Can Irrigant Agitation Lead to the Formation of a Smear Layer?

SIGNIFICANCE

All of the tested irrigant agitation systems led to the formation of a smear layer on the root canal walls of previously cleaned specimens. Further investigation of this finding is warranted to contribute toward establishing a more effective protocol for endodontic cleaning and decontamination.

Methods: Thirteen premolars were instrumented with the Reciproc R40 file (VDW) and embedded in silicone, forming a closed irrigation/aspiration system. The teeth were cleaved, and 4 indentations were made on the inner buccal wall of the canal to standardize the observation sites. All the specimens were cleaned in an ultrasonic bath and evaluated under environmental scanning electron microscopy, thus constituting the control group. The same specimens were reassembled, submitted to final irrigation using UA or the ED or EC systems, and classified using a 4-level scoring system. The data were analyzed using the kappa, Pearson, and Kruskal-Wallis tests (P , .05).
Results: Smear layer formation occurred in all of the experimental groups and at all apical levels. At 3 and 4 mm, all of the experimental groups had significantly higher levels of smear layer formation than the control group. At 2 mm, the level of smear layer formation in the UA group was significantly higher than that of the control group, and there were no significant differences among the EC, ED, and control groups. At 1 mm, there were no significant differences between the ED and control groups, and the levels of smear layer formation in the EC and UA groups were significantly higher than that of the control group. There were no significant differences between the ED and EC groups at any of the apical levels. Conclusions: The smear layer formation occurred in all of the specimens submitted to final irrigation, irrespective of the technique used. (J Endod 2020;-:1–5.)
Root canal instrumentation produces a residual layer comprising organic and inorganic components. This smear layer includes both contaminated and noncontaminated debris, which adheres to the dentinal surface. Its removal is indicated to optimize disinfection and facilitate the penetration of irrigants and intracanal medication as well as to promote better adaptation of the filling materials to the walls of the root canal system (1,2).
The use of ultrasonic methods to activate irrigating solutions promotes biofilm rupture and debris removal by the streaming and acoustic cavitation phenomena (3–5). However, their reach is limited because cavitation occurs only up to 2 mm beyond the ultrasonic tip(6); moreover, this tip must act free of interference inside the root canal to be effective, a condition that hinders its performance in curved canals.
Sonic devices are also used for the same purpose but at a lower frequency. The EDDY system (ED; VDW, Munich, Germany) was created to be used at a maximum frequency of 6000 Hz and is driven by a handpiece and pneumatic motor. This instrument is made of flexible polyamide, has a tip diameter of 0.20 and a taper of 0.04, and should be applied at the working length (WL) level.
Another system developed to agitate irrigating solutions is EasyClean (EC; BassiEndo, Belo Horizonte, BH, Brazil). This system was developed to operate using reciprocating or continuous rotation kinematics and consists of an acrylonitrile butadiene styrene instrument shaped like an airplane wing, with a tip diameter of 0.25 and a taper of 0.047,8.

Studies evaluating the removal of the smear layer typically involve analyses carried out after performing instrumentation and some kind of final irrigant activation technique; in none of them has the complete removal of the smear layer been observed, especially in the apical third of the root canal (7–14). When evaluating the dentinal erosion promoted by irrigant agitation systems using scanning electron microscopy, Simezo et al15 observed the formation of a smear layer on dentinal walls, even when no mechanical
instrumentation procedure had been performed. This observation motivated the present study; our objective was to evaluate whether ultrasonic activation (UA) or the EC or
ED systems used to promote agitation of the irrigating solutions during the final irrigation step can lead to smear layer formation in the apical third of the root canal. The null hypotheses considered were
(1) that neither of the irrigant agitation systems tested would lead to the formation of
the smear layer and
(2) that there would be no difference among the irrigant agitation systems tested in
terms of the amount of smear layer formation associated with their use.

MATERIALS AND METHODS

Sample
This study was approved by the local research ethics committee (register no. 3.194.459). The analysis of variance test was used for sample size calculation in the pilot study. Considering a minimum difference between the medians of the treatments = 1, standard deviation of the error = 0.735, the number of treatments 5 4, the power of the test = 0.80, and alpha = 0.05, it was calculated that a minimum of 13 specimens would be required. Thus, 13 teeth were selected and stored in a 10% thymol solution up to the time of the experiment. The study included mandibular premolars with fully formed roots, a single canal with a curvature between 0 and 15 (16), no cracks or vertical fractures, no internal or external resorption, and no prior endodontic treatment. The teeth were radiographed in the mesiodistal and buccolingual directions and analyzed under an operating microscope (M900; DF Vasconcellos, Sao Paulo, SP, Brazil) with 8 ~ ! magnification to ensure that the inclusion and exclusion criteria were met.

WL Standardization

The root canal of each specimen was accessed with a spherical diamond bur whose
size was compatible with that of the specimen’s pulp chamber. A #10 K-type file (Dentsply Maillefer, Ballaigues, Switzerland) was inserted into the canal until its tip was viewed at the level of the apical foramen. A rubber stop was adjusted to the occlusal edge of the crown, and the real length of the specimen was determined. The crown was then reduced with a steel disc to standardize the specimen size at 10 mm. The WL was established at 1 mm short of the apical foramen.
Each specimen was manually instrumented up to the WL with a #15 K-type file (Dentsply Maillefer) followed by mechanized instrumentation with a Reciproc R40 (40.06) file (VDW) driven by an electric motor (VDW Gold, VDW), and using in-and-out movements with a brushing action on the withdrawal stroke until the WL was reached. At every 3 movements, the canal was irrigated, and foraminal patency was maintained with a #15 K-type file. During instrumentation, the canals were irrigated with 2.5% sodium hypochlorite (NaOCl) using a NaviTip 30-G needle (Ultradent, South Jordan, UT) coupled to a disposable 5-mL plastic syringe positioned 2 mm short of the WL, totaling approximately 15 mL irrigating solution.

Standardization of a Closed Irrigation/Aspiration System

A flask system was set up to simulate a closed irrigation/aspiration system as described by Kato et al7. Longitudinal grooves were created on the buccal and lingual surfaces of each root with a 0.15-mm-thick diamond disc (Komet Brazil, Sao Paulo, SP, Brazil) without reaching ~ the lumen of the canal. The apexes were sealed with wax, placed into 1.5-mL plastic tubes (Eppendorf do Brasil, Sao Paulo, SP, Brazil), and then embedded in a silicone-based impression material (Coltene, Langenau, Germany). After the silicone set, the roots were split with a chisel (Rhosse, Riberao Preto, SP, Brazil) to enable assembly and reassembly of the specimens.

Standardization of Apical Indentations

The buccal half of the root to be used in the experiment was removed with an E5 ultrasonic tip (Helse, Ribeirao Preto, SP, Brazil) coupled to an ultrasonic device (Profi III Bios; Dabi Atlante, Ribeirao Preto, SP, Brazil). Four sequential indentations measuring approximately 0.3 mm in width, height, and depth were then created on the inner wall of the canal, starting at 1 mm short of the WL and separated by 1-mm intervals in order to establish and standardize the sites to be observed later using environmental scanning electron microscopy (ESEM). The observation sites were defined as the areas immediately above the indentations made at the levels of 1 to 4 mm from the apical foramen.

Specimen Cleaning and Environmental Scanning Electron

Microscopic Analysis
The specimens were cleaned in ultrasonic baths with 2.5% NaOCl and 17% EDTA for 1 minute and then washed in running water for 1 minute. They were then dried in an incubator at 38C for a period of 24 hours and subsequently kept in a sealed plastic container to ensure the total absence of debris. Each specimen was then secured onto a metal stub using carbon adhesive tape and submitted to ESEM (Phenom-World, Eindhoven, Netherlands) under 600! magnification. Initial images of the observation sites were captured and digitally stored and became the control group. New images were acquired of these same sites after applying each of the experimental protocols and became the experimental groups. Because ESEM does not require any prior preparation of the specimens, such as sputter coating and critical point drying, the same specimens were used for both the control and the experimental groups, thus enabling standardization. To this end, each specimen was repositioned in its plastic flask after obtaining the initial images (control group) and then successively submitted to the 3 experimental protocols as follows.

Irrigant Agitation Protocols

In the EC group, the irrigating solution was activated with the EC (25/04). The canal was
filled with 2.5% NaOCl using a NaviTip 30-G needle coupled to a 5-mL disposable plastic syringe. The instrument was positioned at the WL and driven by an electric motor (VDW Gold) in Reciproc All mode. Activation was performed in 3 cycles of 20 seconds each, and 2 mL irrigating solution was renewed after each cycle. The irrigant was then aspirated, and the same sequence was performed with the same volume of a 17% EDTA solution followed by another aspiration and 3 more cycles with 2.5% NaOCl. Finally, the canal was washed with distilled water to promote complete removal of the irrigating liquid. On average, 6 mL 2.5% NaOCl and 6 mL 17% EDTA were used in the experimental phase.
After performing the irrigant agitation step, the buccal portion of the root was removed from the flask, dried in an incubator at 37C, and evaluated again under ESEM using the same procedure and capturing images of the same areas targeted for the control group.
Before submitting the specimens to the next experimental protocol, a ProDesign Logic 25/01 rotary file (BassiEndo) was gently applied to the canal walls at a speed of 350 rpm with a torque of 1 Ncm in order to remove possible areas of erosion. The specimens were then submitted to a new ultrasonic bath to promote complete cleaning of the dentinal walls and once again repositioned in their flasks. In the ED group, the irrigating solution was activated with the ED (20/04). The instrument was driven with a pneumatic sonic device (Airscaler Sonic Borden 2000N; KaVo, Biberach, Germany) at a frequency of 6000 Hz, performing in-and-out movements up to the WL. The procedures of agitation, aspiration and irrigant renewal, cleaning and drying of the specimens, and capturing and storing of the images were the same as those performed for the EC group. In the UA group, the irrigating solution was activated with an ultrasonic tip (E1-Irrisonic, Helse) (20/01) coupled to an ultrasound device (Profi III Bios, Dabi Atlante) set to operate at 30% power and positioned 1 mm short of the WL; special care was taken to keep the tip from touching the root canal walls. The procedures of agitation, aspiration and irrigant renewal, cleaning and drying of the specimens, and capturing and storing of the images were the same as those performed for the EC group.

Evaluation of the Images

The images of the experimental and control groups were positioned side by side using
presentation software (PowerPoint; Microsoft, Redmond, WA) and then analyzed and classified by 4 independent and calibrated examiners who were blinded to the study using a 4-level scoring system adapted from the study by Gambarini and Laszkiewicz17 as
follows (Fig. 1):
Score 1: open dentinal tubules with no smear layer
Score 2: open dentinal tubules with the smear layer covering less than 50% of the examined area
Score 3: open dentinal tubules with the smear layer covering more than 50% of the
examined area
Score 4: dentinal tubules with the smear layer covering 100% of the examined area