Archives of Oral Biology 83 (2017) 304–311

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Adiponectin prevents orthodontic tooth movement in rats a

b,c

b,c

MARK b,c

Sigrid Haugen , Kristin Matre Aasarød , Astrid Kamilla Stunes , Mats Peder Mosti , ⁎ Tanya Franzend, Vaska Vandevska-Radunovicd, Unni Syversenb,e, Janne Elin Reselanda, a

Department of Biomaterials, Institute for Clinical Dentistry, University of Oslo, Oslo, Norway Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology, Trondheim, Norway Clinic of Medicine, St. Olav's Hospital, Trondheim, Norway d Department of Orthodontics, Institute of Clinical Dentistry, University of Oslo, Oslo, Norway e Department of Endocrinology, St. Olav’s Hospital, Trondheim, Norway b c

A R T I C L E I N F O

A B S T R A C T

Keywords: Adiponectin Experimental tooth movement

Objective: The objective was to examine the effects of repetitive local administration of adiponectin on experimental tooth movement in rats. Materials and methods: The maxillary right first molar of male Wistar rats (n = 24) was moved mesially for 14 days, with local adiponectin injections (0.2 or 2 μg) every third day. Micro-computed tomography was performed at days 0, 6 and 14 and molar movement, bone density and bone volume fraction were calculated from the scans. Changes in extracellular matrix collagen and cell numbers in the periodontal ligament were analyzed histologically, and levels of circulating cytokines were measured by Luminex and ELISA. Results: Adiponectin injections induced a reduction in tooth movement after 12 and 14 days compared to controls. No tooth movement was observed between days 3 and 14 in the group receiving the highest dosage (2 μg) of adiponectin. Differences in bone density and bone volume fractions between treatment and control groups were not identified. Relative size and morphology of collagen fibrils, and cell number in the periodontal region after adiponectin injections were unchanged compared to controls. Levels of circulating adiponectin or other selected factors in plasma were not influenced by the adiponectin injections. Conclusions: Submucosal injections of adiponectin prevented experimental tooth movement in rats. The effect was dosage-dependent and local. Adiponectin injections caused no detectable changes in bone density, periodontal cell number or collagen content.

1. Introduction Multiple tissues in the body are dependent of mechanical stress for the regulation of normal functions. Endothelial cells, chondrocytes, smooth muscle cells, osteoblasts and fibroblasts are examples of such load-sensitive cells. (Wang & Thampatty, 2006). The periodontal ligament (PDL) is a fibrous connective tissue located between the root of the tooth and the surrounding alveolar bone (Beertsen, McCulloch, & Sodek, 1997; Mariotti, 1993). It is continuously exposed to mechanical stress from occlusal pressure and from orthodontic forces during active orthodontic treatment. Force delivery to the teeth induces certain mechanical, chemical and cellular events resulting in structural changes and, eventually, tooth movement (Krishnan & Davidovitch, 2009). Several substances, e.g. bisphosphonates (Adachi, Igarashi, Mitani, & Shinoda, 1994; Liu et al., 2004), osteoprotegerin (OPG) (Dunn, Park, Kostenuik, Kapila, & Giannobile, 2007), vitamin D3 ⁎

(Kawakami & Takano-Yamamoto, 2004) and anti-inflammatory drugs (Arias & Marquez-Orozco, 2006; Walker & Buring, 2001), have been shown to interfere with this interplay and therefore change the rate of tooth movement. Adiponectin is an abundant adipokine in plasma (0.01% of total plasma proteins) (Arita et al., 1999; Gil-Campos, Cañete, & Gil, 2004), and is involved in a variety of physiological processes including metabolic (Arita et al., 1999; Berg, Combs, Du, Brownlee, & Scherer, 2001) and immune responses (Choi et al., 2007; Engeli et al., 2003; Wolf, Wolf, Rumpold, Enrich, & Tilg, 2004). Adiponectin spontaneously aggregates to form larger assemblies and different structures that are found in plasma including trimeric, hexameric and high molecular weight forms (HMW) (Tsao et al., 2003; Waki et al., 2003; Wang, Lam, Yau, & Xu, 2008). Two receptors and many target tissues have been reported, and adiponectin receptors are found to be expressed on mouse gingival fibroblasts (mMGF), human gingival fibroblasts (hHGF) and

Corresponding author at: Department of Biomaterials, Institute for Clinical Dentistry, P O Box 1109 Blindern, N-0317 Oslo, Norway. E-mail addresses: [email protected] (S. Haugen), [email protected] (K.M. Aasarød), [email protected] (A.K. Stunes), [email protected] (M.P. Mosti), [email protected] (T. Franzen), [email protected] (V. Vandevska-Radunovic), [email protected] (U. Syversen), [email protected] (J.E. Reseland). http://dx.doi.org/10.1016/j.archoralbio.2017.08.009 Received 4 May 2017; Received in revised form 18 August 2017; Accepted 19 August 2017 0003-9969/ © 2017 Elsevier Ltd. All rights reserved.

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human PDL cells (Iwayama et al., 2012; Park et al., 2011; Yamaguchi et al., 2010), in addition to osteoblasts (Berner et al., 2004) and osteoclasts (Pacheco-Pantoja, Waring, Wilson, Fraser, & Gallagher, 2013). Beneficial effects of adiponectin on PDL cells have been demonstrated, including stimulated expression of various growth factors and extracellular matrix (ECM) proteins, enhanced proliferation, mineralization, and in vitro wound healing (Deschner, Eick, Damanaki, & Nokhbehsaim, 2014; Iwayama et al., 2012; Nokhbehsaim et al., 2014). Adiponectin may influence bone metabolism and probably acts via several mechanisms in bone cells. Adiponectin stimulates proliferation and differentiation of osteoblasts (Lin et al., 2014; Luo et al., 2005; Oshima et al., 2005) in addition to favoring differentiation of mesenchymal stem cells towards the osteoblastic lineage (Lee et al., 2009; Pu et al., 2016; Wu et al., 2014). Recently, it was observed that adiponectin might influence bone indirectly by decreasing sympathetic tone (Kajimura et al., 2013; Wu et al., 2014). Most studies demonstrate a suppressive effect of adiponectin on osteoclasts (Luo et al., 2006; Luo et al., 2012; Oshima et al., 2005; Tu et al., 2011; Wang et al., 2014; Williams et al., 2009). Long-term administration of adiponectin has been reported to promote peri-implant osteogenesis in ovariectomized rabbits (Luo et al., 2012) and adiponectin-coated scaffolds induced better bone fill with increased expression of bone markers and enhanced mineralization compared to controls (Hu et al., 2015). Based on the previous observations the hypothesis was that local adiponectin injections could influence the rate of experimental orthodontic tooth movement in rats.

(Fig. 1). Force magnitude was measured using a Correx dynamometer (Haag-Streit, Bern, Switzerland). The animals were randomly divided into three groups; control (PBS), adiponectin 0.2 μg and adiponectin 2 μg. On days 3, 6, 9 and 12 of experimental orthodontic tooth movement (OTM), solutions of recombinant human adiponectin (R & D Systems, MN, USA) dissolved in phosphate buffered saline (PBS) or plain PBS solution were administrated by submucosal injection directly anterior to the first molar. The groups (n = 8), received 10 μl of either 20 μg/ml adiponectin (0.2 μg) or 200 μg/ml adiponectin (2 μg) solutions whilst the control group was given 10 μl PBS. The animals were anaesthetized with 0.35 ml/100 g Hypnorm-Dormicum (fentanyl 0.05 mg/ml, midazolam 1.25 mg/ml, fluanison 2.5 mg/ml) given subcutaneously prior to surgery and microcomputed tomography (μCT), and with isoflurane gas (1% isofluran mixed with 30% O2/70% N2O) during injection of adiponectin and before sacrifice. A feeler gauge (Mitutoyo Co., Kawasaki, Japan) was used to measure tooth movement between the distal surface of the first molar and the mesial surface of the second molar on days 3, 6, 9, 12 and 14. The minimum measurable distance was 50 μm. Weight recordings of the animals were made on the day of appliance insertion and prior to sacrifice. After 14 days, blood samples were collected by cardiac puncture during the final anesthesia before the rats were sacrificed. The animals were decapitated and the heads stored in 70% ethanol until further use.

2. Materials and methods

Live μCT (SkyScan 1176, Bruker, Kontich, Belgium) imaging was performed three times during the study, on days 0, 6 and 14, using two different settings. Scanner settings on days 0 and 14; 65 kilovolt (kV), 384 microampere (μA), 2000 × 1336 pixels (px), 17.66 μm pixel size, 1 mm aluminum (Al) filter, 0.500 rotation step, 360 ° scan. Scanner settings on day 6; 80 kV, 312 μA, 1336 × 2000 px, 17.66 μm pixel size, copper and Al (Cu-Al) filter, 0.500 rotation step, 360 ° scan. Scans were reconstructed by Nrecon 1.6.9.4. (Bruker). Reconstruction settings on days 0 and 14; smoothing 2, ring artifact correction 10, beam hardening correction (%) 20, minimum and maximum for dynamic image range to image conversion 0.000000–0.120000. Reconstruction settings on day 6; smoothing 4, ring artifact correction 20, beam hardening correction (%) 25, minimum and maximum for CS to image conversion 0.000000–0.080000. Co-registration was made by Data Viewer 1.5.0.0 Pre-release (Bruker). The scans were aligned in the frontal/coronal (C) plane using the mid-palatal suture. The third molar was defined as a fixed reference point, and at that specific point transversal (T) and sagittal (S) sections of days 0 and 14 were selected and the 3D dataset aligned accordingly. On the sagittal cut, the occlusal plane on the control side was placed on a virtual straight line. Movement of the first molar between days 0 and 14 was measured using the prior defined third molar as fixed reference. The corner points of an enclosing rectangle around both the first and third molar in the selected slices were marked and various corner coordinates registered (Fig. 2, A–D in the transversal view; 1–4 in the sagittal view). Based on these points, the

2.3. Micro-computed tomography (μCT)

2.1. Animals The experimental protocol was approved by the Norwegian Animal Research Authority (NARA, FOTS ID 5409) and the procedures were conducted in accordance with the Animal Welfare Act. Between 6 and 8 weeks old male Wistar rats (n = 24), weighing 180–200 g, were housed in individually ventilated cages (IVC), four animals per cage, under standard 12 h light/dark cycle and standard conditions. Rat and mouse diet (B & K Universal Ltd, Aldbrough, Hull, UK), along with tap water were provided ad libitum. The animals were acclimatized for one week prior to initiation of the intervention. 2.2. Study design A split mouth design was used, with the right side as the experimental side; the contralateral side functioned as an internal control, as previously described (Franzen, Brudvik, & Vandevska-Radunovic, 2011; Vandevska-Radunovic, Kristiansen, Heyeraas, & Kvinnsland, 1994). On the experimental side a closed coil spring (0.008 × 0.030 inches; Ormco, CA, USA), was ligated to the first molar and the eyelet of an incisor band. The appliance was activated with approximately 0.5 N (N) forces, with no reactivation, during the experimental period of 14 days; this caused a mesial movement of the maxillary right first molar

Fig. 1. Appliance used for orthodontic tooth movement. A closed coil spring was attached to the maxillary right first molar and incisors.

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Fig. 2. Distance of tooth movement. Identification of measurements used to calculate maxillary first molar OTM in μCT scans. The corner points of an enclosing rectangle around both the first and third molar in the selected slices were marked and various corner coordinates registered (A–D in the transversal view; 1–4 in the sagittal view). Based on these points the midpoints of rectangular prisms around the molars were obtained. The length of the vector between these midpoints was calculated. T: transversal, C: coronal, S: sagittal.

One overview image was taken of the PDL on the compression side of the mesial root of the maxillary first molar of each rat. Furthermore, three individual images were taken from the same region showing the upper area, middle area and apical area at higher magnification. Slides were stained in iron hematoxylin (Merck, Darmstadt, Germany) for 8 min, followed by picrosirius red (PSR) (Sigma-Aldrich, St. Louis, MO, USA) staining as described previously (Trombetta, Bradshaw, & Johnson, 2010). Sections were then returned through graded ethanol rinses to xylene and mounted with Entellan (Merck). The sections were exposed to standard white light and polarized white light modes using a Leica DM RBE microscope (Leica, Wetzlar, Germany) and the images analyzed in ImageJ (Wayne Rasband, National Institutes of Health, Bethesda, MD). Collagen fiber relative size and morphology was evaluated from three regions of interest on each of the images. Prior to nuclei counting, slides were treated with 0.1% Triton X100 for 5 min before staining using 20 μg/ml propidium iodide (Sigma Aldrich, St. Louis, MO, USA) for 10 min. Sections were then returned through graded ethanol rinses to xylene and mounted with Prolong Diamond (ThermoFisher, Waltham, MA, USA). Slides were viewed with a 10×/0.40 HCX PL APO CS objective lens in a Leica SP8 confocal microscope (Leica) using 552 nm excitation and 600–650 nm emission filters. Three individual images of each slide were taken as previously described, and the images were manually thresholded and the number of nuclei counted from three regions of interest on each of the images. Sections were stained with tartrate- resistant acid phosphatase (TRAP) as previously described (Brudvik & Rygh, 1993).

midpoints of rectangular prisms around the molars were obtained. The length of the vector between these midpoints was calculated. In each scan, a standardized 3D volume of interest (VOI) was defined based on previous descriptions (Dunn et al., 2007; TrombettaEsilva et al., 2011). Co-registration was made by Data Viewer 1.5.0.0 Pre-release (Bruker). In short, the mesiobuccal and distolingual roots of the first molar were chosen as landmarks where a cube shaped VOI was placed between the roots. From each rat, the VOI with a size of 50/75/ 75 pixels was selected and bone mineral density (BMD) (g/cm3), tissue mineral density (TMD) (g/cm3) and bone volume fraction (BVF) were quantified at days 0 and 14. BMD and TMD were quantified using phantoms with BMD of 0.75 and 0.25 g/cm3 calcium hydroxyapatite (CaHA) and the threshold was set at 44–255. The maxillary first molar furcation area was selected as it was considered a reproducible morphological region. 2.4. Tissue preparation and histological analyses The maxillae were removed, dissected, and fixed in 4% formaldehyde (VWR, Radnor, PA, USA) for 48 h at 4 °C. The tissue was decalcified in 10% ethylene diamine tetra-acetate (EDTA) (VWR) at 4 °C monitored by X-ray and physical methods. The specimens were dehydrated in ascending concentrations of ethanol and embedded in paraffin for histological analysis. Parasagittal sections were cut at 5 μm parallel to the long axis of the first molars and mounted on glass slides. Before staining, sections were de-paraffinized and rehydrated through graded ethanol rinses. 306

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Superimposed μCT images of the same animals at days 0 and 14 showed tooth movement in the control group and reduced tooth movement in the adiponectin (2 μg) group (Fig. 4). No changes in tooth movement were observed on the contralateral side (data not shown).

2.5. Adipokines and cytokines in plasma Multianalyte profiling of the amount of tumor necrosis factor – alpha (TNF-α), interleukin-1 beta (IL-1β), interleukin-6 (IL-6), leptin, monocyte chemoattractant protein-1 (MCP-1) and plasminogen activator inhibitor-1 (PAI-1) in plasma was performed using the Rat Adipocyte Panel – Metabolism Assay (Millipore, Billerica, MA, USA) on the Luminex-200 system (Luminex Corp., Austin, TX, USA). The concentration of adiponectin in rat plasma was determined by enzymelinked immunosorbent assay (ELISA) Kit Rat Adiponectin (Millipore) using BioTek El x 800 plate reader (BioTek Instruments Inc., Winooski, VT, USA). All procedures were performed according to the manufacturers’ instructions.

3.2. Bone effects The bone area close to the injection site without including PDL and tooth structures was chosen for analysis of bone morphometric parameters as the area in front of the moving tooth was considered to be too small for analysis. Therefore, μCT analyses were done on the furcation area between the roots of the maxillary first molar. The local injection of adiponectin could possibly influence the bone at the administration site; however, we were unable to reveal effects on skeletal density of adiponectin. Based on the initial results, we focused on identifying differences between the control and 2 μg adiponectin groups. As expected, BVF was similar in both groups on day 0. The BVF in the high dose adiponectin group (2 μg) as well as the control group was unchanged after 14 days (Table 1 and Fig. 5). No difference in BMD between the groups was identified (Table 1). The resolution of scans was too low to detect any differences in BMD between the actual injection sites exposed to adiponectin and the corresponding control sites.

2.6. Statistical methods Comparisons of maxillary first molar movement, collagen fiber relative size, cell count, μCT BVF, BMD and BMT measurements and plasma analyses were performed, and differences between experimental and control groups were analyzed using independent sample t-test. Values are presented as mean ± SD, if not stated otherwise.When normality and/or equality tests failed, Mann-Whitney U test was used and data presented as median (25–75‰). Changes in body weight were examined by one-way ANOVA with Holm-Sidak post hoc test. A probability value of ≤0.05 was considered significant.

3.3. Histological analysis The relative size and morphology of collagen in the PDL were evaluated by red and green signals in PSR-stained sections by polarized light. Quantification of the red signals in polarized mode revealed no significant difference in collagen between the adiponectin treated (2 μg) and control group (Fig. 6). Weak green signals were visually observed in both groups; however, these could not be reliably evaluated. The collagen fibers appeared to be tighter and in a more organized manner after adiponectin injection, however, there was no difference in the cell number observed in sections from the PDL between the high dose adiponectin group and controls (data not shown). Osteoclast number could not be reliable evaluated in the sections from this study.

3. Results Two animals, one from each adiponectin group (0.2 and 2 μg), were excluded from the study due to lost appliance. There was no significant difference in body weight between day 0 and 14 in either of the groups, or between groups at any time point (data not shown). 3.1. Tooth movement After 14 days, movement of the first maxillary molar measured by feeler gauge was found to be 250 (150–287.5) μm in the control group, 150 (100–225) μm and 100 (100–125) μm in the 0.2 μg adiponectin and 2 μg adiponectin groups, respectively (Fig. 3A). No tooth movement was, however, observed between days 3 and 14 in the rats receiving 2 μg adiponectin (Fig. 3A). The lack of molar tooth movement was found to be significantly different from the control group after both 12 (p = 0.006) and 14 days (p = 0.012) in this group. No significant difference in tooth movement was found by feeler gauge measurement in the group receiving 0.2 μg adiponectin compared to control. Administration of both 0.2 μg and 2 μg adiponectin were found to significantly reduce tooth movement (145 ± 27 and 100 ± 13 μm, respectively, p ≤ 0.001 for both) compared to the control group (274 ± 9 μm) by μCT measurements at day 14 (Fig. 3B).

3.4. Plasma levels of adipokines and bone markers Oral local injections of adiponectin did not result in any significant differences in plasma levels of adiponectin or other adipokines and cytokines compared to controls (Table 2). 4. Discussion Unwanted movement of anchor teeth and relapse of previously moved teeth are clinical problems frequently encountered in

Fig. 3. Measurements of first maxillary molar movement by feeler gauge (A) and μCT (B). Tooth movement measured by feeler gauge (μm) at days 3, 6, 9, 12 and 14 (A) was significantly reduced in the adiponectin group (2 μg) after both 12 (p = 0.006) and 14 days (p = 0.012) compared to the control group. There are otherwise no significant changes between the groups in the experimental period. Relative 3D tooth movement of maxillary first molars measured on the μCT scans between days 0 and 14 (μm) (B). The tooth movement in both adiponectin (0.2 and 2 μg) groups was significantly reduced (p ≤ 0.001 for both) on day 14 compared to the control group. Values are presented as median (25–75‰) in a boxplot for feeler gauge (A) and mean ± SEM for micro CT (B). Median values are highlighted with red dots. Significant difference from control; *, p < 0.05; **, p < 0.01; ***, p ≤ 0.001. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

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Fig. 4. Superimposed 3D μCT reconstructed teeth of the same animals from days 0 and 14 in the control (PBS) (upper panel) and adiponectin (2 μg) group (lower panel). The teeth can be seen from two different perspectives (occlusal and sagittal). Note the visible space between the maxillary first and second molar in the control group, the position of first molar at day 0 is shown in blue. Representative images were taken from selected specimens. Arrow represents direction of OTM.

Table 1 BMD, TMD and BVF determined by μCT in the control and high dose adiponectin (2 μg) groups on days 0 and 14. Adiponectin (2 μg)

Control

BMD TMD BVF

day 0

day 14

p-value

day 0

day 14

p-value

0.73 ± 0.06 0.85 ± 0.03 82.48 ± 6.01

0.67 ± 0.08 0.78 ± 0.06 77.69 ± 7.34

0.306 0.084 0.352

0.75 ± 0.04 0.85 ± 0.03 82.45 ± 4.47

0.72 ± 0.02 0.81 ± 0.01 82.28 ± 3.06

0.160 0.057 0.952

Values are presented as mean ± SD. BMD and TMD (g/cm3 CaHA); BVF (BV/TV) (%). N = 4 in each group.

whereas the feeler gauge measured the distance between the first and second molar. In μCT scans a change in the distance between the midpoint of the first and third molar can be due to lateral movement of the third molar in relation to the first molar or a rotation of the third molar out of its original position. Whilst these two techniques have a different accuracy level and were evaluated by different options, the results at day 14 were comparable for all groups using both methods. OTM is dependent on bone volume (quantity) and density (quality). Various studies have reported changes in microarchitecture of trabecular bone after experimental OTM in rats using a resolution ranging from pixel size of 18–35 μm in live μCT evaluation (Ru, Liu, Zhuang, Li, & Bai, 2013; Vieira, Chaves, Ferreira, de Freitas, & Amorim, 2015) and even higher resolution for post mortem recordings (Franzen, Monjo, Rubert, & Vandevska-Radunovic, 2014; Furfaro, Ang, Lareu, Murray, & Goonewardene, 2014). The area of investigation varies between different studies (Franzen et al., 2014; Furfaro et al., 2014; Ru et al., 2013; Vieira et al., 2015). The first molar has been used as landmark, where a VOI has been chosen for bone evaluations from the areas in front of the first molar. We wanted to evaluate the bone close to the injection site, without including PDL and tooth structures as demonstrated previously using a selected non-extrapolated area in approximation to the mesial tooth surface (Franzen et al., 2014). Since the first molars are influenced by forces and expected to change position into this area between days 0 and 14, we did not use this strategy. Living rats had to be scanned within a short timeframe, which led to low resolution of the scans in this study. For evaluation of bone density, the resolution of the scans was considered to be too low at any bone site to give reliable data. BVF was reduced, but not significant, in the control group, which also experienced most molar movement, whereas the bone volume in the high dose adiponectin treated group was unchanged, and little tooth movement was seen. There are diverging data concerning the effect of adiponectin on osteoclast differentiation. Some report an inhibition of

Fig. 5. Measurements of bone volume fraction. Comparison of bone volume fraction measured from the furcation area of the maxillary first molar by μCT. No significant changes were seen between groups or days. Values are presented as mean ± SEM.

orthodontic practice. A possible strategy to prevent this may be use of biological modulators capable of reducing tooth movement (Adachi et al., 1994; Dunn et al., 2007; Liu et al., 2004). The present study demonstrates that local adiponectin administration significantly reduced experimental orthodontic tooth movement in rats. The injections appeared to have local effects only, as no difference in circulating adiponectin levels was observed between the groups. Dosages of 0.2 and 2 μg were chosen; these concentrations are in the lower level or below dosages of recombinant human adiponectin previous injected in rats without adverse effects (Tullin et al., 2012; Yuan et al., 2016). Molar movement in this study was assessed by two different methods, feeler gauge and μCT. These two procedures have different accuracy levels, and the μCT data included the first and third molar, 308

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Fig. 6. Sections showing collagen in the PDL. Longitudinal sections of the PDL showing collagen stained with PSR in the adiponectin (2 μg) treated (A and B) and the control group (C and D) after 14 days of OTM. The images were taken in standard white light (A and C) and in polarized light (B and D). The selected images are representative for each treatment group. Scale bar: 50 μm. Polarized signal intensity corresponds to the total collagen volume fraction in the ROI (E). Number of nuclei counted in the PDL (F). Values are presented as mean ± SD.

osteoclastogenesis (Luo et al., 2012; Oshima et al., 2005; Tu et al., 2011; Williams et al., 2009; Wu et al., 2014), while others report that adiponectin stimulates RANKL (Luo et al., 2006; Wang et al., 2014). The numbers of osteoclasts were not reliable identified in this study. The frontal resorption of bone may thus cause structural changes at the bone surface, resulting in its increased irregularity. The irregular morphology of this region may influence the identification of osteoclasts (Araujo, Fernandes, Maciel, Netto Jde, & Bolognese, 2015). In addition, the long decalcification time with the chosen protocol caused the sections to be spongy and difficult to cut (González-Chávez, Pacheco-Tena, Macías-Vázquez, & Luévano-Flores, 2013). Periodontal ECM remodeling plays an important role in the process of tooth movement and maintenance of the position of the teeth (Yoshida, Sasaki, Yokoya, Hiraide, & Shibasaki, 1999). Collagen is a dominant component of ECM in the periodontium and periodontal

Table 2 Effect of adiponectin administration on circulating levels of various factors after 14 days.

Adiponectin Leptin IL-1 β PAI-1 Total MCP-1 TNF-α

Control

Adiponectin (0.2 μg)

Adiponectin (2 μg)

86.5 ± 9.2 1.9 ± 0.3 37.4 ± 10.0 17.4 ± 2.7 97.2 ± 8.6 0.4 ± 0.1

75.9 ± 9.1 2.5 ± 0.2 48.3 ± 11.3 27.9 ± 4.7 111.5 ± 10.5 0.4 ± 0.1

66.5 ± 5.2 2.5 ± 0.4 47.1 ± 12.0 31.6 ± 13.3 139.2 ± 22.5 0.5 ± 0.2

IL-1 β: interleukin-1 beta; PAI-1 Total: Plasminogen activator inhibitor-1 Total; MCP-1: Monocyte chemoatractant protein-1; TNF-α: tumor necrosis factor-alpha. Values (pg/ml) are presented as mean ± SEM. n = 7 in both adiponectin groups (0.2 and 2 μg)), n = 8 in the control group. No significant differences between groups were found in the factors tested.

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fibroblast activity during orthodontic tooth movement is characterized by a high collagen turnover (Krishnan & Davidovitch, 2009; Ten Cate, Deporter, & Freeman, 1976). In a wound healing model, adiponectin upregulated keratine gene transcripts and subsequently promoted collagen organization where the collagen fibers were thicker and deposited more orderly compared to control (Salathia, Shi, Zhang, & Glynne, 2013). Adiponectin has been found to enhance the expression of periostin (Nokhbehsaim et al., 2014), which is a matricellular protein (Hamilton, 2008) involved in stiffening of collagen through crosslinking. Adiponectin may improve ECM production by enhancing the POSTN expression, and therefore change the collagen structure in experimental tooth movement. Alterations in collagen size or morphology could, however, not be confirmed in the present study. We observed no change in PDL cell number which is in contrast to a previous in vitro study reporting that adiponectin enhanced the expression of Ki67, a marker of proliferation in cultured PDL cells (Nokhbehsaim et al., 2014).

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Adiponectin prevents orthodontic tooth movement in rats.

The objective was to examine the effects of repetitive local administration of adiponectin on experimental tooth movement in rats...
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