Enhancement of human periodontal ligament by preapplication of orthodontic loading


The quantity of remaining periodontal ligament (PDL) on the root surface of donor teeth is essential for the success of tooth autotransplantation. Preapplication of orthodontic loading increases this quantity on rat tooth root surfaces. However, little is known about the effects of preloading on human PDL or the ease of tooth extraction. This study aimed to determine the optimal duration of preloading for enhanced PDL on the root surface of extracted human premolars and for facilitating extraction.


Sixty patients received orthodontic preloading with a bracket connected to an archwire on one of their maxillary first premolars for 2, 4, 6, or 8 weeks, whereas the contralateral premolar was not loaded as a control. Premolar extractions were performed with a record of the duration and difficulty of extraction. The extracted premolars were fixed and stained with toluidine blue. Digitized images were recorded under a stereomicroscope, and the percentage of stained PDL was analyzed.


Orthodontic preloading for 4, 6, and 8 weeks significantly increased the percentage of stained PDL on the root surface compared with the control ( P <0.05). The duration and difficulty of extraction were significantly less in preloaded than that of unloaded teeth after 4, 6, and 8 weeks ( P <0.05).


A 4-week duration of orthodontic preloading is suggested as the shortest duration to adequately enhance PDL and ease tooth extraction; both outcomes may be beneficial for tooth autotransplantation.


  • Orthodontic preloading for 4 weeks significantly enhanced periodontal ligament (PDL) on the root surface.

  • The preloading for 4 weeks also facilitated tooth extraction.

  • There was a strong correlation between the duration of extraction and the difficulty score.

  • There was an inverse correlation between the duration of extraction and stained PDL.

The use of premolars extracted because of orthodontic treatment in tooth autotransplantation to replace missing or hopeless teeth has been previously reported in the literature. Compared with osseointegrated dental implants, autotransplantation of a natural tooth has several advantages, including better functional adaptation, alveolar ridge preservation, and esthetics, because the transplanted tooth can maintain periodontal ligament (PDL) homeostasis and normal functional proprioception. Thus, tooth autotransplantation is an ideal choice compared with dental implants if combined therapies between orthodontic tooth movement and tooth autotransplantation are designed for the transplanted tooth. , However, some complications after tooth autotransplantation can still occur, such as root resorption, dentoalveolar ankylosis, tooth mobility, and periodontal pocket formation. In monkeys, ankylosis of the donor tooth is often due to PDL injury during extraction. Therefore, the quality and quantity of remaining PDL on the root surface of the extracted donor tooth, as well as the ease of extraction (to avoid possible PDL injury) are important factors for the success of tooth autotransplantation.

PDL is a highly vascular and cellular connective tissue situated between the tooth and alveolar bone that provides supportive, attachment, and sensory functions. Because a principal function of the PDL is to anchor the tooth root to the jaw bone and to distribute multidirectional mechanical stresses, such as masticatory forces, it is, perhaps, not surprising that the PDL exhibits features of a shock-absorbing system. Indeed, some of the most interesting features of the PDL are its adaptability to rapidly applied force levels and its remarkable capacity for repair and regeneration. The PDL that remains attached to the root surface is capable of regenerating new PDL after replantation. Moreover, PDL of the transplanted tooth appears to be able to induce bone production, because PDL cells can be genetically differentiated into 3 distinct types of cells, particularly osteoblasts that generate bone around the transplanted tooth.

A few previous studies in an animal model have suggested preapplication of orthodontic force to increase the width of PDL in order to ease tooth extraction, which may decrease damage to PDL during the extraction. , A previous case report has demonstrated increased mobility of 2 human donor teeth upon orthodontic preloading, as assessed by the Periotest. The increased mobility was shown, immediately, to ease tooth extraction without PDL injury and, after a 1-year follow-up, to have aided in successful tooth autotransplantation. Therefore, it is reasonable to hypothesize that preapplication of orthodontic force to the donor tooth would be useful to increase PDL tissue on the root surface of the preloaded tooth with complete root formation and to reduce PDL injury owing to easier tooth extraction. The objective of this study was to determine an optimal duration for the stimulation of the proliferative activity of PDL tissue on the root surface and the facilitation of extraction after preapplication of orthodontic force for different periods.

Material and methods

Sixty healthy patients (aged 16-37 years; 21 years, 6 months ± 4 years, 8 months [mean ± standard deviation]; 23 male and 37 female) free from any underlying medical conditions, as revealed by their medical history, were recruited from the Postgraduate Clinic, Faculty of Dentistry, Bangkokthonburi University, Thailand with written informed consent. The study protocol was approved by the human ethics committee of Bangkokthonburi University (approval number: 5/2561). The patients were planned for extractions of their maxillary first premolars as part of orthodontic treatment. The inclusion criteria were that patients had a sound first premolar in each quadrant without caries or restorations, were not taking any steroidal or nonsteroidal anti-inflammatory drugs during force application because of their inhibitory effects on PDL and bone remodeling, and had good oral hygiene and healthy periodontium with pocket depths ≤3 mm. The exclusion criteria were an asymmetrical arch form or any root malformations on either side, such as root dilaceration, which might result in a complicated extraction of the maxillary first premolars, as evidenced by intraoral and extraoral radiographs.

By drawing sealed randomization envelopes prepared by a person not involved in this study, 60 patients were randomly assigned to the 4 groups (15 each), which comprised orthodontic preloading on either side of their maxillary first premolars for 2, 4, 6, or 8 weeks, respectively, whereas the contralateral premolar was not loaded as a control. The reason for choosing only maxillary premolars in this study was that local anesthesia of mandibular premolars by inferior alveolar nerve block is considered technically more challenging and less predictable than that of local infiltration for the upper premolars. Hence, the duration and difficulty of extraction between different cases and quadrants could not be controlled. The preloading force was obtained from a 0.016-inch improved superelastic nickel-titanium alloy wire (Sentalloy, Tomy International, Tokyo, Japan) inserted into a slot (Roth prescription; 0.018 × 0.025-inch), attached to a preadjusted edgewise bracket (Tomy International), bonded on the maxillary first premolar using compomer (Grengloo, Ormco, Orange, Calif; Fig 1 , A ) whereas the contralateral premolar was not connected to the wire ( Fig 1 , B ). The inserted wire was tied with the bracket using an elastomeric ring (DynaFlex, Emergo Group, the Hague, the Netherlands; Fig 1 , A ). According to the manufacturer’s instructions (Tomy International), the wire has the shape memory effect that uses body temperature to generate continuous light force (100 g) with a deflection of 0.5-1.8 mm.

Fig 1
Intraoral photographs. A, Preloaded, upper right first premolar (*). B, Unloaded control, upper left first premolar (#).

After orthodontic preloading for the indicated times, the wire was removed, followed by removal of the bracket on the preloaded tooth, using a bracket remover (Tomy International). The tooth surface was cleaned and polished with white stone (Shofu, Kyoto, Japan). Infiltration with 4% articaine with epinephrine 1:100,000 (Septanest SP, Septodont, Saint-Maur-des-Fossés, France) was conducted through the buccal vestibule on both sides. Premolar extractions were performed by the same surgeon, who was blinded to the tooth condition, with a record for the duration of the extraction procedure that began when the forceps (F150, Hu-Friedy, Chicago, Ill) was clamped on the tooth and ended when the tooth was removed out of its socket. After tooth removal, the surgeon was asked to score on a modified 4-point Likert scale (1 = very easy; 2 = easy; 3 = difficult; 4 = very difficult) based on the surgeon’s perception. After extraction, 500 mg of acetaminophen (paracetamol GPO, Government Pharmaceutical Organization, Bangkok, Thailand) was prescribed for the patients every 6 hours as needed for pain.

Staining with toluidine blue was performed to determine the amount of PDL tissue with proliferative cells on the root surface using a protocol modified from Iwata et al. Yee reported that the number of mitotic cells representing PDL cell proliferation was increased after orthodontic loading of rats’ teeth for 24 hours. In addition, cells with increased mitotic activity are usually detected by toluidine blue staining. In brief, the extracted teeth were washed gently in phosphate-buffered saline (PBS), pH 7.2, and fixed within 30 minutes after extraction with 10% buffered formalin solution in 50-ml tubes (Corning, Inc., Corning, NY) for 24 hours. The teeth were then immersed in PBS for 2 days, stained with 0.04% (w/v) toluidine blue (Sigma-Aldrich, St Louis, Mo) for 10 minutes, and destained with 4 ml of PBS, which was changed daily for 14 days. After destaining, the teeth were left to air dry.

The buccal, palatal, mesial, and distal surfaces of the stained roots were observed under a stereomicroscope (Olympus SZX7; Olympus, Tokyo, Japan), and a digitized image of each root surface was recorded perpendicular to the tooth axis using a charge-coupled device (Olympus E-330; Olympus) attached to the stereomicroscope. By using ImageJ software version 1.51r (National Institute of Health, Bethesda, Md), which is commonly used in previous studies to quantify stained images, , , the area of stained PDL in each digital image, represented by blue pixels regardless of their staining intensities, was analyzed and expressed as the percentage of the stained area against the total area, which is a combination of both blue and white pixels, representing the unstained area. The mean percentage of stained PDL in each tooth was derived from an average value of the percentages of stained PDL from all 4 surfaces of that tooth.

Statistical analysis

Differences in the percentage of stained PDL on the root surface, in the duration and ease of tooth extraction between control and preloaded teeth for the various durations of preloading, were analyzed using the paired t -test. Two-way analysis of variance (ANOVA) was used for the analysis of the interactions between preloading or control and the 4 periods of orthodontic preloading. Comparisons between 2 different time points within the same group were analyzed using one-way ANOVA (Duncan’s multiple comparison test). Correlations between these different parameters were analyzed using either the Spearman or the Pearson correlation test. The statistical analyses were conducted using SPSS version 17.0 (IBM Corporation, Armonk, NY). P values of <0.05 were considered statistically significant.


Because of relocation of 2 patients in the 2-week preloading group, as well as root tip fracture of the unloaded control teeth during extraction in 5 patients in the preloading groups for 2 (3 patients) and for 8 (2 patients) weeks, both preloaded and unloaded control teeth of these 7 patients were excluded from analyses of the duration of extraction and PDL staining. Therefore, the number of remaining patients in the preloading groups for 2 and 8 weeks was 10 and 13, respectively. However, from the sample size calculation determined by G*Power software (version; Franz Faul, Universität Kiel, Kiel, Germany) with the effect size = 0.59 derived from the preliminary data, α = 0.05 and 1-β = 0.95, the calculated number of patients in each period of preloading should be ≥10. In addition, no difference in average ages was found among the 4 preloading groups, including 2, 4, 6, and 8 weeks (data not shown).

It was demonstrated that the duration of average extraction time in preloaded teeth determined from all 4 periods of orthodontic preloading (20.78 ± 1.94 seconds) was reduced to nearly 50% compared with that in unloaded control teeth (40.11 ± 5.29 seconds). The mean duration of extraction in preloaded teeth was significantly less than that of unloaded control teeth at 4, 6, and 8 weeks, but not at 2 weeks, of orthodontic preloading ( P <0.05; Fig 2 , A ). Similarly, the mean scores for the difficulty of extraction were significantly lower in preloaded than those of unloaded control teeth at 4, 6, and 8 weeks ( P <0.01; Fig 2 , B ).

Mar 9, 2020 | Posted by in Orthodontics | Comments Off on Enhancement of human periodontal ligament by preapplication of orthodontic loading
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