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Journal of Dental Research
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Biological

Nuclear Factor {kappa}B p65 Phosphorylation in Orthodontic Tooth Movement

J. Zuo, L.A. Archer, A. Cooper, K.L. Johnson, L.S. Holliday and C. Dolce*

Department of Orthodontics, 1600 SW Archer Road, Campus Box 100444, University of Florida College of Dentistry, Gainesville, FL 32610, USA

Correspondence: * corresponding author, cdolce{at}dental.ufl.edu


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS & METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Osteoclasts play a vital role in orthodontic tooth movement. Transactivation of nuclear factor {kappa}B (NF{kappa}B) by phosphorylation of the p65 component of NF{kappa}B at amino acid 536 (p65*536) plays a role in osteoclast differentiation stimulated by receptor activator of nuclear factor {kappa}B-ligand (RANK-L). We hypothesized that this transactivation pathway might be involved in the responses of alveolar bone cells during orthodontic tooth movement. We detected sharp increases in the levels of p65*536 3 and 12 hrs after the application of orthodontic stimuli in rats. In cell culture, osteoclast-like cells displayed no changes in p65*536 in response to RANK-L, but levels rapidly increased after the cells were mechanically scraped. We conclude that p65*536 is produced rapidly in response to orthodontic stimuli and mechanical insults, and may be important in bone remodeling associated with orthodontic tooth movement.

Key Words: osteoclast • bone resorption • RANK • RAW 264.7 • osteoclastogenesis


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS & METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Orthodontic tooth movement requires alterations in the local remodeling of the alveolar bone that underlies teeth (Ren, 2005). This occurs in response to the application of orthodontic force and occurs in three stages: the initial tipping phase (activation of cells), a lag phase that involves the recruitment of osteoclasts and initiation of bone resorption, and the post-lag phase, where tooth movement occurs (Reitan, 1951, 1967).

Nuclear factor {kappa}B (NF{kappa}B) is activated in response to the stimulation of a variety of cell-surface receptors (Siebenlist et al., 1994). This involves translocation of NF{kappa}B from the cytosol to the nucleus, where it functions as a transcription factor (Ghosh and Baltimore, 1990; Hayden and Ghosh, 2004; Ravid and Hochstrasser, 2004; Xiao and Ghosh, 2005). In osteoclasts, activation of NF{kappa}B occurs in response to stimulation by receptor activator of nuclear factor {kappa} B (RANK) by RANK-Ligand (RANK-L) (Lacey et al., 1998; Teitelbaum, 2000). RANK-L is induced in response to bone-modulating factors (Lee and Kim, 2003). Analysis of data suggests that orthodontic force up-regulates RANK-L production, leading to the formation and activation, in the alveolar bone, of osteoclasts that are required to remodel bone to accommodate tooth movement (Ren, 2005; Low et al., 2005).

The p65 component of NF{kappa}B is phosphorylated at several sites (Ghosh and Karin, 2002; Viatour et al., 2005). Phosphorylation of NF{kappa}B p65 at serine 536 (p65*536) has been shown to transactivate NF{kappa}B (Sizemore et al., 1999; Adli and Baldwin, 2006). In a recent report, it was demonstrated that transactivation of NF{kappa}B by the generation of p65*536 in response to RANKL is vital for osteoclast formation (Huang et al., 2006). This led us to examine the role of p65*536 in orthodontic tooth movement. Our findings suggest that transactivation of NF{kappa}B by the generation of p65*536 may play a regulatory role in orthodontic tooth movement. In vitro, we did not detect RANKL-induced changes in the level of p65*536. However, mechanical insult, in the form of scraping the cells from the substrate, triggered rapid increases in p65*536. Because orthodontic tooth movement is characterized by cell damage (Krishnan and Davidovitch, 2006), cell scraping may represent a simple model for the damage incurred during orthodontic procedures. The purpose of this investigation was to characterize pathways leading to the generation of p65*536 in response to an orthodontic force. This should provide fresh insight into the mechanisms of orthodontic tooth movement and bone remodeling.


   MATERIALS & METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Reagents and Antibodies
Unless otherwise noted, all chemical reagents were obtained from the Sigma Chemical Company (St. Louis, MO, USA) and were of the highest grade offered. A polyclonal antibody against p65*536 was obtained from Cell Signaling Technologies (Beverly, MA, USA), and a polyclonal antibody against total p65 was purchased from NeoMarker (Fremont, CA, USA). Secondary antibodies were obtained from Jackson Immunoresearch Labs (West Grove, PA, USA) or Sigma.

Osteoclast Cultures
Osteoclast-like cells were differentiated from RAW 264.7 cells, a mouse hematopoietic cell line, by stimulation with recombinant RANK-L as described previously (Krits et al., 2002). In brief, RAW 264.7 cells were plated at a density of 20,000 cell/cm2 in 6-well plates or on coverslips in 24-well plates and cultured in DME plus 10% BSA. Cells were stimulated with 50 ng/mL GST-RANKL or vehicle for 5 days. The GST-RANKL construct, its expression and purification, were described in detail previously (Krits et al., 2002). After 3 days, the media were replaced, and fresh GST-RANKL or vehicle was added. The formation of osteoclast-like cells was monitored by direct examination of the cultures by light microscopy; RANK-L stimulated cultures were used for experiments if they displayed well-spread giant cells forming a near-monolayer. Vehicle-treated cultures contained mononuclear cells. After 5 days, cells were either prepared for sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting, or were fixed for immunostaining.

Gel Electrophoresis and Immunoblots
Adherent cells in 6-well plates were washed 3 times with PBS, and solubilized directly with Laemmli sample buffer (Bio-Rad, Hercules, CA, USA). Cell remnants were scraped from wells after extraction to ensure complete recovery of cells. The samples were immediately boiled for 5 min and then centrifuged at 200,000 x g for 30 min for removal of nucleic acids and other insoluble materials. Samples were subjected to SDS-PAGE on 12.5% gels, blotted on nitrocellulose, and probed with a 1:250 dilution of a polyclonal antibody specific for p65*536 or a 1:500 dilution of an antibody to total p65. We used the SuperSignal WestDuro system (Pierce, Rockford, IL, USA) to detect interactions on blots, and detected chemiluminescence using a Fluorchem 8000 (Alpha Innatech Corp., San Leandro, CA, USA).

Immunostaining of Osteoclast-like Cells
Osteoclast-like cells on coverslips were fixed with 2% formaldehyde in PBS, permeabilized with PBS plus 0.5% Triton X-100, and blocked with 2% BSA in PBS for 2 hrs, followed by an overnight incubation with a 1:50 dilution of anti- p65*536 in PBS plus 2% BSA at 4°C. Detection was performed by an avidin-biotin immunoperoxidase method (ABC) according to the manufacturer’s protocol (Vector Labs, Burlingame, CA, USA), with 3,3'-diaminobenzidine (DAB) as the chromagen.

Orthodontic Tooth Movement in Rats
Orthodontic tooth movement was performed on male rats as described previously (Dolce et al., 2003; Holliday et al., 2003). Male Sprague-Dawley rats, weighing from 250 to 300 grams each, were purchased from Charles River Breeding Laboratories (Wilmington, MA, USA). The animals were acclimated for 2 wks under experimental conditions. There were preparatory sessions consisting of the following sequence of events: (a) Animal weights were recorded, (b) anesthesia was administered by intra-peritoneal injections of ketamine (87 mg/kg) and xylazine (13 mg/kg), (c) modified orthodontic cleats were bonded bilaterally to the occlusal surfaces of the maxillary first molars, (d) all 4 incisors were pinned to prevent further eruption and minimize movement of the anchorage, (e) the lower incisor edges were reduced 2 mm, and (f) the mandibular first molars were extracted to prevent appliance damage. Rats were allowed to recover for 3 wks, while wound healing and weight gain were monitored. We activated appliances by positioning the rats in a head restrainer and placing orthodontic springs as previously described (King et al., 1991). One end of a nickel titanium closed-coil spring (light; #10-000-06; GAC, Central Islip, NY, USA) was ligated to the molar cleat, while the other was attached to a 40-gram suspended weight. This force was chosen since it has been shown to demonstrate both the typical orthodontic tooth movement kinetics (initial tipping, lag phase, and accelerated tooth movement) and acceptable balance between bone formation and resorption (King et al., 1991). The anterior end of the coil was then bonded (with autocuring methylmethacrylate) to the acid-etched lateral surface of the maxillary incisor, followed by removal of the weight and excess coil spring. This method ensured a precise and reproducible initial orthodontic force designed to tip the maxillary first molars in the mesial direction. The control group was treated identically except for spring placement. These procedures were approved by the University of Florida Institutional Animal Care and Usage Committee.

Immunohistochemistry on Sections
The rats were decapitated, and maxillae were removed. Longitudinal demineralized sections (8 µm thick), demonstrating interradicular alveolar bone immediately adjacent to the periodontal ligament of distobuccal and middle-buccal maxillary first molar roots, were cut from paraffin-embedded tissue blocks. Each section was deparaffinized, then heated for 9 min in a microwave in 10 mM sodium citrate buffer (pH 6.0) for antigen unmasking. Sections were then incubated with blocking buffer (1.5% goat serum in PBS) for 2 hrs prior to exposure to polyclonal antibodies specific for p65*536 (1:50 dilution) and for NF-{kappa}B p65 (1:100 dilution) overnight at 4°C. As a negative control, we used the blocking buffer instead of the primary antibody. Detection was performed by an avidin-biotin immunoperoxidase method (ABC) according to the manufacturer’s protocol (Vector Labs), with 3,3'-diaminobenzidine (DAB) as the chromagen.

Cell Scraping
RAW 264.7 cells were grown for 5 days in the presence of 50 ng/mL GST-RANK-L. Cells were replenished with fresh media, then scraped with a plastic cell scraper. Cells were either harvested immediately in SDS-PAGE sample buffer, or were allowed to settle and then harvested after 1, 3, 6, or 12 hrs. Samples were boiled, and then spun at 200,000 x g for 30 min for the removal of nucleic acids. Samples were maintained at –70°C until they were subjected to SDS-PAGE and blotting.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
p65*536 Levels Were Not Altered During RANK-L-induced Osteoclastogenesis in vitro
RAW 264.7 cells were induced to differentiate into osteoclast-like cells by stimulation with recombinant RANK-L, or were cultured without RANK-L stimulation in 6-well plates, after which whole cells were subjected to SDS-PAGE, blotted, and probed with the anti-p65*536 antibody. A single 65-kDa band was detected, indicating the specificity of the antibodies (Fig. 1AGo). RANK-L-stimulation did not trigger a change in the relative level of p65*536 compared with the level of total p65. In RAW 264.7 osteoclast-like cells, we detected p65*536 associated with nuclei (Fig. 1BGo).


Figure 1
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  		Figure 1. No change in levels of p65*536 was detected in RAW 264.7 cells related to stimulation with RANK-L. (A) RAW 264.7 cells were stimulated with vehicle or recombinant RANK-L for 5 days. Cultured cells that had been treated with RANK-L were visually inspected for the presence of giant multinucleated cells. Whole-cell extracts were then subjected to SDS-PAGE, blotted on nitrocellulose, and probed with antibodies to p65 or p65*536. There was no difference detected in the level of p65*536 staining depending on treatment. (B) p65*536 was detected surrounding and within the nuclei of RANK-L-stimulated RAW 264.7 cells. This experiment was typical of 3 separate experiments. Bar equals 20 µm.

 
Immunostaining of Sections from Rats Subjected to Orthodontic Tooth Movement or Controls
NF{kappa}B is constitutively present, and regulation occurs post-translationally (Ghosh and Karin, 2002). Levels of p65 were similar in the alveolar bone from experimental and control animals (Fig. 2Go). In contrast, very little p65*536 staining was detected in control rats or experimental rats at time 0 (Fig. 3Go, 0 hrs). Three hrs after orthodontic stimuli, the overall level of staining for p65*536 was enhanced (Fig. 3Go, 3 hrs). Levels remained elevated 12 hrs after orthodontic stimuli (Fig. 3Go, 12 hrs). Closer examination of sections of alveolar bone revealed that much of the enhanced staining was associated with osteoclasts (defined as giant, multinuclear cells) lining the alveolar bone (Fig. 3Go, 12 hrs, high mag.).


Figure 2
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  		Figure 2. Orthodontic tooth movement does not alter levels of total p65 in alveolar bone from rats. Representative sections of alveolar bone and periodontium associated with control or orthodontically stimulated incisors from rats stained for the presence of total p65. These are typical of staining detected for all controls and all time-points in 3 separate trials from 6 animals. The root and periodontal ligament (PDL) are indicated. The bars equal 50 µm.

 

Figure 3
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  		Figure 3. Orthodontic tooth movement triggered a large increase in p65*536 in alveolar bone from rats. Representative sections of the alveolar bone underlying incisors orthodontically stimulated for 0, 3, and 12 hrs and stained for the presence of p65*536 are shown. Note the almost complete absence of staining at 0 hrs. Increased staining was detected by 3 hrs, and intense staining was observed at 12 hrs. The arrows point to a section in the 12-hour panel (bottom left) that was magnified in the bottom right panel. At high magnification, giant cells (osteoclasts) were detected and were stained intensely. The bar equals 50 µm in the two top panels and the bottom left panel, and 10 µm in the bottom right panel.

 
p65*536 Levels Rapidly Increased after Osteoblast-like Cells were Mechanically Scraped from Their Substrate in vitro
RAW 264.7 osteoclast-like cells were grown to maturity in 6-well plates and scraped from the surface by means of a cell scraper. Cells were subjected to SDS-PAGE, blotted, and probed for p65 and p65*536. The relative level of p65*536 increased after scraping ceased (Fig. 4Go).


Figure 4
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  		Figure 4. Scraping RAW 264.7 osteoclast-like cells triggers increased levels of p65*536. RAW 264.7 osteoclast-like cells were scraped from their substrate by means of a cell scraper, then prepared immediately for SDS-PAGE (0), or were allowed to settle for 1, 3, 6, or 12 hrs, and prepared for SDS-PAGE. After the proteins were separated, they were blotted on nitrocellulose and probed with either the anti-p65*536 antibody or the anti-p65 antibody. p65*536 was barely detected at time 0, but was present at high levels at subsequent time-points. The levels of total p65 were similar at all time-points.

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
We report here that a rapid increase in p65*536 was associated with orthodontic tooth movement in a rat model and with mechanical scraping of RAW 264.7 osteoclast-like cells. This is the first report suggesting transactivation by generation of p65*536 as a potential regulator of bone remodeling during orthodontic tooth movement.

A recent study reported that transactivation of NF{kappa}B by generation of p65*536 occurred in response to RANK-L treatment in mouse marrow osteoclasts (Huang et al., 2006). This prompted us to examine whether a similar mechanism was in play in a well-characterized rat model of orthodontic tooth movement, and in RAW 264.7 cells. Unlike Huang and colleagues, we did not detect RANK-L stimulation of p65*536 in this model system. We found that scraping these cells free from the substrate and allowing them to settle in the original well triggered rapid increases in levels of p65*536. During orthodontic procedures, cells associated with the periodontium suffered damage as the tooth was tipped (Krishnan and Davidovitch, 2006). We think that it is not unreasonable to consider scraping as a first approximation of the damage inflicted on cells during orthodontic procedures. The fact that we saw rapid increases in p65*536 in response to both cell scraping and the application of orthodontic force supports this idea.

Orthodontic force triggered a large increase in p65*536 in vivo. At least some of the cells which displayed high levels of p65*536 were osteoclasts. In control cultures, staining for p65*536 was barely detected. Staining for total p65 could not be distinguished in controls compared with experimental sections.

In summary, this study provided the first demonstration that p65*536 levels are increased by orthodontic stimuli. We hypothesize that this may be a mechanism through which NF{kappa}B can be rapidly activated. This may contribute to the rapid orthodontic response of rodents. The recent report from Huang and colleagues indicated that phosphorylation of p65 was involved in RANK-L signaling through transforming growth factor β-activated kinase-1, mitogen-activated kinase kinase-3, and mitogen-activated kinase kinase-6 (Huang et al., 2006). It will be of interest to determine if this signaling pathway leading to p65*536 induces transactivation of NF{kappa}B during orthodontic tooth movement.


   ACKNOWLEDGMENTS
 
This work was funded by NIH/NIAMS R01 AR-47959 (to LSH) and 5RO3DE13857-02 (to CD).

Received for publication January 5, 2006. Revision received December 1, 2007. Accepted for publication January 15, 2007.


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS & METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  • Adli M, Baldwin AS (2006). IKK-i/IKKepsilon controls constitutive, cancer cell-associated NF-kappaB activity via regulation of Ser-536 p65/RelA phosphorylation. J Biol Chem 281:26976–26984.[Abstract/Free Full Text]
  • Dolce C, Vakani A, Archer L, Morris-Wiman JA, Holliday LS (2003). Effects of echistatin and an RGD peptide on orthodontic tooth movement. J Dent Res 82:682–686.
  • Ghosh S, Baltimore D (1990). Activation in vitro of NF-kappa B by phosphorylation of its inhibitor I kappa B. Nature 344(6267):678–682.[CrossRef][Medline] [Order article via Infotrieve]
  • Ghosh S, Karin M (2002). Missing pieces in the NF-kappaB puzzle. Cell 109(Suppl):S81–S96.
  • Hayden MS, Ghosh S (2004). Signaling to NF-kappaB. Genes Dev 18:2195–2224.[Abstract/Free Full Text]
  • Holliday LS, Vakani A, Archer L, Dolce C (2003). Effects of matrix metalloproteinase inhibitors on bone resorption and orthodontic tooth movement. J Dent Res 82:687–691.
  • Huang H, Ryu J, Ha J, Chang EJ, Kim HJ, Kim HM, et al. (2006). Osteoclast differentiation requires TAK1 and MKK6 for NFATc1 induction and NF-kappaB transactivation by RANKL. Cell Death Differ 13:1879–1891.[CrossRef][Medline] [Order article via Infotrieve]
  • King GJ, Keeling SD, McCoy EA, Ward TH (1991). Measuring dental drift and orthodontic tooth movement in response to various initial forces in adult rats. Am J Orthod Dentofacial Orthop 99:456–465.[Medline] [Order article via Infotrieve]
  • Krishnan V, Davidovitch Z (2006). Cellular, molecular, and tissue-level reactions to orthodontic force. Am J Orthod Dentofacial Orthop 129:469.e1–32.
  • Krits I, Wysolmerski RB, Holliday LS, Lee BS (2002). Differential localization of myosin II isoforms in resting and activated osteoclasts. Calcif Tissue Int 71:530–538.[CrossRef][Medline] [Order article via Infotrieve]
  • Lacey DL, Timms E, Tan HL, Kelley MJ, Dunstan CR, Burgess T, et al. (1998). Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell 93:165–176.[CrossRef][Medline] [Order article via Infotrieve]
  • Lee ZH, Kim HH (2003). Signal transduction by receptor activator of nuclear factor kappa B in osteoclasts. Biochem Biophys Res Commun 305:211–214.[CrossRef][Medline] [Order article via Infotrieve]
  • Low E, Zoellner H, Kharbanda OP, Darendeliler MA (2005). Expression of mRNA for osteoprotegerin and receptor activator of nuclear factor kappa beta ligand (RANKL) during root resorption induced by the application of heavy orthodontic forces on rat molars. Am J Orthod Dentofacial Orthop 128:497–503.[CrossRef][Medline] [Order article via Infotrieve]
  • Ravid T, Hochstrasser M (2004). NF-kappaB signaling: flipping the switch with polyubiquitin chains. Curr Biol 14:R898–R900.[CrossRef][Medline] [Order article via Infotrieve]
  • Reitan K (1951). The initial tissue reaction incident to orthodontic tooth movement as related to the influence of function; an experimental histologic study on animal and human material. Acta Odontol Scand Suppl 6:1–240.[Medline] [Order article via Infotrieve]
  • Reitan K (1967). Clinical and histologic observations on tooth movement during and after orthodontic treatment. Am J Orthod 53:721–745.[CrossRef][Medline] [Order article via Infotrieve]
  • Ren YJ (2005). Cell biology in orthodontic tooth movement: the known and the unknown. Shanghai Kou Qiang Yi Xue 14:182–189.[Medline] [Order article via Infotrieve]
  • Siebenlist U, Franzoso G, Brown K (1994). Structure, regulation and function of NF-kappa-B. Ann Rev Cell Biol 10:405–455.[CrossRef]
  • Sizemore N, Leung S, Stark GR (1999). Activation of phosphatidylinositol 3-kinase in response to interleukin-1 leads to phosphorylation and activation of the NF-kappaB p65/RelA subunit. Mol Cell Biol 19:4798–4805.[Abstract/Free Full Text]
  • Teitelbaum SL (2000). Bone resorption by osteoclasts. Science 289(5484):1504–1508.[Abstract/Free Full Text]
  • Viatour P, Merville MP, Bours V, Chariot A (2005). Phosphorylation of NF-kappaB and IkappaB proteins: implications in cancer and inflammation. Trends Biochem Sci 30:43–52.[CrossRef][Medline] [Order article via Infotrieve]
  • Xiao C, Ghosh S (2005). NF-kappa B, an evolutionarily conserved mediator of immune and inflammatory responses. Adv Exp Med Biol 560:41–45.[Medline] [Order article via Infotrieve]

Journal of Dental Research, Vol. 86, No. 6, 556-559 (2007)
DOI: 10.1177/154405910708600613
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