To understand the usefulness of β-TCP for development of biomaterials implants, β-TCP was implanted into bone defects of dog mandibles, and gene expression profiles were examined using DNA microarray. An implant drill was used to make bone defects in Beagle dog mandibles, and then β-TCP was filled into bone defects. Total RNA was isolated from all specimens, and mRNA levels were analyzed using Affymetrix GeneChip. Higher mRNA levels of insulin-like growth factors, IGF1 and IGF2, were observed in β-TCP-implanted samples compared with controls. The enhancement of IGF1 and IGF2 mRNA levels by β-TCP was confirmed by RT-PCR and real-time RT-PCR. Immunohistochemical staining revealed increased IGF1 and IGF2 protein expression in β-TCP-implanted bone. Taken together, the stimulation of IGF1 and IGF2 expression by β-TCP might be a mechanism of accelerating bone formation.
We evaluated the properties of porous beta-tricalcium phosphate (β-TCP) coated with a thin layer of hydroxyapatite (HA) using a molecular precursor method. Samples of HA-coated beta-TCP (HATCP) and non-coated β-TCP (TCP) were used for osteoblast culture. The proliferation and function of osteoblasts cultured in the samples were evaluated. Osteoblast proliferation and mineralization activity and secretion of osteoclast activating factors in the samples were observed. The thin HA coating on porous β-TCP was not affected to the three dimensional structures. The calcium concentration with osteoblast was significantly higher in the HATCP group, suggesting the thin HA coating enhanced mineralization activity of osteoblast. Osteoblast proliferation activity was similar in the TCP and HATCP groups. Then, osteoclast activating factors seen in HATCP were more than those in TCP. These results suggest that a thin HA coating on porous β-TCP enhanced osteoblast function activity without reducing proliferation activity.
Streptococcus mutans (S. mutans) is the primary etiological agent of human dental caries. A gene (SMU_1525) encoding MurA was identified in S. mutans UA159 (ATCC700610). The deduced amino-acid sequence has homology (51% identity) with E. coli MurA protein (AAC76221). In this study, we cloned S. mutans murA (SMU_1525) gene, expressed soluble recombinant MurA protein in E. coli BL21(DE3). The recombinant protein was purified by affinity chromatography. The molecular weight of expressed protein was about 45.6 kD. The activity of the protein was identified by high-pressure liquid chromatography (HPLC) and was shown to have UDP-N-acetylglucosamine enolpyruvyl transferase activity. Then one microtiter plate based colourimetric assay for S. mutans MurA activity was developed. The optimal temperature and pH of the purified enzyme were 37°C and 7.5, respectively. The Km for UDP-GlcNAc was 0.120 mM and for PEP was 0.086 mM. The Vmax for UDP-GlcNAc was 0.048 mM min-1 mg-1 and for PEP was 0.098 mM min-1 mg-1. The activity of S. mutans MurA was inhibited by the fosfomycin, known MurA inhibitor.
Alveolar bone defects may result in functional and morphological abnormalities. In such cases, bone can be regenerated by grafting fresh autogenous bone into the defect. Subsequently, we focused on the dentin of third molars, which are usually discarded after extraction, or teeth removed faor orthodontic therapy. As in bone, type I collagen is the main component of the organic matrix in dentin. Also, it has been proven that the organic matrix of dentin contains osteoinductive bone morphogenetic protein (BMP). It would be safe to perform grafting with the dentin of extracted teeth used as an autogenous bone substitute. Therefore, the objective of the present study was to reuse extracted teeth as a bone substitute material. Demineralized dentin matrix (DDM), an organic material derived from the dentine of bovine teeth, was grafted in rat skull defects to determine its bone regeneration ability and the possibility of its use as a bone substitute. In the present study, new bone formed from the defect margin in the control group, but in the graft group, new bone formation occurred from individual DDM granules within the defect, and not just from the margin. At Week 8, defect repair was limited in the control group. In contrast, most of the defect was covered by osseous tissue in the graft group. In conclusion, DDM may be a useful bone substitute that serves as a scaffold for bone regeneration by inducing a high level of new bone formation soon after surgery.
This study investigated the difference in pharyngeal spaces in orthodontic patients with or without setback surgery. The control group consisted of 28 patients who underwent orthodontic treatment alone and a study group consisting of 53 patients that had orthodontic treatment and sagittal split ramus osteotomy. The pharyngeal spaces were compared in both groups using cephalogram before and after treatment. During the initial visit, the middle and lower pharyngeal spaces were significantly larger in the study group than in the control group. However, there were no significant differences in any parameter used to measure the pharyngeal space between the two groups after surgery. The surgery contributed to the reduction in the size of the pharyngeal spaces. The pharyngeal space of patients who need setback surgery may be larger than orthodontic patients who do not need surgery.
Orthodontic mechanical stress was exposed on mouse molars and histopathological changes as well as the expressions of HSP27 and p-HSP27 in the periodontal tissues were examined after removal of mechanical stress. The increased in mechanical stress up to 3 hours led to pathological changes that caused a space in between stretched periodontal ligament fibrous bundles and fibroblasts as well as narrowing of the periodontal ligament space. Degenerative changes were also occurred in the pressure side. Pathological changes did not only occur due to mechanical stress but also at the time of the release of mechanical stress exposure which increased over time. In the control group, both HSP27 and p-HSP27 were negative in the pressure side after mechanical stress was released 3 hours later. On the other hand, the tension side showed a strong positive reaction. The proteins were also expressed after 20 min, 1 hour, 3 hours and 9 hours. The strongest expression was observed 24 hours. A decreased in the intensity of expression was observed 3 days and 1 week later. The results suggest that HSP27 plays an important role in the recovery of injured cells in the periodontal tissues.
Since periodontitis, characterized by the absorption of alveolar bone, is a major cause of tooth loss in adults, many clinical and basic investigators have researched how to regenerate alveolar bone. Recently, from the point of view of regenerating alveolar bone, treatment methods such as guided tissue regeneration and guided bone regeneration have been developed; however, these operations require particular skills and cause great stress to patients. To develop a simple, safe and effective treatment for alveolar bone regeneration, seat-shaped soluble material containing protamine-reduced peptides was applied to carrageenin-induced experimental periodontitis in the mandibular first molar region in rats. After 3D image reconstruction by micro-computerized tomography, the quantity of formation of alveolar bone from the start of the experiment was measured in mesial, middle and distal regions on the buccal side. As a result, after 2 weeks, the volume of alveolar bone had increased from the start of the experiment by the application of protamine-reduced peptides. This result suggested that seat-shaped soluble material containing protamine-reduced peptides may be effective for periodontitis; however, the details of the alveolar bone regenerative mechanism are still unknown, and further research will be necessary in the future.
Formation of nerves and blood and lymph vessels and their remodeling are considered to occur after tooth replantation. Vascular endothelial growth factor (VEGF)-A and VEGF-C are regulatory factors of blood and lymph neovascularization, respectively. Although the expression of VEGF-A in pulp vascular endothelial cells, odontoblasts, and pulp fibroblasts under normal conditions, endothelial cells and fibroblasts in tooth movement has been reported, there have been no previous reports on VEGF-A, VEGF-C, or VEGFR-3 after replantation of extracted teeth in rats. In this study, maxillary right first molars were extracted and replanted immediately after the extraction in rats, and the pulp were immunohistochemically investigated after 1 and 3 days fixed by perfusion with 4% paraformaldehyde solution. The maxilla was excised dissected with teeth from all groups, and 5.0-mm3 blocks were prepared. The blocks were decalcified with 10% EDTA at 4°C for 2 weeks, dehydrated with alcohol by the standard method, and embedded in paraffin. The blocks were then cut into thin sections using a microtome, and VEGF-A and VEGF-C protein expressions were investigated by immunohistochemical staining using antibodies against these proteins and the ABC kit. To clarify the cell nuclei, the sections were stained with methyl green after immunohistochemical staining. The sections were observed under a universal microscope. VEGF-A expression in the pulp was weakly positive in perivascular fibroblasts and odontoblasts in the control group. At 1 day post replantation, VEGF-A was positive in these cells, and strongly positive in the Weil layer. VEGF-C expression was weakly positive in perivascular fibroblasts in the control group, and positive in fibroblast and odontoblast at 1 day post replantation. The VEGFR-3 expression pattern was similar to that of VEGF-C. Based on the above findings, expressions of regulatory factors that increase blood and lymph vessels in dental pulp, VEGF-A and VEGF-C and VEGFR-3, were promoted after replantation of extracted teeth, and odontoblasts were suggested to be related to the production of these factors.
In bone remodeling during orthodontic tooth movement, osteoblasts are closely related to bone matrix formation and osteoclast differentiation. Interleukin (IL)-11 is known to be a regulating factor of bone remodeling. Our previous study showed that compressive force (CF) induced IL-11 production in osteoblastic Saos-2 cells. Therefore, we examined the effect of CF on the expression of IL-11 receptor (IL-11r), IL-11, Runx2, bone morphogenetic protein (BMP)-2, receptor activator of NF-κB ligand (RANKL), macrophage colony-stimulating factor (M-CSF), and osteoprotegerin (OPG) using MC3T3-E1 cells as osteoblasts. To confirm the participation of autocrine mechanisms by CF-induced IL-11, we also examined the effect of CF on expression in the presence and absence of IL-11 antagonist. MC3T3-E1 cells were cultured with or without continuous CF (1.0 or 2.0 g/ cm2) in the presence or absence of IL-11 antagonist for up to 24 h. The expression of the IL-11r, IL-11, Runx2, BMP-2, RANKL, M-CSF, and OPG genes was estimated by determining mRNA levels by real-time PCR; protein expression was examined by ELISA or Western blotting. CF increased the expression of IL-11r and IL-11; the maximum level of these expressions was achieved with CF at 2.0 g/cm2. CF also increased the expression of Runx2 and BMP-2; the maximum level of these expressions was achieved in CF at 1.0 g/cm2. IL-11 antagonist blocked the stimulatory effects of CF at 1.0 or 2.0 g/cm2 on the expression of Runx2, BMP-2, RANKL, and M-CSF but not on the expression of IL-11r, IL-11, and OPG. These results suggest that CF at 1.0 g/cm2 increases the expression of proteins related to bone formation, such as Runx2 and BMP-2, via the autocrine mechanism of CF-induced IL-11 in osteoblasts, whereas CF at 2.0 g/cm2 increases the expression of RANKL and M-CSF that promotes osteoclast formation via CF-induced IL-11. These results also suggest that the balance of bone remodeling changes according to the strength of CF.
The tissue- engineered corneal epithelium can be constructed with cultured limbal stem cells from the cornea of healthy rabbits cornea cultured in vitro, and its transplantation may promote the repair and healing of corneal alkaline burn. The aim of this study was to probe the effect and opportunity of treating corneal alkali burn with a tissue-engineered corneal epithelium transplantation. The tissue-engineered corneal epithelium was prepared in 37 rabbits with limbal stem cells cultured in vitro, for transplantation in a double eye corneal alkali burn model. Autologous or allogenic transplantation was performed at early time points (1, 3, 6, 9 days) or metaphase (14 days) following alkaline burning. 1 week following the burn, large corneal epithelial desquamation occurred, with 72% of the rabbits having epithelial desquamations or corneal ulcers at 2 weeks, continuing to 4 weeks post burn. Only 25% of the rabbits in the transplantation group had these effects 4 weeks post-treatment. Cell infiltration and vascularization of corneal stroma was observed in the metaphase transplantation group, not the early transplantation groups; Within 4 weeks, immunological rejection induced by allogenic transplantation was not greater than that of autologous transplantation. Cultivated autologous or allogenic tissue-engineered corneal epithelium transplantation restored ocular surface integrity, and the effect of early transplantation surpasses that of metaphase.
Giant cell tumors of soft tissue (GCT-ST) are considered to be the soft tissue analogue of giant cell tumors of bone. Although the majority of GCT-ST cases manifest histologically benign features, special consideration is required because GCT-ST can occasionally demonstrate extensive local invasion or distant metastases. Here, we present an extremely rare case of GCT-ST arising from subcutaneous mental soft tissue, rapidly growing over the mandibular gingiva and bone, and lacking overtly malignant features in clinical, radiological and cytological findings.
The purpose of this study was to determine the accuracy of a new high resolution digital camera-based three-dimensional system called Di3D by comparing with traditional direct anthropometry using digital calipers. The study sample consisted of 20 young adults, 10 male and 10 female. Facial soft tissue landmarks were identified and five linear distances were measured between the landmarks for each subject using the two methods; Di3D camera system and a digital caliper. No statistically significant differences (P › 0.05) were found for all five facial soft tissue distances measured by the Di3D camera system and the digital caliper. The magnitude of differences was very small (mean = 0.73 mm). Good congruence was observed between the means derived from the Di3D system and the digital caliper. The overall mean differences of the five measurements with these two methods were small enough to be clinically insignificant. The results show that both methods are concordant and one method can replace the other in clinical use.