RANKL, osteopontin, and osteoclast homeostasis in a hyperocclusion mouse model

The biological mechanisms that maintain the position of teeth in their sockets establish a dynamic equilibrium between bone resorption and apposition. In order to reveal some of the dynamics involved in the tissue responses towards occlusal forces on periodontal ligament (PDL) and alveolar bone homeostasis, we developed the first mouse model of hyperocclusion. Swiss–Webster mice were kept in hyperocclusion for 0, 3, 6, and 9 d. Morphological and histological changes in the periodontium were assessed using micro-computed tomography (micro-CT) and ground sections with fluorescent detection of vital dye labels. Sections were stained for tartrate-resistant acid phosphatase, and the expression of receptor activator of nuclear factor-κB ligand (RANKL) and osteopontin (OPN) was analyzed by immunohistochemistry and real-time polymerase chain reaction (PCR). Traumatic occlusion resulted in enamel surface abrasion, inhibition of alveolar bone apposition, significant formation of osteoclasts at 3, 6 and 9 d, and upregulation of OPN and RANKL. Data from this study suggest that both OPN and RANKL contribute to the stimulation of bone resorption in the hyperocclusive state. In addition, we propose that the inhibition of alveolar bone apposition by occlusal forces is an important mechanism for the control of occlusal height that might work in synergy with RANKL-induced bone resorption to maintain normal occlusion.     

Effects of different force magnitudes on corticotomy-assisted orthodontic tooth movement in rats

Objectives:To investigate the effects of light and heavy forces with corticotomy on tooth movement rate, alveolar bone response, and root resorption in a rat model.Materials and Methods:The right and left sides of 40 male Wistar rats were randomly assigned using the split-mouth design to two groups: light force with corticotomy (LF) and heavy force with corticotomy (HF). Tooth movement was performed on the maxillary first molars using a nickel-titanium closed-coil spring delivering either 10 g (light force) or 50 g (heavy force). Tooth movement and alveolar bone response were assessed by micro–computed tomography (micro-CT) at day 0 as the baseline and on days 7, 14, 21, and 28. Root resorption was examined by histomorphometric analysis at day 28.Results:Micro-CT analysis showed a significantly greater tooth movement in the HF group at days 7 and 14 but no difference in bone volume fraction at any of the observed periods. Histomorphometric analysis found no significant difference in root resorption between the LF and HF groups at day 28.Conclusions:Heavy force with corticotomy increased tooth movement at days 7 and 14 but did not show any difference in alveolar bone change or root resorption.     

The effect of oestrogen deficiency on the alveolar bone resorption caused by traumatic occlusion

This study, using 132 female rats, was designed to investigate whether oestrogen loss facilitates alveolar bone alterations induced by traumatic occlusion. Rats were ovariectomized (OVX) or underwent sham-operation (Sham). Seven days after surgery, half of the rats in each group were subjected to experimental traumatic occlusion (trauma), and the other half were left untreated. Thus, there were four groups: OVX+trauma, Sham+trauma, OVX, and Sham. Rats in each group were killed 1, 3, 5, 7, or 10 days after the introduction of occlusal trauma. The resected mandibles were processed without decalcification, and histomorphometric measurements were performed in the alveolar bone adjacent to the periodontal ligament of the first molar. The statistical assessment of the time- and group-specific differences by analysis of variance revealed significant differences between the OVX+trauma and Sham+trauma groups in the resorption parameters, but not in the formation parameters. The results show that the alveolar bone dynamics induced by traumatic occlusion are enhanced by oestrogen deficiency.     

Histological and histomorphometrical changes in rat alveolar bone following antagonistic tooth extraction and/or ovariectomy

OBJECTIVE: To examine changes appearing in the alveolar bone following the removal of the mechanical stress of occlusal loading, as well as the added influence of estrogen deficiency on such changes. DESIGN: The right mandibular molars of female rats were extracted. After 8 weeks, 12 animals were ovariectomized (OVX), and the other 12 were subjected to sham surgery (sham). Four weeks after surgery, all rats were sacrificed. The left-half and right-half maxillas of the sham group (the sham-occluded side and the sham-extruded side, respectively) and right-half maxilla of OVX group (the OVX-extruded side) were examined by histological observation and bone histomorphometry. RESULTS: The vertical height of alveolar bone in the sham-extruded and the OVX-extruded sides increased as compared with that of the sham-occluded side. In both extruded sides, active bone formation occurred on the surface of the alveolar bone facing the periodontal ligament, but the bone marrow was expanded and the bone volume had decreased in the internal area of the alveolar bone. In the OVX-extruded side, the bone marrow expanded more remarkably than that of the sham-extruded side, and the highest percentage of osteoclast surface was detected. CONCLUSIONS: Around the extruded teeth, there were regional differences in bone dynamics between the internal area of the alveolar bone and the bone surface facing the periodontal ligament, and estrogen deficiency seems to have caused further loss of bone volume in the interior of the alveolar bone supporting the extruded tooth.     

牙齿移动过程中牙槽骨显微结构动态变化的微型CT研究

目的 通过微型CT探讨正畸牙齿移动过程中矫治力对牙槽骨骨小梁的影响规律,以期为正畸临床提供参考.方法 选择10周龄SD雄性大鼠10只建立实验动物模型,施加0.196 N矫治力,近中移动上颌第一磨牙,分别于加力前、加力后第3、7、14 d对大鼠上颌牙槽骨骨小梁进行微型CT扫描,计算骨体积分数、骨表面积体积比、结构模型指数...     

单根牙牙槽骨吸收度与最大（牙合）力关系的研究

目的研究单根牙牙槽骨吸收程度与最大（牙合）力的关系.方法选择37例牙周病患者,710个单根牙,记录其前牙超覆（牙合）情况,拍摄牙片,测量每个受试牙的牙槽骨吸收程度和最大（牙合）力.结果最大牙合力值因个体间和牙位不同存在明显差异.当左右同名牙牙槽骨高度相近时,最大（牙合）力值相近.最大（牙合）力值与牙槽骨吸收程度有着明显的相关性,牙槽骨吸收程度增加,最大（牙合）力降低.牙周病易于导致前牙覆（牙合）深、超（牙合）大的（牙合）型.结论牙槽骨高度影响最大（牙合）力,牙周病将导致牙合的异常及咀嚼功能受到损害.     

Influence of chronic alcoholism and estrogen deficiency on the immunohistochemical expression of regulatory proteins of the bone resorption process in the periodontium of Wistar rats

ObjectiveThe aim of this study was to investigate possible changes in immunohistochemical expression of proteins regulating the bone resorption process in the periodontium of rats subjected to alcoholism and/or estrogen deficiency. The investigated proteins were receptor activator of nuclear factor-kappa β ligand (RANKL), a protein that stimulates bone resorption, and osteoprotegerin (OPG), a protein that inhibits bone resorption. At the molecular level, decreased OPG expression and/or increased RANKL expression are consistent with a greater predisposition to bone resorption.DesignWistar female rats were divided into ovariectomized (ovx) and non-ovariectomized (sham) groups, and subdivided into ad libitum diet (free diet), alcoholic diet (20% solution), and isocaloric diet (diet with a similar amount of calories as compared with groups ingesting an alcoholic diet). The alveolar bone crest and adjacent tissues were evaluated by immunohistochemical analyses for detection of OPG and RANKL.ResultsA significant decrease in OPG expression and a significant increase in RANKL expression were observed in ovariectomized animals which received alcohol as compared with non-ovariectomized animals which received isocaloric diet (experimental control). When estrogen deficiency was evaluated independently of the diet type, a significant decrease in OPG expression and a significant increase in RANKL expression were observed in ovariectomized animals as compared with non-ovariectomized animals.ConclusionsEstrogen deficiency associated with alcoholic diet, as well as estrogen deficiency (analyzed independently of diet type), decreased the immunostaining for OPG and increased the immunostaining for RANKL in the periodontium of rats.     

Effects of occlusal stimuli on alveolar/jaw bone formation

Occlusion is known to influence the growth and development of the craniofacial complex. However, the consequences of occlusal hypofunction, or its recovery, on the amount of formation and development of alveolar bone and the jaw are not fully understood. Therefore, the present study was designed to elucidate the relationship between the occlusal stimuli and alveolar and jaw bone growth by the use of a hypofunction/recovered occlusal function model in growing rats. Bone histomorphometric analyses, including bone apposition rate and mineral apposition rate, were evaluated in double-labeled frontal sections of mandibular second molars. Results showed that occlusal hypofunction significantly suppressed alveolar and jaw bone formation compared with that in animals growing normally (p < 0.05). However, recovered occlusal function induced an enhancement in jaw bone formation. These results indicate the influence of occlusal function on alveolar and jaw bone formation during the growth period.     

A histological and micro-CT investigation in to the effect of NGF and EGF on the periodontal,alveolar bone,root and pulpal healing of replanted molars in a rat model - a pilot study

Background

This study aims to investigate, utilising micro-computed tomography (micro-CT) and histology, whether the topical application of nerve growth factor (NGF) and/or epidermal growth factor (EGF) can enhance periodontal, alveolar bone, root and pulpal tissue regeneration while minimising the risk of pulpal necrosis, root resorption and ankylosis of replanted molars in a rat model.

Methods

Twelve four-week-old male Sprague-Dawley rats were divided into four groups: sham, collagen, EGF and NGF. The maxillary right first molar was elevated and replanted with or without a collagen membrane impregnated with either the growth factors EGF or NGF, or a saline solution. Four weeks after replantation, the animals were sacrificed and the posterior maxilla was assessed using histological and micro-CT analysis. The maxillary left first molar served as the control for the corresponding right first molar.

Results

Micro-CT analysis revealed a tendency for all replanted molars to have reduced root length, root volume, alveolar bone height and inter-radicular alveolar bone volume. It appears that the use of the collagen membrane had a negative effect while no positive effect was noted with the incorporation of EGF or NGF. Histologically, the incorporation of the collagen membrane was found to negatively affect pulpal, root, periodontal and alveolar bone healing with pulpal inflammation and hard tissue formation, extensive root resorption and alveolar bone fragmentation. The incorporation of EGF and NGF did not improve root, periodontal or alveolar bone healing. However, EGF was found to improve pulp vascularisation while NGF-improved pulpal architecture and cell organisation, although not to the level of the control group.

Conclusions

Results indicate a possible benefit on pulpal vascularisation and pulpal cell organisation following the incorporation of EGF and NGF, respectively, into the alveolar socket of replanted molars in the rat model. No potential benefit of EGF and NGF was detected in periodontal or root healing, while the use of a collagen membrane carrier was found to have a negative effect on the healing response.     

The use of live-animal micro-computed tomography to determine the effect of a novel phospholipase A2 inhibitor on alveolar bone loss in an in vivo mouse model of periodontitis

Background and Objective:  Live-animal micro-computed tomography is a new and promising technique that can be used to quantify changes in bone volume for periodontal disease models. The major aim of this study was to develop the methodology of live-animal micro-computed tomography and to determine the effect of a novel secretory phospholipase A₂ inhibitor on alveolar bone loss.
Material and Methods:  Periodontitis was induced in mice by oral infection with Porphyromonas gingivalis over a period of 13 wk, and live-animal micro-computed tomography scans were taken at different time-points to determine bone volume changes with disease progression. This enabled conclusions to be made as to when treatment was most likely to be effective. In addition, the model was used to investigate a novel drug, the secretory phospholipase A₂ inhibitor, KHO64, and its potential ability to inhibit osteoclast bone resorption and treat periodontitis.
Results:  The results from live-animal micro-computed tomography scans revealed greater, statistically significant, bone volume loss in diseased mice compared with normal mice ( p  < 0.05). This corresponded to a larger area from the cemento–enamel junction to the alveolar bone crest, as assessed by stereo imaging ( p  < 0.001). These techniques can therefore detect and quantify alveolar bone loss. Both methods revealed that KHO64 had no significant effect on the volume of bone resorption.
Conclusion:  Live-animal micro-computed tomography is a robust, reproducible technique that clearly demonstrates significant time-dependent changes in alveolar bone volume in a small-animal model of periodontitis.     

Alveolar bone loss induced by the orthodontic tooth movement under hypofunctional conditions in rats

PurposeTo examine the effect of occlusal hypofunctional conditions on orthodontic tooth movement and its relation to the structure and quality of alveolar bone using the rat model.Materials and methodsTwelve-week-old male Sprague-Dawley rats were divided into 4 groups of 8 animals each: normal occlusion (N) group, normal occlusion with tooth movement (M) group, occlusal hypofunction (H) group, and occlusal hypofunction with tooth movement (HM) group. In H and HM groups, the anterior bite plate and metal cap were attached to the maxillary and mandibular incisors using a light-curing composite resin to induce the occlusal hypofunctional condition. In M and HM groups, an orthodontic force was applied in a palatal direction to the buccal surface of the maxillary first molar (M1) using a nickel–titanium alloy wire. Micro-CT imaging and histomorphometric analysis using fluorescent bone labeling of the alveolar bone surrounding the M1s were performed in each group.ResultsTooth movement of M1 in HM group, was rather accelerated with enhanced tipping than in M group. Micro-CT analysis revealed significant decrease in bone volume fraction, bone mineral density and trabecular thickness of the interradicular bone in HM group among the experimental groups. The fluorescent labeling lines in the interradicular bone were decreased in number in H and M groups compared with N group. A few discontinuous irregular dotted lines-like labeling was observed in HM group.ConclusionThe occlusal hypofunctional condition accelerates orthodontic tooth movement of the respective teeth, while it results in severe bone loss in the surrounding alveolar bone.     

Influence of different durations of estrogen deficiency on alveolar bone loss in rats

The purpose of this study was to morphometrically evaluate the influence of different durations of ovariectomy-induced estrogen deficiency on alveolar bone loss associated with ligature-induced bone loss in rats. Sixty female Wistar rats were randomly assigned to ovariectomy (OVX test group) or sham operation (SHAM control group). The OVX and SHAM groups were each distributed into three subgroups of ten rats each according to the duration of estrogen deficiency (30, 60 and 90 postoperative days). In all groups, for the last 30 days of the experimental period, cotton ligatures were placed around the cervix of the right upper second molar; the contralateral tooth was left unligated to serve as a control. The maxillary bones were removed, and the alveolar bone loss was analyzed by measuring the distance between the cementoenamel junction and the alveolar bone crest at the buccal site of the right upper second molar. A comparison between the ligated and unligated groups verified the presence of ligature-induced alveolar bone loss (p < 0.05). No significant differences were observed among the unligated groups (p > 0.05). A significant increase in bone loss was observed when ligation occurred 90 days after ovariectomy compared with the sham group (p < 0.05). These results demonstrate that long-term estrogen deficiency affects ligature-induced alveolar bone loss.     

Diagnostic accuracy of digitized periapical radiographs validated against micro-computed tomography scanning in evaluating orthodontically induced apical root resorption

The aim of this study was to validate the use of digitized periapical radiographs in evaluating orthodontically induced apical root resorption against micro-computed tomography (micro-CT) scanning as a criterion standard test. In a standardized experimental protocol, 29 premolars in 16 subjects were tipped buccally for 8 wk. Nineteen contralateral premolars not subjected to orthodontic movement served as controls. Standardized periapical radiographs were taken before and after the experimental period (Rx method). These teeth were extracted and scanned using a micro-CT technique with a 9 mum resolution. Two calibrated examiners assessed blindly the presence or absence of apical root resorption on digitized radiographs and three-dimensional reconstructions of the scans. Significant differences were detected between the orthodontically moved teeth and controls: 86% of the orthodontically moved teeth and 21% of the control teeth showed apical root resorption when using micro-CT as a validation method. A total of 55% of the experimental teeth and 5% of the control teeth showed resorption when assessed using Rx method. The Rx method showed a specificity of 78% and a sensitivity of 44%, which means that less than half of the cases with root resorption identified using a CT scanner were identified by radiography. Nearly all the orthodontically moved teeth showed apical root resorption. Apical root resorption may be underestimated when evaluated using digitized periapical radiographs.     

Three-dimensional micro-computed tomographic evaluation of periodontal regeneration: a human report of intrabony defects treated with Bio-Oss collagen

This study utilized three-dimensional micro-computed tomography (micro-CT) to evaluate the regenerative response to Bio-Oss Collagen when used alone or in combination with a Bio-Gide bilayer collagen membrane for the treatment of four intrabony defects (5 to 7 mm) around single-rooted teeth. The micro-CT observations are compared to the clinical, radiographic, and histologic results, which have been previously reported. After reflecting a full-thickness flap, thorough degranulation and root planing were accomplished. Bio-Oss Collagen was then used to fill the defects, and in two cases a Bio-Gide membrane was placed over the filled defect. Radiographs, clinical probing depths, and attachment levels were obtained before treatment and immediately preceding en bloc resection of teeth and surrounding tissues 9 months later. A mean pocket depth reduction of 5.75 mm and mean clinical attachment level gain of 5.25 mm were recorded. The histologic evaluation demonstrated the formation of a complete new attachment apparatus with new cementum, periodontal ligament, and alveolar bone at the level of and coronal to the calculus reference notch. Micro-CT evaluation confirmed the histologic results and demonstrated the absence of ankylosis or root resorption for all specimens. This human histologic study demonstrated that Bio-Oss Collagen has the capacity to facilitate regeneration of the periodontal attachment apparatus when placed in intrabony defects. Micro-CT observations confirmed the histologic results and enhanced the three-dimensional understanding of periodontal wound healing. The results indicate that micro-CT may be useful for three-dimensional evaluation of periodontal regenerative procedures.     

尼古丁对实验性牙周炎大鼠牙槽骨影响的显微CT观察

目的 建立牙周炎大鼠的牙槽骨三维模型,采用显微CT观察尼古丁对大鼠牙槽骨的影响.方法 36只SD大鼠,丝线结扎上颌右侧(实验侧)第二磨牙颈部,左侧不予结扎,作为自身对照(对照侧),使用完全随机分组方法分为对照组(A)及尼古丁注射低剂量(B)和高剂量(C)组,每组12只.分别给予生理盐水和尼古丁0.83、1.67 mg·kg-1·d-1腹腔注射.每组分别于给药后第14、28天各处死6只,取双侧上颌磨牙区牙体牙周复合组织,行显微CT扫描、重建、测最和分析.结果 随尼古丁给药剂量增加,双侧牙槽骨骨密度、骨体积分数、骨小梁厚度逐渐降低,牙槽骨高度丧失与骨小梁间隙逐渐升高.28 d时C组牙槽骨高度丧失[对照侧和实验侧分别为(0.61±0.14)、(1.39±0.09)mm]显著高于B组[对照侧和实验侧分别为(0.39±0.10)、(1.31±0.06)mm]和A组[对照侧和实验侧分别为(0.30±0.06)、(0.94±0.07)mm];C组牙槽骨骨密度[对照侧和实验侧分别为(617.86±34.27)、(572.46±31.62)mg/cm3]显著低于B组[对照侧和实验侧分别为(660.04±36.73)、(604.97±32.59)mg/cm3]和A组[对照侧和实验侧分别为(709.15±34.95)、(657.04±30.06)mg/cm3].结论 尼古丁可加重丝线结扎造成的大鼠牙槽骨骨量丧失和骨质微观结构的变化,导致牙槽骨骨质疏松.     

Micro-computed tomography for evaluating alveolar bone resorption induced by hyperocclusion

Purpose

Occlusal trauma, resulting in the destruction of alveolar bone, is a form of periodontal disease caused by excessive mechanical stress (MS) during hyperocclusion. Previously, we showed that CC chemokine ligand (CCL) 2/CCR2 receptor axis plays a crucial role in MS-dependent osteoclastogenesis. However, in the previous work, we were unable to precisely measure changes in alveolar bone profiles. In the present study, we sought to establish a precise method for evaluating alveolar bone resorption induced by hyperocclusion using micro-computed tomography.

Effects of bone marrow transplantation on impacted dental germs in osteopetrotic op/op rats

The osteopetrotic op/op rats suffered from a complete failure of tooth eruption in association with a reduced bone resorption. An unresorbed occlusal bone overlaid the germs which remained embedded in bone. The process of prefunctional eruption in limited growing teeth depends on the resorption of the alveolar bone covering the germs. The aim of this study was to investigate the effects of hematopoietic grafts in newborn op/op rats on the process of prefunctional eruption. Fifteen op/op treated rats were used for radiographic, microradiographic and histologic examinations. After transfusion of bone marrow cells, we observed the restoration of bone resorption and a delayed eruption of the molars. The incisors never erupted. They developed odontome-like tumors near their apices. These results confirm the dependence of tooth eruption on bone resorption.     

High resolution image in bone biology I. Review of the literature

Bone microstructure has usually been assessed by obtaining samples invasively and analyzing them with conventional histomorphometric methods. Improvements in high-resolution image acquisition systems have enabled non-invasive assessment of bone morphology and a more precise 3-D evaluation by means of "virtual biopsies", permitting bone assessment in regeneration or remodeling processes. This review describes the characteristics and limitations of bone assessment using different high-resolution image systems (synchrotron-radiation computed tomography, micro-computed tomography, acoustic scanning microscope; micro-magnetic resonance imaging). Morphometric variables that can be obtained from these images are reported and compared with conventional histomorphometric variables.     

Influence of occlusal hypofunction on the elastic property and bone formation of rat alveolar bone

The purpose of this study was to investigate the influence of occlusal hypofunction on the maintenance of the elastic property and alveolar bone formation using an occlusal hypofunction model. Analyses of the elastic property and bone formation of the alveolar bone were carried out by the compression test and the bone histomorphometry in the two directions (the tooth axis direction and the bucco-lingual direction). The compression test was done with the texturometer to analyse the springiness of bone specimens of the mandibular first molar. Bone histomorphometric analyses were evaluated using double labeled frontal section. Results showed that occlusal hypofunction significantly reduced the springiness and suppressed the alveolar bone formation in the bucco-lingual direction. These results suggested that occlusal function plays an important role in maintenance of the elastic property and bone formation of the alveolar bone.     

Preliminary Clinical Study to Evaluate the Relationship between Systemic Bone Turnover and the Microstructure of the Alveolar Bone

The objective of this study was to assess the possibility of developing a clinical minimally invasive and standardized method to evaluate the relationship between the microstructure of the jaw bone and systemic bone turnover. For this purpose, we performed standardized bone biopsy of the alveolar bone, and compared the 3D bone microstructure using micro-computed tomography (micro-CT) with bone mineral density (BMD) of the lumbar spine and biochemical markers of bone turnover. We evaluated a total of 9 samples taken from 6 patients by standardized biopsy using a trephine bur. BMD was evaluated using dual energy X-ray absorptiometry (DXA). Regarding the biochemical markers of bone turnover, serum bone-specific alkaline phosphatase (BAP) and serum osteocalcin (OC) were used as bone formation markers, and urinary cross-linked N-telopeptides of type I collagen (NTx) and urinary deoxypyridinoline (DPD) were selected as bone resorption markers. We scanned micro-CT images of these samples. Bone volume fraction (BV/TV), trabecular thickness (Tb.Th), trabecular number (Tb.N), trabecular spacing (Tb.Spac), fractal dimension, trabecular bone pattern factor (TBPf) and node-strut (Nd.Nd/TV, TSL/TV) were measured. Regarding the correlations between the parameters of bone microstructures, TB/TV, Tb.N, fractal dimension, and node-strut seemed to be positively correlated and Tb.Spac and TBPf seemed to be negatively correlated with each other, but Tb.Th seemed to have a low correlation with other parameters. OC and/or BAP showed a significantly high correlation with many structural parameters (p < 0.05%). In conclusion, some microstructural parameters may change according to the systemic bone turnover.