Korean J Orthod.  2020 Nov;50(6):391-400. 10.4041/kjod.2020.50.6.391.

Antifibrotic effects of sulforaphane treatment on gingival elasticity reduces orthodontic relapse after rotational tooth movement in beagle dogs

• KMID: 2508447
• DOI: http://doi.org/10.4041/kjod.2020.50.6.391

Abstract

Objective
Increased gingival elasticity has been implicated as the cause of relapse following orthodontic rotational tooth movement and approaches to reduce relapse are limited. This study aimed to investigate the effects of sulforaphane (SFN), an inhibitor of osteoclastogenesis, on gene expression in gingival fibroblasts and relapse after rotational tooth movement in beagle dogs.
Methods
The lower lateral incisors of five beagle dogs were rotated. SFN or dimethylsulfoxide (DMSO) were injected into the supra-alveolar gingiva of the experimental and control group, respectively, and the effect of SFN on relapse tendency was evaluated. Changes in mRNA expression of extracellular matrix components associated with gingival elasticity in beagles were investigated by real-time polymerase chain reaction. Morphology and arrangement of collagen fibers were observed on Masson’s trichrome staining of buccal gingival tissues of experimental and control teeth.
Results
SFN reduced the amount and percentage of relapse of orthodontic rotation. It also decreased the gene expression of lysyl oxidase and increased the gene expression of matrix metalloproteinase (MMP) 1 and MMP 12, compared with DMSO control subjects. Histologically, collagen fiber bundles were arranged irregularly and were not well connected in the SFN-treated group, whereas the fibers extended in parallel and perpendicular directions toward the gingiva and alveolar bone in a more regular and well-ordered arrangement in the DMSO-treated group.
Conclusions
Our findings demonstrated that SFN treatment may be a promising pharmacologic approach to prevent orthodontic rotational relapse caused by increased gingival elasticity of rotated teeth in beagle dogs.

Keyword

Rotation; Relapse; Gingival elasticity; Sulforaphane

Figure

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  Figure 1 A, Occlusal view of a beagle’s lower teeth and orthodontic buttons with elastic chains. Mandibular lateral incisors (I2 teeth) of beagle dogs were rotated for 6 weeks by applying rotational couple force using a power chain. B, C, The needle injection points of gingiva of experimental teeth. After the needle was inserted at right angles to the attached gingiva area located directly below the boundary between free gingiva and attached gingiva, vertically, and at both centers of half of the buccal gingiva of the experimental teeth, horizontally, sulforaphane (SFN) and dimethylsulfoxide (DMSO) were injected into each side of 3 units (UI) (B, frontal view; C, sagittal view).

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  Figure 2 Schema. The experimental schedule for the beagle dogs. The experimental teeth were rotated by applying rotational couple force with a power chain for 6 weeks. During a retention period of 4 weeks, SFN (experimental group) and DMSO (control group) were injected into the supra-alveolar gingiva of the buccal side of the rotated teeth once per week, four times total. One week after the last injection, the appliances were removed to allow relapse of the rotated teeth. An intraoral impression was obtained to measure the extent of tooth rotation and relapse. SFN, Sulforaphane; DMSO, dimethylsulfoxide; T0, initial; T1, immediately after retention and SFN treatment for 4 weeks; T2, after relapse for 2 weeks; T3, after relapse for 4 weeks.

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  Figure 3 The extent of tooth rotation and relapse. Measurements were made using the Geomagic Control XTM (3D SYSTEMS, Seoul, Korea) software. The degree of rotation and amount of relapse after 2 and 4 weeks, and percentage of relapse after 2 and 4 weeks were calculated. A, Scanning image of T1 model from beagle no. 1. B, Scanning image of T2 model from beagle no. 1. T1, Immediately after retention and SFN treatment for 4 weeks; T2, after relapse for 2 weeks.

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  Figure 4 Relative mRNA expression of genes encoding COL1A1, LOX, ELN, MMP 1, MMP 12, and TIMP 1 in gingival tissue of beagle dogs after sulforaphane treatment. COL1A1 (A), LOX (B), ELN (C), MMP 1 (D), MMP 12 (E), and TIMP 1 (F) mRNA levels in gingival tissue were quantified by real-time polymerase chain reaction. The symbol (*) indicates statistical significance between the control group and experimental group (*p < 0.05, Wilcoxon signed rank test). DMSO, Dimethylsulfoxide; COL1A1, collagen type I alpha 1; LOX, lysyl oxidase; ELN, elastin; MMP, matrix metalloproteinase; TIMP, tissue inhibitor of metalloproteinase.

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  Figure 5 Histologic appearance of collagen fibers in buccal supra-alveolar gingiva. Massonʼs trichome staining of the buccal-lingual longitudinal sections of the lower lateral incisors of beagle dogs injected with either dimethylsulfoxide (DMSO) or sulforaphane (SFN) for 4 weeks. The control and experimental samples were from same beagle subject. Collagen fibers bundles were longer and more connected in the DMSO-treated group than in the SFN-treated group. A, C, DMSO group. B, D, SFN group. Scale bar = 1 mm (A, B), 400 μm (C, D). Higher magnification views of the box areas can be seen Figure 6.

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  Figure 6 Morphology and arrangement of principal gingival fibers in the buccal supra-alveolar gingiva. The higher magnification views of dentogingival fibers (black arrows indicate dentogingival fibers) and dentoperiosteal fibers (white arrows indicate dentoperiosteal fibers) showed that collagen fiber bundles connected to form the principal gingival fibers were irregularly and not well connected in the sulforaphane (SFN)-treated group compared to the dimethylsulfoxide (DMSO)-treated group. Circular fibers (indicated in black circle dotted lines) of the SFN-treated group were arranged in a thinner and less compact manner compared to the DMSO-treated group. A, C, E, DMSO group. B, D, F, SFN group. Scale bar = 200 μm.

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