Osteoclasts are multinucleated cells that differentiate from hematopoietic cells and possess characteristics responsible for bone resorption. To study the involvement of mitogen-activated protein kinases (MAPKs) in osteoclastogenesis of the murine monocytic cell line RAW264.7, which can differentiate into osteoclast-like cells in the presence of the receptor activator of nuclear factor kappa B ligand (RANKL), we treated the cells with specific inhibitors of p38 MAPK, PD169316 and SB203580, and specific inhibitors of MAPK extracellular signaling-regulated kinase (ERK) kinase (MEK), U0126 and PD98059. Each inhibitor blocked differentiation into osteoclast-like cells when the cells were plated at the standard cell density (2000 -4000 cells per well (96-well)). However, the effect of MEK inhibitors on osteoclastogenesis varied according to the initial cell density during culture, because cell growth was clearly inhibited by them. When the cells were plated at more than 8000 cells per well, marked enhancement and acceleration of the differentiation were observed. In addition, immunoblot analysis revealed that phosphorylation of ERK was increased by treatment with the p38 inhibitors, whereas the MEK inhibitors increased phosphorylation of p38, which implies a seesaw-like balance between ERK and p38 phosphorylation. We suggest that osteoclastogenesis is regulated under a balance between ERK and p38 pathways and that the MEK/ERK pathway negatively regulates osteoclastogenesis while the p38 pathway does so positively. This is the first report that an inhibitor of signal transduction enhanced osteoclastogenesis.

Objective: To test the hypothesis that there is no difference in the effect of different continuous moderate to very heavy forces on root resorption or amount of tooth movement. Materials and Methods: In the study, 10, 25, 50 and 100 g mesial force were applied to the maxillary first molars of rat using nickel titanium closed-coil springs for 3 days, 14 days, and 28 days. The molars were extracted and the surface areas of the root resorption craters were measured using scanning electron microscope. The depths of the root resorption craters were measured using a threedimensional laser scanning microscope. Tooth movement of the maxillary first molar was measured in relation to the maxillary second molar on digitized lateral cephalometric radiographs. Results: Three days after force application, the tooth movement was not proportionally related to force magnitude. However, 14 days of force application resulted in significantly more tooth movement in the 10, 25, and 50 g force groups than in the 100 g force group. A force application of 10 g produced significantly more tooth movement at 28 days than all the other three force applications. The largest and deepest resorption craters were observed in the disto-buccal root followed by distopalatal, middle-buccal, middle-palatal, and mesial root. Root resorption and tooth movement increased over time from 3 to 28 days. As heavier forces were applied, greater root resorption occurred. Conclusion:The hypothesis is rejected. The light mesially oriented forces, as applied in this study, produced more tooth movement and less root resorption compared with heavier forces.

Focusing on the final step of osteoclastogenesis, we studied cell fusion from tartrate-resistant acid phosphatase (TRAP)-positive mononuclear cells into multinuclear cells. TRAP-positive mononuclear cells before generation of multinuclear cells by cell fusion were differentiated from RAW264.7 cells by treatment with receptor activator of nuclear factor kappa B ligand (RANKL), and then the cells were treated with lipopolysaccharide (LPS), followed by culturing for further 12 h. LPS-induced cell fusion even in the absence of RANKL. Similarly, tumor necrosis factor (TNF)-alpha and peptidoglycan (PGN) induced cell fusion, but M-CSF did not. The cell fusion induced by RANKL, TNF-alpha, and LPS was specifically blocked by osteoprotegerin (OPG), anti-TNF-alpha antibody, and polymyxin B, respectively. LPS- and PGN-induced cell fusion was partly inhibited by anti-TNF-alpha antibody but not by OPG. When TRAP-positive mononuclear cells fused to yield multinuclear cells, phosphorylation of Akt, Src, extracellular signal-regulated kinase (ERK), p38MAPK (p38), and c-Jun NH2-terminal kinase (JNK) was observed. The specific chemical inhibitors LY294002 (PI3K), PP2 (Src), U0126 (MAPK-ERK kinase (MEK)/ERK), and SP600125 (JNK) effectively suppressed cell fusion, although SB203580 (p38) did not. mRNA of nuclear factor of activated T-cells c1 (NFATc1) and dendritic cell-specific transmembrane protein (DC-STAMP) during the cell fusion was quantified, however, there was no obvious difference among the TRAP-positive mononuclear cells treated with or without M-CSF, RANKL, TNF-alpha, LPS, or PGN. Collectively, RANKL, TNF-alpha, LPS, and PGN induced cell fusion of osteoclasts through their own receptors. Subsequent activation of signaling pathways involving PI3K, Src, ERK, and JNK molecules was required for the cell fusion. Although DC-STAMP is considered to be a requisite for cell fusion of osteoclasts, cell fusion-inducing factors other than DC-STAMP might be necessary for the cell fusion.

The maxillary dental casts can be reliably superimposed on the medial points of the third palatal rugae and the palatal vault as reference landmarks.

Despite inherent errors, cephalometric superimpositions are currently the most widely used means for assessing sagittal and vertical tooth movements. The purpose of this study was to compare three-dimensional (3D) digital model superimposition with cephalometric superimposition. The material was collected from initial and final maxillary casts and lateral cephalometric radiographs of 30 patients (6 males, 24 females, mean age 17.7 years) who underwent orthodontic treatment with extraction of permanent teeth. Each pair of cephalograms was traced and superimposed according to Ricketts' four-step method. 3D scanning of the maxillary dental casts was performed using INUS dental scanning solution(R), which consists of a 3D scanner, an autoscan system, and 3D reverse modelling software. The 3D superimposition was carried out using the surface-to-surface matching (best-fit method) function of the autoscan system. The antero-posterior movement of the maxillary first molar and central incisor was evaluated cephalometrically and on 3D digital models. To determine whether any difference existed between the two measuring techniques, paired t-tests and correlation analysis were undertaken. The results revealed no statistical differences between the mean incisor and molar movements as assessed cephalometrically or by 3D model superimposition. These findings suggest that the 3D digital orthodontic model superimposition technique used in this study is clinically as reliable as cephalometric superimposition for assessing orthodontic tooth movements.