buehler Ltd, Lake Bluff, Illinois, USA), approximately 1.5 mm of the tooth structure was removed under water cooling from the occlusal plane of each tooth to expose dentin surface. The teeth were randomly seperated into three groups (n = 13), based on the surface treatment type: (1) The control group, which received no surface treatment. (2) The Er: YAG laser treatment group: An Er: YAG laser (Fidelis Plus III, Ljubljana, Slovenia) with a wavelength of 2940 nm was applied using a handpiece (R02). A laser optic fiber (0.9 mm) was placed perpendicularly to the dentin surface at 1 mm, and the laser applied to dentin area with water irrigation and air cooling for 20 s. The laser parameters were as follows: 200 mJ, 20 Hz, and 50 µs (short pulse mode). (3) The femtosecond laser treatment group: A femtosecond amplifier (Quantronix Integra C-3.5, NY, USA) was used to apply laser pulses at a wavelength of 800 nm and a pulse duration of 90 femtosecond for dentin conditioning. The other laser parameters were as follows: power, 400 mW; marking speed, 30 mm/s; skip speed, 1250 mm/s; pulse repetition rate, 1 kHz; focal length, 11 cm; and focal spot diameter, 28.06 μm.

Morphologic analysis

One sample from each surface-treated group was evaluated by atomic force microscope (AFM) and scanning electron microscope (SEM) both before and after surface treatments to determine the morphologic effects of surface treatments on dentin surfaces. These specimens were not used for other experiments.

AFM analysis

A specimen from each group was evaluated both before and after surface treatments by AFM (NT-MDT NTEGRA Solaris, Moscow, Russia) to see the roughness and morphologic effect of surface treatments on dentin surface. The dentin surface of the same tooth sample was examined before and after laser application. Digital images were taken in air using non-contact mode at a frequency of 240 kHz. Changes in vertical position provided the height of the images, registered as bright and dark regions. A 25 × 25 μm digital image was taken for each specimen and recorded with a slow scan rate (1 Hz).

SEM analysis

The samples used in AFM analysis were also evaluated by SEM (EVO LS10; Zeiss, Cambridge, United Kingdom) both before and after surface treatments to determine the morphologic effects of surface treatments on dentin surfaces. The SEM images were obtained at ×1500 magnification.

Energy dispersive X-ray spectroscopy analysis

After the surface treatment, the dentin area was marked in each tooth. Each specimen underwent analysis by energy dispersive X-ray spectroscopy (EDX) (EVO LS10; Zeiss) both before and after bonding application to evaluate the elemental compositional changes in the dentin surface. In this analysis, the weight% of calcium (Ca) and phosphorus (P), and the Ca/P ratio were detected.

Each surface-treated group was separated into two subgroups, based on which two-step self-etching adhesive was applied [Clearfil SE Bond (Kuraray Noritake, Okayama, Japan) or Clearfil SE Protect (Kuraray Noritake) (n = 6)]. The composition, pH value and manufacturers of the tested two-step self-etch adhesive systems are given in Table 1. The two-step self-etch adhesives were exerted according to the manufacturer’s instructions. The Clearfil SE Bond primer was exerted to dentin surface and air dried after 20 s. The bonding agent was applied to dentin surface, and then air dried and light polymerized for 10 s. The Clearfil SE Protect Bond primer was exerted to dentin surface and air dried after 20 s. This bonding agent was also applied to dentin surface, and then air dried and light polymerized for 10 s. After bonding application, dual-cure resin cement (Panavia; Kuraray Medical Inc., Okayama, Japan) was applied to dentin surfaces with a special mold (Fig. 1). The diameter of the teflon mold was 3 mm and the height of the teflon mold was 3 mm. Therefore, the cements applied to the dentin was 3 mm high. The resin cement was light polymerized (Bluephase; Ivoclar Vivadent, Schaan, Liehtenstein) for 40 s on each surface of the samples for a total of 200 s. All samples were then kept in distilled water for 24 h at 37ºC.

Table 1 The composition and manufacturers of two-step self-etch adhesive systems.
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Fig. 1
figure 1

The special mold (3 × 3 mm).

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Shear bond strength test

Shear bond strength test was executed in a universal testing machine (Shimadzu AGS-X; Shimadzu Corporation, Tokyo, Japan) at a crosshead speed of 0.5 mm/min. The shear force was applied at the resin cement-dentin interface. A chisel-shaped tip was used in the shear bond strength test to apply the separating force. This tip was positioned as close as possible to the bonding surface. Bond strength values were obtained according to the formula: BS = F/A, where BS is the bond strength, F is the force required to debond the cement, and A is the area of the adhesive interface. The measurement unit of megapascals is used for bond strength value.

Each elemental (Ca, P, and Ca/P ratio) weight% values in the surface-treated groups, and the difference of each elemental weight% value before and after bonding application were compared by one-way analysis of variance (ANOVA) and two-way ANOVA, respectively. In addition, for each surface-treated group, a paired t-test was executed to compare the percentage by weight of each element before and after bonding application. Shear bond strength data were analyzed by two-way ANOVA. Post hoc tests were executed using Tukey’s honest significant difference test (P=0.05).

Results

Microscope analysis

AFM results

The AFM images of all specimens both before and after different surface treatments are given in Fig. 2. Er-YAG laser group increased the roughness of the dentin surface with holes and valleys (Fig. 2C). On the other side, femtosecond laser roughened the dentin surface more homogenously than the Er: YAG laser treated group (Fig. 2C, E), but lowered the roughness of the dentin surface as compared to the control group. (Fig. 2A, E).

Fig. 2
figure 2

Atomic force microscope images of dentin surfaces. (A) Control. (B) Before erbium: yttrium aluminum garnet (Er: YAG) laser treatment. (C) After Er: YAG laser treatment. (D) Before femtosecond laser treatment. (E) After femtosecond laser treatment (25 × 25 μm).

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SEM results

The SEM images of surface treated dentin surfaces are given in Fig. 3. The Er: YAG (Fig. 3C), and femtosecond laser-treated dentin (Fig. 3E) showed a rough surface morphology than the control specimen (Fig. 3A). The dentin tubules were opened in both Er: YAG and femtosecond laser-treated dentin surface. The femtosecond laser-treated dentin showed more homogenous surface structure than Er-YAG laser treated dentin.

Fig. 3
figure 3

Scanning electron microscope images of dentin surfaces. (A) Control. (B) Before erbium: yttrium aluminum garnet (Er: YAG) laser treatment. (C) After erbium: yttrium aluminum garnet (Er: YAG) laser treatment. (D) Before femtosecond laser treatment. (E) After femtosecond laser treatment (magnification, ×1500).

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Elemental and bond strength analyses

The results of statistical analyses were given in Tables 2, 3 and 4. Based on one-way ANOVA, the laser-treated dentin surfaces (Er: YAG and femtosecond lasers) revealed similar elemental compositions with regard to the percentage by weight of Ca, P, and the Ca/P ratio. Both of laser treatments significantly increased the Ca and Ca/P ratio content of dentin, compared to the control group (no surface treatment) (P<0.05). In addition, the P content of dentin was higher in the Er: YAG laser-treated group than in the control group (P=0.001) (Table 2).

Table 2 Weight% values of the elemental composition of dentin after different surface treatments.
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Table 3 Mean difference of the elemental weight% values of groups before and after bonding application.
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Table 4 Shear bond strength values of groups.
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Based on two-way ANOVA, the surface treatment and bonding type significantly affected the difference of each element’s weight% value before and after bonding application (P<0.05). The paired t-test analysis revealed that the Ca and P content of dentin decreased after the bonding application compared to the surface-treated dentin. In laser-treated dentin groups, the application of Clearfil SE Protect led to a significant decrease in Ca, P, and Ca/P ratio (Table 3). Based on bond strength analysis, neither the surface treatment nor the bonding type influenced the bond strength values (P>0.05) (Table 4).

Discussion

For the long-term endurance of prosthetic restorations, the bond between dentin and the resin cement is important and is affected by mechanical and chemical factors. Most prosthetic restorations cover the dentin surface instead of enamel; therefore, the present study examined the impact of surface treatments and bonding type on mineral content and bond strength of dentin.

In the current study, different laser treatments (Er: YAG and femtosecond laser) were applied to dentin surfaces to obtain mechanical retention because of the advantage of creating microretentive surfaces with minimal injury to the ambient tissues22,24. The AFM and SEM images of Er: YAG and femtosecond laser-treated dentin showed a rough surface morphology (Figs. 2 and 3), which was similar to the SEM results of previous studies6,18,24,28,29,30,31,32. Er: YAG laser-applied dentin showed the lack of a smear layer, whereas the femtosecond laser-treated dentin showed a debris-like surface structure. The application of the femtosecond laser without water irrigation may have caused this issue. In addition, AFM and SEM images revealed that femtosecond laser treatment roughened the dentin surface more homogenously than Er: YAG laser-treated surface. The AFM images also revealed that femtosecond laser roughened the dentin surface, however lowered the roughness value of dentin surface. This might be due to the short pulse duration of femtosecond laser as compared to the Er-YAG laser.

In addition to morphologic alterations, it is necessary to consider the chemical changes produced by lasers in the irradiated tissue because diversities in the quality and quantity of the smear layer could affect the bonding ability of adhesives33. Several studies6,20,29,34,35,36 have investigated the chemical alterations of dentin after Er: YAG laser application. Some authors8,20,29,34 reported that Er: YAG laser application did not alter the mineral composition of dentin; however, other authors6,35,36 reported that chemical alterations occurred in the organic and inorganic composition of dentin. Ji et al.10 investigated the effect of femtosecond laser ablation on mineral content of dentin and reported that the P content of the tooth surface did not change in the ablated area. To detect elemental changes in dentin in our study, the same dentin area of each sample was examined by EDX analysis both after the surface treatment and after the bonding application. The analysis of laser-treated (Er: YAG and femtosecond laser) dentin revealed significantly higher Ca, P and Ca/P ratio values, compared to the control group. The reason for this result may be the fact Hossain et al.37 also found out that during the Er: YAG laser radiation, it is likely that the elevation in temperature in the lased area caused a relative increase in Ca or P elements due to the reduction of the organic components38.

Apart from the extent of the researchers’ knowledge, no standard adhesive application technique has so far been decisive for clinicians for better durability of resin–dentin bonds of adhesive systems39. For prosthetic restorations, bonding agents have an important role in the chemical adhesion of resin cements to the tooth surface. Except for self-adhesive resin cements, most resin cements require bonding agents for resin cementation. These bonding agents are categorized as etch-rinse adhesives or self-etch adhesives. Etch-rinse adhesives consist of an acid, primer, and bonding agent. Self-etch adhesive systems are categorized as one- or two-step self-etch adhesives. One-step self-etch adhesive systems are all-in-one adhesives, which consolidate acid etching, priming, and bonding. They are also entitled “universal” or “multi-mode adhesives”. Two-step self-etch adhesive systems include of a seperate primer and bonding agent13,14,40. Primer treatment in either etching mode before cementation may induce chemical interaction because of functional monomers reaching the intact dentin through the smear layer or the hybrid layer41. Most studies21,24,25,26 using Er: YAG laser treatment reported that bond strength of two-step self-etch adhesive systems on dentin was better than that of etch-rinse adhesive systems. In addition, it was reported that the hybrid layer created by a self-etching system would contain minerals because self-etching systems demineralize dentin and do not use a rinsing step15,42. Reincorporation of minerals into the dentin is noteworthy because the deposited minerals may repair nanometer-sized voids and may be resistant to deterioration in the mouth15,42. It seems that previous reports suggested that higher bond strength was achieved when a two-step self etch adhesive system was used. This could be explained by the complete dissolution of the smear layer into the adhesive. Based on the results of former studies21,24,25,26, phosphate-containing 10-methacryloyloxydecyl dihydrogen phosphate (MDP)-based two-step self-etch adhesive systems (Clearfil SE Bond and Clearfil SE Protect) were used in our study to determine whether the P-containing self-etch adhesives would reincorporate P into the dentin. The current study revealed that, after the bonding application, the mineral content of dentin decreased, compared to the surface-treated dentin. This finding may be because of the acidic content (pH 2) of the self-etch adhesive systems. The acidic monomer found in self-etch adhesives demineralizes the superficial dentin surface by partially dissolving minerals around the collagen fibrils and simultaneously allowing infiltration of resin monomers43,44. Because both surface treatments and the bonding type affected the mineral content of dentin, the first hypothesis was accepted.

In the literature, the bond strength of dentin was investigated from different perspectives. Studies on Er: YAG laser-treated dentin investigated effect of different adhesives, irradiation distance26 and different energies24,25 on the bond strength of dentin. Some authors21,24,25,26 reported that the bond strength values of Er: YAG laser-treated dentin were higher with two-step self-etch adhesives than with total-etch systems. Shirani et al.26 reported no significant difference in the bond strength values between control dentin and Er: YAG laser-treated dentin after the execution of two-step self-etch adhesives. However, Ramos et al.25 pointed out that Er: YAG laser treatment decreased the bond strength of dentin, compared to the control group, when used with two-step self-etch adhesives.

Moreover, studies focusing on femtosecond laser-treated dentin researched the effect of surface shape22, and different adhesives24 on the bond strength of dentin. Gerhardt-Szep et al.22 pointed out no significant difference in bond strength values of femtosecond laser-treated dentin groups for primer application or for primer + bond application. Portillo et al.24 reported that Er: YAG laser treatment and femtosecond laser treatment decreased the bond strength of dentin, compared to the control group, when used with two-step self-etch adhesives.

According to the bond strength results of this research, as similar to the findings of other studies22,26, different surface treatments and bonding types did not affect the bond strength values (P>0.05). Based on these findings, the second hypothesis was rejected.

Therefore, based on the findings of this research, either surface treatment type (no treatment, Er: YAG, or femtosecond laser) can be used on the dentin surface before resin cementation. Despite the bond strength values were not significantly different among the groups, after laser application (Er: YAG or femtosecond laser), Clearfil SE Bond should be recommended instead of Clearfil SE Protect, because in both laser-treated groups, Clearfil SE Protect significantly decreased the mineral content of dentin in comparison to Clearfil SE Bond (P<0.005) (Table 3). Waidyasekera et al.45 pointed out that dentin decalcified by acids in self-etch adhesives is more sensitive to react with fluoride due to the increased porosity. They also stated that as Clearfil Protect Bond is a fluoride-ion-releasing adhesive system, fluoride ions are reported to increase the rate of calcium phosphate crystallization compound and decrease the rate of apatite dissolution and mineral content of dentin45.

This study had some limitations. The elemental composition of dentin was investigated after both the surface treatment, and the bonding application; however, the elemental composition of dentin before the surface treatment and the impact of aging on the bond strength of dentin were not investigated. In addition, the failure types after the bond strength test were not investigated. In this study, the mineral content of dentin decreased after the bonding application, compared to the surface-treated dentin. Therefore, to provide favorable interaction with the laser-modified dentin surfaces, future research should focus on developing Ca- and P-containing adhesive systems that contain no acidic agents. Further, the effect of aging on ion elution from these bonding agents to dentin and on dentin bond strength should also be investigated.

Conclusion

  1. 1.

    Based both on the mineral content and bond strength analysis of this study, either surface treatment type (control, Er: YAG laser or femtosecond laser) can be used on dentin surface before resin cementation.

  2. 2.

    After laser treatment (Er: YAG or femtosecond laser), Clearfil SE Bond is recommended instead of Clearfil SE Protect, because it preserves more of the mineral content of dentin (P<0.005).