Yuji Tsuka

Comparison of the bone regeneration ability between stem cells from human exfoliated deciduous teeth, human dental pulp stem cells and human bone marrow mesenchymal stem cells

Cleft lip and palate is the most common congenital anomaly in the orofacial region. Autogenous iliac bone graft, in general, has been employed for closing the bone defect at the alveolar cleft. However, such iliac bone graft provides patients with substantial surgical and psychological invasions. Consequently, development of a less invasive method has been highly anticipated. Stem cells from human exfoliated deciduous teeth (SHED) are a major candidate for playing a significant role in tissue engineering and regenerative medicine. The aim of this study was to elucidate the nature of bone regeneration by SHED as compared to that of human dental pulp stem cells (hDPSCs) and bone marrow mesenchymal stem cells (hBMSCs). The stems cells derived from pulp tissues and bone marrow were transplanted with a polylactic-coglycolic acid barrier membrane as a scaffold, for use in bone regeneration in an artificial bone defect of 4 mm in diameter in the calvaria of immunodeficient mice. Three-dimensional analysis using micro CT and histological evaluation were performed. Degree of bone regeneration with SHED relative to the bone defect was almost equivalent to that with hDPSCs and hBMSCs 12 weeks after transplantation. The ratio of new bone formation relative to the pre-created bone defect was not significantly different among groups with SHED, hDPSCs and hBMSCs. In addition, as a result of histological evaluation, SHED produced the largest osteoid and widely distributed collagen fibers compared to hDPSCs and hBMSCs groups. Thus, SHED transplantation exerted bone regeneration ability sufficient for the repair of bone defect. The present study has demonstrated that SHED is one of the best candidate as a cell source for the reconstruction of alveolar cleft due to the bone regeneration ability with less surgical invasion.

Role of Articular Disc in Condylar Regeneration of the Mandible

The articular disc in the temporomandibular joint plays an important role in mandibular growth. Functional appliances induce regeneration of the condyle even after condylectomy. The aim of this study was to examine the role of the articular disc in regeneration of the condyle after unilateral condylectomy with use of a functional appliance in growing rats. Fifty growing rats were subjected to unilateral condylectomy and then half of them underwent discectomy. The functional appliance was applied to half of the rats in each group to induce regeneration of the condyle. Four and eight weeks later, morphometric and histologic analyses of the mandible were performed. Regeneration of the condyle was demonstrated in the two condylectomy groups. In the condylectomy+appliance group, the shape and cartilage of the condyle were equivalent to a normal condyle. However, regeneration of the condyle was not observed in the two discectomy groups even with the use of the functional appliance. The articular disc appears to be crucial in the regeneration of a damaged condyle, suggesting that defects or damage to the articular disc may influence mandibular growth and regeneration or repair of the condyle.

Effects of Neodymium-Doped Yttrium Aluminium Garnet (Nd:YAG) Laser Irradiation on Bone Metabolism During Tooth Movement

INTRODUCTION The aim of this study is to evaluate the effects of low-level neodymium-doped yttrium aluminium garnet (Nd:YAG) laser irradiation on orthodontic tooth movement and histological examination. METHODS Eleven male Wistar rats (aged 10 weeks) were included. To produce experimental tooth movement in rats, 10 g force was applied to maxillary first molars with nickel titanium closed coil springs. Right molars were irradiated with Nd:YAG laser on days 0, 1, 2, 3, 7, 10, 14, 17, 21 and 24, while un-irradiated left molars were used as control. Distance between mesial side of second molar and distal side of first molar was measured on μCT image during tooth movement and the rats were sacrificed 4 weeks after the initiation of tooth movement. RESULTS The amount of tooth movement was significantly greater in the irradiation group (0.20 ± 0.06) than in the control group (0.14 ± 0.03) during the first week (P < 0.05). However, no statistically significant difference was found afterwards. There was a tendency of higher tartrate-resistant acid phosphatase (TRAP)-positive nuclei count in the pressure zones of the laser irradiation group, but it was not statistically significant. In immuno-histological examination, expressions of alkaline phosphatase (ALP) and receptor activator of nuclear factor kappa-B ligand (RANKL) were higher at the pressure site of the laser irradiation group than the control group, whereas there was no difference in osteoprotegerin (OPG) expression. CONCLUSION The results suggest that low-level Nd:YAG laser may stimulate osteoclast and osteoblast activation and accelerate bone metabolism during tooth movement.

Regeneration of condyle with a functional appliance.

Condylar regeneration with the use of functional appliances after condylectomy has been validated. However, the process during treatment remains unclear. In this study, we evaluated the condylar regeneration process and then examined mandibular growth and masticatory muscle activity after regeneration in growing rats. Seventy-five male Wistar rats aged 4 weeks were equally divided into 3 groups: unilateral condylectomy group, unilateral condylectomy + appliance group, or control group. The use of a functional appliance following condylectomy promoted mandibular growth and regeneration of the condyle 1 week after condylectomy. Condyle regeneration showing normal morphology was finally achieved 8 weeks after condylectomy. Asymmetrical masticatory muscle activity was observed after condylectomy. However, the use of a functional appliance produced symmetrical masticatory muscle activity. These results indicate a favorable regeneration process in the condylectomized area due to the use of a functional appliance. In addition, due to condylar regeneration, symmetrical masticatory muscle activity was achieved.

The effect of mesenchymal stem cells on chemotaxis of osteoclast precursor cells

Regeneration of tissue, including bone, using mesenchymal stem cells (MSCs) has been progressing rapidly. Regeneration of bone requires the presence of an appropriate environment and efficient chemotaxis of cells to the target site. Differentiation of MSCs into mesenchymal cells has received considerable attention, but the effect of MSCs on chemotaxis is not well understood. In this study, we investigated the effect of MSCs on chemotaxis of RAW264 cells via C-C motif chemokine ligand 2 (CCL2). Balb/c mouse bone marrow-derived MSCs and RAW264 cells, which are osteoclast precursor cells, were co-cultured without cell contact. The gene expression of CCL2 in MSCs and CC-chemokine receptor 2 (CCR2) in RAW264 cells was determined using quantitative real-time PCR. Analysis of RAW264 cell chemotaxis was performed using the Boyden chamber assay. mRNAs for CCL2 and CCR2 were significantly upregulated upon co-culture in comparison to culture of either cell type alone, and the number of chemotactic RAW264 cells was significantly increased by co-culture. MSCs enhanced the chemotaxis of RAW264 cells, possibly via CCL2-CCR2 interaction, suggesting the potential utility of MSCs for tissue regeneration.

Effect of high-frequency near-infrared diode laser irradiation on periodontal tissues during experimental tooth movement in rats: HIGH FREQUENCY LASER ACCELERATES TOOTH MOVEMENT

Tooth movement during orthodontic treatment is associated with bone neoplasticity and bone resorption on the tension and pressure sides. Previous clinical studies have suggested that low‐power laser irradiation can accelerate tooth movement during orthodontic treatment, although the underlying mechanism remains unclear. In this study, we used a high‐frequency near‐infrared diode laser that generates less heat and examined the histologic changes in periodontal tissue during experimental tooth movement with laser irradiation.

Dynamic imaging of the effect of mesenchymal stem cells on osteoclast precursor cell chemotaxis for bone defects in the mouse skull

Background/purpose Mesenchymal stem cells (MSCs) transplantation has previously been used in the field of regenerative medicine. Although bone regeneration is known to occur through the interaction between osteoblasts and osteoclasts, the effect of MSCs on osteoclasts is unknown. Therefore, the purpose of this study was to investigate the effect of MSCs on the chemotaxis of osteoclast precursor cells (RAW264 macrophage cells). Materials and methods Bone defects were created in mice skulls, and MSCs and a scaffold of carbonated hydroxyapatite were transplanted into the bone defects. RAW264 cells were then transplanted into the mouse tail vein, and their dynamics were observed by an in vivo imaging system. Results The fluorescent intensity of the MSCs transplant group at the bone defect region was significantly higher on days 3, 5, and 7 compared with the MSCs non-transplant group. Conclusion Increased RAW264 chemotaxis to the bone defect region occurred following the simultaneous implantation of MSCs in the skull defect.

A comparison of proliferation of human mesenchymal stem cells derived from adipose tissue treated with human full-length amelogenin and C-terminal amelogenin peptide

Amelogenins are enamel matrix proteins that play crucial roles in enamel formation. Previous studies have indicated that amelogenin and amelogenin C-terminal peptides have cell-signaling functions. Recently, adipocyte-derived mesenchymal stem cells (ADSCs) have received attention as a potential source of stem cells for use in regeneration therapy. In this study, we examined the effects of human full-length amelogenin (rh174) and amelogenin C-terminal peptide (amgCP) on the proliferation of ADSCs. ADSCs were cultured in the presence of amgCP or rh174. Cell proliferation was analyzed using BrdU immunoassay and MTS assay. Cell migration was evaluated by ELISA. The MAPK-ERK pathway was examined by phospho-p44/42 MAPK (Thr202/Tyr204) sandwich ELISA and western blotting. A specific MAPK inhibitor, U0126, was used to block ERK activity. ADSC proliferation and migration were significantly (P < 0.05) increased in the presence of rh174 or amgCP compared to non-treated control cells. The increased proliferation of ADSCs induced by rh174 or amgCP was significantly (P < 0.05) inhibited in the presence of 2 µg/ml U0126. The pERK/tERK ratio was significantly (P < 0.05) increased upon treatment with rh174 or amgCP compared to non-treated ADSCs, while this increase was significantly (P < 0.05) suppressed by the addition of U0126. Similar results were found by western blot analysis. In conclusion, amgCP and rh174 increase ADSC proliferation via the MAPK-ERK signaling pathway, and ADSCs may be useful for tissue regeneration in the orofacial region.

Role of articular disc in cartilaginous growth of the mandible in rats

Displacement of the temporomandibular joint (TMJ) disc causes a lateral shift of the mandible and less-developed and/or distally located mandible unilaterally and bilaterally, respectively, if occurred in growing individuals. The purpose of this study was to evaluate mandibular condylar growth in growing rats after TMJ discectomy and to explore a certain significant role of articular disc in the TMJ in mandibular or cartilaginous growth. Eighteen 4-week-old Wistar strain male rats were divided into two groups with nine in each group, i.e., rats with TMJ discectomy (discectomy group) and only sham operation (control group). Four weeks after initiating the experiment, morphometric analyses of the mandible were performed using a rat and mouse cephalometer and micro-computed tomography. Then, the mandibular condyles were subjected to histomorphometric analyses. Condylar and mandibular growth was reduced significantly in the discectomy group than in the control group. In the discectomy group, the condyle also became flatter and smaller. In addition, the 4-layer structure of condylar cartilage was unclear with thicker fibrous and thinner lower hypertrophic layers in the discectomy group when compared to the controls. It is shown that resection of the articular disc substantially affects condylar and mandibular growth in terms of the cartilaginous growth, suggesting that TMJ disc is indispensable for maintaining normal growth of the condyle and mandible, leading to optimal development of the TMJ and the entire mandible.