ORIGINAL ARTICLE

Effect of low-frequency mechanical vibrationon orthodontic tooth movement

Sumit Yadav,a Thomas Dobie,b Amir Assefnia,c Himank Gupta,c Zana Kalajzic,d and Ravindra Nandae

Farmington, Conn

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Introduction: Our objective was to investigate the effect of low-frequency mechanical vibration (LFMV) on therate of tooth movement, bone volume fraction, tissue density, and the integrity of the periodontal ligament. Ournull hypothesis was that there would be no difference in the amount of tooth movement between different valuesof LFMV. Methods: Sixty-four male CD1 mice, 12 weeks old, were used for orthodontic tooth movement. Themice were randomly divided into 2 groups: control groups (baseline; no spring 1 5 Hz; no spring 1 10 Hz;and no spring 1 20 Hz) and experimental groups (spring 1 no vibration; spring 1 5 Hz; spring 1 10 Hz; andspring 1 20 Hz). In the experimental groups, the first molars were moved mesially for 2 weeks using nickel-titanium coil springs delivering 10 g of force. In the control and experimental groups, LFMV was applied at 5,10, or 20 Hz. Microfocus x-ray computed tomography analysis was used for tooth movement measurements,bone volume fraction, and tissue density. Additionally, immunostaining for sclerostin, tartrate-resistant acidphosphatase (TRAP) staining, and picrosirius red staining were used on the histologic sections. Simpledescriptive statistics were used to summarize the data. Kruskal-Wallis tests were used to compare theoutcomes across treatment groups. Results: LFMV did not increase the rate of orthodontic tooth movement.Microfocus x-ray computed tomography analysis showed increases in bone volume fractions and tissue den-sities with applications of LFMV. Sclerostin expression was decreased with 10 and 20 Hz vibrations in boththe control and experimental groups. Additionally, the picrosirius staining showed that LFMV helped in maintain-ing the thickness and integrity of collagen fibers in the periodontal ligament. Conclusions: There was no signif-icant increase in tooth movement by applying LFMV when compared with the control groups (spring 1 novibration). (Am J Orthod Dentofacial Orthop 2015;148:440-9)

Tooth movement involves both remodeling andmodeling of bone.1,2 Remodeling and modelinginvolve a coordinated action of osteoclasts and

osteoblasts in response to mechanical loading.3 More-over, inflammatory mediators (interleukin [IL]-1, IL-2,IL-6, IL-8, and tumor necrosis factor-alpha) are releasedafter mechanical stimulus or injury, triggering the bio-logic process associated with orthodontic tooth move-ment (OTM).4–7

the Health Center, University of Connecticut, Division of Orthdontics,ngton, Conn.tant professor.ing assistant professor.ent.arch associate.ssor and head.thors have completed and submitted the ICMJE Form for Disclosure of Po-l Conflicts of Interest, and none were reported.rted by the American Association of Orthodontists Foundation and theon of Orthodontics of the University of Connecticut.ss correspondence to: Sumit Yadav, Division of Orthodontics, University ofecticut Health Center, 263 Farmington Ave, Room L7063 MC1725, Farm-n, CT 06030; e-mail, yadav_sumit17@yahoo.com.itted, November 2014; revised and accepted, March 2015.5406/$36.00ight � 2015 by the American Association of Orthodontists./dx.doi.org/10.1016/j.ajodo.2015.03.031

Currently, orthodontic treatment requires approxi-mately 24 to 30 months of intervention to completethe treatment.8–10 The longer duration of treatment isa great concern and poses high risks for caries, rootresorption, and decreased patient compliance andsatisfaction.8,10,11 Thus, accelerating OTM andshortening the total treatment duration is a primarygoal of the orthodontist, and it can preventdetrimental effects of longer treatment time andincrease patient satisfaction.

Nishimura et al have shown that the application ofcyclical forces (60Hz) on themaxillaryfirstmolar increasesthe rate of OTM.12 However, the main drawback of theNishimura study was the method of force application(transpalatal expansion spring). The force was applied toaccelerate tooth movement in the first order (buccolin-gually) rather than in the second order (mesiodistally),which comprises the majority of OTM. Moreover, it mayconfound the actual tooth movement because of itsskeletal effects.12 Studies have shown that whole bodyvibration (30, 45, and 90 Hz) may have an anabolicresponse on bone mass and architecture.13–15 Mileset al,16 in their randomized controlled trial, showed that

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application of 111 Hz of vibrational frequency for 20 mi-nutes per day did not speed tooth movement whencompared with the controls. Kalajzic et al17 showed theinhibitory effect of cyclical forces (30 Hz and 40 g of forceapplied with an electromechanical actuator) on OTM inrats; moreover, they showed the deleterious effects ofthe cyclical forces on the periodontal ligament (PDL).The major drawback of their model was higher cyclicalforce (40 g). The effect and mechanism of low-frequency mechanical vibrations (LFMV) (#20 Hz) onOTM and paradental tissues still remain unclear. To ourunderstanding, this is the first in-vivo study regardingthe effect of LFMV (frequency,#20 Hz) on OTM.

Recently, LFMV has gained interest in acceleratingOTM by increasing alveolar bone turnover. The osteo-cytes in the bone tissue are thought to orchestrate “me-chanotransduction” by reacting to different forms ofmechanical loading through biologic signals.18,19 Therole of osteocytes in bone remodeling and modelinghas been well documented.18,19 It has been shown thatosteocytes are the major source of sclerostin (productof SOST gene), and they antagonize the canonical Wntsignaling pathway, thus exhibiting an inhibitory effecton bone formation.20,21 Matsumoto et al22 demon-strated the role of osteocytes in resorption modelingduring OTM (mechanotransduction) using osteocyte-ablated mice.

Our null hypothesis was that there would be no dif-ference in the amount of tooth movement betweendifferent amounts of LFMV. We had 3 specific aims:(1) to determine the effect of LFMV on the rate of toothmovement; (2) to quantify bone modeling and remodel-ing in both the control and experimental groups usingmicrofocus computed tomography (micro-CT) and im-munostaining; and (3) to determine the effect ofLFMV on the PDL.

MATERIAL AND METHODS

The Institutional Animal Care Committee of the Uni-versity of Connecticut Health Center approved thisstudy, which conformed to the ARRIVE guidelines.Data were obtained from 64 male CD1 mice (CharlesRiver Laboratories, Wilmington, Mass; body weight,24-30 g). The 12-week-old mice were randomly dividedinto 2 groups: control and experimental. The controlgroup was further subdivided into 4 groups; each con-trol group had 5 mice: (1) group 1 (baseline), no springand no mechanical vibration; (2) group 2, no orthodon-tic spring, but 5 Hz vibration was applied to the maxillaryfirst molars; (3) group 3, no orthodontic spring, but10 Hz vibration was applied to the maxillary first molars;and (4) group 4, no orthodontic spring, but 20 Hz

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vibration was applied to the maxillary first molars. Theexperimental group was also subdivided into 4 groups,each with 11 mice: (1) group 5, orthodontic springonly but no vibration; (2) group 6, orthodontic springand 5 Hz vibration applied to the maxillary first molars;(3) group 7, orthodontic spring and 10 Hz vibrationapplied to the maxillary first molars; (4) group 8, ortho-dontic spring and 20 Hz vibration applied to the maxil-lary first molars. The animals were allowed at least aweek of acclimatization at our Health Center to compen-sate for their different origins.

The animals were placed under general anesthesiawith xylazine (13 mg/kg) and ketamine (87 mg/kg). Acustom mouth-prop was fabricated from 0.036-instainless steel wire and placed between the maxillaryand mandibular incisors to hold the mouth open. A0.004-in stainless steel ligature wire was passedbeneath the contact between the maxillary first andsecond molars and threaded to the maxillary first molar(Fig 1, A). A low force/deflection rate nickel-titaniumcoil spring (Ultimate Wireforms, Bristol, Conn) deliv-ering 10 g of force was attached to the 0.004-in stain-less steel ligature around the first molar, and the otherend of the spring was attached to the incisors with0.004-in stainless steel wire. The force/deflection ratefor the spring was determined to calibrate the amountof force produced by activation of the nickel-titaniumcoil spring.

Additionally, grooves 0.5 mm from the gingivalmargin were prepared on the facial, mesial, and distalsurfaces of the maxillary central incisors to preventthe ligatures from dislodging from the incisor becauseof their lingual curvature and eruption pattern. Self-etching primer (Transbond Plus; 3M Unitek, Monrovia,Calif) and light-cured adhesive resin cement (Trans-bond; 3M Unitek) were applied to the lingual surfacesof the maxillary first molars and incisors to secure theligature wire. Moreover, to minimize the distal move-ment of the right incisor and to reinforce the anterioranchorage, the right and left incisors were joinedtogether to act as a unit (Fig 1, A). After applianceinsertion, the mice were allowed to recover with an in-candescent light for warmth; they were returned totheir cages once full ambulation and self-cleansing re-turned. The appliances were checked every other day,and additional light-cured bonding material was addedif necessary.

After anesthesia with ketamine (87 mg/kg) and xyla-zine (13 mg/kg), a custom mouth-prop fabricated from0.017 3 0.025-in titanium-molybdenum alloy wire wasplaced between the maxillary and mandibular incisors tohold the mouth open. A feedback loop–controlled elec-tromechanical actuator (model 3230; Bose Enduratec,

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Fig 1. A, Inserted nickel-titanium coil spring; B, application of LFMV on the maxillary right molar (theelectromechanical actuator applies mechanical vibration at 5, 10, and 20 Hz at a compression force of1 g);C, schematic of the tooth movement model with the spring attached to the maxillary first molar andmaxillary incisors; D, intermolar distance (M1-M2) at day 14 in the experimental groups; E-H, sagittalmicro-CT sections used for measuring intermolar distance in the control groups (E, baseline; F, nospring 1 5 Hz vibration; G, no spring 1 10 Hz vibration; H, no spring 1 20 Hz vibration); I-L, sagittalmicro-CT sections used for measuring intermolar distance in the experimental groups (I, spring 1 novibration; J, spring 1 5 Hz vibration; K, spring 1 10 Hz vibration; L, spring 1 20 Hz vibration).

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Minnetonka, Minn) was used to apply unilateral LFMVto the occlusal surface of the maxillary right first molaralong its long axis (Fig 1, B). Loading protocols foreach animal consisted of 15 minutes of LFMV at 1 cNof force with the electromechanical actuator at a fre-quency of 5, 10, or 20 Hz (cycles/second) dependingon the mouse's group. LFMV was applied to the maxil-lary right first molar at 3-day intervals (days 1, 4, 7,10, and 13).

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After the 14 days of the experimental period, the an-imals were killed by inhalation of carbon dioxide fol-lowed by cervical dislocation.

The maxilla was hemisected, and the attached softtissues and muscles were removed. Subsequently, thehemisected maxilla was placed in 10% formalin for5 days at 4�C. The samples were then decalcified in14% ethylenediaminetetraacetic acid for 3 weeks andthen processed for standard paraffin embedding. Serial

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Fig 2. Micro-CT data showing BVF and tissue density. Each value in each graph represents themean 6 the standard deviation: A, region of interest where the bone parameters (BVF and tissuedensity) were measured; B, BVF values in the control groups (significantly [*] higher BVF in theno spring 1 5 Hz vibration group compared with the no spring 1 20 Hz vibration group); C, BVFvalues in the experimental groups; BVF decreases with tooth movement; baseline had a significantlyhigher BVF compared with the spring 1 no vibration group, the spring 1 5 Hz vibration group, thespring 1 10 Hz vibration group, and the spring 1 20 Hz vibration group; moreover, thespring 1 no vibration group had significantly less BVF than did the spring 1 5 Hz vibration (*)and the spring 1 10 Hz vibration (#) groups. D, Tissue density in the control groups; note the trendfor an increase in tissue density with vibration; the spring 1 5 Hz group had significantly higher bonedensity compared with baseline (*); a,b,c,d signifies P \ 0.5. E, tissue density in the experimentalgroups.

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Fig 3. Sclerostin expression in the control and experimental groups on the compression and tensionsides in alveolar bone. Upper panel, control groups; lower panel, experimental groups. A-D, Tensionside in the control groups (A, baseline; B, no spring 6 5 Hz; C, no spring 6 10 Hz; D, no spring6 20 Hz);E-H, tension side in experimental groups (E, spring1 no vibration; F, spring15 Hz;G, spring110 Hz;H, spring120 Hz); I-L,compression side in the control groups (I, baseline; J, no spring6 5 Hz;K, no spring 6 10 Hz; L: no spring 6 20 HzA); M-P, compression side in the experimental groups(M, spring 1 no vibration; N, spring 15 Hz; O, spring 1 10 Hz; P, spring 120 Hz). Note the decreasein sclerostin expression in the 10 Hz (C, G, K, and O) and 20 Hz (D, H, L, and P) vibrations in both thecontrol and experimental groups. Q, with no expression of sclerostin, is a negative control. The doublewhite arrows signify the cells positive for sclerostin.

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sagittal sections (5 mm) were obtained from the mesialand distal roots of the maxillary right first molar.

All the animals in the different control (5 in eachgroup) and experimental (11 in each group) groupswere used to measure tooth movements. Three-dimensional image arrays of the hemisected rightmaxillae were collected using micro-CT. OTM wasdefined and measured as the distance between themaxillary first and second molars at the most mesialpoint of the second molar crown and the most distalpoint of the first molar crown.17,23 The distance at day0 was 0 mm in all groups (ie, the convex crownsurfaces were touching). The OTM measurements were

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done in the sagittal plane, locating the image planethat showed the most root structure. The original 2-dimensional image was then magnified 10 times formore precise line drawings, which were made at theclosest proximity of the 2 convex molar crown surfaces.The OTM was measured in the 3 sagittal sections of eachanimal in all groups.

Micro-CT images were used for quantitative ana-lyses of bone changes occurring in the region of themaxillary first molar. Changes in the alveolar bonewere studied by analyzing the furcation area of themaxillary first molar. The region of interest for thealveolar bone analysis was defined vertically as the

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Fig 4. Histologic examination of PDL collagen fibers on the tension side by polarized light microscopy.The sirius red stained PDL shows a more organized PDL in the control groups (A, baseline; B, nospring 1 5 Hz; C, no spring 1 10 Hz; D, no spring 1 20 Hz) compared with the experimental groups(E, spring 1 no vibration; F, spring 1 5 Hz vibration; G, spring 1 10 Hz vibration; H, spring 1 20 Hzvibration). Note that vibration only does not lead to disorganization of PDL collagen fibers. Thespring 1 no vibration group had disorganized and thin PDL fibers. Note that the spring 1 vibration(F, G, and H) had more thickened and organized fibers (shown by the red staining) compared withthe spring 1 no vibration group. A.B., Alveolar bone; P, periodontal ligament; De, dentin.

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most occlusal point of the furcation to the apex of themaxillary first molar roots; transversely, it was definedas the space between the buccal and lingual corticalbone; sagittally, it included 50 sections (10 mm) begin-ning at the mesial root and continuing distally (Fig 2,A). The parameters studied and measured using the es-tablished algorithms were bone volume fraction (BVF)and tissue density.

Histology, immunohistochemistry, tartrate-resistantacid phosphatase (TRAP), and picrosirius red staininghistology were performed on each section. After block-ing for 10 minutes at room temperature with 1x univer-sal blocking reagent (HK085-5k; BioGenex, Fremont,Calif), deparaffinized histologic sections were incubatedwith goat polyclonal anti-SOST antibody (AF1589; R&DSystems, Minneapolis, Minn) overnight at 140�C at aconcentration of 5 mg per milliliter. Subsequently, thesections were washed with phosphate-buffered salineand incubated with Alexa Fluor 594 donkey anti-goatIgG (A-11058; Life Technologies, Grand Island, NY) at

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a concentration of 1:300 for 1 hour at room temperatureand mounted with a suspension of 50% glycerol inphosphate-buffered saline containing the nuclear stain(Hoechst H3570; Life Technologies) at a concentrationof 1:5000.

TRAP staining was performed using a leukocyte acidphosphatase (TRAP) kit (386-1 KT; Sigma-Aldrich, StLouis, Mo) according to the manufacturer's instructions.TRAP positive, multinucleated cells were counted on thealveolar bone surfaces on the mesial sides of the disto-buccal roots. The area for quantification included asquare with 1 side extending from the apex to the bifur-cation, and the other side extending 200 mm from thePDL border inside the alveolar bone. The osteoclastnumbers were counted in 6 sections from 4 mice ineach group, and the values were then averaged foreach animal to run a statistical test.

Deparaffinized sections were also stained with 1%picrosirius red for 1 hour, washed with acidified water(0.5% acetic acid water), and dehydrated with serial

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=

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ethanol washes before mounting. The sections wereexamined with a fluorescent microscope (Carl Zeiss,Thornwood, NY).

Statistical analysis

Simple descriptive statistics were used to summarizethe data. The outcomes examined included intermolardistance, BVF, and tissue density. Because of the samplesize, nonparametric tests were used to examine the asso-ciation between the outcome variables and the treat-ment groups (spring 1 no vibration, spring 1 5 Hz,spring 1 10 Hz, and spring 1 20 Hz). TheKolmogorov-Smirnov test was used to examine the dis-tribution of the outcome variables. Kruskal-Wallis testswere used to compare the outcomes across treatmentgroups. Pairwise comparisons between different groupswere conducted with the Mann-Whitney U test. All sta-tistical tests were 2 sided, and a P value of\0.05 wasdeemed to be statistically significant for the Kruskal-Wallis test. Because of the multiple pairwise comparisonsused, to minimize type 1 errors, Bonferroni correctionswere used.

RESULTS

All mice in the study remained healthy and had aslight increase in body weight. The delta displacementof the molar was the same throughout the vibration tomaintain 1 g of force with the vibrator tip.

The distance between the first and second molars inthe control groups was 0 mm (Fig 1, C), whereas therewas no statistically significant difference in the experi-mental groups after 14 days of orthodontic force appli-cation (Fig 1, D). However, the maximum toothmovement was observed in the spring 1 10 Hz group(0.248 6 0.074 mm), and the least was in thespring 1 20 Hz group (0.200 6 0.075 mm).

For the bone parameters, the micro-CT analysisshowed a statistically significant (P\0.05) decrease inBVF when the control groups were compared with theexperimental groups (baseline vs spring 1 no vibration

Fig 5. Quantification and histologic examination of oosteoclast numbers in A, different control groups andhigher osteoclast numbers in the spring1 no vibrationtologic examinations in: C, the different control groups10 Hz; F, no spring 1 20 Hz; and experimental groupbration; I, spring1 10 Hz vibration; J, spring1 20 HzPDL and alveolar bone in the spring 1 no vibration gament; De, dentin.

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[P \0.05]; no spring 1 5 Hz vs spring 1 5 Hz[P \0.05]; no spring 1 10 Hz vs spring 1 10 Hz[P\0.05]; and no spring 1 20 Hz vs spring 1 20 Hz[P \0.05]) (Fig 2, C and D). Moreover, in the controlgroups, the no spring 1 5 Hz group had a significantlygreater BVF than did the no spring 1 20 Hz group. Inthe experimental groups, the BVF was significantly(P \0.05) lower for the spring 1 no vibration group,the spring 1 5 Hz group, the spring 1 10 Hz group,and the spring 1 20 Hz group, when compared withthe baseline (Fig 2, B and C).

In the control groups, there was a trend for an in-crease in tissue density with mechanical vibration; how-ever, the no spring 1 5 Hz group had significantly(P\0.05) more tissue density than did the baseline con-trol. Similarly, in the experimental groups, there was atrend for an increase in bone density with mechanical vi-bration, but it was not statistically significant among theexperimental groups (Fig 2, D and E).

Our data show reductions in sclerostin expression onthe tension and compression sides with 10 and 20 Hz inboth the control (no spring 1 10 Hz, and nospring 1 20 Hz) and the experimental (spring 1 10 Hz,and spring 1 20 Hz) groups (Fig 3). However, there wasno decrease in sclerostin expression in either the controlor the experimental group with 5 Hz vibration (Fig 3).

Our data show that LFMV in the control group at5, 10, and 20 Hz did not affect the quality of thecollagen fibers. However, orthodontic loading doesmake a huge impact on the integrity and the qualityof the fibers (Fig 4, E). Our experimental groupsshowed that LFMV at 5, 10, and 20 Hz brings backthe integrity and the thickness of collagen fibersthat were lost due to orthodontic loading. Thespring-only group (orthodontic loading) had the leastthickened and wavy fibers, whereas the other experi-mental groups (spring 1 5 Hz, spring 1 10 Hz, andspring 1 20 Hz) had thick collagen fibers, as shownby the dark red stain.

Our experimental groups showed significantlyincreased numbers of osteoclasts when compared with

steoclast numbers at day 14: quantification ofB, different experimental groups; *significantlygroup compared with the baseline control. His-at baseline; D, no spring1 5 Hz; E, no spring1

s: G, spring 1 no vibration; H, spring 1 5 Hz vi-vibration. TRAP positive cells were higher in theroup (G). A.B., Alveolar bone; P, periodontal lig-

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the control groups (spring 1 no vibration [P\0.05] vsbaseline; spring 1 5 Hz [P \0.05] vs nospring 1 5 Hz; spring 1 10 Hz [P \0.05] vs nospring 1 10 Hz; and spring 1 20 Hz [P\0.05] vs nospring1 20 Hz [P\0.05]) (Fig 5). However, in the con-trol groups, there were no significant differences in oste-oclast numbers, implying that LFMV does not affect theosteoclast numbers in molars not subjected to toothmovement. In the experimental groups, there was atrend toward a decrease in osteoclast numbers withLFMV, although it was not statistically significant.

DISCUSSION

Our null hypothesis that there would be no differencein the amount of tooth movement between the experi-mental groups (spring1 no vibration, spring1 5 Hz vi-bration, spring 1 10 Hz vibration, and spring 1 20 Hzvibration) was accepted. We selected LFMV (5, 10, and20 Hz) for our study because Kalajzic et al17 showedthat higher cyclical forces inhibit OTM and are delete-rious to the PDL. Our results of OTM with LFMV werecontrary to those of Liu et al24 and Nishimura et al12;a plausible reason was a different tooth movementmodel. Nishimura et al and Liu et al used transpalatalexpansion springs (orthodontic load in the first order),whereas we used nickel-titanium coil springs (orthodon-tic load in the second order), and the orthodontic forcewas in the mesial direction. In the transpalatal model,the increase in OTM can be due to both skeletal anddental effects, whereas in our model, the OTM was pri-marily due to dental effects because we used adultmice, in which growth of the alveolar bone was com-plete.

OTM primarily depends on bone volume and bonedensity (quantity and quality of bone). In this study,there was a trend toward an increase in tissue densitywith LFMV in both the control and experimentalgroups. Moreover, a similar trend toward an increasein BVF in the experimental group was noted after theapplication of LFMV. Orthodontic loading decreasedthe BVF (baseline, 79.24%; spring 1 no vibration,53.45%), which was recovered by LFMV(spring 1 5 Hz, 64.45%; spring 1 10 Hz, 66.28%; andspring 1 20 Hz, 59.54%). Moreover, the spring 1 novibration group had significantly less (P \0.05) BVFthan did the spring 1 5 Hz and the spring 1 10 Hzgroups. Similarly, the results regarding BVF were shownby Kalajzic et al,17 and probably the reason could be in-hibition of osteoclastogenesis with LFMV. Furthermore,Vij and Mao25 showed that cyclical loading (4 Hz and300 mN) can cause sutural growth, and Alikhaniet al26 showed that application of high-frequency

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acceleration significantly increases the rate of alveolarbone formation. Moreover, our study showed that therewere decreased osteoclast numbers with LFMV(spring 1 no vibration [4.658] was greater thanspring 1 5 Hz [1.233], spring 1 10 Hz [3.875], andspring 1 20 Hz [2.738]). To our knowledge, this is thefirst study to show a decrease in osteoclastic activity(although it was not statistically significant) in the alve-olar bone with LFMV on the maxillary first molar. How-ever, similar results were obtained by Rubin et al27 andXie et al28 with whole body vibration in tibiae, andthey attributed this to kinase-dependent inhibition ofRANKL expression in the bone stromal cells.

The sclerostin expression was decreased on both thetension and compression sides with 10 and 20 Hz vibra-tions in the control and experimental groups (Fig 3). Tuet al29 showed that sclerostin is secreted by osteocytesand inhibits bone formation through Wnt signaling.Moreover, they showed that the loss of sclerostin expres-sion shows a high bone mass phenotype. Our experi-mental group showed a trend toward an increase inBVF with mechanical vibration, but for an unknownreason the sclerostin expression was not decreased inthe 5 Hz group in both the control and experimentalanimals (Fig 3).

The microscopic observation of the collagen fibersshowed no detrimental effect in the control group withdifferent LFMV (Fig 4). However, the collagen fiberslook thin and wavy after orthodontic loading in thespring 1 no vibration group. Moreover, we found thatLFMV (spring 1 5 Hz, spring 1 10 Hz, andspring1 20 Hz) after orthodontic loading was beneficialfor the fibers, and they look organized and thick (Fig 4).

Because this was an animal study, extrapolationof our findings to the clinical situation must bedone with caution as there is no osteonal remodeling(secondary remodeling) in mice, unlike in humans.Moreover, a frictionless space closure mechanismwas used in this study. Nevertheless, this in-vivostudy helped us to understand the effect of LFMVon the surrounding alveolar bone. Our future studieswill focus on understanding the signaling pathwaysassociated with LFMV and in-vitro gene expressionof the vibrated osteoblasts, osteoclasts, and cemen-toblasts.

CONCLUSIONS

1. There was no difference in the rate of tooth move-ment between the different experimental groups. How-ever, the maximum tooth movement was observed in thespring1 10 Hz group, and the least was in the spring120 Hz group.

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2. Tooth movement significantly decreased theBVF. The baseline control group had significantlymore BVF compared with the spring 1 no vibrationgroup.

3. LFMV at 5, 10, and 20 Hz had no deleterious effecton the integrity of the PDL. LFMV helped in maintainingthe thickness and integrity of the PDL after applicationof the orthodontic load.

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September 2015 � Vol 148 � Issue 3