Impact of Bioactivity on Push-Out Bond Strength of AH Plus Bioceramic versus BC Bioceramic Root Canal Sealers

Abstract

This study compared the push-out bond strength and adaptation of the recently developed AH Plus bioceramic (AHP-Bio) root canal sealer with Bioceramic Endosequence (BC) and AH Plus (AHP) sealers when exposed to simulated body fluid for inducing bioactivity. Cross-section discs of 1 mm thick slices from obturated root canals were prepared and either kept dry or immersed in serum for 30 days. All discs were evaluated using scanning electron microscopy (SEM) and then subjected to a push-out test. The failure modes were also determined. The data were statistically analyzed using an ANOVA test at p < 0.05. In both environments, the BC sealer recorded the greatest bond strength, while the AHP-Bio sealer recorded the lowest mean values. However, bond strength was significantly improved after immersion in serum (p < 0.001). The chi-square test and Fisher’s exact test revealed a significant difference in failure mode among the tested groups at p < 0.001. The predominant failure mode was cohesive failure in both bioceramic sealers, with the greatest value for AHP-Bio (70%), and adhesive failure for AHP/gutta-percha (60%, 80%) in both environments. SEM revealed good dentin adaptation of the three sealers, with marked decreases in gaps at the bioceramic/dentin interface after immersion in serum. In conclusion, although BC exhibited greater push-out bond strength than AHP-Bio, the latter achieved good displacement resistance that increased when the sealer was exposed to simulated body fluid (serum).

1. Introduction

For full guarantee of successful endodontic treatment, the formation of a single adhesive obturation unit is a biological rationale for the treatment [1]. The use of gutta-percha alone, without root canal sealer, fails to obtain the optimum bacteria-tight seal for the root canal system [2]. As claimed by Grossman [3], sealer is critical to achieving a proper fluid-tight seal obturation with good adaptation to radicular dentin. An ideal root canal sealer should offer an adhesion between gutta-percha and the root canal walls, resulting in a gap-free obturation to discourage bacterial growth [3,4]. This type of adhesion is considered an indication of proper sealing ability in root canal spaces [5].

In the endodontic field, a broad variety of root canal sealers have been proposed, each with different compositions and physico-chemical properties. Although no sealer could fulfill all of the requirements mentioned by Grossman [3], epoxy resin-based sealers, such as AH Plus (AHP), have proved to be the gold standard [6]. AHP is an insoluble sealer offering sufficient adaptability and resisting material dislodgment from radicular dentin [7]. Regarding sealing ability, the property termed “bioactivity” is the mechanism that causes a sealer containing calcium to create a biological response by forming an apatite layer when exposed to tissue fluid. Bioactivity can improve sealing and reduce microleakage by increasing the adaptability at the sealer/dentin contact [8]. However, AHP does not demonstrate bioactivity and has been shown to fail to form an apatite layer [9].

Bioceramic Endosequence (BC) has been extensively used since 2009 because of its biological and bioactivity characteristics [9,10]. As a premixed, injectable paste composed of calcium silicate, calcium phosphate, calcium hydroxide and zirconium oxide [9,11,12], BC is a hydraulic cement that uses the fluid inside the dentinal tubules to harden [10,11]. Although it has superior antibacterial, biocompatibility, tissue inductivity, bioactivity and flowability properties [9,10,13], the high solubility of BC is its main drawback compared with epoxy resin sealers [11,14]. To improve the performance of bioceramic sealers, the manufacturer Dentsply Sirona launched a new type of AHP bioceramic (AHP-Bio) root canal sealer containing both bioactive bioceramic compounds and AHP resin particles [15]. AHP-Bio is a premixed injectable paste composed of tricalcium silicate, dimethyl sulfoxide, lithium carbonate, zirconium oxide and thickening agents. Similar to all bioceramics, AHP-Bio is a hydrophilic sealer that solidifies in the presence of a moist environment [15,16]. As claimed by the manufacturer, this new sealer offers adequate adhesion and bioactivity when the sealer contact simulates tissue fluids [15,16]. The manufacturer also reports that AHP-Bio has improved solubility [15].

Good adhesion and adaptation are important properties for root canal obturation, which depends mainly on the sealer. The assessment of interfacial bond strength between the sealer and radicular dentin is clinically relevant to the adhesive quality of the root canal sealer. Push-out bond strength is regarded as a significant diagnostic factor for evaluating sealer/dentin adhesion [17]. Scanning electron microscopy is the best method to evaluate the adaptation of root canal obturation. It is effective for detecting the degree of sealer penetration within dentinal tubules, the formation of tag-like structures and the presence or absence of voids at the sealer/dentin interface that compromise the degree of adaptation, and can be responsible for easy obturation displacement [18]. There is a lack of information about the adhesion ability of the AHP-Bio sealer to radicular dentin. Therefore, the aim of this study was to evaluate the push-out bond strength and adaptation of the AHP-Bio sealer when exposed to simulated tissue fluid as compared with regular BC and AHP epoxy resin root canal sealers. The specific hypothesis is that there would be no difference between the tested sealers.

2. Materials and Methods

King Abdulaziz University’s ethical approval research committee approved the procedures of this study (#57678). Single-rooted mandibular mature premolars that were extracted for periodontal, prosthetic or orthodontic purposes were collected from the maxillo-facial department and used for this study. Mesio-distal and labio-lingual radiographs were taken for the collected teeth. Teeth with previous endodontic treatment, root resorption, open apex or severe root curvature were excluded. The teeth were autoclaved and stored in a 1% thymol solution until used [19].

2.1. Specimen Preparation

The specimens were prepared in the Advanced Technology Dental Research Laboratory at the Faculty of Dentistry, King Abdulaziz University. Using a diamond disc under a cooling system, the crowns of the selected teeth were trimmed. The working length was determined to be 1 mm short of the visible k-file #10 from the apical foramen. The root canals were then chemo-mechanically prepared using a F5 ProTaper Gold rotary file (Dentsply, Ballaigues, Switzerland) accompanied by 5.25% sodium hypochlorite irrigation. Finally, the root canals were irrigated with 5 mL of 17% ethylenediamine tetra-acetic acid (EDTA) and then with 5 mL of distilled water. The prepared roots were obturated with single-matched gutta-percha cones, size F5 (Dentsply, Ballaigues, Switzerland). According to the sealer used, the root canals were randomly divided into three main groups: Bioceramic Endosequence (BC, Brasseler Boulevard, Savannah, GA, USA), AH Plus bioceramic (AHP-Bio, Dentsply Sirona, Tulsa Dental, Johnson, Republic of Korea) or AH Plus epoxy resin (AHP, Dentsply, Maillefer, Switzerland). All obturated roots were incubated at 37 °C and 100% humidity for a week until complete setting of the sealer.

The roots were vertically embedded in blocks made of clear self-cure acrylic resin. Using a precision cutting machine (Buehler, Lake Bluff, IL, USA), the roots were trimmed perpendicularly to their long access into 1 mm thick slices. To standardize the diameter of the tested discs, three discs were obtained from the middle region of each root (n = 20), each with nearly identical diameters [20]. To guarantee consistency in the specimens’ dimensions, the thickness of each slice and canal diameter were verified using a digital caliper (RexBeti, China).

Each main group was divided into two subgroups: they were either kept dry or kept in vials containing 5 mL of phosphate-containing tissue fluid (Serum, Gibco, Life technologies, Carlsbad, CA, USA) for 30 days (n = 10) [21,22]. The serum was refreshed every 3 days.

2.2. Push-Out Test

After the incubation period, the specimens were subjected to a push-out bond strength test using a universal testing machine (MicroTester Precision Instruments, Model 5944, Instron, Norwood, MA, USA). Each tested disc sample was positioned on a metal slab with a central hole, allowing for a free-motion 0.5 mm diameter plugger. Under continuous vertical downward pressure at a constant speed of 0.5 mm/min, the test was applied until the canal filling was displaced. The maximum force of the filling displacement was recorded in newtons. The bond strength of failure in megapascals (MPa) was calculated according to Equation (1), where r1 and r2 are the radius of the coronal and the apical root canal, respectively, and h is the thickness of the dentin disc sample [23]. As s (sine theta) = h, sine theta (s) was used.

$$Bond strength \left(\mathrm{MPa}\right)={\displaystyle \frac{Maximum faliure bond \left(\mathrm{N}\right) }{\mathsf{\pi }\left(\mathrm{r}1+\mathrm{r}2\right)\mathrm{s}}}$$

(1)

2.3. Failure Mode

At the end of the push-out test, each sample was analyzed under stereomicroscopy (Meiji Techno Co., Ltd., Tokyo, Japan) at 50× magnification. The mode of failure (Figure 1) was categorized as one of the following {Amara, 2012 #27}:

• Adhesive failure at the sealer/dentin (S/D) interface. This occurred when there was complete dislodgment of the obturation at the S/D interface, without evidence of sealer on the dentin surface, i.e., all canal walls were free of sealer.
• Adhesive failure at the sealer/gutta-percha (S/GP) interface. This occurred when there was complete dislodgment of the gutta-percha at the S/GP interface, without evidence of sealer at the surface of the gutta-percha, i.e., all gutta-percha walls were free of sealer, while all canal walls were covered with sealer.
• Cohesive failure of the sealer. In this type of failure, a fracture occurred within the sealer layer, as the sealer was visible all over the circumference of both the dentin surface and the gutta-percha surface, i.e., both the dentin and gutta-percha surfaces were covered with sealer.
• Mixed failure. This occurred when all of the previous categories were detected in a single sample, i.e., cohesive failure was detected in one area of the sample and adhesive failure was detected in another area.

2.4. Surface Microstructure Analysis

Two samples of each group were analyzed using scanning electron microscopy (SEM) (Seron AURA 100, Seron Technologies Inc., Uiwang-si, Republic of Korea). The samples were sputter-coated with gold and analyzed at ×500, ×1000 and ×2000 magnification. All of the specimens, both in the dry condition and after immersion in serum, were analyzed to determine the status of the S/D and S/GP interfaces. The quality of contact adaptation and the presence or absence of voids/gaps at both the S/D and S/GP interfaces was evaluated. At the sealer layer, homogeneity or the presence of voids inside the sealer itself was also evaluated.

2.5. Statistical Analysis

The data of the push-out test were statistically analyzed using one-way ANOVA and post hoc (Tukey) tests based on the normality test (Kolmogorov–Smirnov, p > 0.05). The data of the failure modes were analyzed using nonparametric statistical tests (chi-square and Fisher’s exact tests). The significant value was set at 0.05 using the Statistical Package for the Social Sciences (SPSS) version 20 (SPSS Inc., Chicago, IL, USA).

3. Results

3.1. Push-Out Bond Strength

Based on the normality test (Kolmogorov–Smirnov, p > 0.05), the data were analyzed using ANOVA and post hoc Tukey tests. Figure 2 represents the mean ± standard deviation values of the push-out bond strength for all experimental groups, both in the dry condition and after immersion in serum. In the dry condition, BC recorded the greatest mean bond strength (1.49 ± 0.45 MPa), followed by AHP (1.29 ± 0.38 MPa), while AHP-Bio recorded the lowest mean values (1.07 ± 0.36 MPa). There was no significant difference between the three sealers at p > 0.05. After the sealer was exposed to the serum, the bond strength of BC was significantly increased (2.87 ± 0.54 MPa, p < 0.001). Although the bond strength was increased for AHP-Bio and AHP (1.39 ± 0.37 and 1.82 ± 0.55 MPa, respectively), there was no significant difference between the resin sealers (AHP-Bio and AHP) in either the dry condition or with serum immersion (p > 0.05).

3.2. Failure Mode

Figure 3 describes the frequency (%) of the different categories of failure in all tested groups after the push-out test. The chi-square test and Fisher’s exact test revealed a significant difference among the tested groups at p < 0.001. The BC samples, in both the dry condition and after immersion in serum, displayed cohesive sealer failure (40%) followed by adhesive S/D failure (30% in the dry condition) and mixed failure (30% in serum). Cohesive failure (70%) was predominant in AHP-Bio, in both the dry condition and in serum. For AHP, the predominant failure mode was adhesive S/GP (80% in the dry condition and 60% after immersion in serum). When AHP was immersed in serum, the second most predominant failure mode was mixed (40%) (Figure 3).

3.3. Scanning Electron Microscopy (SEM) Analysis

In the dry condition, all obturated canals showed gaps at the S/D interface to some degree, regardless of the sealer used. For the BC group, although close contact was generally detected at the BC/D (thin white arrow) and BC/GP (thin orange arrow) interfaces, large gaps (14.2–28.4 µm) were detected in some areas (Figure 4A). Large voids were also detected within the sealer layer. For AHP-Bio (Figure 4B), a thin gap was detected at the AHP-Bio/D interface ranging in size from 1.59 to 5.31 µm, with evidence of voids within the sealer layer. There was also contact at the AHP-Bio/GP interface and small gaps in certain areas (range 1.34–3.26 µm). However, for the AHP group (Figure 4C), there were thin gaps at the AHP/D and AHP/GP interfaces (range of 1.64–3.28 and 0.64–2.15 µm, respectively), with few voids within the sealer layer.

After immersing the samples in serum, the gaps at the BC/D interface were markedly decreased (range of 1.68–3.86 µm) (Figure 4D). Close contact was detected with no line of demarcation at the BC/GP interface (thin orange arrows). However, there were a few voids within the BC layer and in certain areas of the BC/GP (wide orange arrow) interface (range of 1.51–4.25 µm). For AHP-Bio samples, close contact was detected at the AHP-Bio/D interface, with a few voids in certain areas ranging from 1.97 to 4.21 µm. Voids of varied diameters within the sealer layer were detected as well. There was evidence of close contact, with small gaps in certain areas of the AHP-Bio/GP interface (Figure 4E). For the AHP group (Figure 4F), there was a thin gap at the AHP/D interface (range of 0.71–1.14 µm) and a large gap at the AHP/GP interface (range 2.64–6.89 µm). A few voids within the sealer layer were also detected.

4. Discussion

Endodontic treatment aims to create 3D obturation with adequate lateral and apical seals [24] to discourage bacterial growth, which can compromise the treatment outcome [3]. In order to form a monoblock, which is defined as a gap-free single-unit obturation, the sealer is necessary to bind and/or adhere to the dentin from one side and to the gutta-percha core from the other side [25]. It has been proposed that a mechanical interlocking force increases the sealer’s adherence ability to radicular dentin [26]. The push-out test is the most common technique for assessing how well the sealer adheres to radicular dentin [7,27,28]. The push-out test is the most effective method for determining bond strength, which is the force needed to separate the adhesive filling material from the dentin [26]. The current study evaluated the push-out bond strength and thus the adhesion of the recently developed resin-modified bioceramic sealer AHP-Bio compared with the gold standard epoxy resin (AHP) and traditional bioceramic (BC) root canal sealers. While some studies have evaluated the push-out bond strength of a sealer when the root canal is filled with the sealer only [29,30], most studies obturate the root canal with sealer and gutta-percha cones. In the current study, obturation with sealer and gutta-percha cones was applied, as that best simulates the clinical situation [28,31,32]. In the current study, the samples were selected from the middle third of obturated root canals with a nearly identical diameter for standardization, as described in a previous study [20]. Furthermore, Silva et al. [17] confirmed this selection, as the significant difference in push-out bond strength between bioceramic and AH Plus was only detected in the middle third of the root canal. So, this region is considered a critical area for evaluating the push-out bond strength of root canal sealers.

The current study tried to determine the predictable reason for push-out bond strength using failure mode analysis examined by stereo microscopy and sealer/dentin adaptation using SEM. The failure mode analysis evaluates the definitive area of failure either at the sealer/dentin or sealer/gutta percha interface, or at the sealer itself. Meanwhile, SEM evaluates the predictable reason for failure either due to the sealer’s failure to penetrate into dentinal tubules, adherence failure between the sealer and gutta-percha, or the presence of voids/gaps in the sealer layer itself.

In the dry condition, the present findings show that the lowest bond strength values were recorded by AHP-Bio (1.07 ± 0.36 MPa), while the greatest values were obtained by BC (1.49 ± 0.45 MPa), with no significant difference between the three sealers (p > 0.05). This finding may be attributed to the percent of calcium silicate content. BC sealers are composed mainly of calcium silicate, while AHP-Bio is composed of calcium silicate and resin, as its content of calcium silicate is lower than that of BC. It was reported that the calcium silicate content of the sealer was responsible for a micromechanical interaction between the sealer and the dentin wall, and promoted a chemical interaction producing a mineral infiltration zone [20]. Echoing this finding, Pawar et al. [28] previously determined a greater push-out bond strength for BC than AHP. Conversely, some studies have found that the bond strength of AHP is greater than that of the BC sealer [20,30], with no significant difference between them [17]. Volumetric changes during sealer setting could affect the sealer’s bond to dentin and/or gutta-percha. It has been suggested that BC sealers exhibit slight expansion during setting [17], which promotes good adaptation to dentin, while AHP undergoes a chemical bond to dentin collagen fibers [33,34], with low polymerization shrinkage [35,36] at the AHP/GP interface, creating a gap and adhesive failure at this interface. It is possible that the adhesion of AHP to dentin is stronger than its adhesion to gutta-percha, which could explain our results of 80% adhesive failure at the AHP/GP interface. The current study’s finding of a lower bond strength for AHP-Bio, with 70% cohesive failure, may be attributed to its porous structure, as described by de Souza et al. [37], which may result in a fragile material.

Different categorization systems for the failure modes of root canal filling materials have been used in previous studies. The most commonly used system includes three categories: cohesive failure of the sealer, adhesive failure at the S/D interface and mixed cohesive/adhesive failure [5,35,38]. In another study, a five-category system was used, adding adhesive failure at the S/GP interface and mixed adhesive failure (at both the S/D and S/GP interfaces) [27,39]. However, the current study used a system of four categories, excluding mixed adhesive failure, as that type was limited to only one case of BC sealer and could have affected the statistical analysis.

In the dry condition, the predominant failure modes of both bioceramic (BC and AHP-Bio) sealers were cohesive failure and mixed failure, with greater cohesive failure values in AHP-Bio (70%). In support of these findings, the iRoot Bioceramic sealer has previously displayed greater cohesive failure within the sealer layer [35]. This result may be attributed to the nature of the material’s particle size and/or the presence of voids within the sealer layer. BC has a small particle size, allowing the particles to coalesce with each other and contain fewer voids [9,40]. Interestingly, AHP-Bio was found to have a matrix of elongated and rounded larger particles containing more pores between them [37]. Furthermore, it has been suggested that the large particle ratio within the sealer layer may yield a weak bond [29]. Adding to these previous findings, the current study’s SEM images revealed voids within both the BC and AHP-Bio sealers. The large porous structure of the AHP-Bio sealer could yield a fragile material and be responsible for the lower bond strength and 70% cohesive failure. No adhesive failure at the AHP-Bio/D interface was detected. This may be attributed to the presence of dimethyl sulfoxide, which promotes good adhesion to dentin [41].

SEM images of the dry condition revealed close contact at the BC/D interface in most cases. However, there were varying degrees of gaps in certain areas of the S/D interfaces of all groups, particularly in certain areas of the BC samples (14.2–28.4 µm). Both AHP and AHP-Bio exhibited thin gaps at the S/D interface, ranging from 1.64 to 3.28 and from 1.59 to 5.31 µm, respectively. Voids were also detected within the AHP and AHP-Bio sealer layers. The great voids detected in BC may be attributed to the setting of the BC sealer in the dry condition, as a poor crystalline porous structure has been previously found for this type of sealer [9].

In the BC group, due to the presence of voids both within the sealer layer and in some areas of the BC/D interface, both cohesive failure (40%) and adhesive failure (30%) were detected at the S/D interface. The voids detected in all sealer groups may be attributed to the matched cone technique used in this study. Similarly, Kim et al. [42] found voids within the obturation when using the single-cone technique with both BC and AHP. Furthermore, the use of sealer with gutta-percha results in a thinner sealer film compared to a canal obturated with sealer only. This technique usually exhibits cohesive failure within the thin sealer film [29]. It has been reported that the single-cone technique shows greater leakage than the continuous wave technique [43]. This may be attributed to the relatively large sealer volume associated with the single-cone technique, which encourages the development of voids within the sealer layers [44,45]. However, it has been suggested that the use of matching cones that are calibrated to the canal preparation could minimize the sealer volume [45].

The BC sealer has previously been reported to be gap-free or to have a small gap at the S/D interface [45]. However, previous research has found little adaptation at the AHP/GP interface and good adaptation at the BC/GP interface [45]. One factor that influences the adhesion ability of sealers to dentin is the chemical composition of the sealer and its interaction with radicular dentin [24]. It has been suggested that the high alkalinity of bioceramic sealers is responsible for the denaturation of the collagen fibers of radicular dentin, which can penetrate the dentinal tubules and improve the adaptation of the sealer to the dentin wall [33,34]. On the other hand, the good dentin adaptation of AHP may be attributed to the chemical covalent bond between the dentinal collagen fibers and the amino groups of epoxy resin [46]; however, its polymerization shrinkage may lead to adhesive AHP/GP failure [35,36]. AHP-Bio seems to possess properties of both AHP and bioceramics; however, it contains a thickening agent that may be responsible for gaps within the sealer layer, possibly resulting in cohesive failure.

Bioactivity is the characteristic property of calcium silicate-based sealers, as their bioactive and/or calcium particles react with the phosphorus of simulated body fluid (SBF) to form a mineral structure similar to that of dentin apatite. It is an important property for improving the adaptation and sealing ability of the sealer [8,9], as it may help prevent the filling material from dislodging from radicular dentin. Serum can be used similarly to SBF because it is typically made of protein, growth factors, hormones and minerals, including phosphorus [22,47]. Additionally, it has been shown to successfully create an apatite layer over bioceramic materials [21,22]. To create an apatite layer, the specimens used in the current study were soaked in serum for 30 days.

Overall, the current bond strength values in all tested sealer groups were improved after immersion in serum compared to the sealers in the dry condition (Figure 2). This finding is supported by previous research showing that the displacement resistance of calcium silicate-based materials increased after immersion in SBF due to bioactivity and apatite layer formation at the S/D interface [30]. It has been suggested that these materials release calcium ions that react with the phosphorus in SBF, triggering the biomineralization process on the interfacial dentin surface. Calcium silicate-based materials provide mineral deposition similar to tooth structure, leading to the formation of tag-like structures that provide interfacial adaptation and reduce bacterial leakage [48]. This previous finding was confirmed in the current study’s SEM analysis, which found a marked decrease in gaps in both bioceramic (BC and AHP-Bio) sealers (Figure 4D and Figure 4E, respectively), which improved displacement resistance. However, due to AHP’s lack of bioactivity, large voids were detected within the sealer layer and at the AHP/GP interface after immersion in serum (Figure 4F).

5. Conclusions

Upon completion of the study, based on the current findings, although BC exhibited greater push-out bond strength than AHP-Bio, AHP-Bio achieved good displacement resistance that increased when the sealer was exposed to simulated body fluid (serum). Therefore, it was concluded that the recently developed resin-modified bioceramic sealer AHP-Bio appears to possess the properties of both resin, as it adhered well to radicular dentin, and bioceramics such as BC sealer, as its adaptation and displacement resistance were improved after exposure to SBF. However, its main drawback was that the most common occurring adhesive failure modes were at the sealer/gutta-percha interface or cohesive failure with the presence of voids within the sealer layer.