Assessment of IL-1β and VEGF concentration in a rat model during orthodontic tooth movement and extracorporeal shock wave therapy

ObjectiveThis study aimed to investigate PDL's cytokine concentration fluctuations after induction of orthodontic force with and without extracorporeal shock wave therapy in a rat model.Materials and methodsAn orthodontic appliance was fabricated and applied between the molars and the incisors of rats. The rats were treated by a single episode of 1000 shock waves and gingival crevicular fluid was collected for 3 days. The expression and concentration of IL-1β and VEGF were evaluated by ELISA assay. On day 3 all rats were sacrificed and histologic and immunohistochemical assays were applied.ResultsIL-1β concentration rose in both the treated and non treated shockwave groups on the first day, however it was statistically significantly higher in the treated group on day 2. No statistically significant difference was detected between the groups on day 3. The number/area of TRAP positive cells was higher in the non shockwave group than in the treated group. The percentage of cells expressing VEGF displayed the opposite trend. The findings regarding the immunohistochemical assay for IL-1β corresponded with those of the ELISA assay on day 3.ConclusionThe application of shockwaves during orthodontic tooth movement influences the expression of IL-1β and VEGF and may alternate the periodontal remodelling expected rate.     

Human interleukin-1 beta and interleukin-1 receptor antagonist secretion and velocity of tooth movement

The cytokines interleukin-1 beta (IL-1 beta) and IL-1 receptor antagonist (IL-1RA) probably play a part in orthodontic tooth movement. Here, the force magnitudes and the area of force application in the compressed periodontal ligament (PDL) were controlled and the velocity of tooth movement correlated with concentrations of IL-1 beta and IL-1RA in the gingival crevicular fluid (GCF). Seven individuals undergoing orthodontic treatment involving maxillary first premolar extractions and distal movement (bodily retraction) of the maxillary canines participated in the 84-day study. For each participant, continuous retraction forces were applied so that they received equivalent PDL stresses of 13 kPa for one canine and 4 kPa for the other. GCF cytokine concentrations from experimental and control teeth were expressed relative to total protein in the GCF and compared using an 'Activity Index' (AI)=Experimental (IL-1 beta/IL-1RA)/Control (IL-1 beta/IL-1RA). The results showed that the velocity of tooth movement in an individual was related to their AI. The correlation between AI and tooth movement was stronger from the distal (R(d)=0.78) than from the mesial (R(m)=0.65) of retracted teeth. The results demonstrate that equivalent force systems produce individual differences in cytokine production, which correlate with interindividual differences in the velocity of canine retraction.     

Relationship between substance P and interleukin-1beta in gingival crevicular fluid during orthodontic tooth movement in adults

Metabolism by peptidases plays an important role in modulating the levels of biologically-active neuropeptides, while that of substance P (SP), a component of gingival crevicular fluid (GCF), may potentiate the inflammatory process in orthodontic tooth movement. The aim of this study was two-fold: (1) to investigate GCF levels of SP and interleukin-1beta (IL-1beta) during human orthodontic tooth movement, and (2) to determine the correlation coefficients between SP and IL-1beta levels in the GCF. The subjects were 3 males, with a mean age of 21.3 +/- 2.8 years old, and 6 females, with a mean age of 23.1 +/- 2.4 years, undergoing orthodontic movement of a single tooth, with the contralateral tooth used as the control. GCF was sampled at the control and treatment (compression) sites before and 1, 4, 8, 24, 72, 120, and 168 hours after initiation of orthodontic treatment. Prevention of plaque-induced inflammation allowed assessment of the dynamics of mechanically stimulated SP and IL-1beta levels in the GCF, which were determined using enzyme-linked immunosorbent assay (ELISA) kits. GCF levels of SP and IL-1beta for the treated teeth were significantly higher (P < 0.001) than for the corresponding control teeth from 8 to 72 hours, and peaked at 24 hours. These results show that the amounts of SP and IL-1beta in GCF increase with orthodontic tooth movement, and indicate that such increases may be involved in inflammation in response to mechanical stress.     

Interleukin-1alpha and tumor necrosis factor-alpha expression during the early phases of orthodontic tooth movement in rats

Remodelling of the periodontium after application of mechanical forces constitutes the basis of clinical orthodontics and various immunoregulatory molecules are involved in this process. The aim of this study was to investigate the expression of the cytokines interleukin-1alpha (IL-1alpha) and tumor necrosis factor-alpha (TNF-alpha) in dental tissues during the early phases of orthodontic tooth movement. Eightteen male Wistar rats were used. All maxillary right first molars were moved orthodontically, with a force of 0.5 N, for 3 h, 1 d, and 3 d. The contralateral sides served as untreated controls. Parasagittal sections of the maxillary molars and the surrounding tissues were subjected to immunohistochemical staining for IL-1alpha or TNF-alpha, and were evaluated with light microscopy. IL-1alpha and TNF-alpha were expressed in the bone and periodontal ligament (PDL) along the roots of the orthodontically moved molars and in the gingiva. Increased expression of both cytokines was observed in the aforementioned areas after 1 and 3 d of tooth movement. The pulp tissue exhibited only minor changes in cytokine expression during tooth movement. The results suggest that mechanical stress results in almost immediate inflammatory reactions in various dental tissues.     

Neurotransmitters, cytokines, and the control of alveolar bone remodeling in orthodontics

This article described research aimed at testing the hypothesis that tissue remodeling during orthodontic tooth movement is modulated, at least in part, by factors derived from the nervous and vascular (immune) systems. Specifically, the neurotransmitters SP and VIP and the cytokines IL-1 alpha and IL-1 beta were localized immunohistochemically in paradental tissues of cat canines that had been treated by the application of an 80 g tipping force for 1 hour to 14 days. Increased staining (concentrations) of these agents were found in areas of PDL tension and compression at different time periods. Moreover, administration of SP and IL-1 beta to human PDL fibroblasts in vitro for 1 to 60 minutes resulted in significant increases in the levels of the intracellular "second messenger" cAMP, as well as of PGE2, a plasma membrane-associated fatty acid believed to serve as a local regulator of bone cell activity. Taken together, these results tend to support the hypothesis that neurotransmitters and cytokines play a regulatory role in orthodontic force-induced alveolar bone remodeling. Consequently, determination of the cytokine synthetic activity by leukocytes of orthodontic patients may inform about their alveolar bone remodeling potential.     

Soluble cytokine receptor treatment in experimental orthodontic tooth movement in the rat

Pro-inflammatory cytokines, such as interleukin-1 (IL-1) and tumour necrosis factor-alpha (TNF-alpha), are believed to play a role in the biological processes involved in the course of orthodontic tooth movement and especially in root resorption. The inhibition of cytokine activity, e.g. by soluble receptors, could be beneficial in reducing this unwanted side-effect.The aim of this study was to investigate the role of cytokines IL-1 and TNF-alpha in the course of experimentally induced tooth movement. The upper left first molar was moved orthodontically in 80 male Wistar rats using a coil spring with a force of 0.5 N. Starting at day -1, three groups of 20 animals each received daily intraperitoneal injections (ip) of 2 ml of 1 mug/ml soluble receptors (a) to IL-1(sIL-RII), (b) to TNF-alpha (sTNF-alpha-RI) and (c) a combination of (a) and (b). Twenty animals served as the control. After 3, 6, 9 and 12 days, the animals were killed in groups of five. The amount of tooth movement was registered and the maxillae were prepared for histological and histomorphometric analysis. Osteoclasts and odontoclasts were identified using tartrate-resistant acid phosphatase (TRAP) histochemistry.The amount of tooth movement was reduced in all receptor-treated groups by approximately 50 per cent. At the same time, the number of TRAP-positive cells on the desmodontal bone surface and on the surface of the roots was reduced. Thus, systemic application of soluble receptors to IL-1 and TNF-alpha following experimental induction of tooth movement in the rat reduced the number of osteoclasts as well as odontoclasts.     

Impact of extracorporeal shock wave therapy (ESWT) on orthodontic tooth movement—a randomized clinical trial

Objectives

This randomized clinical trial investigated the effect of extracorporeal shock waves on the amount of orthodontic tooth movement and periodontal parameters.

Material and methods

Twenty-six adult orthodontic patients participated in this clinical trial; all of them receiving lower second molar mesially directed movement. The fixed orthodontic device included superelastic coil springs (200 cN) and miniscrews as temporary anchorage device. The active treatment group received a single shock wave treatment with 1,000 impulses in the region of tooth movement. The placebo group was treated with deactivated shock wave applicator with an acoustic sham. The study period lasted 4 months with a monthly data exploration.

Results

No statistically significant difference in posterior-anterior tooth movement between the treatment and placebo group was seen during observation period. Gender had no significant influence on tooth movement in either group. No significant difference occurred in mesio-distal tipping and rotation, but a significant difference (p?=?0.035) in bucco-lingual tipping of the molars was found. Periodontal status of the patients (sulcus probing depth, gingival index) did not significantly differ in both groups. The plaque index showed a significant difference (p?=?0.003).

Conclusions

Single application of extracorporeal shock wave treatment was associated neither with a statistically significant acceleration of tooth movement nor with an altered periodontal status in vivo.

Clinical relevance

Shock waves showed no harmful effects in the investigated area. Their clinical use for lithotripsy during orthodontic therapy might be permitted.     

Orthodontic tooth movement and de novo synthesis of proinflammatory cytokines.

Interleukin-1beta (IL-1beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) are proinflammatory cytokines that are thought to play a role in bone remodeling, bone resorption, and new bone deposition. In the present work, in situ hybridization was performed to measure the messenger RNA expression of IL-1beta, IL-6, and TNF-alpha at 3, 7, and 10 days after the application of orthodontic force on the maxillary first molars of 12 rats. The contralateral side and 3 untreated rats served as controls. Measurements of the messenger RNA expression were selected as the means to investigate the role of orthodontic force in de novo synthesis of proinflammatory cytokines. After the application of force, the induction of IL-1beta and IL-6 was observed to reach a maximum on day 3 and to decline thereafter. No messenger RNA induction of either cytokine was measured in the control teeth. The messenger RNA expression of TNF-alpha was not detected at any time point of this study in the experimental or contralateral sides or in the control animals. Our data support the hypothesis that these proinflammatory cytokines may play important roles in bone resorption after the application of orthodontic force.     

Composition changes in gingival crevicular fluid during orthodontic tooth movement: comparisons between tension and compression sides

The aim of this study was to evaluate whether the application of tension or compression forces exerted on the periodontium during the early phase of orthodontic tooth movement is reflected by differences in the composition of the gingival crevicular fluid (GCF), at the level of interleukin-1beta (IL-1beta), substance P (SP), and prostaglandin E(2) (PGE(2)). Eighteen children (mean age 10.8 yr) starting orthodontic treatment were included in the study. Molar elastic separators were inserted mesially to two first upper or lower molars. One of the antagonist molars served as the control. GCF was collected from the mesial and distal sites of each molar, before (-7 d, 0 d) and immediately after (1 min, 1 h, 1 d, and 7 d) the placement of separators. The levels of IL-1beta, SP, and PGE(2) were determined by enzme-linked immunosorbent assay. At the orthodontically moved teeth, the GCF levels of IL-1beta, SP, and PGE(2) were significantly higher than at the control teeth in both tension and compression sides, and at almost all occasions after insertion of separators. The increase, relative to baseline values, was generally higher in tension sides. For the control teeth, the three mediators remained at baseline levels throughout the experiment. The results suggest that IL-1beta, SP, and PGE(2) levels in the GCF reflect the biologic activity in the periodontium during orthodontic tooth movement.     

Cytokine profiles in crevicular fluid during orthodontic tooth movement of short and long durations

BACKGROUND: Orthodontic treatment induces a distortion of the extracellular matrix of the periodontium, resulting in alterations in cytoskeletal configuration. Cytokines are known to facilitate this process by inducing cellular proliferation, differentiation, and stimulation of periodontal remodeling. The aim of the present study was to measure a panel of proinflammatory cytokines (PICs) in crevicular fluid (GCF) samples during tooth movement of short and long durations. METHODS: Twelve patients (11 to 27 years of age) participated in this study: six patients each for tooth movement of short and long duration. GCF sampling was done at different times, ranging from 24 hours to 4 months after force application. The profiles of PICs were analyzed with a multiplex technique. RESULTS: PICs were elevated significantly in the early stage of tooth movement but at different time points. Interleukin (IL)-1beta and -6 and tumor necrosis factor-alpha (TNF-alpha) reached significant levels at 24 hours; IL-8 reached a significant elevation at 1 month. During the linear stage of tooth movement, all cytokines were diminished to their baseline levels. The results demonstrated that IL-1beta, -6, and -8 and TNF-alpha play a significant role during the early stage of tooth movement but not during the linear stage. CONCLUSIONS: Once the microenvironment of periodontal tissue is activated by an orthodontic force, several key PICs are produced to trigger a cascade of cellular events. The periodontal system stabilizes at a new physiological homeostasis as indicated by the downregulation of the early-phase PICs.     

实验性牙移动兔牙周组织中VEGF受体1（VEGFR-1）的变化

目的：观察兔实验性正畸牙齿移动过程中VEGFR-1在牙周组织中的表达。方法：选用体重在2.0 kg左右的日本大耳白兔35只,分为正常组与实验1、3、5、7、14、21天组。建立兔正畸牙齿移动模型,对实验标本进行VEGFR-1免疫组化染色。通过组织学观察,计算机图像分析系统对兔牙周组织中VEGFR-1的表达变化进行平均灰度分析,进行统计学处理与分析。结果：在正常牙周组织中,VEGFR-1有所表达,局限分布于新生血管内皮细胞。牙周组织受到正畸压力后,压力区1-7 d VEGFR-1的表达与正常牙周组织相比,差异显著（P〈0.01）;在张力区在3-14 d时,VEGFR-1阳性反应灰度积分与正常牙周组织相比,差异显著（P〈0.01）。高峰值压力区出现在牙齿受力后第5天,张力区出现在第5天。结论：正常兔牙周组织中存在VEGFR-1;VEGFR-1参与了兔实验性正畸牙齿移动牙周组织的改建过程。     

大鼠正畸牙移动过程中牙周组织内转化生长因子β1的变化

目的 了解大鼠正畸牙移动过程中转化生长因子β１（ｔｒａｎｓｆｏｒｍｉｎｇ ｇｒｏｗｔｈ ｆａｃｔｏｒβ１,ＴＧＦ－β１）在牙周组织内的变化,探讨ＴＧＦ－β１在正畸牙移动过程中的作用。方法 对大习正畸牙移动不同阶段的牙及牙周组织联合切片,进行免疫组织化学研究。结果 正畸牙移动过程中,牙周膜内ＴＧＦ－β１表达增加。强力第５～１０天,张力区ＴＧＦ－β１表达增加的幅度明显大于压力区。结论 ＴＧＦ－β１在正     

中草药灯盏花对兔正畸牙移动过程中牙周组织血管内皮生长因子表达的影响

目的 检测中草药灯盏花对正畸牙牙周组织中血管内皮生长因子（VEGF）表达的影响,推测其加速正畸牙移动的可能作用机理。方法 45只兔分为A、B、C组,均以双侧下颌第一磨牙为实验牙。A组右侧第一磨牙离子导入灯盏花（A-R组）,左侧导入生理盐水作为对照（A-L组）。B组安装使双侧下颌第一磨牙近中移动的正畸加力装置,同时右侧离子导入灯盏花（B-R组）,左侧导入生理盐水作为对照（B-L组）。C组为空白对照组。A、B组分别于实验的1、3、7、14 d处死,每次处死5只兔。取所有动物的双侧下颌骨标本,B组测量牙移动距离后,所有标本制成第一磨牙的牙周组织切片,免疫组化法检测VEGF的表达。结果 B组第一磨牙移动距离在加力1、3、7、14 d时依次递增,B-R组在3、7、14 d的移动距离较B-L组大（P<0.01）。免疫组化染色表明C组和A-L组VEGF表达阴性,
A-R组、B-L组和B-R组表达较强,其中B-R组表达最强;A-R组和B-R组表达高峰在实验第3天,而B-L组表达高峰出现于第7天。结论 灯盏花可能通过上调正畸牙加力初期牙周组织中VEGF的表达而加速其移动。     

Effect of orthodontic force on expression levels of ten cytokines in gingival crevicular fluid

Various types of inflammatory mediators are involved in the cascade of biological events behind tissue remodeling allowing orthodontic tooth movement. This split-mouth longitudinal study aimed to evaluate the gingival crevicular fluid (GCF) levels of ten cytokines, IL-6, IL-8, IL-10, IL-13, IL-17, IFN-γ, GM-CSF, MCP-1, MIP-1β and TNF-α, during initial orthodontic treatment. The sample comprised 15 healthy patients (9 males and 6 females, 13.9 ± 2.5 years). The lower (test) incisors were moved using fixed appliance carrying a 0.014-inch nickel titanium wire, whereas the upper (control) incisors were bonded without any force. The GCF was collected from the test and control teeth before fixed appliance mounting (baseline) and after 1, 7 and 21 days. In 6 sites per tooth, from canine to canine, periodontal conditions were defined as the percentage of sites with visible plaque and bleeding on probing. The total GCF cytokines levels were quantified using multianalysis Luminex technology. Throughout the experimental term, and for both test and control teeth, the mean percentage of sites with visible plaque and bleeding on probing were generally below 25% and 15%, respectively, although variability was also seen. In the test teeth, the GCF levels of all the cytokines remained constant throughout the experimental term. On the contrary, significant reductions were seen in the control teeth for each cytokine. Moreover, significantly greater levels of IL-6, GM-CSF, MCP-1 and TNFα were seen in the test teeth as compared to the control teeth at 7 days. The reasons for the differential behavior in the levels of all the investigated cytokines between the test and control teeth may be related to the presence of orthodontic forces and/or subclinical tissue inflammation. Further investigation is needed to elucidate potential roles for these biomarkers in the tissue remodeling incident to orthodontic tooth movement.     

Immunocompetent cells and cytokine expression in the rat periodontal ligament at the initial stage of orthodontic tooth movement

Objective

The aim of the present study is to investigate the involvement of immunocompetent cells, pro-inflammatory cytokines and HSP, to evaluate a change of periodontal ligament during the initial stage of orthodontic tooth movement.

Design

In the present study, we investigated the distributional density of immunocompetent cells, the localisation of cytokines, and the expression levels of their mRNA in the periodontal ligament during the initial stage of orthodontic tooth movement, using immunohistochemistry and real-time PCR.

Results

Orthodontic tooth movement led to significant recruitment of OX6⁺ cells and ED1⁺ cells in the rat PDL. Double-immunofluorescence staining showed that some ED1⁺ cells expressed pro-inflammatory factors of IL-1β and TNF-α in the PDL during orthodontic tooth movement. Real-time PCR analysis revealed that the expression levels of IL-1β (Il1b) and TNF-α (Tnf) mRNA gradually increased following its decrease after 1 h of orthodontic tooth movement. These findings suggest that ED1⁺ cells are involved in the expression of TNF-α and IL-1β and the subsequent regulation of bone resorption on pressure side. HSP27 (Hspb1) mRNA levels were significantly increased as compared with the control at 1 h of the initial stage of treatment.

Conclusion

ED1⁺ cells involved in the expression of TNF-α and IL-1β may play an important role in the initial reaction of the PDL and in the induction of the osteoclastic bone resorption during orthodontic tooth movement.     

大鼠牙齿移动过程中牙周组织中基质金属蛋白酶—2的表达和分布

目的：观察大鼠牙齿正畸形移动过程中牙周组织中基质金属蛋白酶－2（MMP－2）在受力下的表达和分布。方法：选取雄性SD大鼠30只,分为空白对照组、牙齿移动1、3、6和10d组。用原位杂交染色方法观察并比较各组中第一磨牙牙周组织中MMP－2的表达和分布。结果：MMP－2基因在受力牙齿压力侧牙周组织的破骨细胞和张力侧牙周组织的成骨细胞和成纤细胞中的阳性表达明显强于对照组,且随受力时间的增加而增强。结论：本实验证明MMP－2参与了正畸牙齿移动过程中牙周组织的改建。     

矫治力对兔牙周组织中Akt蛋白表达的影响

目的 探讨矫治力对兔牙周组织中Akt蛋白表达的影响。方法 选用24只日本大耳白兔建立正畸牙移动的动物模型,将实验动物上颌右侧戴矫治器,作为实验组;左侧未戴矫治器,作为对照组。分别在戴矫治器后3、5、7、14d各处死6只实验动物。用Western blot免疫印迹分析方法 对Akt蛋白表达的变化进行检测。结果 Western blot免疫印迹结果显示,加力3d后牙周组织中Akt蛋白表达增强,7d后牙周组织中Akt蛋白明显增强,随后缓慢下降,与对照侧相比,差异有统计学意义(P<0.01)。结论 矫治力作用下兔牙周组织中Akt蛋白的表达变化明显,提示Akt参与牙周组织改建。     

犬牙周膜牵张快速移动牙压力侧牙周组织中RANKL和OPG表达的研究

目的观察Beagle犬牙槽隔减阻牙周膜牵张(periodontal ligament distraction,PDLD)快速移动牙齿压力侧牙周组织中RANKL和OPG的表达变化。方法 8只纯种Beagle犬随机分为四组,每只犬下颌左右第一前磨牙随机分为PDLD侧和常规方法正畸牙齿移动对照侧。测量相同加力时间内移动牙的移动距离;取标本,HE染色观察移动牙压力侧牙周组织的形态学变化;TRAP染色观察压力侧牙周组织中破骨细胞数目的变化;免疫组化染色观察移动牙压力侧牙周组织中RANKL、OPG的变化。结果加力后,PDLD侧移动牙压力侧牙周组织中TRAP阳性破骨细胞数明显增加,在加力2周时达峰值。PDLD侧移动牙压力侧牙周组织中RANKL和OPG含量均有升高,RANKL/OPG含量比值在加力2周时最高。结论与常规正畸牙齿移动相比,在牙槽隔减阻牙周膜牵张移动牙齿过程中RANKL、OPG的表达和RANKL/OPG含量比值变化更显著,与移动牙压力侧牙周组织的快速改建有关。     

Force-induced IL-8 from periodontal ligament cells requires IL-1beta

During orthodontic tooth movement, mechanical stresses induce inflammatory reactions in the periodontal ligament (PDL). We hypothesized that chemokines released from PDL cells under mechanical stress regulate osteoclastogenesis, and investigated the profiles and mechanisms of chemokine expression by human PDL cells in response to mechanical stress. In vitro, shear stress and pressure force rapidly increased the gene and protein expressions of IL-8/CXCL8 by PDL cells. Consistently, amounts of IL-8 in the gingival crevicular fluid of healthy individuals increased within 2 to 4 days of orthodontic force application. The PDL cells constitutively expressed low levels of IL-1beta, which were not further increased by mechanical stress. Interestingly, neutralization of IL-1beta abolished IL-8 induction by mechanical stresses, indicating that IL-1beta is essential for IL-8 induction, presumably though autocrine or paracrine mechanisms. Finally, experiments with signal-specific inhibitors indicated that MAP kinase activation is essential for IL-8 induction.     

Extracorporeal shock wave therapy induces alveolar bone regeneration

Periodontal inflammation with alveolar bone resorption is a hallmark of periodontitis. We hypothesized that extracorporeal shock wave therapy (ESWT) could promote the regeneration of alveolar bone following Porphyromonas gingivalis-induced periodontitis in rats. Rats were infected with P. gingivalis for 10 wks, which caused alveolar bone resorption. The rats were then treated with a single episode of 100, 300, or 1000 impulses of shock wave on both cheeks at energy levels 0.1 mJ/mm(2). Alveolar bone levels were determined at 0, 3, 6, and 12 wks following ESWT and compared with those in untreated controls. Infected rats treated with 300 and 1000 impulses demonstrated significantly improved alveolar bone levels at 3 wks compared with untreated controls, and the improved levels remained for at least 6 wks in most rats. The results demonstrated effective regeneration of alveolar bone by ESWT and suggested that ESWT should be evaluated as an adjunct in the regeneration of periodontal tissues following periodontal disease. Abbreviations: ESWT, extracorporeal shock wave therapy; PCR, polymerase chain-reaction.