Extracellular enzymatically synthesized glycogen promotes osteogenesis by activating osteoblast differentiation via Akt/GSK-3ß signaling pathway.

Glycogen is the stored form of glucose and plays a major role in energy metabolism. Recently, it has become clear that enzymatically synthesized glycogen (ESG) has biological functions, such as the macrophage-stimulating activity. This study aimed to evaluate the effect of ESG on osteogenesis. MC3T3-E1 cells were cultured with ESG, and their cell proliferative activity and osteoblast differentiation were measured. An in vivo study was conducted in which ESG pellets with BMP-2 were grafted into mouse calvarial defects and histomorphometrically analyzed for the new bone formation. To confirm the effect of ESG on bone growth in vivo, ESG was orally administered to pregnant mice and the femurs of their pups were examined. We observed that ESG stimulated cell proliferation and enhanced messenger RNA expression of osteocalcin and osteopontin in MC3T3-E1 cells. ESG was taken up by the cells associated with GLUT-1 and activated the Akt/GSK-3ß pathway. In vivo, the new bone formation in the calvarial defect was significantly accelerated by ESG and the maternal administration of ESG promoted fetal bone growth. In conclusion, ESG stimulates cell proliferation and differentiation of preosteoblasts via the activation of Akt/GSK-3ß signaling and promotes new bone formation in vivo, suggesting that ESG could be a useful stimulant for osteogenesis.

The accuracy of quantifying the degree of hard tissue calcification using an electron probe micro analyzer, micro-focus X-ray computed tomography, and tissue sectioning methods.

OBJECTIVES: Micro-focus X-ray computed tomography (µCT) helps evaluate specimens without destroying it. However, its accuracy of quantifying bone mineral density remains to be fully elucidated. We aimed to verify the accuracy of calcification assessed by µCT, by comparing the images of identical specimens obtained via different methods such as µCT and electron probe micro analyzer (EPMA) analyses. METHODS: The maxillae, mandibles, and tibiae of five-week-old male mice were analyzed. Calcification density was analyzed using µCT. The right sides of the specimens were decalcified and processed for Azan staining. The left side of the specimens underwent elemental mapping for Ca, Mg, and P using EPMA. RESULTS: µCT revealed a significant increase in calcification levels in the following order: enamel, dentin, cortical bone, and trabecular bone. These results reflected the Ca and P levels observed in the EPMA analyses. µCT demonstrated significant differences in the degree of calcification among the enamel tissues or dentin tissues, except for dentin in the maxillary incisors and molars. However, EPMA analysis did not demonstrate significant differences in the Ca and P levels among the same tissue samples. CONCLUSIONS: EPMA elemental analysis can be used to measure Ca and P levels for evaluating the calcification rate of hard tissues. Additionally, the study results validate the evaluation of calcification density via µCT. Furthermore, µCT can evaluate even minute differences in calcification rates compared with EPMA analysis.

Practical Preparation of Elastomer-Immobilized Nonclose-Packed Colloidal Photonic Crystal Films with Various Uniform Colors.

Colloidal photonic crystals, which are three-dimensional periodic structures of monodisperse submicron-sized particles, are expected to be suitable for novel photonic applications and color materials. In particular, nonclose-packed colloidal photonic crystals immobilized in elastomers exhibit significant potential for use in tunable photonic applications and strain sensors that detect strain based on color change. This paper reports a practical method for preparing elastomer-immobilized nonclose-packed colloidal photonic crystal films with various uniform Bragg reflection colors using one kind of gel-immobilized nonclose-packed colloidal photonic crystal film. The degree of swelling was controlled by the mixing ratio of the precursor solutions, which used a mixture of solutions with high and low affinities for the gel film as the swelling solvent. This facilitated color tuning over a wide range, enabling the facile preparation of elastomer-immobilized nonclose-packed colloidal photonic crystal films with various uniform colors via subsequent photopolymerization. The present preparation method can contribute to the development of practical applications of elastomer-immobilized tunable colloidal photonic crystals and sensors.

Angle-Independent Color Change in Thermoresponsive Gel-Immobilized Colloidal Amorphous Film Attached to PET Substrate.

Gel-immobilized colloidal amorphous structures comprise short-range-ordered monodisperse submicrometer particles embedded into a soft polymer gel. They exhibit an angle-independent structural color that is tunable in response to external stimuli via a volume change in the gel, which has significant potential for the development of sensors that respond to stimuli via angle-independent color changes. In this study, the amorphous structure of a charged colloidal suspension in water was immobilized in a thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) gel film and simultaneously attached to a polyethylene terephthalate (PET) substrate. The gel film exhibited a uniform angle-independent color that changed in response to changes in temperature (i.e., thermosensitivity). Attachment to the PET substrate suppressed changes in the gel film area and film distortion, despite significant volume changes in the gel. Consequently, the degree of thermosensitivity was enhanced. The PET-attached gel-immobilized colloidal amorphous film was easy to handle and had excellent flexibility, allowing it to wrap around the surfaces of curved objects. These features are advantageous for sensor applications.

Sequential analysis of head movement during mandibular open-close movements in TMD patients with disc displacement with reduction.

This study examined relationships between starting points of head and mandibular movements in 11 female control subjects and 10 TMD patients showing disc displacement with reduction during consecutive open-close movements, using a six-degree-of-freedom measuring device. During the first mandibular opening cycle, in the TMD group, head movement was significantly preceded in relation to mandibular movement when compared with the control group, and major differences in onset were seen between maximum and minimal values at the beginning of mandibular movements. After the second cycle, significant differences in starting points were not evident. In TMD patients showing disc displacement with reduction, in the first cycle, at the commencement of opening movements, it is proposed that the head precedes the mandible in order to compensate for irregular condylar movements, and the degree of the condylar movement irregularity might affect the onset relation between the head and the mandible.

Mast4 determines the cell fate of MSCs for bone and cartilage development.

Mesenchymal stromal cells (MSCs) differentiation into different lineages is precisely controlled by signaling pathways. Given that protein kinases play a crucial role in signal transduction, here we show that Microtubule Associated Serine/Threonine Kinase Family Member 4 (Mast4) serves as an important mediator of TGF-ß and Wnt signal transduction in regulating chondro-osteogenic differentiation of MSCs. Suppression of Mast4 by TGF-ß1 led to increased Sox9 stability by blocking Mast4-induced Sox9 serine 494 phosphorylation and subsequent proteasomal degradation, ultimately enhancing chondrogenesis of MSCs. On the other hand, Mast4 protein, which stability was enhanced by Wnt-mediated inhibition of GSK-3ß and subsequent Smurf1 recruitment, promoted ß-catenin nuclear localization and Runx2 activity, increasing osteogenesis of MSCs. Consistently, Mast4-/- mice demonstrated excessive cartilage synthesis, while exhibiting osteoporotic phenotype. Interestingly, Mast4 depletion in MSCs facilitated cartilage formation and regeneration in vivo. Altogether, our findings uncover essential roles of Mast4 in determining the fate of MSC development into cartilage or bone.

Posterior superior alveolar nerves contribute to sensation in the anterior teeth.

BACKGROUND: There is no available data on the occurrence rate of a converged alveolar canal, the detailed three-dimensional (3D) courses of alveolar canals/grooves (ACGs), or the contribution of each superior alveolar nerve to each area in the maxilla. This study aimed to clarify the 3D courses of ACGs, the relationship between ACGs and superior alveolar nerves, and the contribution of posterior superior alveolar nerves (PSANs) using computed tomography (CT) with histological analysis. METHODS: During the gross anatomy course at Niigata University, we investigated nine human cadavers. RESULTS: All anterior and posterior ACGs converged into the common alveolar canal, which contained blood vessels and several nerve bundles surrounded by perineurium, located at the nasal floor near the pyriform aperture. Histometrical analysis clarified that 16.3% of the nerve bundles in this canal were derived from PSANs, and 67% of the bundles were dispersed while they coursed down to the nasal floor. There seems to be no relationship between the density of nerve bundles in the canal and the number of remaining anterior teeth. CONCLUSIONS: Data obtained from observing the detailed 3D courses of anterior and posterior ACGs, and their relationship with superior alveolar nerves, suggest that PSANs partially contribute to the nociception of the anterior teeth.

Trabecular bone microstructure and mineral density in human residual ridge at various intervals over a long period after tooth extraction.

BACKGROUND: Long-term changes of trabecular microstructure in human tooth extraction socket have not been investigated. PURPOSE: To examine the trabecular microstructure of human residual ridges at various intervals following tooth extraction, and to determine whether bone remodeling activity can attain points of relative stability and when such points are reached. MATERIALS AND METHODS: Forty-four bone biopsy specimens were obtained from lower molar or premolar regions of residual ridges. Postextraction times ranged from 1.6 to 360 months. Samples were analyzed using micro-computed tomography and three-dimensional bone morphometry with histological analyses. Trabecular bone parameters were plotted against postextraction times, and a stepwise piecewise linear regression analysis was performed to determine at which points of time these parameters either increased or decreased. RESULTS: Using piecewise linear regression, "inflection points" were found in most trabecular bone parameters between 7 and 12 months postextraction. Among the residual ridge samples, woven trabecular structure became mature, consisting of thick lamellar trabeculae with sufficient bone density, under dynamic bone remodeling until the 7th to 12th month post-tooth extraction. After this period, the mature network structure remained stable with low remodeling activity. CONCLUSION: Bone remodeling of trabecular structure in human residual ridge after tooth extraction had a stabilization period.

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.

Long-term changes in trabecular structure of aged rat alveolar bone after ovariectomy.

OBJECTIVE: Trabecular structural changes in the jaw after long-term estrogen deficiency are not well understood. Therefore, we sought to observe the changes in rat alveolar bone for 1 year. METHODS: Six-month-old female rats were ovariectomized (OVX) or underwent a sham operation. After 1 year, bone histomorphometry and a node-strut analysis were performed on the interradicular septum of the mandibular first molar by using micro computed tomography and confocal laser scanning microscopy. Statistical analysis was carried out by using analysis of variance. RESULTS: The alveolar trabeculae of rats in the sham group had network structures, whereas the trabeculae of rats in the OVX group became fragmented. The trabecular bone volume, number, and thickness in the OVX group were significantly lower than those found in the sham group, and the trabecular separation was 4-fold higher in the OVX group than in the sham group. Bone resorptive and formative activity appeared to be moderately higher in the OVX group than in the sham group, but only the difference in bone formation was of statistical significance. CONCLUSION: By 1 year after ovariectomy, bone loss and trabecular fragmentation had occurred in the rat mandibular alveolar bone.