Introduction

It is known that the root canal system is complex, exhibiting many anatomical variations, such as lateral canals, isthmuses, and intercanal communications [1]. Disintegration of the epithelial root sheath before dentin formation, or due to a lack of dentin formation, around a blood vessel may result in lateral canal and apical ramification formation [2]. These branch canals are present in dentin partitions formed by the physiological growth of dentin, or are present in the periradicular connective tissue, and hence influence the number and form of root canals [3]. Branch canals are often difficult to detect, poorly accessible to root canal instruments, irrigants, and medications, and may serve as tissue and bacterial reservoirs. They allow potential two-way communication between the main canal and the periodontal ligament (PDL), which might account for failure of nonsurgical or surgical therapy in some cases [2].

Knowledge of branch canal morphology and configuration is essential for successful endodontic therapy. The aim of this study was to analyze and detect the incidence and distribution of such branch canals in 300 mandibular anterior teeth by means of micro-computed tomography (micro-CT), using a three-dimensional (3-D) visualization and reconstruction technique, and to investigate the regularity of branch canals in the mandibular anterior region.

Materials and methods

Collection and preparation of samples

After approval from the Ethics Committee of School and Stomatology Wenzhou Medical University, 300 extracted mandibular anterior teeth, comprising 100 central incisors, 100 lateral incisors, and 100 canines, were randomly obtained from the dental outpatient clinic of the Hospital of Stomatology, Wenzhou Medical University. No information regarding the cause of extraction, age, or sex of the patients was available. We included extracted unrestored teeth that had intact crowns or where any carious defects present did not infringe on the pulp cavity. Teeth that had undergone endodontic treatment or had immature root formation were excluded.

After tooth extraction, the remaining soft tissue was removed using standard periodontal scalers; calculus was debrided with an ultrasonic scaler (SUPRASSOM P5; Satelec Co, Merignac, France). The teeth were then stored in 10% neutral-buffered formalin solution prior to further study.

Micro-CT scanning and 3-D visualization

The specimens were scanned from the crown to the apex, using a micro-CT system (mCT-80; Scanco Medical AG, Brüttisellen, Switzerland) with an isotropic voxel size of 30 μm. Raw data were then reconstructed into 3-D images using VGStudio MAX reconstruction software (Volume Graphics GmbH, Heidelberg, Germany). After image filtration and noise reduction, a region-growing algorithm, based on gray contrast, was used for selecting the edge of the root canal system. Volume rendering was performed for reconstruction of the tooth contour and for 3-D visualization of the root canal system.

Incidence of branch canals

In this study, canals branching from the main canal to the periodontal ligament were set as primary branch canals, while branches extending from these branch canals were regarded as secondary branch canals. From the 3-D visualization model of the branch canals and the cross-sectional image of the root canal, the number of branch canals in each sample was recorded, and the incidence of branch canals was calculated at various tooth locations.

Vertical distribution of branch canals

The method used for assessing the vertical distribution of branch canals is shown in Fig. 1. A cross-section of the anatomical apex was set as a starting plane, and each cross-section was calculated and marked at an average distance of 1 mm from the starting plane. The 3-D visualization model of each sample was analyzed and the start and stop points of the primary branch canals in cross-sectional images were recorded; from this record, the distance from the branch canal to the anatomical apex was determined.

Fig. 1
figure 1

Schematic diagram of the method used for investigating the vertical distribution of branch canals

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Orientation and distribution of branch canals

Each branch canal orientation was classified according to criteria set forth by Yoshiuchi et al. [4]. In brief, the plane was divided into eight sections on an actual cross-section of a root containing a branch canal; the 12 o’clock position was assigned to the middle of the labial root surface, and 6 o’clock was assigned to the center of the lingual surfaces. The area between 11 and 1 o’clock was defined as labial (buccal) surfaces (B), that between 1 and 2 o’clock as the mesiolabial (BM or distolabial, BD), and that between 2 and 4 o’clock as the mesial (M, or distal D) regions. Branch orifice orientations were then determined according to these divisions (Fig. 2).

Fig. 2
figure 2

Schematic diagram of the method used to investigate the horizontal orientation of branch canals

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Statistical analysis

Statistical analyses were performed using SPSS19.0. Differences between the vertical distributions of branch canals were assessed using the Kruskal−Wallis test, while the chi-square test was used to compare the root canal orientation and distribution among different teeth. Statistical significance was set at P < 0.05.

Results

Incidence of branch canals

Of the 300 mandibular anterior teeth, 102 (34%) had branch canals. The highest incidence of branch canals (50%) was seen in mandibular canines, followed by lateral incisors (29%). Of the mandibular anterior teeth with branch canals, 36.30% had no less than two primary branches, while 18.63% had at least one secondary branch canal. A total of 153 primary branch canals and 35 secondary branches were observed in all specimens. The incidence and number of branch canals in different anterior tooth locations are summarized in Table 1.

Table 1 Incidence of branch canals in mandibular anterior teeth
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Vertical distribution of branch canals

As shown in Table 2, of the 153 primary branch root canals, about 82.35% were located in the apical 3 mm of the root. No significant differences in incidence of branch canals were observed in terms of the vertical distribution at different tooth locations (P > 0.05).

Table 2 Vertical distribution of branch canals in mandibular anterior teeth
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Horizontal orientation of branch canals

The orientation of branch canals at different tooth locations is shown in Table 3. Of the 153 branch canals, 71.90% were located on the labial and lingual aspects. The incidence was as high as 81.48% in mandibular central incisors, followed by 74.42 and 67.47% in mandibular lateral incisors and mandibular canines respectively. There were no significant differences in the orientation and distribution at different locations on the teeth (P > 0.05).

Table 3 Horizontal orientation of branch canals in mandibular anterior teeth
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Discussion

In the present study, micro-CT was used to investigate branch canals in mandibular anterior teeth. Root canal systems in all specimens were reconstructed three-dimensionally for visualization, and the distance from 153 primary branch canals to the anatomical apex was also accurately measured. Cross-sections of each branch canal were analyzed to determine the orientation. This approach enhanced the ease and precision of evaluating the detailed morphology of the root canal system.

In the past, the dye-and-clearing technique, an inexpensive experimental procedure, which affords a 3-D view of the root canal system, and which allows processing large numbers of samples, has been used extensively in laboratory studies of the configurations of branch canals [5,6,7,8,9,10]. However, the accuracy of and results obtained with this technique are easily affected by a number of factors, including the staining stage, canal contents, and method of decalcification. This has led to significant variation in the reported incidence of branch canals in mandibular anterior teeth [7,8,9, 11]. In addition, it is difficult to measure the vertical distribution of branch canals accurately; thus, the whole root was roughly divided into three different locations: the coronal third, the middle third, and the apical third [12,13,14,15,16,17]. A hypothetical cross-section of the root was used in investigating the direction of branch canals, and the direction could only be determined relative to the orifice on the surface of the periodontal ligament [4, 18, 19]. Consequently, the dye-and-clearing technique presents inherent limitations to its usage.

Micro-CT was first used in the field of endodontics by Nielsen, in 1995 [20]. Since then, its use has gradually increased, and it has been employed to evaluate the configuration of the root canal system, given its high resolution, high-quality imaging capability, more accurate 3-D reconstruction, excellent repeatability of its results, and its usefulness for quantification. In 2006, Cheung et al. [21] investigated the incidence of branch canals and apical deltas in the apical 5 mm of mandibular second molars using visualization of 3-D models obtained using micro-CT. Verma et al. [22] and Somma et al. [23] both studied the mesiobuccal root canal morphology of maxillary first molars using micro-CT. Somma et al. [23] reported that the prevalence of accessory canals in the coronal, middle, and apical thirds of the mesiobuccal roots of maxillary first molars was 8.3, 50.0, and 91.6%, respectively. All of these results confirmed the accuracy and reliability of micro-CT as a method for studying the ultrastructure of entities such as branch canals. Nevertheless, the complexity of the equipment, the high cost, and the time required to perform micro-CT investigations have restricted the application of micro-CT to small sample sizes, making it unsuitable for analyzing the surface morphology of large numbers of samples.

Current data on the incidence of branch canals in the mandibular anterior teeth are sparse and diverse [17,18,19,20]. Apart from differences in race, study design, and sample size, the inconsistency in the definitions used has also influenced the final results reported in these studies. In the present study, branch canals were defined as canals, including lateral canals, which communicated with the periodontal ligament, as well as bifurcated structures (not including accessory root canals deriving from the pulp chamber). Branches deriving from the main root were defined as primary branch canals, while those extending from lateral canals were designated as secondary branch canals. This classification allows a more accurate evaluation of complicated root canal systems.

Nearly all previous studies have confirmed that the incidence of branch canals was highest in the apical third of roots [12,13,14,15,16,17]. Research on the vertical distribution of branch canals in mandibular anterior teeth appears to be rare, and to date, there have been no reports on the distribution and orientation of branch canals in mandibular anterior teeth. Miyashita et al. [19] investigated 1047 transparent specimens of root canals of extracted mandibular incisors. They found that 88.6% of branch canals were located in the apical third of the root. Al-Qudah et al. [24] and Kartal et al. [25] also reported that 64.3 and 78.3% of branch canals were located in the apical third of the roots of mandibular incisors. Our study demonstrated that 82.35% of branch canals are found within 3 mm of the root apex, with 91.5% of these canals found within 4 mm of the anatomical apex. These results were consistent with previous findings. In order to remove as much of the tissue, bacteria, and debris harbored within branch canals in the apical region as possible, several devices and disinfection techniques have been developed. Tulus et al. [26] recommended the use of a dental microscope, NaOCl irrigant combined with ultrasonic activation, pre-bent preparation instruments, specially designed handpieces, and thermoplastic obturation technique to perform therapy of branched root canal systems. De Gregorio et al. [27] evaluated different irrigation activation systems and found that passive ultrasonic irrigation facilitated better penetration of irrigant into the lateral canals, whereas apical negative pressure irrigation facilitated efficient irrigation of the root canal system up to the working length.

As the demand to save and maintain natural teeth has increased, more endodontic procedures are being performed, with a consequent increased risk for failure of root canal therapy or retreatment, and periradicular endodontic surgery. The success rate of endodontic surgery in the literature ranges from 59.1 to 96.8% [28, 29]. Most failures were due to inadequate root canal sealing. Access, visibility, and instrumentation are easier in anterior tooth sextants. Thus, endodontic surgery involving anterior teeth will be the preferred choice for endodontists and general practitioners. Most scholars have suggested that optimal resection of apical root tissue should be within 2–3 mm [30,31,32]. Kim et al. [33] found that, when the root resection level approached 3 mm, it resulted in elimination of 93% of branch canals, thus facilitating improved canal obturation, and apical sealing. Degerness et al. [34] conducted research on the mesiobuccal roots of maxillary molars and found that 80% of branch canals were located within 3.64 mm of the anatomical apex; they therefore recommended a minimum mesiobuccal root resection level for maxillary molars of 3.6 mm. The results of our experimental study showed that, when root resection was performed at 3 mm, 82.35% of branch canals were removed in mandibular anterior teeth, while resection at 4 mm yielded a 91.5% rate of branch canal removal. Therefore, after completing a root resection procedure, some branch canals remained in the tooth, and a surgical microscope plus auxiliary equipment, combined with methylene blue staining, was necessary to aid investigation of resected roots [29].

Additionally, our results showed that 71.90% of branch canals were located on the labial and lingual aspects of mandibular anterior teeth. That may be because the root canals of mandibular anterior teeth are usually broad buccolingually. The deposition of secondary dentine in ribbon-shaped canals results in the formation of partitions, which may cause extensive differentiation in root canals, finally forming branch canals [35]. Based on the findings of our research, we recommend that, when performing surgical endodontic therapy for mandibular anterior teeth, greater emphasis should be placed on locating branch canals on the labial and lingual aspects of these teeth. Furthermore, no significant difference was found among the mandibular anterior teeth in terms of the incidence of branch canals, in either vertical or horizontal orientation. Therefore, equal emphasis should be placed on the branch canals among the three mandibular anterior teeth. This will help to reduce the number of missed branch canals and improve complete root canal obturation and apical sealing.