Effect of Systemic Delivery of Substance P On Experimental Tooth Movement in Rats

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ORIGINAL ARTICLE

Effect of systemic delivery of Substance P


on experimental tooth movement in rats
Shu An,a Yueling Zhang,a Qian Chen,b Bin Xiong,c Jin Hao,d Yingcheng Zheng,a Xueman Zhou,a and Jun Wanga
Chengdu, Luzhou, and Nantong, China, and Boston, Mass

Introduction: The purpose of this study was to investigate the effect of systemic delivery of Substance P (SP) on
experimental tooth movement. Methods: Forty-eight adult Sprague-Dawley rats were randomly divided into 2
groups and their maxillary first molars were mesially moved with the use of closed-coil springs. The
experiment group received systemic injection of SP and the control group received phosphate-buffered saline
solution. Transportation distances of first molars were measured. Hematoxylin and eosin staining, tartrate-
resistant acid phosphatase staining, and immunohistochemistry staining were performed to evaluate alveolar
bone remodeling. Then the interferon (IFN) g and tumor necrosis factor (TNF) a concentrations in peripheral
blood and local periodontal tissue were measured. Finally, the effects of SP on bone marrow–derived stem
cell (BMSC) proliferation and migration were tested in vitro. Results: Systemic delivery of SP significantly
increased the distance of tooth movement and stimulated both osteoclast and osteoblast activities. The concen-
trations of IFN-g and TNF-a increased in peripheral blood at early phases of the experiment and decreased in
periodontal tissue at late phases. In vitro, the proliferation and migration of BMSCs were promoted by SP.
Conclusions: Systemic delivery of SP can accelerate orthodontic tooth movement and promote alveolar bone remod-
eling potentially through immunomodulation and mobilizing endogenous mesenchymal stem cells. (Am J Orthod
Dentofacial Orthop 2019;155:642-9)

O
rthodontic tooth movement is a process that involves peripheral nerves and immune cells, which regulates
force-induced bone metabolism and aseptic inflam- immune cell functions through autocrine and paracrine
mation, which is accompanied by minor reversible mechanisms via the high-affinity neurokinin 1 receptor.7
injury to the tooth-supporting tissue.1 Sufficient studies Studies have shown increased levels of SP in periodontal tis-
have confirmed that inflammatory cytokines, such as tumor sues during orthodontic tooth movement, indicating that SP
necrosis factor (TNF) a and interferon (IFN) g, play important may be associated in some way with the altered inflamma-
roles in the process of orthodontically induced tooth move- tory microenvironment and consequently influence tooth
ment.2-6 There have been lesser known mediators, such as movement.8,9 SP acts as a chemoattractant for monocytes
Substance P (SP), an undecapeptide produced from and lymphocytes and can enhance lymphocyte
proliferation and immunoglobulin production.10-15 In a
recent study, researchers demonstrated that SP acted as a
From
a
State Key Laboratory of Oral Diseases, Department of Orthodontics, West China
systemic messenger of injury and that systemic
School of Stomatology, West China Hospital of Stomatology, Sichuan University, administration of SP could successfully mobilize
Chengdu, People's Republic of China.
b
mesenchymal stem cells (MSCs) into peripheral blood in
Department of Orthodontics, Hospital of Stomatology Southwest Medical Uni-
versity, Luzhou, People's Republic of China.
both injured and uninjured animals.16 Because these mobi-
c
Nantong Stomatological Hospital, Nantong, People's Republic of China. lized cells possess pluripotent capacities for bone remodeling
d
Harvard School of Dental Medicine, Harvard University, Boston, Mass. and immunomodulation, how they would affect orthodon-
Shu An and Yueling Zhang are joint first authors and contributed equally to this
work.
tic tooth movement remains unpredictable and un-
All authors have completed and submitted the ICMJE Form for Disclosure of Po- tested.17-19 Therefore, we conducted the present study to
tential Conflicts of Interest, and none were reported. investigate the effect of systemic delivery of SP on
Funding: National Natural Science Foundation of China (grant numbers
81771114 and 81470776).
experimental orthodontic tooth movement in rats, which is
Address correspondence to: Jun Wang, State Key Laboratory of Oral Diseases, probably one of the first studies to test the systemic role of
West China School of Stomatology, West China Hospital of Stomatology, Si- SP in orthodontic treatment.
chuan University, 14#, 3rd Section, Renmin South Road, Chengdu 610041, Peo-
ple's Republic of China; e-mail, wangjunv@scu.edu.cn.
Submitted, January 2018; revised and accepted, May 2018. MATERIAL AND METHODS
0889-5406/$36.00
Ó 2019 by the American Association of Orthodontists. All rights reserved. All experimental procedures performed in this study
https://doi.org/10.1016/j.ajodo.2018.05.026 were based on a protocols approved by the Animal
642
An et al 643

Experiment Ethics Committee of the State Key Labora- protein samples were separated by means of 15% so-
tory of Oral Diseases of Sichuan University in China. dium dodecyl sulfate–polyacrylamide gel electrophoresis
All animals were obtained from the Sichuan University's and electroblotted onto nitrocellulose membrane. After
experimental animal center. Six-week-old Sprague- blocking with 5% nonfat dry milk, the membrane was
Dawley rats were used for cell culture and 10-week-old incubated with primary antibodies for TNF-a and INF-
Sprague-Dawley rats were used for establishing the an- g (Abcam, Cambridge, U.K.) followed by incubation
imal model of experimental tooth movement. with secondary antibody (1:5000; ZSGB-Bio, Beijing,
Forty-eight male Sprague-Dawley rats (10 weeks old, China). The signals were then visualized by means of
200 g) were randomly divided into 2 groups: the SP chemiluminescence detection.
group and the control group. A nickel-titanium For enzyme-linked immunosorbent assay (ELISA)
closed-coil spring (Grikin Advanced Materials, Beijing, analysis, peripheral blood samples harvested from rats
China) was fixed between the maxillary first molars killed on days 3, 7, and 14 were used for evaluating
and incisors. The maxillary first molars were moved serum concentrations of TNF-a and IFN-g with the
mesially with an orthodontic force of 40 g. The appli- use of ELISA kits (R&D Systems, Minneapolis, Minn).
ances were activated immediately after insertion, and The samples were set stable for 30 minutes. After centri-
no reactivation was performed in the following experi- fugation at 3000 rpm at 4 C for 10 minutes, the super-
ment period. The rats in the SP group received 3 intrave- nate was collected and assayed by means of ELISA for
nous injections of SP (5 nmol/kg; Sigma, St Louis, Mo) IFN-g and TNF-a (Abcam).
respectively at 0, 24, and 48 hours after the installation For cell counting, bone marrow cells were flushed out
of the orthodontic appliance. Correspondingly, the rats from bone cavity of femurs and tibias of 6-week-old
in control group received 3 intravenous injection of Sprague-Dawley rats. A single-cell suspension was ob-
phosphate-buffered saline solution. All rats were able tained by passing all bone marrow cells through a 70-
to survive during the experiment and maintain appro- mm cell strainer (BD Bioscience, Franklin Lakes, NJ). All
priate body weight. of the single cells were seeded at 1 3 106 onto 100-
Eight rats from each group were killed on days 3, 7, mm culture dishes (Corning, Inc., Corning, NY) and incu-
and 14 after application of orthodontic forces, and bated in a-MEM medium supplemented with 10% fetal
1-1.5 mL peripheral blood was taken from each killed bovine serum, 100 U/mL penicillin, and 100 mg/mL
rat. Then the maxilla was dissected and divided into streptomycin (all from Gibco, Grand Island, NY) at
halves. 37 C and 5% CO2. Medium was changed every 3 days.
The tooth movement was determined by measuring We seeded 1 3 106 bone marrow–derived stem cells
the distance between the first and second maxillary mo- (BMSCs) on 100-mm cell culture plates and cocultured
lars with the use of a feeler gauge on the dissected max- them with different doses of SP (0 mol/L, 1 3
illas from days 3, 7, and 14. 10 10 mol/L, 1 3 10 8 mol/L, and 1 3 10 6 mol/L). Af-
For histologic staining, the left half of the maxilla of ter 48 hours of treatment, cell counting was performed.
each rat was fixed in 4% paraformaldehyde for 24 hours Cell Counting Kit 8 (CCK-8) was also carried out. We
and decalcified in 10% EDTA solution (pH 7.4) at 4 C for seeded 4 3 103 BMSCs in each well of a 96-well plate in
at least 4 weeks. After dehydration, the hemimaxillas 100 mL culture medium and preincubated the plate for
were paraffin embedded and cut into 7-mm serial sec- 24 hours at 37 C and 5% CO2. Then the cells were
tions in a mesiodistal direction parallel to the long axis treated with 1 3 10 10 mol/L, 1 3 10 8 mol/L, and
of the distal root of the first molar. Selected sections 1 3 10 6 mol/L SP in 3 experiment groups. After
were investigated with hematoxylin and eosin staining, 48 hours of treatment, 10 mL CCK-8 solution (Dojindo,
tartrate-resistant acid phosphatase (TRAP) staining, Tabaru, Japan) was added to each well of the plate. After
and immunohistochemical staining. The primary anti- 2-hour incubation at 37 C and 5% CO2, the optical den-
body for osteocalcin (Santa Cruz Biotechnology, Santa sity value of each well was measured at 450 nm with the
Cruz, Calif) was used at a dilution of 1:100. use of a microplate reader (Bio-Rad, Hercules, Calif).
For Western blots, the maxillary first molar together Cell migration was evaluated with the use of a scratch
with its periodontal tissue was collected from the right assay. We seeded the third passage of BMSCs in 100-mm
half of the maxilla. The collected tissue was immediately plates and incubated the plates at 37 C and 5% CO2.
frozen with the use of liquid nitrogen and then ground When cell confluence reached 90%, a scratch was created
with mortars and pestles. The tissue powder was lysed with the use of a P200 pipette tip in all groups. In 3 exper-
with ice-cold lysis buffer (Keygen Biotech, Nanjing, iment groups, the cells were treated with 1 3 10 10 mol/
China) and then centrifuged at 10,000 rpm for 20 mi- L, 1 3 10 8 mol/L, and 1 3 10 6 mol/L of SP. At 0, 9, and
nutes at 4 C before quantification. Equal amounts of 18 hours after the treatment, the cells in the 4 groups

American Journal of Orthodontics and Dentofacial Orthopedics May 2019  Vol 155  Issue 5
644 An et al

were photographed with the use of a phase-contrast mi- Systemic delivery of SP altered both local and sys-
croscope. temic inflammatory states during orthodontic tooth
movement. As described earlier, orthodontic tooth
Statistical analysis movement is accompanied by aseptic inflammation,
SPSS 11.5 software (IBM, Chicago, Ill) was used for and proinflammatory cytokines such as TNF-a and
the statistical analyses. Significance was assessed with IFN-g are important in alveolar bone remodeling dur-
the use of 1-way analysis of variance, least square differ- ing tooth movement. We assessed serum concentra-
ence test, and t test. P \0.05 was considered to be tions of IFN-g and TNF-a in peripheral blood.
significant. ELISA showed that the serum concentrations of
proinflammatory cytokines increased after systemic
RESULTS delivery of SP (Figs 3, A and B). On days 3 and 7,
IFN-g was 1.4-fold greater in the SP group than in
Substance P accelerated orthodontic tooth move- the control group (P \0.01), and TNF-a was 1.3-
ment. During the first 3 days, the tooth movement under fold greater in the SP group (P \0.05; Figs 3, A
orthodontic forces was limited in both SP and control and B). Then we evaluated local inflammatory states
groups (Fig 1, C). In addition, our data showed that dur- of periodontal tissue by means of Western blot anal-
ing the early phase of the experiment, there was no sig- ysis. The data suggest that systemic delivery of SP
nificant difference of total tooth movement between could reduce the IFN-g and TNF-a expression in
groups (P .0.05; Fig 1, C). However, during the late periodontal tissues, especially on days 7 and 14
phase, the differences became significant (Fig 1). On (P \0.05; Fig 3, C).
Days 7 and 14, systemic delivery of SP increased total Substance P promoted the proliferation and migra-
tooth movement by, respectively, 1.3-fold (P \0.01) tion of rat BMSCs in a dose-dependent manner. To
and 1.5-fold (P\0.01) compared with the control group find out the effects of SP on the proliferation of BMSCs,
(Fig 1, C). we seeded BMSCs in 100-mm plates and treated them
Systemic delivery of SP promoted alveolar bone re- with 1 3 10 10 mol/L, 1 3 10 8 mol/L, and
modeling during tooth movement. Histologic staining 1 3 10 6 mol/L of SP respectively in three experiment
showed that after application of orthodontic forces, groups. After 24 hours and 48 hours, the experiment
the periodontal ligament on the compression side of group under treatment with 1 3 10 6 mol/L SP reached
the first molar became compressed whereas on the ten- higher cell confluences than the control group (Fig 4, A).
sion side the tissue expanded (Fig 2, A). Multinucleated And viable cell counting assay showed that the group
clastic cells were observed on the compression side, and with 1 3 10 6 mol/L SP showed the largest number of
these cells formed resorption lacunae around themselves viable cells (P \0.01) whereas the other 2 SP groups
on the surface of alveolar bone (Fig 2, B). Meanwhile, (1 3 10 8 mol/L and 1 3 10 10 mol/L) had similar
new bone deposition and many cuboidal osteoblasts numbers of viable cells to the control group (P .0.05;
were detected on the surface of alveolar bone on the ten- Fig 4, B), which was consistent with the results of
sion side (Fig 2, C). CCK-8 assay (Fig 4, C).
According to TRAP staining, TRAP-positive osteo- Then we applied a wound scratch assay to determine
clasts appeared on day 3 and increased on days 7 and the effects of SP on migration of BMSCs. The results
14 (Fig 2, D). In addition, quantitative analyses revealed showed that SP promoted BMSC migration in a dose-
that there were more osteoclasts in the periodontal tis- dependent manner after 9 hours and 18 hours of treat-
sue of the SP group than the control group on days 7 ment (Fig 4, D).
and 14 (P \0.05; Fig 2, D).
Immunohistochemical staining was performed to
investigate expression of osteocalcin to evaluate new DISCUSSION
alveolar bone formation. The signals of osteocalcin Various approaches have been reported to accelerate
were detected mainly in periodontal ligament cells, the orthodontic tooth movement, such as laser therapy,
osteoblast-like cells, and extracelluar matrix (Fig 2, E). corticotomy, infusion of parathyroid hormone, and local
Statistical analyses have shown that the expression of receptor activator of nuclear factor kB ligand gene
osteocalcin within each group increased on days 7 and transfer.20-22 Acceleration of efficient tooth movement
14 compared with day 3 (Fig 2, E). In addition, according has become the focus of orthodontic research.22 These
to quantitative analyses the SP group showed a signifi- techniques will shorten the time of orthodontic treat-
cantly higher expression of osteocalcin than the control ment sharply and subsequently benefit both patients
group at each time point (P \0.05; Fig 2, E). and orthodontists.

May 2019  Vol 155  Issue 5 American Journal of Orthodontics and Dentofacial Orthopedics
An et al 645

Fig 1. Effects of systemic delivery of substance P on orthodontically induced tooth movement in rats.
A, Representative pictures of intraoral molar movement for both groups on days 7 and 14 (occlusal
view of maxilla). The short blue lines indicate the spaces between the first and second molars which
was created by the mesial movement of maxillary first molar. B, Representative pictures of molar move-
ment on each side of the maxilla with a feeler gauge on days 3, 7, and 14 (occlusal and lateral views of
maxilla). The red arrows point to the spaces between first and second molars created by mesial move-
ment of the first molar. C, Time course of orthodontic tooth movement in rats treated with phosphate-
buffered saline solution (Vehicle) injection and substance P (SP) injection mean 6 SD. **P \0.01.
Each group at each time point: n 5 8.

Substance P, as a neuropeptide, can be released from late stages of orthodontic treatment compared with
both nervous system and immune cells.7 Its receptor neu- control (Fig 1, A-C). In addition, the number of TRAP-
rokinin 1 receptor can be found on both osteoclast and positive osteoclasts on the alveolar bone surface of the
immune cells, both of which are involved in alveolar compression side increased significantly in the SP group
bone remodeling during orthodontically induced tooth (Fig 2, D), which was consistent with some previous
movement.23,24 In addition, some studies have shown in vitro studies.25 These results indicate that SP could
that SP, as an injury-inducible factor, increased in peri- enhance osteoclast activities, which were associated
odontal tissue during orthodontic treatment and had with alveolar bone remodeling during tooth movement.
the potential to regulate inflammation during this pro- In addition, osteocalcin levels increased on the tension
cess.8,9 To find out whether SP could also be a potential side of alveolar bone after systemic administration of
target for accelerating orthodontically induced tooth SP, suggesting the potential capacity of SP in osteoblast
movement, we used the animal model of experimental stimulation (Fig 2, E). Some previous researchers have
tooth movement to test the systemic role of SP. This shown similar results although other researchers have re-
study is probably one of the first studies to investigate ported that SP could inhibit osteogenetic activities.26,27
the effects of SP on orthodontic tooth movement. This discrepancy could be explained by the different
The results regarding transportation distances of first inflammatory states within different experiment
molars showed that SP facilitated tooth movement at the models. In a periodontal disease model, SP together

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646 An et al

Fig 2. Effects of systemic delivery of substance P on alveolar bone (AB) remodeling during orthodontic
tooth movement. A, Hematoxylin and eosin staining of periodontal tissue of maxillary first molar. PDL,
periodontal ligament. Scale bar 5 200 mm. B, Magnification of the compression side of the distal root of
maxillary first molar. The red arrows indicate the osteoclasts and the resorption lacunae around them.
Scale bar 5 100 mm. C, Magnification of the tension side of the distal root of maxillary first molar. The
black arrows indicate the cuboidal osteoblast. Scale bar 5 100 mm. D, TRAP staining of periodontal
tissue of maxillary first molar on days 3, 7, and 14 for both groups. The black arrows indicate the
TRAP-positive osteoclasts. Scale bar 5 100 mm. E, Immunohistochemical staining for osteocalcin of
periodontal tissue of maxillary first molar on days 3, 7, and 14 for both groups. Scale bar 5 50 mm.

Fig 3. Effects of systemic delivery of substance P on local and systemic inflammatory states during
orthodontic tooth movement. A, B, Time course of proinflammatory cytokine levels in peripheral blood
of rats treated with phosphate-buffered saline solution (Vehicle) injection and substance P (SP) injec-
tion respectively, mean 6 SD. *P\0.05. Each group at each time point: n 5 8. C, Western blot analysis
of IFN-g and TNF-a in periodontal tissue of maxillary first molar on days 3, 7, and 14.

May 2019  Vol 155  Issue 5 American Journal of Orthodontics and Dentofacial Orthopedics
An et al 647

Fig 4. Effects of substance P on the proliferation and migration of rat BMSCs. A, Representative pic-
tures of cell counting assay for control group and SP group (1 3 10 6 mol/L) at 0, 24, and 48 hours. B,
Statistical analysis of viable cell numbers within 4 groups after 48-hour treatment. **P \0.01. C, CCK-8
analysis of 4 groups after 48-hour treatment. *P \0.05; **P \0.01. D, Migration assay of 4 groups at 0,
9, and 18 hours. The space between the two red dotted lines indicates the initial scratch created with the
use of a P200 pipette tip.

with lipopolysaccharide (LPS) could potentially inhibit effect on periodontal tissue as well as tooth roots if
osteoblast differentiation.26 However, in our orthodon- not well controlled.4,29 At early stages of
tic aseptic inflammatory model without LPS, SP could orthodontically induced tooth movement, TNF-a and
potentially promote osteoblast activities.27 Furthermore, IFN-g can be essential triggers of the remodeling of
considering the concept that mechanical loading may alveolar bone.3 However, it has also been demonstrated
alter the effects of proinflammatory substances on resi- that long-lasting inflammation can impair tooth move-
dent cells, the mechanobiology of osteoblasts might be ment, damage the periodontal tissue and even lead to
changed to another situation during orthodontic treat- unfavorable root resorption.4 In this study, we showed
ment, in which osteoblasts might increase their re- that local inflammation can be efficiently controlled
sponses to the stimulation of SP.28 Further studies with systemic delivery of SP from day 7 after surgery
need be conducted to find out the exact effects of SP (Fig 3, C). Correspondingly, tooth movement increased
on osteoblasts under different inflammatory and me- from day 7 after surgery (Fig 1). Taking all of this
chanical conditions. together, SP could indirectly promote tooth movement
In addition to the direct roles of SP on osteoclasts and through reducing inflammatory states at late stages of
osteoblasts during orthodontically induced tooth orthodontic treatment.
movement, our studies also demonstrated its capacity In our studies, SP altered not only the local inflam-
for immunomodulation. The reduced levels of proin- matory states but also systemic inflammation (Fig 3).
flammatory factors such as TNF-a and IFN-g suggest By days 3 and 7 after surgery, the serum concentrations
that SP could also alter the inflammatory state during of IFN-g and TNF-a were significantly greater in the SP
the orthodontic treatment (Fig 3, C). It is worth noting group than in the control group (Fig 3, A and B). This
that inflammation is “a double-eged sword”: It can phenomenon could be associated with the proinflamma-
promote alveolar bone remodeling and accelerate the tory roles of SP.7 However, the increased concentrations
tooth movement, and it may also have a destructive of these proinflammatory cytokines in the circulatory

American Journal of Orthodontics and Dentofacial Orthopedics May 2019  Vol 155  Issue 5
648 An et al

system were restricted to early stages of orthodontic ment during orthodontic tooth movement in humans. Eur J Oral
treatment, and within 2 weeks after the treatment the Sci 2007;115:355-62.
4. Ren Y, Hazemeijer H, de Haan B, Qu N, de Vos P. Cytokine profiles
inflammation had been brought back to levels similar
in crevicular fluid during orthodontic tooth movement of short
to that of control group (Fig 3, A and B), which suggests and long durations. J Periodontol 2007;78:453-8.
that systemic delivery of SP is safe for the overall health 5. Yamaguchi M, Kasai K. Inflammation in periodontal tissues in
of the animals. response to mechanical forces. Arch Immunol Ther Exp (Warsz)
Recent researchers have reported that systemic deliv- 2005;53:388-98.
6. Hazan-Molina H, Reznick AZ, Kaufman H, Aizenbud D. Peri-
ery of SP could mobilize MSCs from the bone marrow
odontal cytokines profile under orthodontic force and extracorpo-
and accelerate wound healing.16 Considering this capac- real shock wave stimuli in a rat model. J Periodontal Res 2015;50:
ity of SP, it is possible that SP could accelerate rat tooth 389-96.
movement through mobilizing endogenous MSCs. 7. Suvas S. Role of Substance P Neuropeptide in inflammation, wound
Endogenous MSCs mobilized by systemic delivery of healing, and tissue homeostasis. J Immunol 2017;199:1543-52.
8. Nicolay OF, Davidovitch Z, Shanfeld JL, Alley K. Substance P
SP might contribute to speeding tooth movement by
immunoreactivity in periodontal tissues during orthodontic tooth
differentiating into osteoblasts and then participating movement. Bone Miner 1990;11:19-29.
in alveolar bone remodeling.16,17 These cells could not 9. O’Hara AH, Sampson WJ, Dreyer CW, Pierce AM, Ferguson IA.
only have multipotent differentiation ability but also Immunohistochemical detection of nerve growth factor and its re-
play an important role in immunoregulation. ceptors in the rat periodontal ligament during tooth movement.
Arch Oral Biol 2009;54:871-8.
Considering the reduced expression of TNF-a and IFN-
10. Ho WZ, Kaufman D, Uvaydova M, Douglas SD. Substance P aug-
g (Fig 3, C), it is also possible that recruited MSCs ments interleukin-10 and tumor necrosis factor-alpha release by
together with SP help to control the local inflammatory human cord blood monocytes and macrophages. J Neuroimmunol
state and subsequently promote tooth movement. In this 1996;71:73-80.
way, the role of systemic delivery of SP may be attributed 11. Maggi CA. The effects of tachykinins on inflammatory and im-
mune cells. Regul Pept 1997;70:75-90.
to the concept of in situ tissue regeneration, which
12. Santoni G, Perfumi MC, Spreghini E, Romagnoli S, Piccoli M. Neu-
means using bioactive molecules to recruit endogenous rokinin type-1 receptor antagonist inhibits enhancement of T cell
stem cell homing to the site of injury.30,31 functions by substance P in normal and neuromanipulated
To find out the effects of SP on BMSCs, we carried capsaicin-treated rats. J Neuroimmunol 1999;93:15-25.
out in vitro culture of rat BMSCs and administered 13. Hood VC, Cruwys SC, Urban L, Kidd BL. Differential role of neuro-
kinin receptors in human lymphocyte and monocyte chemotaxis.
different concentrations of SP. The results showed that
Regul Pept 2000;96:17-21.
SP could stimulate BMSC proliferation and migration, 14. O’Connell TM, O’Connell J, O'Brien DI, Goode T, Bredin CP,
which further indicated that SP might promote tooth Shanahan F. The role of substance P in inflammatory disease.
movement through mobilizing endogenous MSCs in J Cell Physiol 2004;201:167-80.
this experimental tooth movement model (Fig 4). 15. Schratzberger P, Reinisch N, Prodinger WM, et al. Differential
chemotactic activities of sensory neuropeptides for human pe-
ripheral blood mononuclear cells. J Immunol 1997;158:
CONCLUSIONS 3895-901.
Our results show that systemic delivery of SP can 16. Hong HS, Lee J, Lee E, et al. A new role of substance P as an injury-
inducible messenger for mobilization of CD29(1) stromal-like
potentially promote orthodontic tooth movement and
cells. Nat Med 2009;15:425-35.
stimulate alveolar bone remodeling. This phenomenon 17. Bernardo ME, Fibbe WE. Mesenchymal stromal cells: sensors and
could be explained in part by the potential roles of SP switchers of inflammation. Cell Stem Cell 2013;13:392-402.
in increasing osteoclast proliferation initially and con- 18. Burr SP, Dazzi F, Garden OA. Mesenchymal stromal cells and reg-
trolling inflammation at later stages of orthodontic ulatory T cells: the Yin and Yang of peripheral tolerance? Immunol
Cell Biol 2013;91:12-8.
tooth movement. In addition, our results suggest that
19. Hall SR, Tsoyi K, Ith B, Padera RF Jr, Lederer JA, Wang Z, Liu X,
SP can promote tooth movement through mobilizing Perrella MA. Mesenchymal stromal cells improve survival during
endogenous MSCs, which may differentiate into osteo- sepsis in the absence of heme oxygenase-1: the importance of
blasts and then contribute to alveolar bone remodeling. neutrophils. Stem Cells 2013;31:397-407.
20. Kanzaki H, Chiba M, Arai K, et al. Local RANKL gene transfer to the
periodontal tissue accelerates orthodontic tooth movement. Gene
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