MTA and Biodentine exhibit similar immediate microleakage regardless of irrigant.
Biomineralization does not prevent the increase in leakage with hCSC materials.
The effect of irrigant on long-term leakage and bond strength is material-dependent.
To evaluate the long-term effect of chlorhexidine (CHX) and dimethyl sulfoxide (DMSO) on the sealing ability and biomineralization of two different calcium silicate cements (CSC) in root canal.
Sixty human third molar root canals were obturated with ProRoot MTA or Biodentine. Before obturation the canals were irrigated with saline (control), 2% CHX or 5% DMSO. Microleakage was tested after three days and after six months. After additional six months (12 months after root filling) the roots were cut into 2 mm thick dentine discs. The discs were stored in artificial saliva for one year. The bond strength was measured with the push-out method, and the failure mode was evaluated with a stereomicroscope. The most apical disc of each tooth was used for Vickers hardness test.
No significant differences between the groups was found in initial microleakage. The leakage increased significantly during the 6-month storage in all groups except in Biodentine-CHX group and Biodentine-DMSO group. CHX and DMSO irrigation significantly increased the leakage with ProRoot MTA with time, but there was no statistically significant difference compared to the ProRoot MTA-control group at six months’ time point. CHX significantly reduced the push-out bond strength of ProRoot MTA. With Biodentine irrigation with CHX or DMSO resulted with significantly higher push-out strength compared to the Biodentine control group. Fracture analysis showed statistically significant difference in the distribution of the fractures between the groups, but neither CHX nor DMSO change the fracture pattern statistically significantly. With Vickers hardness test ProRoot MTA with and without DMSO as the final irrigant showed significantly higher dentin hardness than any Biodentine-group.
Considering that aging increased the leakage in all groups except with Biodentine-DMSO and the differences in the push-out strength and surface microhardness data, it appears that the time-related biomineralizing effect of MTA and Biodentine does not improve sealing to dentin. CHX significantly reduced ProRoot MTA bond strength and increased pure adhesive failures with both cements.
Calcium silicate-based cements, so-called mineral trioxide aggregate (MTA) and materials modified from it are commonly used when tight but non-resin-based seal between dental pulp tissue and restorations or root canal and periradicular tissue is needed. One common feature to all the products on the market is that they all contain tricalcium silicate (Ca 3 SiO 5 ) and radiopacifying agent [ ]. The first commercially available hydraulic calcium-silicate cement (hCSC), ProRoot MTA (Dentsply Maillefer, Ballaigues, Switzerland), is composed of Portland cement and bismuth oxide as radiopacifying agent. Biodentine (Septodont, Saint-Maur-des-Fossés Cedex, France) is another commonly used hCSC, with zirconium dioxide to increase radio-opacity. These materials are widely used for repairing root and furcal perforations, in ortho- and retrograde root end fillings, pulp capping and even to obturate the whole root canal. They can be used in wet environment, and some even require wet environment for setting [ , ]. The bioactivity of hCSC, if considered as a property to induce mineral precipitation to the adjacent tissues, is an important issue in the ability of possible remineralization of the dentin [ ]. Calcium silicate based materials can nucleate apatite on their surface and possibly stimulate tissue repair [ ] and even remineralize dentin [ ]. However, biological and other fluids may have different effects on the setting of the materials [ ]. Considering the widespread clinical use of hCSCs in endodontics, the information of the effect of different root canal irrigants on them is surprisingly scarce.
Chlorhexidine (CHX) is commonly used final irrigant in endodontics because of its antimicrobial properties and adhesion into root canal dentin (substantivity) [ ]. CHX also decreases the activity of collagenolytic enzymes, matrix metalloproteinases (MMPs) in radicular dentin [ , ]. This effect could have a positive influence on the sealing ability and adhesion of the root canal filling material. CHX improves the long term adhesion to dentin with composite fillings [ ] and it may also at least moderately improve the immediate [ ] and the long-term [ ] post adhesion to root dentin. CHX is said to improve the sealer wettability [ , ]. On the other hand, CHX does not seem to affect the immediate or long-term sealing ability of endodontic sealers [ , ]. The data on the effect of CHX irrigation on the behavior of hCSC in root canals is inconsistent [ ].
Dimethyl sulfoxide (DMSO) is a solvent that has long history in industry and pharmacology. Recent studies show that even low DMSO concentration improves both immediate and long-term adhesive bond strength in dentin [ ]. The effect may be related to the increased dentin wettability even up to 36% [ ]. DMSO acts as a MMP-enzyme inhibitor [ ] and by that may preserve the bond strength [ , ]. DMSO also decreases the immediate and long-term microleakage of endodontic sealers, and although the difference is minor [ ] it may increase the long-term bond strength of root canal post [ ]. The influence of DMSO to hCSC is not known.
The aim of this study was to determine the effect of CHX and DMSO on the immediate and long-term sealing ability and adhesion of ProRoot MTA and Biodentine by measuring the microleakage, push-out bond strength and fracture modes in root canals. In addition, the potential effect of hCSCs on root canal dentin after aging was evaluated using Vickers hardness test. We hypothesize that neither CHX nor DMSO would affect the immediate or long-term microleakage or adhesion of either cement. The second hypothesis was that the hCSCs do not affect the microhardness of the dentin.
Materials and methods
Sixty human third molars extracted as a part of normal treatment in the University Student Health Care Centre in Tampere and Oulu, and the Unit of Specialised Oral Care in the City of Helsinki, Finland were used for the study with the patients’ consent and approval from the Ethical Committee, Faculty of Medicine, University of Oulu. Teeth were stored in 0.2% sodium azide at 4 °C until used. Only molars with one straight separate root were selected. Crowns were removed at the cemento-enamel junction and the selected root was separated with a diamond disc. The length of the roots was adjusted to 9−10 mm. The root canals were prepared with Gates burs (Dentsply Maillefer, Ballaigues, Switzerland) no 3, 4 and 5 to a final 1.3 mm diameter through the apex. After preparation the roots were irrigated with 3 ml of 3% sodium hypochlorite (ChlorCid, Ultradent, Salt Lake City, UT, USA) followed by 3 ml of 18% ethylenediaminetetraacetic acid, (EDTA, Ultradent) to remove the smear layer. The roots were randomly divided to two groups (30 roots for each group). Both groups were further randomly divided to three subgroups, two experimental groups and a control group (10 roots for each group). Before obturation the canals of the experimental groups were irrigated either with 2% CHX (Consepsis, Ultradent) indicated for the final irrigation of root canal, or 5% DMSO (Sigma-Aldrich, St Louis, MO, USA) for 60 s and dried with paper points. 5% DMSO concentration was chosen because of its superior wettability effect [ ] and the best preservation of dentin adhesive bond strength and interface integrity after aging [ ]. The canals of control groups were irrigated with sterile saline for 60 s and dried with paper points before obturation. The apex was closed with Coltoflax impression material (Coltoflax, Colténe/Whaledent, Altstätten, Switzerland) during the irrigation. The roots were filled either with ProRoot MTA (Dentsply Maillefer, Ballaigues, Switzerland) or Biodentine (Septodont, Saint-Maur-des-Fossés Cedex, France) using amalgam carrier and a plugger, holdig the root against a glass sheet, leaving 1 mm of the coronal part of the canal unfilled for temporary filling material (Cavit G, 3 M ESPE, Neuss, Germany). The materials were mixed and used according to each manufacturer’s protocols. After filling the roots with ProRoot MTA a cotton pellet moistened with saline was placed on the coronal 1 mm of the canal and sealed with Cavit G. The roots filled with Biodentine were left on the table for 12 min to set and then sealed with Cavit G. All samples were stored at 100% humidity at 37 °C until tested. The microleakage was initially tested after 3 days. The temporary fillings were removed before the leakage was tested. After initial fluid filtration test, the temporary filling was replaced, and the specimen were further stored at 100% humidity at 37 °C until the leakage was tested again after 6 months. After additional 6 months in 100% humidity (12 months after root filling), the roots were cut with an IsoMet Low Speed Saw (Buehler, Lake Bluff, IL, USA) into 2 mm discs. Three discs per tooth were obtained. The discs were stored in artificial saliva at 37 °C for one week, and then the push-out test was performed for two discs of each tooth. The most apical disc of each tooth was used for Vickers hardness test.
Fluid filtration test
Microleakage was measured by using a fluid filtration method as described by Bouillaguet et al. [ ]. The apical part of the root was glued with cyanoacrylate glue (Flex Gel, LOCTITE Super Glue, Henkel, Düsseldorf, Germany) into a silicone tube connected to the device recording the fluid flow (Flodec, De Marco Engineering, Geneva, Switzerland). The tube was filled with distilled water under constant hydrostatic pressure of 6.89 kPa (10 psi) [ , ]. If any tube leakage were observed during testing, the measurement was stopped, the leaks were sealed and the measurement was repeated. The water pressure was applied to each root for 30 min and the fluid flow was recorded constantly with three seconds time interval.
The push-out test for measuring shear strength of the materials to dentin was performed by pushing the obturation material from the apical side of each disc using a universal testing machine (Lloyd LRX, Lloyd Instrument Ltd., Fareham, UK) with a custom-made jig and 1.0 mm/min cross-head speed. The force required to debond or break the material was registered and the point of failure was observed from the loading curve. To analyse the fracture modes, all discs were examined from both sides with a stereomicroscope (magnification 16 x), and digitally photographed.
The effect of obturation materials on dentin hardness was evaluated using surface microhardness test (Vickers hardness, VHN) with automated tester (Shimadzu Micro Hardness Tester HMV-G21, Shimadzu Corp., Tokyo, Japan). The most apical sections, not used for the push-out tests, of five randomly selected teeth in each group were tested with 1.961 N (200 g) force and 10 s holding time. Six to eight points were tested immediately (30−50 μm) under the dentin-obturation material border and at the respective normal reference dentin at about midroot. For each sample, a mean value of material-affected and reference dentin was calculated. Since the sections represented apical thirds of the teeth and the differences in apically advancing sclerosis between the samples could not be completely ruled out, the relative hardness was calculated for the material-affected dentin of each tooth using the normal dentin as reference (100%), and these relative hardness values were used to represent the sample.
Statistical analyses were performed with SPSS 22.0 for Windows (SPSS Inc., Chicago, IL, USA). Kolmogorov-Smirnov and Shapiro-Wilk tests demonstrated that all groups did not follow the normal distribution, and Levene test indicated that the data was heteroscedastic for all the measured parameters (leakage, push-out strength and microhardness). Therefore, non-parametric Kruskal-Wallis and Mann-Whitney tests were used to analyse the significance of the differences between the groups. Wilcoxon Signed Ranks test was used to analyse the significance of the differences in leakage between the immediate and 6 months aged samples within the groups, and the differences in Vickers hardness values between the material-affected dentin and control dentin. Pearson Chi-Square test was used to analyse the differences in fracture modes. The statistical unit for all measurements was the root, and the level of significance was set into p < 0.05.
The immediate and 6-month leakage values are presented in Fig. 1 . The immediate leakage was low (between 394 and 681 nl/30 min) in all groups, with no significant differences between the groups. The leakage increased significantly during the 6-month storage compared to the immediate values in all groups except in Biodentine-CHX group and Biodentine-DMSO group. The lowest leakage after 6 months was observed in the Biodentine-DMSO group, which was comparable to the ProRoot MTA-control group (p > 0.05). Even though both CHX and DMSO irrigation significantly increased the leakage with ProRoot MTA with time, there was no statistically significant difference compared to the ProRoot MTA-control group at six months’ time point, mainly due to high variation in leakage in both experimental irrigant groups.