Phototherapy is unable to exert beneficial effects on orthodontic tooth movement in rat molars
To investigate the effects of irradiation lasers and light-emitting diode (LED) light on root resorption in rat molars during orthodontic tooth movement (OTM). Twenty-one 12-week-old Wistar male rats were divided into three groups: OTM only (control [CG]), OTM and LED irradiation (DG), and OTM and low laser irradiation therapy (LG). The distance between the first and second molars was used to evaluate the amount of tooth movement. The mesial surfaces of the distopalatal roots were analyzed by scanning electron microscopy, and the area of the resorption was calculated. Statistical analysis showed a decreased amount of tooth movement in the exposed DG or LG compared with CG, which was statistically significant (P = .031 and P = .004, respectively). However, when the DG and LG groups were compared, no statistically significant differences were found (P = .504). The root resorption areas were similar between CG and DG. However, statistically significant differences were found between LG and CG (P = .014) and LG and DG (P = .038). Phototherapy did not enhance tooth movement, while infrared laser irradiation did increase root resorption.ABSTRACT
Objectives:
Materials and Methods:
Results:
Conclusions:
INTRODUCTION
Root resorption during orthodontic tooth movement (OTM) is an unavoidable pathological process that affects cementum and root dentin as a consequence of compression of the periodontal ligament during orthodontic treatment.1 Although orthodontic force causes local inflammation on the periodontal ligament, this process is essential to tooth movement because it triggers osteoclastogenesis for the necessary resorption of the alveolar wall. However, it is this fundamental component that may yield areas of root resorption as well.2
There are different degrees of root resorption severity: only the cementum is resorbed; cementum and the root dentin are resorbed; and the root apex is resorbed, resulting in root shortening.1 Some degree of root resorption may be found in approximately 80% to 90% of orthodontic patients, and 0.5% of subjects present apical root resorption affecting more than one-third of root length.3,4 Although root resorption is evident at a high prevalence in orthodontic clinics, there is no effective therapy for avoiding root resorption areas.
Photobiomodulation employing light-emitting diodes (LEDs) or low-level laser in experimental studies has been shown to enhance and accelerate wound healing and offer anti-inflammatory action.5–7 Lasers and LEDs have been used in human and animal studies during OTM to observe the rate of tooth movement, orthodontic postadjustment pain, and bone remodeling. However, these results are, in general, controversial.8–13
To date, there is no study evaluating root resorption during orthodontic movement under these two different infrared light therapies (laser and LED). The aim of this study, therefore, was to investigate the effects of irradiation with coherent light (laser) and noncoherent light (LED) on OTM in rat molars. The hypothesis was that infrared light therapy might accelerate the rate of movement while reducing root resorption lacunae.
MATERIALS AND METHODS
Animals
Twenty-one 12-week-old Wistar male rats weighing 250–300 g were employed in this study. During the experiment, the rats remained inside appropriate cages in rooms with controlled temperature (25°C), 12/12 h light cycles and water and chow ad libitum. Principles of laboratory animal care (NIH publication 85–23, 1985) and national laws on animal use were observed for the present study, which was authorized by the Ethical Committee for Animal Research of the University São Paulo, Brazil (004/2016).
Tooth Movement
All procedures were carried out under general anesthesia using an intramuscular injection of 12 mL/100 g body weight ketamine hydrochloride and 6 mL/100 g body weight xylazine hydrochloride mixed. Experimental OTM was performed using the modified method described by Heller and Nanda,14 in which a closed-coil spring was bonded to the first upper left molar cleat by a stainless steel ligating wire (wire size: 0.008″, Morelli®, SP, Brazil). The other side of the coil spring was bonded to the upper left incisor exerting 25 cN force to move the molar mesially, as described previously.15,16 The teeth were covered with light-activated resin to improve coil spring retention. The lower first molar tooth was extracted to eliminate occlusal contacts that could interfere with biomechanics. The coil was maintained on the teeth for 14 days.15–17 The rats were randomly allocated into three groups of 7 rats each, according to the phototherapy protocol used: control group (CG): tooth movement only, the rats were not subjected to any therapy; laser group (LG): tooth movement + laser therapy; and LED group (DG): tooth movement + LED therapy.
Phototherapy
An LED device with a wavelength of 850 nm was used in the present study. Irradiation was performed with continuous waves, and the power density was 30 mW/cm2. Irradiation was performed under a lower dose of general anesthesia (3 mL/100 g body weight ketamine hydrochloride and 1.5 mL/100 g body weight xylazine hydrochloride) to ensure animal immobility. Before irradiation, the animals were trichotomized on the left cheek area that covers the upper molars. The LED device was extraorally positioned close to the left cheek, and the 2 cm × 1 cm area was irradiated for 4 minutes per day for 5 days (Table 1).
LASER Therapy XT (DMC Equipamentos, São Carlos, Brazil) at a wavelength of 808 nm and continuous waves at 100 mW output power were used. The laser spot area was 0.028 cm2, and irradiation was administered, under anesthesia by placing the end of the laser beam at three points (in contact with the mesial buccal, mesial palatal, and distal palatal sides of the gingiva, located in the area of the moved upper left first molar). Irradiation was performed for 20 seconds at each point (total 1 minute) once a day for 5 consecutive days. The energy density and energy applied per point was approximately 70 J/cm2 and 2 J, respectively.
Measurement of Tooth Movement
On days 0 (TI) and 14 (TF), the amount of tooth movement was evaluated, under anesthesia by measuring the distance between the mesial surface of the upper first molar and the palatal point between incisors at the gingival level (incisive papilla). All measurements were made with an electronic digital caliper. The measurements were repeated three times by the same investigator; the mean value was used as the final measurement.
Root Resorption
The rats were euthanized after 14 days, and the maxillary left first molars were carefully extracted to avoid root fracture. The distal roots were separated using a diamond disc; only the distopalatal roots were used in this study (Figure 1). To eliminate all of the periodontal ligament remnants, the extracted molars were submerged in 1% sodium hypochlorite with ultrasound for 10 minutes, washed with distilled water, transferred to 30% ethanol and dehydrated in increasing concentrations of ethanol. The roots were mounted on aluminum stubs with their mesial surfaces facing up using a colloidal silver adhesive and sputter-coated with gold in a Bal-Tec SDC-050 apparatus (Balzers, Liechtenstein). The specimens were examined under a Leo 430i scanning electron microscope operating at 15 kV (Figure 2). The area of the resorption was calculated using Image J-1.50b software (National Institutes of Health, Bethesda, Md) (Figure 3). The same investigator performed all measurements, which were repeated three times. The mean value was used as the final measurement.
Citation: The Angle Orthodontist 89, 6; 10.2319/101518-745.1
Citation: The Angle Orthodontist 89, 6; 10.2319/101518-745.1
Citation: The Angle Orthodontist 89, 6; 10.2319/101518-745.1
Statistical Analysis
All statistical analyses were performed with the Statistical Package for Social Sciences version 24 (SPSS Inc, Chicago, IL). Descriptive statistics were calculated for each group. The normality of data distribution was analyzed with the Kolmogorov-Smirnov test. The Kruskal-Wallis test, followed by Student-Newman-Keuls test, was used to compare root resorption between groups. The differences between the groups were considered significant when P < .05.
RESULTS
Amount of Tooth Movement
Measurements after 14 days showed that the amount of tooth movement of the upper first molar was 0.289 mm in the CG, while it was notably less in the DG and in the LG (0.064 mm and 0.047 mm, respectively) (Table 2). The statistical analysis showed that the decreased amount of tooth movement in the exposed DG or LG, compared to the CG, was statistically significant (P < .05). However, when the DG and LG were compared, no statistically significant differences were found (Figure 4).
Citation: The Angle Orthodontist 89, 6; 10.2319/101518-745.1
Root Resorption
Root resorption lacunae were noted on the mesial surface of all distopalatal roots after 14 days of tooth movement in all studied groups. The predominant location of the resorption area was identified as the cervical root half. Although the severity of root resorption varied in each root, most resorption lacunae exhibited a wide shallow surface that resorbed the root dentin; few lacunae were very deep, frequently reaching the root canal, while few areas showed smooth resorbed cementum (Figures 5A through C).
Citation: The Angle Orthodontist 89, 6; 10.2319/101518-745.1
The average area of distopalatal root resorption of the upper first molar during experimental OTM was 66,756.41 μm2 in CG, 73,786.46 μm2 in DG, and 219,168.26 μm2 in LG (Table 3). The root resorption areas were similar between the CG and DG. However, statistically significant differences were found between the LG and CG (P = .014) and the LG and DG (P = .038) (Figure 6).
Citation: The Angle Orthodontist 89, 6; 10.2319/101518-745.1
DISCUSSION
The present scanning microscopy results revealed that infrared LED was not able to reduce root resorption areas, while laser therapy increased root resorption during experimental OTM in the rat upper first molar. LED and laser therapies did not accelerate the applied orthodontic movement. Based on these results, the hypothesis is rejected.
This study employed a widely used experimental model based on a closed-coil steel spring (25 cN) that mesially moves the maxillary first molar by using the upper incisor as anchorage.15 An experimental time (14 days) was chosen with the aim of observing possible areas of root resorption. Indeed, this period was adequate since bone remodeling events take place within the first 5 days; therefore, some resorbed lacunae could be filled with newly formed bone.16,17
The energy used in this study was similar to that proposed by Shirazi et al.,18 but the frequency of applications was different. In the present study, LED and laser therapies were applied for 5 consecutive days in order to attenuate the inflammatory process, which can be observed in the periodontal ligament during the initial phase of OTM. In this regard, irradiation should be applied during the initial phase of the acute inflammatory response and during the recruitment and differentiation of osteoclasts.19
Considering the amount of tooth movement in all groups, less OTM was detected under LED and laser irradiation in relation to the control. When both irradiated groups were compared, no significant differences were detected between them. In contrast, other studies that employed low-level laser therapy18,20–23 and infrared LED24 have reported that OTM was enhanced when irradiation was applied. However, the different dosimetry employed in these previous studies could explain the opposite findings.
As a local inflammatory response occurs during OTM, the clastic cells are believed to take part in the removal of the hyaline area in the periodontal ligament before being activated for resorption of the adjacent alveolar bone wall. LED and laser phototherapies cause changes in intracellular mechanisms5–7 that may both increase tooth movement and stimulate root resorption.25 Therefore, some studies have confirmed the acceleration of tooth movement stimulated by laser therapy,18,20,23,26 and other studies have showed reduced resorption in relation to root resorption.19,24,27 The root resorption areas on the mesial surface of the distal palatal root in both the laser and LED groups presented similar effects, although the specimens from the LG exhibited slightly more resorption area than that of the DG. However, while the extent of resorption area varied in each group, extensive lacunae were observed mostly in the LG. The difference found between these irradiation groups could be due to differences in the distance between the equipment and the target tissue. The laser beam was delivered in contact with the gingiva, located in the area of the moved upper left first molar.20,21 However, since the rat intraoral space is too small to accommodate the probe of the LED instrument, irradiation of the facial skin area adjacent to the upper molars was performed.19,24
The severity of the resorption areas on the root surface was a somewhat surprising finding because the same was not observed at the alveolar bone surface, which did not allow the rate of tooth movement to increase. Although phototherapy has often been related to stimulation of osteoblast proliferation and activity (alkaline phosphatase expression) in vivo and in vitro,28 some studies suggest that osteoclasts may be influenced by phototherapy as well.20,26,29 Indeed, Kawasaki and Shimizu20 reported that the number of multinuclear osteoclasts in the laser irradiation group increased compared with the control group. They suggested that lasers may stimulate the fusion of mononuclear macrophages to mature osteoclasts in vitro. In this context, irradiation may enhance both remodeling events as a whole, as cementum does not remodel as bone does; this process could explain the presence of resorbed areas in the cementum surface.
In summary, although phototherapy has been clinically employed with the purpose of accelerating tooth movement in orthodontic treatments, this animal study provided evidence of no benefits on acceleration, and, more importantly, it showed adverse effects of phototherapy on root resorption.
CONCLUSIONS
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According to the parameters used in the present study, laser and LED irradiation did not enhance tooth movement, while infrared laser irradiation increased root resorption.
Rat upper first molar. (a) mesial root; (b) distal root.
Scanning electron micrograph of the distopalatal root from a mesial view.
The area of resorption (Image J software).
Amount of tooth movement (mm) of the upper first molar during 14 days. Statistically significant differences were found between CG and DG (P < .05) and CG and LG (P < .05) using Kruskal Wallis and post hoc test.
Scanning electron micrographs (×60; ×100) showing different types of root resorption lacunae: (A) root surface covered by cementum with a characteristic smooth surface; (B) wide shallow lacunae; and (C) deep lacunae.
Root resorption lacunae (μm2) of root surface on upper first molar without irradiation (control), exposed to infrared LED therapy (LED), and exposed to infrared laser (Laser). Data are expressed as mean (column). Statistically significant differences were found between LG and CG (P < .05) and LG and DG (P < .050) using Kruskal-Wallis and post hoc test.
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