Search

The objective of this study was to identify the differences between osteoblasts derived from normal adult rat mandibles and osteoporotic adult rats. An osteoporotic animal model was established by performing a bilateral ovariectomy (ovx group). The proliferation and differentiation abilities of osteoblasts were determined by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-2 H-tetrazolium bromide), alkaline phosphatase (ALP) and osteocalcin release (OC) assays. Transmission electron microscopy (TEM) was performed to assess differences in the ultrastructure. Proliferating cell nuclear antigen (PCNA) and uncoupling protein 2 (UCP2) protein concentrations were analyzed by Western blot. In addition, UCP2 protein in osteoblasts was assessed by immunohistochemistry staining . ATP and reactive oxygen species (ROS) concentrations were analyzed separately with ATP and ROS quantification kits. At four and 12 weeks after the operation, osteoblasts of the ovx group showed earlier attachment, fewer dead cells and faster growth compared with cells in the sham group. TEM showed that osteoblasts of the ovx group had fewer folds, lysosomes, peroxisomes and less rough endoplasmic reticulum. The results of the MTT, ALP activity and OC assays were all higher in osteoblasts from the ovx group at four or 12 weeks postsurgery than osteoblasts from the sham group. PCNA protein concentrations in the ovx group increased significantly compared with those of the sham group at four or 12 weeks after the operation, but UCP2 concentrations decreased over the same time period. UCP2 immunohistochemical staining of osteoblasts showed that the protein was concentrated in the cytoplasm and that the osteoblasts from the sham group had higher expression than those from the ovx group. The ATP and ROS concentrations of the ovx groups were significantly higher than the sham groups at four or 12 weeks postsurgery. Therefore, we concluded that there are differences in cell ultrastructure, proliferation, differentiation, ATP and ROS concentrations, and PCNA and UCP2 protein expression levels in osteoblasts from the mandibles of rats of the ovx group compared with those from the sham group.

Glimepiride, a third-generation sulfonylurea, has also been reported to have extrapancreatic functions including activation of PI3-kinase (PI3K) and Akt in rat adipocytes, skeletal muscle and endothelial cells. It is tempting to speculate that glimepiride would improve bone-implant contact in diabetic patients by mediating the activity of GLUT1 and 3 via the PI3K/Akt pathway. In this study, we investigated the effects of glimepiride on rat mandible osteoblasts cultured under two different levels of glucose. Cell proliferation was determined by the MTT assay. The supernatant was used to measure alkaline phosphatase (ALP) activity. Glucose uptake was determined by measuring the rate of 2-deoxy-d-glucose (2-DG) uptake. Western blotting was performed used to determine collagen I and PI3K/Akt expression. RT-PCR was performed used to determine osteocalcin (OCN) mRNA expression. We found that hyperglycemia down-regulated proliferation, ALP activity, OCN mRNA and GLUT3 protein expression in rat osteoblasts, and upregulated collagen I and GLUT1 protein expressions. Glimepiride enhanced the proliferation, ALP activity and OCN mRNA levels, and upregulated collagen I and GLUT1 and 3 protein expressions of rat osteoblasts at two different glucose concentrations. This study also provides the first evidence that glimepiride stimulates the phosphorylation of PI3K/Akt in osteoblasts and ameliorated the damage caused by high concentrations of glucose through the PI3K/Akt pathway.

To explore the effect of high glucose on proliferation of bone marrow stromal stem cells through Wnt/Β-catenin pathway.Bone marrow stormal cells were obtained from the mandible of Wistar rats and stimulated with different concentrations of glucose (5.5 and 16.5 mmol/L). Cell proliferation was evaluated with methyl thiazolyl tetrazolium assay (1, 3, 5, and 7 d)and cell cycle analysis by flow cytometry (5 d). Β-catenin and cyclin D1 protein levels were determined by Western blot. The mRNA expression of lymphoid enhancer binding factor-1 (LEF-1) and cyclin D1 were tested by real-time polymerase chain reaction.The results of methyl thiazolyl tetrazolium assay indicated that the optical density values of two different concentrations of the glucose had no statistical difference on day 1 (P=0.700). On days 3, 5, and 7, the optical density values of the 16.5 mmol/L group were significantly lower than those in the 5.5 mmol/L group (P=0.006, P=0.002, and P=0.003). Cell cycle analysis indicated that high glucose concentration could reduced the progression from phase G1 to S, and the proliferation index values of the 16.5 mmol/L group were significantly lower than those of the 5.5 mmol/L group (P=0.014). The Β-catenin and cyclin D1 levels were lower in the 16.5 mmol/L group when compared with the 5.5 mmol/L group. High glucose condition also reduced the mRNA expressions of LEF-1 and cyclin D1.High glucose can inhibit the proliferation of bone marrow stormal cells by suppressing the expressions of Β-catenin, LEF-1, and cyclin D1 in the Wnt/Β-catenin pathway.

OBJECTIVE: To study the effect of peri-implantitis inflammatory microenvironment on the biological function of jaw bone osteoblasts. METHODS: Primary mandible osteoblasts from peri-implantitis and normal tissue were isolated and cultured. Third-generation purified osteoblasts were identified and detected. The proliferative activity of osteoblasts was evaluated through MTT assay. Osteocalcin (OCN), Runx2, and collagen Ⅰ (Col Ⅰ) mRNA levels were examined by real-time quantitative polymerase chain reaction. OCN protein levels were determined by Western blot. RESULTS: After 4 d of culture, the proliferative activity of osteoblasts from peri-implantitis became lower than that of normal tissue (P

To evaluate the effects of hyperglycemia and glimepiride on proliferation, differentiation and mineralization of rat mandibular osteoblasts to verify the hypothesis of dental implant administration.Primary osteoblasts were isolated and cultured. Then the cells were placed in an osteogenic medium, containing 2 different concentrations of glucose (5.5 mmol/L and 16.5 mmol/L), with or without glimepiride (10 μmol/L). Cell proliferation was evaluated through MTT assay. Alkaline phosphatase (ALP) activity was determined by biochemistric method. Col I protein levels were determined by Western blot. OCN mRNA levels were tested by RT-PCR. SPSS 13.0 software package was used for statistical analysis.Hyperglycemic conditions interfered with the proliferation, ALP activity and OCN mRNA expression of rat osteoblasts, but improved the expression of Col I on day 14. Glimepiride stimulated rat osteoblast proliferation, ALP activity and OCN mRNA expression. The addition of glimepiride to normoglycemic (5.5 mmol/L) cultures registered a significant increase of Col I expression at 7 d and 14 d. Glimepiride significantly increased Col I expression in cells cultured with 16.5 mmol/L glucose for 7 days, but failed to increase at 14 d.Hyperglycemic conditions interfered with the proliferation, differentiation and mineralization of osteoblasts in rats; however, glimepiride improved the proliferation, differentiation and mineralization of osteoblasts in rats.

To investigate the differentially expressed proteins in mandible between normal rats, and type 2 diabetic rats.Wistar rats and Goto-Kakizaki (GK) rats were selected as normal controls and experimental rats respectively. Weight and blood glucose were measured before sacrificed. The mandibles of each group were removed at the same time. After protein extraction, differentially expressed proteins were separated by two-dimensional gel electrophoresis (2-DE) and identified by MALDI-TOF/TOF. Statistical analysis was performed using SPSS 15.0 software package.Blood glucose levels of the experimental group were significantly higher than that of the control group(P0.05). The weights of the two groups were not significantly different. 20 proteins were differentially expressed at least 3-fold between GK rats and Wistar rats. Of the 20 proteins, 5 proteins were more than 20-fold different between the two groups. The 5 proteins fell into 3 functional categories as follows: metabolism, binding proteins, and signal transduction.The differentially expressed proteins of mandible between GK rats and Wistar rats may be new targets for investigating the mechanisms of the effect of type 2 diabetes on mandible.

To explore the effect of insulin on the expression of prohibitin in osteogenic differentiation of rat's mandibular bone marrow stromal cells(BMSC) in high glucose.Bone marrow stromal cells were obtained from the mandible of Wistar rats and stimulated in three glucose concentrations mineral medium(5.5, 25, 45 mM) with or without insulin(10-5 M) for 1, 3, 7,14 and 21 days. The expression of prohibitin was quantified via enzyme-linked immunosorbent assays (ELISA). The mineralization nodules assessment was performed at day 21 by alizarine red staining. The statistics analysis was performed using SPSS 15.0 software package.High glucose of 45 mM inhibited mineralization of rBMSC and insulin could improve the mineralization in high glucose. The expression of prohibitin of 45 mM group decreased significantly compared with 5.5 mM group and 25 mM group. The expression of prohibitin of each group achieved maximum at day 3. Insulin (10-5 M) increased the expression of prohibitin of 25 mM group and 45 mM group in osteogenic differentiation of rBMSC.High glucose inhibited the expression of prohibitin in osteogenic differentiation of rBMSC, and insulin can improve the effect. Prohibitin may play an important part in early stage in osteogenic differentiation of rBMSC.

The purpose of this study was to investigate the cortical bone thickness of the inter-dental area of both jaws for orthodontic miniscrew placement. The cone-beam computerized tomography images of 32 non-orthodontic adults with normal occlusion were taken to measure the cortical bone thickness in both jaws. One-way analysis of variance (ANOVA) was used to analyze the differences in cortical bone thickness. Buccal cortical bone in the mandible was thicker than that in the maxilla. In the maxilla, cortical bone thickness was thicker in the buccal side than in the palatal side. Buccal cortical bone thickness in the mandible was thickest at the site distal to the first molar, and in the maxilla it was thickest at the site mesial to the first molar, while in the palatal side of maxilla it was thickest at the site mesial to the second premolar. The changing pattern of cortical bone thickness varies at different sites. In the buccal side of maxilla, the thinnest cortical bone thickness was found to be at 4 mm level from the alveolar crest, while the thickest was at 10 mm level (except for the site mesial to the first premolar). The buccal cortical bone thickness at the sites mesial or distal to the first molar in the mandible and palatal cortical bone thickness of maxilla tended to increase with increasing distance from the alveolar bone.

To observe the intake of minocycline and its amount in mature rat mandibular osteoblasts (MRMOB) in vitro, and to identify the feasibility of intracellular anti-bacterial activity of minocycline.Four groups of MRMOB were incubated in 100 mg/L minocycline for 15, 30, 45 and 60 minutes respectively, and the accumulation of minocycline within MRMOB was measured using a fluorescence spectrophotometer.The intracellular accumulation amount of minocycline in the four groups of MRMOB was (17.29 +/- 1.49), (16.87 +/- 1.57), (16.96 +/- 1.67) and (17.94 +/- 1.63) mg/g respectively after osteoblasts were cultured in Dulbecco's modified Eagle's medium (DMEM) which contained minocycline for 15, 30, 45 and 60 min. There was no significant difference in amount of minocycline among the four groups of MRMOB.The mature rat mandibular osteoblasts can ingest minocycline, and the accumulation amount of minocycline in MRMOB is irrelevant with the exposure time of MRMOB to minocycline.

To investigate an efficient method of primary mandibular osteoblasts (MO) culture.The mandible harvested from 24 h SD rats was stripped off all soft tissues including the periosteum, rinsed and cut to trivial bone block under sterile condition. Then the bone block was subcultured in culture flask after digested with modified repeating enzymatic digestion-adherent explants method. The proliferation of the acquired cells were examined with assay of methyl thiazolyl tetrazolium (MTT) and identified with invert microscope, immunohistochemistry stain of osteocalcin, alkaline phosphorase (ALP) stain and stain of calcified nodules.MTT assay showed that the cells grew slowly in 1-3 days of post-inoculation, it was cell growth adaptation period. At seventh day, the cells growth reached highest, then the proliferation of the cells was slow gradually. The cells were identified to be osteoblasts by invert microscope, immunohistochemistry stain of osteocalcin, ALP stain and alizarin bordeaux stain of calcified nodule.The modified repeating enzymatic digestion-adherent explants method is an ideal method to obtain and culture neonatal rat's MO having typical characteristics.

To evaluate the quality of the alveolar ridge augmented by Ti-Ni shape memory alloy (Ti-Ni SMA) distractor.All of the mandibular premolars and the first molars were extracted from 12 adult mongrel canines. One month later, box-shaped osteotomy was carried out in the left mandibles and Ti-Ni SMA distractors were imbedded. The right sides served as blank control. The canines were killed 3 or 6 months later and specimens were taken from corresponding area of the experiment sides and the control. Bone mineral density and biomechanical function were measured.Bone mineral density, ultimate compressive strength and Young's Modules of the augmented ridge were lower than the blank control three months after distraction. But the indexes increased and showed no significant difference with the control six months after distraction.The canines mandibular posterior alveolar ridge could be augmented by distraction osteogenesis using Ti-Ni SMA distractor, and the new bone is strong enough for implant placement.

To investigate the feasibility of using natural poritos as scaffolds in bone tissue engineering (TE) and repair of caprine mandibular segmental defect with titanium reticulum reinforced.Natural poritos with a pore of 190-230 microm in size and porosity of about 50percent-65percent was molded into the shape of granules 5 mm x 5 mm x 5 mm in size. Expanded autologous caprine marrow mesenchymal stem cells were induced by recombinant human morphogenetic protein-2 (rhBMP2) to improve osteoblastic phenotype. Then marrow derived osteoblasts were seeded into poritos in density of 4 x 10(7)/ml and incubated in vitro for 48 hours prior to implantation. Then osteoblastic cells/poritos complexes were implanted into mandibular defect and the defect was reinforced by titanium reticulum. Implantation of poritos alone acted as the control. Bone regeneration was assessed 4, 8, 16 weeks after implantation using roentgenographic analysis and histological observation was done after 16 weeks.New bone could be observed histologically on the surface and in the pores of natural coral in all specimens in the cell-seeding group, whereas in the control group there was no evidence of osteogenesis process in the center of the construction. The results showed that new bone grafts were successfully restored 16 weeks after implantation.This study suggests the feasibility of using porous coral as scaffold material transplanted with marrow derived osteoblasts by TE method. By means of titanium reticulum reinforcement, mandibular defect could be successfully restored. It shows the potentiality of using this method for the reconstruction of bone defect in clinic.

To accelerate distraction osteogenesis by utilizing guided bone regeneration technique, and to improve the automatic process of canine segmental mandibular reconstruction by elasticity distraction osteogenesis using shape memory metal alloy distractor.Adult hybrid canines were used. Osteotomy was performed to remove a bone segment of 2.5-4.0 cm in one side of the mandible. Mandibular fixation devices and shape memory metal alloy distractor were secured according to the principles of bi-focal distraction technique. A piece of ePTFE membrane was sutured to cover the buccal side of bone defect and osteotomy gap of transport disc, where the periosteum was peeled for exposing mandible in operation. Then the incisions were totally closed. The canines were sacrificed 3 months later to harvest the mandibles for morphological observation and measurement of bone density and intensity.The shape memory metal alloy distractor performed distraction osteogenesis automatically and reconstructed the mandibles with bone defect. A regenerated bone segment of 1.5-2.0 cm was formed in the compression region between transport disc and contralateral mandibular end. The new bone in both of distraction and compression regions had similar height and thickness with normal mandible. Moreover the new bone density and intensity were rather satisfied.Membrane guided bone regeneration can avoid bone synthesis commonly caused by cicatrization of soft tissue in the bone defect, and accelerate ossification and maturation of new generated bone.

To improve the design of the devices used for autoatic elasticity distraction osteogenesis, and to preliminarily testify their feasibility for clinical application.The internal automatic distraction devices were constructed by three parts, including sinusoid shape memory alloy distractor, reconstructive plate and mini-plate. The animal model of trifocal bone transport reconstruction of canine mandibular mentis was duplicated, in which a 6 cm mandibular mentis segment was removed and the transport disc was created by osteotomy in both ends of the mandibular defect. The plates were used for internal fixation and shape reconstruction of the mandible. They also served as transport distraction guidance for the elasticity distractor.The new devices could complete bone transport as expected. Regenerated bone segments were formed in two months and both sides of the mandibular body were lengthened. Because of the narrow angle and obstruction from the thick soft tissues in the mentis area, the ultimate symphysis mentalis was not reconstructed.The design of the internal elasticity distraction devices showed potential feasibility for clinical practice. But further studies are still needed for better control of the strength and balance of distraction forces.

To discuss the effects of titanium nickel shape memory alloy (TiNi-SMA) distractor with different stretching force and different distance between two fixed ends on the height of alveolar ridge augmentation.The mini distractors were all made of TiNi-SMA wire. The maximum stretching force and the distance between two fixed ends according to different distractors (A, B and C) were 20 mm, 700 g; 15 mm, 1000 g; 20 mm, 1000 g. 15 adult mongrel canines were divided into three groups randomly and extracted the mandibular premolars and first molar. One month later, box-shaped osteotomies of the alveolar bone were carried out in the left mandible and the TiNi-SMA distractors were buried. The bone height was measured and the radiographs were taken before and five weeks post operation continuously.One week post operation, the vertical ridge augmentation of group A, B and C averaged (7.55 +/- 1.02) mm, (4.97 +/- 0.34) mm, (8.54 +/- 0.82) mm and the height remained (7.20 +/- 1.05) mm, (4.93 +/- 0.21) mm, (8.04 +/- 0.78) mm five weeks post operation. There was no statistic difference in the height of alveolar ridge augmentation between Group A and C, but Group B showed significant difference with the other two groups. In each group, the height did not changed significantly with the time. The radiographs showed obvious distraction gap post operation, and the new bone regenerated five weeks later.It suggests that in the scope of suitable stretching force, the height of alveolar ridge augmentation was determined by the size of TiNi-SMA distractor, that is to say, determined by the distance between the two fixed ends.

The purpose of the present study was to investigate effects of developmental lateral jaw deformity (DLJD) on masticatory movement.Three coordinates of mandibular movement signals were recorded in 35 young healthy adults with normal occlusion and 28 patients with DLJD by means of a Sirognathography System during gum chewing movement. Paired and unpaired t-tests were used to compare significant difference of intra-group and inter-groups in displacement and speed. P0.05 was chosen as significance level.(1) The rhythm and directional sequence of chewing trajectory in both frontal and sagittal planes were normal in control group, while anomalies of rhythm and distribution (n = 9) and reversed sequencing (n = 7) were found in DLJD group. (2) The maximum magnitude of jaw retrusion and average speed were significant higher in DLJD group than those in control group (P0.01). The spatial displacement and average speed in horizontal plane and in chewing phase were also higher in DLJD group compared with control group (P0.05).DLJD patients showed anomalies in chewing pattern as well as chewing speed and displacement in horizontal plane.

To investigate the stress distribution in the mandibular fixed partial denture (FPD) by changing the length of pontics.Based on a 3-DFE model of the mandibular FPD, we analyzed the stress distribution in the mandibular FPDs by loading the same level force at pontics with different lengths that were 2 to 4 times of normal length.The distribution trends of the stresses in the mandibular FPDs with differently elongated pontics were similar, but the highest von Mises stress in the bridge was enlarged.Damages may develop in the mandibular FDP, when the length of the pontic is longer than 3 times of normal length.

To establish a continuous and automatic distraction osteogenesis technique in reconstruction of segmental mandibular defect by using embedded titanium-nickel alloy distractor with characteristics of shape-memory and super-elasticity.Adult hybrid canines were used as the animal model. Segmental defects of 1-3 cm in the body of mandible were created by surgical osteotomy. Bi-focal distraction osteogenesis was applied using embedded titanium-nickel distractor designed by the authors. The canines were sacrificed 3 months after the operation and the mandibles were harvested to examine the results of bone regeneration.The histocompatibility of titanium-nickel distractors was good. Distraction osteogenesis was completed automatically and the defects were elementarily restored. Radiological and histological examination showed well bone regeneration in distraction area.Distraction osteogenesis using embedded self-loading titanium-nickel distractor could be a hopeful and useful technique. It might help to solve the problems of functional mandibular reconstruction in the near future.