Local delivery of osteoprotegerin inhibits mechanically mediated bone modeling in orthodontic tooth movement

Abstract

Introduction: The RANKL-OPG axis is a key regulator of osteoclastogenesis and bone turnover activity. Its contribution to bone resorption under altered mechanical states, however, has not been fully elucidated. Here we examined the role of OPG in regulating mechanically induced bone modeling in a rat model of orthodontic tooth movement.

Methods: The maxillary first molars of male Sprague-Dawley rats were moved mesially using a calibrated nickel-titanium spring attached to the maxillary incisor teeth. Two different doses (0.5 mg/kg, 5.0 mg/kg) of a recombinant fusion protein (OPG-Fc), were injected twice weekly mesial to the first molars. Tooth movement was measured using stone casts that were scanned and magnified. Changes in bone quantity were measured using micro-computed tomography and histomorphometric analysis was used to quantify osteoclasts and volumetric parameters. Finally, circulating levels of TRAP-5b (a bone resorption marker) was measured using enzyme-linked immunosorbent assay.

Results: The 5.0 mg/kg OPG-Fc dose showed a potent reduction in mesial molar movement and osteoclast numbers compared to controls (p<0.01). The molar movement was inhibited by 45.7%, 70.6%, and 78.7% compared to controls at days 7, 14, and 21 respectively, with the high dose of OPG. The 0.5 mg dose also significantly (p<0.05) inhibited molar movement at days 7 (43.8%) and 14 (31.8%). While incisor retraction was also decreased by OPG-Fc, the ratio of incisor to molar tooth movement was markedly better in the high-dose OPG group (5.2:1, p<0.001) compared to the control group (2.3:1) and the low-dose OPG group (2.0:1).

Conclusions: Local delivery of OPG-Fc inhibits osteoclastogenesis and tooth movement at targeted dental sites.

Fig. 1

(A) Intraoral photographs of orthodontic appliances in place at day 21 prior to sacrifice. Animals received twice weekly injections of vehicle, 0.5 mg/kg OPG-Fc, or 5.0 mg/kg OPG-Fc into the mucosa just mesial to the maxillary first molars. Note the amount of space visible between the first and second molar teeth. (B) Three-dimensional coronal and sagittal micro-computed tomography views of the same animals shown in panel A. Note the interdental distance visible between the first and second molar teeth as well as the differences in bone quantity around the first molars. Arrows indicate 2-D force direction.

Fig. 2

Inhibition of tooth movement by local delivery of OPG-Fc. (A) Molar tooth movement over the course of time in vehicle, 0.5 mg/kg OPG-Fc, and 5.0 mg/kg OPG-Fc injected groups. No tooth movement was noted in sham control animals (inactive spring) (data not shown). (B) Incisor retraction measured at day 21. (C) Ratio of incisor retraction to molar tooth movement at day 21. All results are expressed as the mean±SEM, n = 10: *p<0.05; **p<0.01; ***p<0.001. Comparisons made versus time-matched vehicle groups.

Fig. 3

TRAP stained histological sections (4–5 μm) counterstained with hematoxylin taken from the mesial tooth root of the maxillary first molar. (A) TRAP-positive multinucleated osteoclasts (40× magnification); (B) control group (4× magnification); (C) low-dose OPG group (4× magnification); (D) high-dose OPG group (4× magnification), which lacks osteoclasts. MR, mesial root; Co, compression side alveolar bone; T, tension side alveolar bone; OC, osteoclast. Arrows indicate osteoclasts.

Fig. 4

Osteoclasts per root surface on the tooth compression surface (A), tension side (B), and total osteoclasts (C) along the alveolar bone adjacent to the mesial root of the maxillary first molar. All results are expressed as the mean±SEM, n = 10 (vehicle), 9 (low-dose OPG, high-dose OPG): *p<0.05; ***p<0.001. Comparisons made versus time-matched groups.

Fig. 5

Comparison of bone volume fraction measured from the furcation area to tooth root apex of the maxillary first molar with micro-computed tomography. Specimens were placed on a rotating stage allowing polychromatic X-rays to penetrate the sample, pass through an image intensifier, and be captured by a CCD camera, producing 2-D slices of 18 μm thickness. These cross-sectional images were then reconstructed into a 3-D structure by GE Healthcare reconstruction utility. Results are expressed as the mean±SD, n=3 (baseline), n=10 (vehicle, high-dose OPG), n = 9 (low-dose OPG): *p<0.05; **p<0.01; ***p<0.001. Comparisons made versus time-matched groups.

Fig. 6

Concentrations of TRAP-5b in serum as measured by ELISA. Blood was drawn from the lateral tail vein at days 0, 7, 14, and 21 prior to the next administration of OPG-Fc or vehicle. Results are expressed as the mean±SEM, n = 10; *p<0.05. Comparisons made versus time-matched groups.