Introduction

Silver diamine fluoride (SDF) treatment has gained much attention in the past decade due to its simplicity and affordability [1, 2]. Clinical trials have shown that SDF can be used to prevent pit and fissure caries [3] and to arrest coronal caries in the primary [4] and permanent [5] teeth of children. SDF is also used to prevent root caries in the elderly [6]. Laboratory studies have found that SDF prevents dental hard tissues from demineralising due to acid production from biofilms [1, 7, 8] or acid challenge [9, 10]. SDF-treated carious dentine also represented a biologically acceptable pulpal response [11]. However, the calcium fluoride deposits generated during treatment can be dissolved after washing with water [10, 12], and an increase in the fluoride concentration in saliva is evident within 12 h of the topical application of fluoride [13]. Therefore, methods are required to increase the fluoride uptake to improve the long-term effectiveness of the treatment.

Topical fluoride therapy is often performed on exposed dentine for caries prevention [14]. Studies have shown that dentine becomes more acid resistant if its fluoride content is increased [15, 16]. However, the amount of fluoride taken up is low and limited to the surface layer [10]. Laser irradiation can be used to promote fluoride uptake by dental hard tissue. Bahar and his co-workers reported neodymium-doped yttrium aluminium garnet (Nd:YAG) laser irradiation to be effective in increasing fluoride uptake into the pit and fissure enamel [17]. Another study found that CO2 promotes the fluoride uptake of enamel treated with sodium fluoride [18]. The use of a diode laser has been shown to increase the fluoride uptake of enamel and to protect the enamel surface from acid attack [19]. An erbium-doped yttrium aluminium garnet (Er:YAG) laser was also found to increase the fluoride uptake of enamel treated with acidulated phosphate fluoride gel [20]. Following laser treatment, fluoride is not only deposited on the enamel and dentine but is also incorporated into the enamel and dentine crystalline structure [18, 21].

At present, there are no standardised parameters for the use of lasers for the prevention of dental caries in dentine. Even manufacturers often do not provide guidelines for the use of their dental laser products for caries prevention. Moreover, a literature search in PubMed found no studies comparing the effect of different types of laser on the fluoride uptake of SDF-treated dentine for caries prevention. Thus, the objective of the study was to compare the fluoride uptake of 38 % SDF-treated dentine using four common dental laser irradiations at sub-ablative energy levels.

Materials and methods

Sample preparation

This study was approved by the Institutional Review Board of the University of Hong Kong/Hospital Authority Hong Kong West Cluster (UW08-052). A flow chart of the study is shown in Fig. 1. Extracted sound human molars were collected with the patients’ consent. One section is taken from one tooth and the dentine is taken from the crown. Fifteen 3-mm-thick dentine slices were prepared from the molars. The surfaces of the slices were polished using micro-fine 4,000-grid sanding paper. The polished slices were examined using a stereomicroscope to exclude samples with cracks, hypoplasia or white spot lesions. The examined slices were treated with 1 % citric acid for 1 min and ultrasonically washed with deionised water to eliminate the smear layer. After autoclaving, each slice was divided into four parts or samples for different treatments; thus, 60 samples were prepared from 15 dentine slices. There were two parts to the experiment. In the first part, 48 samples from 12 of the dentine slices were used to study the fluoride uptake of 38 % SDF-treated dentine using four common dental laser irradiations at sub-ablative energy levels. In the second part, 12 samples from the remaining three dentine slices were used to compare the fluoride uptake of 38 % SDF-treated dentine by the four laser irradiations.

Fig. 1
figure 1

Flowchart of the experiment

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Part 1—determining the best settings for the four dental laser irradiations

A 38 % (w/v) SDF solution (Saforide; Toyo Seiyaku Kasei Co. Ltd., Osaka, Japan) was topically applied to the specimens using a micro-brush (Micro applicator—regular, Premium Plus International Ltd., Hong Kong, China). The mean (±SD) amount of SDF applied was 0.22 ± 0.07 mg (or 8.8 ± 2.8-μg fluoride), estimated by calculating the difference in the gravimetric micro-brush before and after application. The specimens were gently blown dry with a 3-in-1 syringe. Sub-ablative low-energy laser irradiation was then applied to the surface of the specimens. Four common types of laser used in dentistry, namely, CO2, Er:YAG, Nd:YAG and diode, were assessed. A CYMA dental laser (Bison Medical Co., Seoul, South Korea) was used to deliver the CO2 laser (10,600 nm); a Fidelis Plus III dental laser (Fotona Co., Ljubljana, Slovenia) was used to deliver the Er:YAG laser (2,940 nm); a Fidelis Plus III dental laser (Fotona Co., Ljubljana, Slovenia) was used to deliver the Nd:YAG laser (1,064 nm); and an Elexxion Claros dental laser (Elexxion AG Co., Radolfzell am Bodensee, Germany) was used to deliver the diode laser (810 nm). As no parameters have been specified for the use of these laser machines for fluoride treatment for caries prevention, three parameters at sub-ablative energy levels were chosen for each laser after consultation with the manufacturers. The output energy of the laser was validated by a laser power metre (Nova Handheld Laser Power Meter, Ophir Optronics, Utah, USA). Three samples from each dentine slice received the laser irradiation at the three selected settings, and deionised water was applied to the remaining sample as a control (Table 1).

Table 1 The four lasers and their settings used in the study
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After the application of SDF followed by laser irradiation, the samples were sectioned vertically for assessment. The fluoride uptake of the samples was examined using energy dispersive X-ray spectrometry (EDX) under scanning electron microscopy (SEM) (Hitachi S-4800 FEG Scanning Electron Microscope, Hitachi Ltd., Tokyo, Japan). An elemental assessment was performed by measuring five areas 5 × 5 μm2 at the surface and at 5, 10, 15 and 20 μm below the surface [22]. The mean weight percentage of fluoride uptake was calculated. The setting with the highest fluoride uptake was chosen as the selected parameter [23].

Part 2—comparison of the fluoride uptake of dentine treated by different dental lasers

Twelve samples from three dentine slices were prepared and treated with a 38 % SDF solution, as described for experiment 1. The four samples from each dentine slice received CO2 irradiation (group 1), Er:YAG irradiation (group 2), Nd:YAG irradiation (group 3) and diode irradiation (group 4), respectively, using the best parameters obtained in the first experiment. The samples were sectioned vertically for elemental assessment. The fluoride uptake of dentine was conducted using EDX under SEM. Elemental assessment was performed by measuring five areas 5 × 5 μm2 at the surface and at 5, 10, 15 and 20 μm from the surface. The five areas were randomly selected. The mean weight percentage of fluoride uptake was calculated.

Statistical analyses

The Shapiro–Wilk test of normality (p > 0.05) was used to assess whether the data had a normal distribution. A separate two-way mixed model analysis of variance was used to compare the mean weight percentage of fluoride among the groups and the mean weight percentage of fluoride among each treatment among the five depths, with sample as a random effect. All of the analyses were conducted using IBM SPSS Version 2.0 software (IBM Corporation, Armonk, New York, USA). The cut-off level of significance was taken as 5 % for all of the analyses.

Results

Part 1—determining the best settings for the four dental laser irradiations

The weight percentages of fluoride uptake at different depths of dentine using different settings for the CO2, Er:YAG, Nd:YAG and diode lasers are summarised in Table 2. The fluoride uptake of dentine using setting P3 was significantly higher at all depths than for other settings after CO2, Nd:YAG and diode laser irradiation. The fluoride uptake of dentine using setting P1 was significantly higher at all depths than for other settings after Er:YAG laser irradiation. Therefore, the selected parameters for the second experiment were 1.0 W, 50 Hz, 8 ms pulse for the CO2 laser; 0.5 W, 10 Hz, 1 ms pulse for the Er:YAG laser; 2.0 W, 10 Hz, 0.2 ms pulse for the Nd:YAG laser; and 3.0 W continuous wave for the diode laser.

Table 2 Fluoride content in weight % (±SD) of SDF-treated dentine with different laser irradiation settings
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Part 2—comparison of the fluoride uptake of dentine treated by different dental lasers

The weight percentages of fluoride uptake at different depths of dentine after CO2, Er:YAG,Nd:YAG and diode laser irradiation are summarised in Table 3. The results demonstrate that the CO2 laser, followed by the Er:YAG laser, resulted in a higher fluoride uptake at all depths (p < 0.05) than the Nd:YAG and diode lasers. No significant difference in weight percentages of fluoride were detected among different depth in Er:YAG and Nd:YAG groups. While in CO2 and Diode groups, weight percentages of fluoride at 5 μm seemed to be higher than some other levels.

Table 3 Fluoride content in weight % (±SD) of SDF-treated dentine at various depths after laser irradiation
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Discussion

Various studies have suggested that laser irradiation promotes fluoride uptake and prevents caries [14–16]. However, there are currently no standardised parameters for the use of laser irradiation with fluoride agents for the prevention of dental caries. Even the manufacturers of the laser machines used in this study did not provide clear guidelines for the use of their dental laser products for caries prevention. This study provides useful information to help dentists select the best laser types and parameters to promote fluoride uptake in exposed dentine, which is at risk of developing root caries. The use of different lasers in combination with fluoride has been shown to have a synergistic effect on the prevention of caries by increasing fluoride uptake in the enamel or root surface [20–23]. Laser-tissue interactions are mainly controlled by laser parameters such as the wavelength, energy density, pulse duration, exposure time, emission mode and repetition rate [24]. However, these factors vary across laser types, and different studies have used different parameters for the same type of laser. Therefore, it is important to develop a set of guidelines on the best laser parameters to use for dental caries prevention.

This was an in vitro study, and the experimental conditions were simplified to compare the results with different types of laser. Because the elemental components of teeth vary, four samples from the same dentine slice were allocated to the four experimental groups in the first and second parts of the experiment to allow the comparison of the results among groups [23]. Therefore, the percentages for fluoride content in Table 2 are somewhat different from those at the same depth and same settings in Table 3 due to different specimens. Furthermore, the variation within a dentine slice was small because of the small standard deviation. It should be noted that fluoride uptake is only one of the factors that influences the prevention effect, and the parameters chosen were regulated by the available settings of the laser machines used in the study. We measured both loosely bound and firmly bound fluoride, and we did not wash the slices after the laser irradiation. Previous study suggested that laser treatment seemed to be more effective in enhancing loosely bound fluoride uptake in the dentine than in enamel due to its porous surface [21]. In addition, accuracy of EDX spectrum can be affected by nature of the samples and amount and density of materials. Inhomogeneous and rough samples could also adversely affect accuracy. Furthermore, there could be variations among the areas even within the same slice and this is considered a limitation of and discussed in this study. Therefore, caution should be exercised in the interpretation of the results.

Higher-energy laser treatment has been used to melt dental tissue and seal the dental surface for caries prevention [22, 25]. However, clinicians are concerned that high-powered lasers may potentially damage gingival or pulpal tissues. Therefore, we used a sub-ablative energy laser to treat the dentine tissues in this study. SEM observation confirmed that the laser-treated dentine surface suffered no ablation (data not shown). Sub-ablative energy has been shown to have an organic blocking effect in caries prevention, in contrast to the inorganic blocking caused by high-energy laser therapy [24, 26]. Some study indicates even that when a sub-ablative energy was used, Er,Cr:YSGG laser can alter the micro-structure of radicular dentine [27]; future study may investigate the effect of alteration in micro-structure on surface roughness changes. To choose an appropriate irradiation condition for each laser, the effects of different wavelengths on certain components of the target tissue must be considered. CO2 and Er:YAG lasers are both absorbed in water and hydroxyapatite. Nd:YAG and diode lasers exhibit high absorption peaks for coloured tissue such as melanin and haemoglobin, but have limited interactions with water and hydroxyapatite [28]. The laser machines used in this study had different sized laser beams, which could not be standardised. The diameter of the laser beam, whether delivered in contact or non-contact with the tissue, creates a certain energy density: the smaller the beam, the greater the energy density. Thus, the use of a small spot greatly increases thermal transfer from the laser to the tissue, with a corresponding increase in absorption due to heating a small area. The amount of time that the beam is allowed to strike the target tissue affects the rate at which the tissue temperature increases. The time can also be regulated by the repetition rate of the pulsed laser emission mode [27]. Therefore, the settings selected in part 1 were based on the characteristic properties of each laser. Selecting an energy level that would not cause thermal damage to the pulp and periodontal tissues was an important consideration, and we also referred to related studies and asked the manufacturers for advice.

A previous study showed that the depth of fluoride penetration was within 20 μm of the root surface after laser treatment [22]. We therefore chose to measure fluoride uptake up to 20 μm below the dentine surface. The results showed that the fluoride uptake in the samples treated by the CO2 and Er:YAG lasers was higher than that in the samples treated by the Nd:YAG and diode lasers at all depths measured. Previous studies have suggested several general mechanisms. Heat has been found to enhance the uptake of fluoride to form fluoride apatite, and the thermal effect of the laser was speculated to be the main factor in promoting fluoride uptake [18]. Another possible mechanism may be the laser-induced alteration of the surface. Surface changes such as an increase in cracks and roughness may play a role in increasing the fluoride uptake [22]. A recent study indicated that sub-ablative low-energy laser irradiation following fluoride treatment increased fluoride deposition and transformed hydroxyapatite into fluoridated hydroxyapatite [24]. However, the exact mechanism is not known. It is noticeable that the fluoride uptake of the dentine after SDF treatment without laser irrigation (represented by the values in control group) might also account for the variation in fluoride uptake across the different dentine slices in different treated areas of the slices.

The different tissues targeted by different types of laser also play an important role in the fluoride uptake effect. Each laser wavelength affects the interrelated components of the target tissue, such as the water content, chemical composition, vascularity and colour. The CO2 and Er:YAG lasers have relatively long wavelengths that are readily absorbed by water and hydroxyapatite, and thus, minimal energy is transmitted to the adjacent tissues. In contrast, light in the visible and near-infrared spectrum is negligibly absorbed and moderately scattered by enamel and dentine. The Nd:YAG and diode lasers are so-called colour dependency lasers that mainly interact with coloured tissues such as melanin, and are only slightly absorbed by water. As human dentine is a light-coloured tissue, dentine may not interact with these two lasers as effectively as with the CO2 and Er:YAG lasers. In addition, although SDF can cause the staining of dentine, the effect is not significant on sound dentine. Although staining can occur, it takes a while for SDF to darken the tissue after application. This may be one of the reasons why SDF was found to be less effective than black ink in enhancing the absorption of Nd:YAG laser energy [25].

Conclusion

Among the four commercially available dental lasers assessed in this study, the CO2 and Er:YAG lasers resulted in a higher fluoride uptake in the SDF-treated dentine than the Nd:YAG and diode lasers.