Rat Mandible Research Articles

Diffusion patterns of anesthetic solution in the infiltration anesthesia depending on injection sites in the mandible of rats

PurposeThis study was conducted for the morphological and quantitative evaluation of the diffusible patterns of local anesthetic depending on the site of injection in infiltration anesthesia. ExperimentsThe mandibular first molars of rats were anesthetized by the local infiltration of 30 μl of 2% lidocaine containing trypan blue at a distance of 2 mm from the gingival margin on either the lingual or buccal side over the distal root apex of the first molar. Frozen sections were prepared to measure the volume of the infiltrated anesthetic using volume-calculating software. ResultsOn the lingual side, the local anesthetic infiltrated into the mouth floor and spread backward in a sagittal direction at an infiltration volume of 8.36 ± 7.34 mm3. On the buccal side, the anesthetic solution was restricted to the injected area and the infiltration volume was 2.22 ± 1.62 mm3.Conclusion The present results indicate that the infiltration volume of local anesthetics is very different between the lingual and buccal sides.

Characterization of Oxidative Stress Status during Diabetic Bone Healing

Early events associated with bone healing in patients with type 2 diabetes mellitus appear to be delayed. Hyperglycaemia and an associated increase in oxidative stress are cited as potential factors leading to a change in cellular behaviour. Using an in vivo model monitoring bone formation around implants placed into rat mandibles, we have previously identified that the onset of cell proliferation and osteoblast differentiation are delayed and subsequently prolonged compared with normal bone. This study used the same implant model to characterize oxidative stress biomarkers and primary antioxidant enzyme profiles during diabetic bone healing in vivo. Implants were placed into the sockets of incisors extracted from the mandibles of normal Wistar and diabetic Goto-Kakizaki rats for 3 and 9 weeks after implant insertion. Histochemical analysis confirmed a delay in bone healing around implants in diabetic animals. Immunohistochemical localization of peri-cellular staining for protein carbonyl groups, as a biomarker of oxidized protein content, was slightly higher in diabetic granulation tissue compared with normal tissue. However, no differences were observed in the staining patterns of advanced glycation end products. Minimal differences were observed in the number of cells positive for cytoplasmic superoxide dismutase (SOD)1 or mitochondrial SOD2. Significantly, catalase was absent in diabetic tissues. The results suggest that the oxidative environment in healing bone is differentially affected by hyperglycaemia, particularly in relation to catalase. The significance of these observations for diabetic bone healing is discussed.

Development of Distraction Osteogenesis Model of the Inferior Mandibular Border and Alveolar Bone

The aims of this study were to establish an experimental rat model for distraction osteogenesis of the mandible and alveolar bone for the investigation of histologic and radiographic analysis and to develop the device for various distraction models. For osteotomy location and distraction device design, micro-computed tomographic analysis of the structure of rat mandible was performed. Twenty mandibles of Sprawl-Dawley rats were osteotomized based on the analysis, and newly developed custom-made devices were applied with bone screws. After a 5-day latency period, a distraction of 0.2 mm at a single session, and a 4-week consolidation period, animals were killed for histologic samples. The rat distraction osteogenesis model for the inferior mandibular border and alveolar bone was successfully established based on the protocols used in this study. New bone formation occurred at both body and alveolar bones of the mandible by the combination of 2 modes: intramembranous ossification was predominant at the end of distraction period, whereas endochondral ossification was observed at the center of the distraction gap at the end of the 4-week consolidation period. This model can be applied to various conditions related to distraction osteogenesis and provide information for the development of improvements in clinical treatments using distraction osteogenesis.

Effects of risedronate on cortical and trabecular bone of the mandible in glucocorticoid-treated growing rats

This study was performed to estimate the effects of risedronate on mandibular bone density, bone structure, and bone metabolism in established glucocorticoid-induced osteoporosis in growing rats. The rats were given oral risedronate at 0, 0.5, or 1.0 mg per kilogram per day for 4 weeks after the administration of oral prednisolone at 30 mg per kilogram per 2 days for 6 weeks. Trabecular and cortical bone masses were analyzed by using peripheral quantitative computed tomography, and bone structure and bone formation were measured by using static and dynamic histomorphometry. In trabecular bone, risedronate improved the prednisolone-induced decreases in bone cross-sectional area and bone mineral content. Risedronate increased bone density and also formed dense bone microarchitecture by reducing the bone turnover rate. In cortical bone, risedronate improved the prednisolone-induced decreases in bone cross-sectional area and bone mineral content without affecting bone density by increasing the mineralizing surface. Risedronate improved prednisolone-induced retardation of trabecular and cortical bone growth, but the bone turnover in these 2 sites was regulated differently in the growing rat mandibles.

Applicability of equine hydroxyapatite collagen (eHAC) bone blocks for lateral augmentation of the alveolar crest. A histological and histomorphometric analysis in rats

This study assessed the mechanical characteristics, biocompatibility and osteoconductive properties of an equine hydroxyapatite collagen (eHAC) bone block when applied as a bone substitute for lateral augmentation of rat mandible. 96 rats underwent lateral augmentation of the mandible, using two substitute bone blocks (eHAC or Bio-Oss spongiosa) or autologous bone grafts. Signs of inflammation, amount of bone formation and ingrowth of bone into the bone blocks were assessed at 1 and 3 months. eHAC blocks were mechanically rigid and could be fixed firmly and easily. Bio-Oss spongiosa blocks were brittle and fixation was difficult. eHAC and Bio-Oss spongiosa blocks were biocompatible and induced few or no signs of inflammation. Inflammation prevalence between the groups was not statistically different. Bone formation and bone growth into the blocks was significantly higher in eHAC than Bio-Oss spongiosa blocks, but lower than in autologous bone grafts (after 1 and 3 months). Regression analysis showed that the autologous bone graft predicted new bone formation at both time points. The eHAC block was only a predictor at 1 month; a trend was found at 3 months. The application of biodegradable membranes was not related to more bone ingrowth.

Early alveolar bone regeneration in rats after topical administration of simvastatin

The aim of this study was to ultrastructurally examine the influence of simvastatin on bone healing in surgically created defects in rat mandibles. Bone defects 0.8 mm in diameter were created in the buccal aspect of first mandibular molar roots and filled with 2.5% simvastatin gel, while the controls were allowed to heal spontaneously. The rats were humanely killed 7, 9, 11, or 14 days postoperatively, and the specimens were processed for scanning and transmission electron microscopy, as well as for colloidal gold immunolabeling of osteopontin. The regenerated alveolar bone in the simvastatin-treated defects presented smaller marrow spaces, and the collagen fibrils were regularly packed exhibiting a lamellar bone aspect. Osteopontin was present through the bone matrix during the wound healing and alveolar bone regeneration. The present study provides evidence that a single topical application of 2.5% simvastatin gel improves the quality of the new bone and decreases bone resorption.

Electroporation to deliver plasmid DNA into rat dental tissues

Delivery of DNA into the target tissues is an important technique in gene function studies and gene therapy. Surgical treatment of tooth eruption disorders, such as impacted third molars, is a major healthcare cost. Because the dental follicle (DF) is essential for regulating tooth eruption, establishment of local gene transfer protocols is needed to determine the effect of various genes on eruption and to develop gene therapy approaches for inducing the eruption of impacted molars. Plasmids containing lacZ reporter gene were injected into rat mandibles and then electroporated at the designated settings. Mandibles were collected 24 h after electroporation for X-gal staining to evaluate the transfection efficiency. Tissues were collected at various days post-electroporation to determine the expression of the transgene. For the DF, depth of injection and pulse number appear to be important. Six pulses can achieve above 80% transfection of the DF at 50 V or 120 V. For alveolar bone (AB) transfection, voltages are important, with 120 V being optimal. Regarding pulse durations, we determined that durations of 20 and 30 ms achieve the maximum transfection in AB and DF, respectively. The present study demonstrates for the first time the feasibility of electroporation to locally deliver plasmids into dental tissues. Parameters affecting electroporation to deliver plasmids into the dental tissues were optimized. This protocol could be used to deliver short hairpin RNA or genes of interest into the dental tissues to regulate tooth eruption. Thus, it may be possible to develop nonsurgical treatments for inducing the eruption of impacted teeth.

Delayed osteoblast differentiation and altered inflammatory response around implants placed in incisor sockets of type 2 diabetic rats

Central to the process of osseointegration is the recruitment of mesenchymal progenitor cells to the healing site, their proliferation and differentiation to bone synthesising osteoblasts. The process is under the control of pro-inflammatory cytokines and growth factors. The aim of this study was to monitor these key stages of osseointegration and the signalling milieu during bone healing around implants placed in healthy and diabetic bone. Implants were placed into the sockets of incisors extracted from the mandibles of normal Wistar and diabetic Goto-Kakizaki rats. Mandibles 1-12 weeks post-insertion of the implant were examined by histochemistry and immunocytochemistry to localise the presence of Stro-1- positive mesenchymal progenitor cells, proliferating cellular nuclear antigen proliferative cells, osteopontin and osteocalcin, macrophages, pro-inflammatory cytokines interleukin (IL)-1β, IL-6, tumour necrosis factor (TNF)-α and tumour growth factor (TGF)-β1. Image analysis provided a semi-quantification of positively expressing cells. Histological staining identified a delay in the formation of mineralised bone around implants placed in diabetic animals. Within the diabetic bone, the migration of Stro-1 mesenchymal cells in the healing tissue appeared to be unaffected. However, in the diabetic healing bone, the onset of cell proliferation and osteoblast differentiation were delayed and subsequently prolonged compared with normal bone. Similar patterns of change were observed in diabetic bone for the presence of IL-1β, TNF-α, macrophages and TGF-β1. The observed alterations in the extracellular presence of pro-inflammatory cytokines, macrophages and growth factors within diabetic tissues that correlate to changes in the signalling milieu, may affect the proliferation and differentiation of mesenchymal progenitor cells in the osseointegration process.

Biocompatibility of Accelerated Mineral Trioxide Aggregate in a Rat Model

Introduction The purpose of this study was to evaluate the biocompatibility of gray mineral trioxide aggregate (MTA) mixed with selected accelerants by examining the inflammatory response through histological analysis after implantation into rat mandibles. Methods Sixty rats were randomly divided into experimental groups of calcium chloride (CaCl 2), calcium nitrite/nitrate (CaN/N), or calcium formate (CaF) mixed with MTA and compared with controls of MTA mixed with sterile water (MTA control) and unfilled osteotomies. Material was implanted into an osteotomy prepared between the roots of the mandibular molars and the incisors in the mandible. After 2 or 8 weeks, tissues were collected and processed for hematoxylin and eosin staining and light microscopic evaluation. Blinded evaluators graded the inflammatory response along the MTA-bone interface on an ordinal scale. Data were analyzed using the Kruskal-Wallis test, and interobserver agreement was determined by the kappa analysis. Results Interobserver agreement was good with κ = 0.72. No statistically significant differences were noted between experimental and control groups at the 2-week time point ( p > 0.05). At this time, all groups displayed a range of inflammatory responses from none to severe with mostly mild to moderate reactions. At the 8-week time point, the inflammatory reaction of CaF mixed with MTA was statistically different from the controls ( p = 0.03). CaCl 2 and CaN/N were not statistically different from the controls, and the MTA control displayed no inflammation at this time point. Conclusions The findings of this study indicate that MTA mixed with accelerants may be a biocompatible alternative when a rapid set is indicated clinically.

Osteogenic Potential of Mandibular vs. Long-bone Marrow Stromal Cells

Although fundamentally similar to other bones, the jaws demonstrate discrete responses to developmental, mechanical, and homeostatic regulatory signals. Here, we hypothesized that rat mandible vs. long-bone marrow-derived cells possess different osteogenic potential. We established a protocol for rat mandible and long-bone marrow stromal cell (BMSC) isolation and culture. Mandible BMSC cultures formed more colonies, suggesting an increased CFU-F population. Both mandible and long-bone BMSCs differentiated into osteoblasts. However, mandible BMSCs demonstrated augmented alkaline phosphatase activity, mineralization, and osteoblast gene expression. Importantly, upon implantation into nude mice, mandible BMSCs formed 70% larger bone nodules containing three-fold more mineralized bone compared with long-bone BMSCs. Analysis of these data demonstrates an increased osteogenic potential and augmented capacity of mandible BMSCs to induce bone formation in vitro and in vivo. Our findings support differences in the mechanisms underlying mandible homeostasis and the pathophysiology of diseases unique to the jaws.

片側側頭筋切除がラット下顎骨に及ぼす影響 第1報 : 形態学的検討

片側側頭筋切除がラット下顎骨に及ぼす影響 第1報 : 形態学的検討

Rehabilitation of masticatory function improves the alveolar bone architecture of the mandible in adult rats

Masticatory functional changes have been shown to influence the quantity and quality of the alveolar bone during growth. This study was designed to investigate the effect of masticatory function rehabilitation on the morphology and the trabecular architecture of the mandibular alveolar bone after cessation of growth. Forty-four Sprague–Dawley male rats received soft diet in order to develop masticatory muscle hypofunction. After 21 weeks, after cessation of growth, the animals were divided into two groups: the first group continued receiving soft diet for six more weeks ( hypofunction group), while the second group changed to ordinary (hard) diet with the aim to restore a normal masticatory function ( rehabilitation group). A third group of 16 male rats ( normal group) received ordinary (hard) diet during the whole experimental period and served as control. Micro-tomographic histomorphometry was used to evaluate the architecture of the mandibular alveolar bone (e.g. bone volume fraction, trabecular thickness, trabecular separation, etc.) at the end of the experiment (27 weeks). The height and width of the alveolar process were measured as well. The alveolar process trabecular bone volume fraction (BV/TV) was lower for the animals of the hypofunctional group as compared to those of the normal ( p < 0.01) and the rehabilitation ( p < 0.05) groups. Despite the significant improvement observed in the rehabilitation group, their BV/TV was lower in comparison to the normal group ( p < 0.05) at the end of this experiment. All the other micro-tomographic parameters followed the same pattern of change between groups; values of the rehabilitation group were between the values of the two other groups, differing significantly from both of them. The alveolar process was significantly shorter in the normal group in comparison to both the hypofunctional and rehabilitation groups ( p < 0.05). On the other hand, both the normal and rehabilitation groups were had a wider alveolar process than the hypofunctional group ( p < 0.05). Both alveolar height and width were significantly correlated with all micro-tomographic parameters under study. The rehabilitation of masticatory function led to a significant improvement of alveolar bone architecture in adult rats, although the negative effects of hypofunction were not totally reversed during the period under study.

Study on neo-natal rat mandible in organ culture (Part1)

Study on neo-natal rat mandible in organ culture (Part1)

Elastic system fibers in rat incisor periodontal ligament – immunohistochemical study using sections of fresh-frozen un-demineralized tissues

The aim of this study was to examine the localization and distribution of the components of elastic system fibers in the periodontal ligament of continuously erupting rat incisors in an effort to understand the mechanism of the eruption of the tooth. Sections of fresh-frozen, un-demineralized incisors of the rat mandible were prepared for immunohistochemical localization of elastin, fibrillin-2 and microfibril-associated glycoprotein-1 (MAGP-1). The structure of the periodontal ligament was well preserved in sections of fresh-frozen tissues. At the basal region of the ligament, intense immunolabelling for fibrillin-2 and MAGP-1 was observed as dot-like structures (transversely sectioned fibers) mainly on the tooth side of the ligament close to the cementum. These dot-like structures gradually increased in number towards the incisal area and were distributed throughout the tooth side of the ligament. This pattern of distribution was the same as that of reported oxytalan fibers. Elastin-immunopositive fibers were also detected in the ligament, although the labelling was limited and distribution was sparse. In conclusion, both fibrillin-2 and MAGP-1 immunopositive fibers may serve as a scaffold for deposition of tropoelastin during elastogenesis in the periodontal ligament. They may also provide guidance for the migration of fibroblasts to the occlusive side, which generates contractile forces for the movement of the tooth for continuous eruption of incisors.

Distribution of Small Integrin-Binding LIgand, N-linked Glycoproteins (SIBLING) in the condylar cartilage of rat mandible

The Small Integrin-Binding LIgand, N-linked Glycoprotein (SIBLING) family is one category of non-collagenous proteins closely related to osteogenesis. In this study, the authors systematically evaluated the presence and distribution of four SIBLING family members, dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP1), bone sialoprotein (BSP) and osteopontin (OPN), in rat mandibular condylar cartilage using protein chemistry and immunohistochemistry. For protein chemistry, SIBLING proteins in the dissected condylar cartilage were extracted with 4M guanidium-HCl, separated by ion-exchange chromatography, and analyzed by Western immunoblotting. Immunohistochemistry was employed to assess the distribution of these four SIBLING proteins in the condylar cartilage of 2-, 5- and 8-week-old rats. Results from both approaches showed that all four members are expressed in the condylar cartilage. DSPP, unlike that observed in dentin and bone, exists as a full-length form (uncleaved) in the condylar cartilage. The NH(2)-terminal fragment of DMP1 is mainly detected in the matrix of the cartilage while the COOH-terminal fragment is primarily localized in the nuclei of cells in the chondroblastic and hypertrophic layers. The data obtained in this investigation provide clues about the potential roles of these SIBLING proteins in chondrogenesis.

ラット下顎骨延長モデルにおけるSmadsの動態

ラット下顎骨延長モデルにおけるSmadsの動態

Experimental Results of the Fibrin Clot Use to Accelerate the Regeneration of Damaged Bone in the Rat Lower Jaw

Morphological and radiological methods were used to study regeneration of the damaged bone of rat mandibles after application of plateletenriched fibrin clot. A bone hole was artificially created, and in the natural course of regeneration, the hole was immediately filled with blood and there a blood clot formed. After one week of healing, separate islands of young bone tissue appeared. After two to three weeks, the opening in the mandible was completely replaced by the young bone tissue. When a similar bone hole was filled with autological fibrin clot, the blood clot did not form. But after one week the entire hole was filed with newlyformed fused bone tissue. By the second week after the use of fibrin clot, the bone hole had further healed and bone callus was formed.

Expression of nitric oxide synthases in the mandibular condyle of anterior repositioned rat mandibles

Expression of nitric oxide synthases in the mandibular condyle of anterior repositioned rat mandibles

Validation of histologic changes induced by external irradiation in mandibular bone. An experimental animal model

The present experimental study sought to determine the effect of high-dose irradiation on the rat mandible in order to establish an experimental model of radiogenic bone damage. The left mandibles of 20 adult Wistar rats were irradiated (single fraction 1500cGy, total dose 60Gy) by means of a hypofractionated stereotactic radiotherapy (hfSRT) over a period of 6 weeks. Follow-up was 6 weeks (group 1, n=10) and 12 weeks (group 2, n=10). The contralateral mandibles as well as 5 non-irradiated animals served as controls. Primary endpoints were fibrosis, loss of cell count, decreased immunohistochemical labelling for bone morphogenetic protein-2 (BMP-2) and osteocalcin as well as increased expression of transforming growth factor (TGF-beta). Cell loss, progressive fibrosis, and focal necrosis were detected in all irradiated sites. Quantitative measurement revealed 32.0+/-8.7% and 37.3+/-9.5% empty osteocyte lacunae for groups 1 and 2 resp., compared to 16.3+/-4.7% and 18.9+/-4.9% on the contralateral side and 7.9+/-1.7% for unirradiated controls (Mann-Whitney U test; p<.01). BMP-2 and osteocalcin labelling showed a marked decrease in irradiated and contralateral sides while TGF-beta was expressed strongly in irradiated sites only (for all p<.05). External hypofractionated irradiation with a total dose of 60Gy is feasible in rats and yields all histologic changes attributed to osteoradionecrosis (ORN) after a follow-up of 6 weeks. The irradiation protocol is suitable for an assessment of regenerative options in severe radiogenic bone damage. As a split mouth design entails major inaccuracies healthy animals have to be used as controls.

Effect of recombinant human bone morphogenetic protein‐2 on bone formation in the acute distraction osteogenesis of rat mandibles

Distraction osteogenesis (DO) is a method of producing new bone directly from the osteotomy site by gradual traction of the divided bone fragments. The purpose of the present study was to evaluate histomorphometrically whether acute DO would constitute a viable alternative to the conventional continuous distraction treatment and also to verify the capacity of a recombinant human BMP (rhBMP-2) associated with monoolein gel to stimulate bone formation in the acute distraction process. Forty-eight Wistar rats were assigned to three groups: Group 1, treated at a conventional continuous distraction rate (0.5 mm/day), Group 2, treated with acute distraction of 2.5 mm at the time of the surgical procedure, and Group 3, subjected to acute distraction associated with rhBMP-2. The animals from each experimental group were killed at the end of the second or fourth post-operative weeks and the volume fraction of newly formed bone trabeculae was estimated in histological images by a differential point-counting method. The results showed that after 2 and 4 weeks, bone volumes in the rhBMP-2 group were significantly higher than in the other groups (P<0.05), but no significant difference was observed in the volume fraction of newly formed bone between the continuous and acute DO groups. In conclusion, the study indicates that rhBMP-2 can enhance the bone formation at acute DO, which may potentially reduce the treatment period and complications related to the distraction procedure.