Impact of Clear Aligners vs Conventional Brackets on Oxidant and Antioxidant Levels: A Case-Control Study

Introduction

The reduction in molecular oxygen in biological systems is accompanied by a large release of free energy giving rise to reactive oxygen species (ROS) or free radicals [1]. ROS, regarded as being highly destructive in nature, play an important role in cell signaling and metabolic processes and contribute to pathogenic processes in a variety of inflammatory disorders [2]. ROS cause tissue damage via multiple mechanisms, such as DNA damage, lipid peroxidation, protein damage, enzyme oxidation, and the stimulation of pro-inflammatory cytokine release. In healthy organisms, ROS are produced as a normal product of cellular metabolism, and the level of ROS can be stabilized by an antioxidant defence mechanism [3].

Antioxidants, many of which are released locally at sites of inflammation by polymorphonuclear leucocytes and other cells, are present in all body fluids and tissues [4]. Under normal conditions, there is a dynamic balance between ROS production and tissue concentrations of antioxidants in the body, and when the normal balance shifts in favor of ROS production, a situation arises from either an excess of ROS or a reduction of antioxidants [5]. Excess production of ROS and the resultant oxidative stress contribute significantly to tissue damage in many diseases, such as rheumatoid arthritis, diabetes, AIDS, and cancer. It is also known that there are significant interactions between oxidant status and periodontal status. Antioxidants change the progress of oral problems, such as periodontitis and gingivitis, by compromising antioxidant capacity of crevicular fluid and plasma [6].

Dental plaque is the primary etiological cause of gingival inflammation and periodontitis [7]. The impact of orthodontic treatment in relation to supragingival plaque accumulation and gingival inflammation has already been reported by several studies. Increased accumulation of dental plaque and inflammatory response can be explained by the higher number of plaque-retentive sites and impaired mechanical plaque removal [8]. As the design and the material characteristics of orthodontic bracket types vary considerably, plaque adhesion and, therefore, the induction of gingivitis might differ among currently used bracket types. Some orthodontic treatment tools cause brushing difficulty of the patients and, most importantly, a response to tooth movement [9]. A study by van Gastel et al [10] evaluated the plaque formation on teeth bonded with different types of orthodontic brackets (conventional vs self-ligating brackets) over a 7-day period and concluded that conventional brackets (CBs) could have a significant impact on bacterial load and on periodontal parameters.

Clear aligners (CAs) have been gaining popularity over the past several decades. In the Invisalign system, which is the pioneer in CA technology, 3-dimensional models can be obtained by use of a computer-aided design (CAD/CAM technology) instead of by setting up a new measurement during every appointment [11]. Even though these systems, which are available in the market with several different trademarks, are similar in general aspects, they can differ by their production methods and by their professional approach to dentists and patients. Among the advantages of treatment with CAs include having a better appearance, being easy to use, requiring a shorter appointment time, causing less pain, having no difficulty in eating, and having less dental plaque, less microbiota, and less oral hygiene [12–14]. There is a higher risk of decay in orthodontic treatment with CBs. There is a lower risk of decay in orthodontic treatment with CAs. Proper oral hygiene and correct treatment selection cause fewer problems [15].

Given that dental plaque can affect the equilibrium between ROS production and tissue concentrations of antioxidants, and that CAs provide better oral hygiene, the aims of this study were to analyze the total antioxidant status (TAS) level and total oxidant status (TOS) level induced by CBs and CAs in vivo, over a 30-day period.

Material and Methods

PARTICIPANTS:

The study group consisted of 34 patients that were divided in 2 groups. The CB group included 10 women and 7 men, aged 20 to 25 (mean 22.4±2.4 years), and the CA group (Align Technology, Santa Clara, CA, USA) included 9 women and 8 men, aged 19 to 25 (mean 22.8±2.1 years). The number of patients included in the study was decided using power analyses (G Power 3.1.3. version). The approval from the Ethics Committee of Izmir Katip Çelebi University was granted in accordance with the principles of the Declaration of Helsinki, and informed consent was obtained prior to sample collection.

The inclusion criteria of the study were as follows: no systemic disease, no use of any medicine in the last 6-month period, no substance addiction, non-smoking, favorable co-operation with the study, appropriate level of oral hygiene, pubertal growth spurts that were about to be completed or that were completed, and need for orthodontic treatment. It was ensured that the individuals participating in the study had at least 14 teeth in their mandible and maxilla.

After clinical evaluation, the patients underwent diagnosis of periodontal disease (gingivitis), according to the new classification [16].

A total of 34 patients were included in the study. The patients were provided with training on oral hygiene at certain intervals before starting treatment and during treatment. Gingival crevicular fluid (GCF) samples were taken from the patients before treatment (T0), at the end of the first week (T1) of treatment, and at the end of the first month (T2) of treatment. Clinical periodontal parameters were measured at T0 and at T2. After initial measurements prior to treatment, one group was treated with CAs and the other group was treated with CBs.

Following the clinical periodontal assessment, the clinical examinations of all of the patients who participated in the study were conducted. In this regard, the plaque index [17] and gingival index [18] were used to assess the periodontal condition of each patient. Also, pocket depth, clinical attachment loss, and bleeding on probing were recorded from the patients.

The periodontal status of the patients was determined by the expert periodontist (ASE), according to the new periodontal classification, before treatment began. The oral hygiene habits and methods of the patients were determined. According to this evaluation, the oral hygiene methods suitable for the patient were recommended. The oral hygiene application methods of the patients at the beginning of the study were followed throughout the study.

GCF VOLUME MEASUREMENTS:

All of the clinical assessments except the plaque index were conducted after obtaining GCF samples in an attempt to protect GCF volume during sampling of GCF. Before obtaining GCF samples, the teeth were isolated with dental cotton rolls. Current plaques were removed by means of cotton pellets after the surfaces of teeth were examined for any supra-gingival plaques. Air-water spray was positioned vertically on the teeth, and saliva and blood, if available, was removed by spraying air for 5 s. When obtaining GCF samples from the mesial and distal surfaces of maxillary incisor and canine teeth, a deep intra-crevicular method was applied. Standard paper strips (Periopaper Oraflow, Smithtown, NY, USA) were gently pushed onto the gingival sulcus of the teeth, from which the GCF sample was to be taken, until any resistance was encountered, and they were left for 30 s. The paper strips contaminated by blood or saliva were discarded. After sampling, to avoid evaporation, the paper strips were scanned with a Periotron 8000 device (Periotron Oraflow, Smithtown, NY, USA) within 5 s, and then the measurement values obtained were converted to microliters. The paper strips were placed into Eppendorf tubes. The samples that were collected were stored at −80 ºC until biochemical analysis was conducted [19].

ENZYME-LINKED IMMUNOSORBENT ASSAY MEASUREMENTS:

The TAS and TOS analysis of the GCF samples was conducted by using an enzyme-linked immunosorbent assay (ELISA) kit (DIAsource kit DIAsource ImmunoAssays SA, Belgium). All of the assay procedures were conducted according to the manufacturer’s instructions. The minimum detection limit was 2 ng/mL for the TAS ELISA kit, and the minimum detection limit for the TOS ELISA kit was 200 pg/mL. Then, 100 μL of the standard or sample was added to each well, and the plates were incubated for 1 h at 37°C. A total of 100 μL prepared detection antibody was added, and the plates were incubated for 1 h at 37°C, aspirated, and washed 3 times. Prepared detector (100 μL) was added, and the plates were incubated with this solution for 60 min at 37°C, aspirated, and washed 5 times. After the washes, 100 μL of the substrate solution was added, and the plates were incubated 15 to 25 min at 37°C. The ELISA plates were then assessed at 450 nm immediately in an ELISA reader (Microplate Reader Biotek, VT, USA). The TAS and TOS amount collected in 30 s was (pg/30 s) determined.

STATISTICAL ANALYSIS:

This was a single-blind study. Normal distribution compatibility of the biochemical and clinical variables was tested through the use of the Kolmogorov-Smirnov test. According to the results of the analysis, as most of the parameters examined did not exhibit normal distribution, statistical analyses were conducted using non-parametric tests. Descriptive statistical analyses, including mean and standard deviations, were conducted. The Wilcoxon test was performed to determine which group caused a difference. Different inter-groups of patients were compared using the non-parametric test (Mann-Whitney U with Bonferroni correction). Correlations between the clinical parameters and laboratory parameters were examined with the Spearman rank correlation analysis. Packaged software (SPSS for Windows version 22, IBM Corp, Armonk, NY, USA) was used for analyzing data. For power analyses, G Power 3.1.3. version (Kiel, Germany) was used. Three publications that were related to the publication and had common data were identified [20–22]. The value P<0.05 was accepted as statistically significant in the analyses.

Results

The average age of 34 patients in total, consisting of 14 men and 16 women, included in our study was 22.54±1.47 years. No statistically significant difference was found between the 2 groups in terms of age and sex.

The periodontal clinical data (plaque index, gingival index, pocket depth, clinical attachment loss, and bleeding on probing) in the CA group and the CB group were significantly decreased at T2, relative to T0 (P<0.05). The decrease was greater in the CA group. When the periodontal clinical data were compared at T2 in both groups, the periodontal data of the CA group were lower than that of the CB group (P<0.05) (Table 1).

When the TAS data in GCF were examined in the CA group, the TAS level was highest in T1 and lowest in T0 (P>0.05). In the CB group, the TAS level was highest in T2 and lowest in T0 (P<0.05). The change in TAS level in the CB group was statistically significantly higher than that in the CA group (P>0.05) (Table 2, Figure 1).

When the TOS data in GCF were examined in the CA group, the TOS level was highest in T1 and lowest in T0 (P>0.05). In the CB group, the TOS level was highest in T2 and lowest in T0 (P<0.05). The change in TOS level in the CB group was statistically significantly higher than that in the CA group (P>0.05) (Table 2, Figure 2).

The correlations between TAS and TOS levels in GCF are shown in Figure 3. The GCF and TAS levels were positively correlated with the GCF TOS level in the CA and CB groups. Correlation between bleeding on probing and GCF TOS and TAS data are shown in Table 3.

Discussion

Today, the developments in orthodontics are particularly concentrated on the need for treatment for improving appearance. As the orthodontic treatment attracts great attention from every age group, it requires treatments that can ensure a favorable appearance and a smooth operational process for patients. Production of appealing orthodontic appliances has resulted in a situation in which patients do not only desire to have healthier teeth, but also request for orthodontic treatment to improve the appearance of their teeth [23]. The individuals who have aesthetic concerns do not like seeing the brackets on their teeth during orthodontic treatment; therefore, they are not in favor of using conventional fixed appliances [15]. In light of the importance of the aesthetic appearance of CAs, we aimed to examine the level of tissue damage that is caused by CAs. In the results of the present study, we found that CAs cause less oxidative damage than do CBs, and they provide favorable data for oral hygiene relative to conventional fixed orthodontic treatment.

The search for an aesthetic and comfortable treatment has caused patients to head toward orthodontic treatment methods that promise better aesthetic appearance, and to this end, as an alternative to metal brackets, ceramic, vinyl, polycarbonate, plastic, zirconia, and Teflon-coated arch brackets have been produced [11]. As the patients have not been satisfied with these materials produced in the color of natural teeth in terms of appearance, dentists have become more focused on aesthetic orthodontic mechanics, such as lingual brackets that can be adhered to the inner surfaces of teeth, or CAs, which are less visible from outside and that can be easily worn and taken out by the patient. The patients that have received such treatment models have not felt any concerns for their appearance and, thus, have more self-confidence [12]. Orthodontic treatment with CAs has been a favorable treatment alternative for patients who seek a charming smile without having fixed mechanics on their teeth. Among the advantages of this treatment are better appearance, ease of use, shorter appointment sessions than with treatment with conventional fixed appliances, less pain and discomfort, convenience for eating, comfort, and lack of limitations during oral hygiene [13,14]. In light of this information in the literature, in this study, we used CAs, which have been welcomed by patients with concerns for the appearance of their teeth, to compare with conventional fixed orthodontic treatment. Because the CAs caused less oxidative damage, in addition to improved appearance, the hypothesis of our study was supported.

The forces having an impact on teeth during orthodontic teeth movements cause distortion of the periodontal ligament and changes in the extracellular matrix and local tissue injuries, and thus lead to vasodilatation and an increase in the permeability in capillaries of the periodontium. Thus, an aseptic inflammatory table that is characterized by the migration of inflammatory cells and the release of remarkable cytokines, such as TAS and TOS, emerged [20]. Cytokines are proteins that carry signals between the immune system cells. These proteins cause the development of periodontium cells, including their transformation, activation, and apoptosis, in the aseptic inflammatory period during orthodontic teeth movements. In vivo and in vitro studies show that, due to the functions of some cytokines such as TAS and TOS in bone resorption and bone deposition, they are important in the remodeling of the bone, and the studies also show that these cytokines are increased in the early stages of treatment, depending on the aseptic inflammatory occurring with orthodontic treatment [21]. Another source of these cytokines being increased in orthodontic treatment is periodontal inflammation, which increases plaque retention of the materials used in orthodontic treatment and affects the oral microbiological state, which thus causes periodontal inflammation. Because of this inflammation that occurs depending on the plaque, it has been reported that the TAS and TOS levels in GCF escalates [22]. In dentistry, particularly in the field of periodontology, studies that examine the correlation between oxidative stress and periodontal inflammation have been conducted. In orthodontics, indeed, in vitro studies [20–22] that are associated with oxidative stress created by the materials used have been conducted; however, we found no study conducted on humans concerning oxidative stress and oxidative damage that is likely to occur depending on the inflammation observed in orthodontic treatment with CA.

Some conventional orthodontic treatments can be conducted by applying more force. Some treatment methods also cause more dental plaque accumulation. The presence of more dental plaque can lead to increased inflammation. Increased force-dependent rapid remodeling and predisposition to inflammation can cause more cytokine secretion. These effects can be observed with CBs. With CAs, this effect can be less noticeable [23]. One of the incidents occurring in tissues due to oxidative stress is DNA damage. Today, TAS and TOS are the most utilized molecules in determining oxidative DNA damage [20,21]. We found no studies that examined the oxidative DNA damage that is likely to be created by an aseptic inflammation caused by orthodontic treatment. Moreover, it should be considered that some adjunctive treatments, such as ozone [24], photobiomodulation [25], and probiotics [26], showed a significant effect on clinical parameters. Therefore, it can be interesting in the future to test oxidative stress in orthodontic patients who are undergoing these preventive treatments

We found no studies in the literature that examined TAS and TOS data related to orthodontic treatments with CA. Our study is the first with this distinct property. However, this study had some limitations. First, the number of patients was limited to 30. Second, although the TAS and TOS data in GCF were assessed, the TAS and TOS data in blood was not assessed. Finally, we were unable to use additional markers in determining tissue inflammation and oxidative stress.

Conclusions

The significant increase in TOS levels that were observed after 1 week of orthodontic treatment relative to baseline and the decrease of these levels to baseline levels after 1 month of treatment support our hypothesis that the free radical level in the periodontium remains within physiological limits, depending on the materials used and teeth movements provided with the orthodontic treatment using CA. Moreover, these results can be interpreted as CAs causing far less oxidative damage than CBs. In our study, no significant change was observed in the TAS and TOS levels during the first month of treatment in the CA group. This suggests that the orthodontic movements made with CAs, and the aseptic inflammation resulting from the treatment, did not cause oxidative damage to the DNA of the periodontium. This can be interpreted as an indication that periodontal inflammation did not develop in patients who received orthodontic treatment with CAs. For patient groups in which higher oxidative damage is a concern, CAs may be preferred over conventional fixed orthodontic treatment. Further studies are needed to fully understand the effects on tissues of orthodontic treatment using CAs.