Hitoshi Hotokezaka

Correlation between cortical plate proximity and apical root resorption

Root resorption is one of the most common iatrogenic sequelae of orthodontic treatment. Recently, root contact with the labial or palatal cortical plate at root apex level during orthodontic tooth movement was reported to be related to root resorption, and dentofacial morphology was suggested to predispose certain persons to root contact with the cortical plate. In this study, we constructed a best-fit straight line for the maxillary palatal cortical plate and set a line for the labial cortical plate from A point to Prosthion point in order to obtain measurements of proximity of root apices with the cortical plates of the maxillary alveolus. We investigated the correlation between apical root resorption and the measured variables. Our findings suggest that root approximation to the palatal cortical plate during orthodontic treatment could explain approximately 12% of the variance observed in the level of root resorption and the maxillary alveolar bone width about 2%. Tooth extrusion and crown lingualization also contributed to root resorption. We concluded that maxillary central incisor apical root resorption is influenced by root approximation to the palatal cortical plate during orthodontic treatment.

Force Magnitude and Duration Effects on Amount of Tooth Movement and Root Resorption in the Rat Molar

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 three-dimensional 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 disto-palatal, 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.

Molecular analysis of RANKL-independent cell fusion of osteoclast-like cells induced by TNF-α, lipopolysaccharide, or peptidoglycan

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)‐α and peptidoglycan (PGN) induced cell fusion, but M‐CSF did not. The cell fusion induced by RANKL, TNF‐α, and LPS was specifically blocked by osteoprotegerin (OPG), anti‐TNF‐α antibody, and polymyxin B, respectively. LPS‐ and PGN‐induced cell fusion was partly inhibited by anti‐TNF‐α 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‐α, LPS, or PGN. Collectively, RANKL, TNF‐α, 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. J. Cell. Biochem. 101: 122–134, 2007.

Effect of IL-12 on TNF-α-Mediated Osteoclast Formation in Bone Marrow Cells: Apoptosis Mediated by Fas/Fas Ligand Interaction

Recently, it has been found that differentiation into osteoclasts is induced by TNF-α. In this study, we investigated the effect of IL-12 on TNF-α-mediated osteoclastogenesis. When mouse bone marrow cells were cultured with TNF-α, osteoclast-like cells were formed. When they were cultured with both TNF-α and IL-12, the number of adherent cells in the bone marrow cells decreased in an IL-12 dose-dependent manner. A combination of IL-12 and TNF-α was necessary to induce death of the adherent cells in this culture system. Apoptotic alterations, which were indicated by morphological changes such as cellular atrophy, nuclear and cellular fragmentation, and biochemical changes such as DNA fragmentation, were observed in the adherent cells. Apoptosis of the adherent cells was markedly inhibited by anti-Fas ligand (FasL) Ab. RT-PCR and FACS analyses revealed that TNF-α up-regulated Fas transcription to lead to Fas expression on the surfaces of the adherent cells, whereas IL-12 could not induce Fas on the cells. In contrast, IL-12 induced FasL transcription to lead to FasL expression on the surfaces of nonadherent bone marrow cells, whereas TNF-α could not induce FasL on the cells. These results implied that apoptosis of the adherent cells in bone marrow cells might be caused by interaction between TNF-α-induced Fas on the adherent cells and IL-12-induced FasL on the nonadherent cells.

Characterization of the gene encoding the MPB51, one of the major secreted protein antigens of Mycobacterium bovis BCG, and identification of the secreted protein closely related to the fibronectin binding 85 complex

The secreted protein MPB51 is one of the major proteins in the culture filtrate of Mycobacterium bovis BCG (BCG) and is a protein immunologically cross‐reacting with the fibronectin binding 85 complex secreted by this bacterium. The gene encoding MPB51 (mpb51) was cloned, sequenced, and expressed in Escherichia coll. The mpb51 gene was mapped downstream of the gene for 85A component with 179 bp spaces. The mpb51 gene encoded 299 amino acids, including 33 amino acids for the signal peptide, followed by 266 amino acids for the mature protein with a molecular mass of 27807.37 Da. This is the first complete sequence of MPB51. MPB51 showed 37–43% homology to the components of 85 complex. Two‐dimensional electrophoresis of culture fluids of BCG and Western blotting indicated the existence of the other novel protein(s) which strongly cross‐reacted with the α antigen (85B) and MPB51.

Effects of Steroidal and Nonsteroidal Drugs on Tooth Movement and Root Resorption in the Rat Molar

OBJECTIVE To test the hypothesis that the administration of aspirin, acetaminophen, meloxicam, celecoxib, and prednisolone have no effect on root resorption and tooth movement. MATERIALS AND METHODS A mesial force of 50 g was applied to the left maxillary first molars of sixty 10-week-old male Wistar rats using nickel titanium closed coil springs attached to the cervical area of the incisors. The rats were randomly divided into 12 groups of 5 each. High and low doses of aspirin, acetaminophen, meloxicam, celecoxib, and prednisolone were administered via drinking water for 2 weeks. The experimental control group had tooth movement but received no drug. The negative control group received neither tooth movement nor drugs. The amount of tooth movement was measured on digitized lateral cephalometric radiographs. Rats were sacrificed after 2 weeks. Mesial and distal roots (distobuccal and distopalatal) were examined using scanning electron and three-dimensional (3D) scanning laser microscopes. The surface area, depth, volume, and roughness of the root resorption craters were measured. RESULTS When compared with experimental control rats, only prednisolone- and high-dose celecoxib-treated groups showed significantly less root resorption and less tooth movement. Although low dose celecoxib-treated group significantly decreased the tooth movement, root resorption was similar to the control group. Furthermore, resorption craters showed a smoother surface in the prednisolone-treated rats. CONCLUSIONS The hypothesis was rejected. Administration of prednisolone and high-dose celecoxib reduces root resorption and interferes with tooth movement in rats. Both drugs may interfere in the arachidonic acid cascade depending on dose thresholds.

Cloning and sequencing of a unique antigen MPT70 from Mycobacterium tuberculosis H37Rv and expression in BCG using E. coli-mycobacteria shuttle vector.

MPB70 is known to be an immunogenic mycobacterial protein secreted in large amounts from Mycobacterium bovis BCG (BCG) Tokyo. The analogous gene for MPT70 was cloned from Mycobacterium tuberculosis H37Rv which produces this protein in only a small amount. The gene encoding 193 amino acid residues inciuding 30 amino acids for the signal peptide. the promoter‐like sequence, and the ribosome‐binding site, was completely identical to that of BCG Tokyo. Computer analysis revealed that the carboxy‐terminal half of MPT70 was homologous to amino acid sequences of fasciclin 1, osteoblast‐specific factor 2 (OSF‐2), and human transforming growth factor‐beta induced gene product (βIG‐H3). Escherichia coli (E. coli) ‐mycobacteria shuttle vectors containing mpt70 or mpb70 genes 0.7kbp upstream of the 5′ end of them were able to be expressed in BCG Pasteur which is a MPB70 low‐producer, but the extent of the expression was not that of a high‐producer.

Severe Dental Open Bite Malocclusion With Tongue Reduction After Orthodontic Treatment

We treated a 21-year-old woman with a severe open bite and macroglossia with a standard edgewise appliance and without partial glossectomy. This was followed by retention using a Begg-type plate retainer for the upper dental arch and a fixed canine-to-canine for the lower arch. A crib was added to the upper plate retainer for suppression of a tongue thrust. The lower arch relapsed during the retention period, with a widening of the intermolar distance, flaring of the anterior teeth, and increased mobility of the teeth. We chose tongue reduction to resolve these problems and one-third of the middle dorsal part of the tongue was excised. After the tongue reduction, the patient experienced no functional problem in mastication, swallowing, and gustation, but she complained of mild speech difficulty and slight pain on the dorsal portion of her tongue. These symptoms disappeared 6 months after surgery. At this time, the mandibular dental arch was markedly improved. The flared lower dental arch had returned to an upright position and the tooth mobility reduced to normal. No appliance was used after surgery. Most of the recovery changes occurred within 4 months. This case highlights the importance of the teeth tending to move toward a balance between the tongue pressure from the inside and labio-buccal pressure from the outside.

An In Vivo 3D Micro-CT Evaluation of Tooth Movement After the Application of Different Force Magnitudes in Rat Molar

OBJECTIVE To investigate the precise longitudinal change in the periodontal ligament (PDL) space width and three-dimensional tooth movement with continuous-force magnitudes in living rats. MATERIALS AND METHODS Using nickel-titanium closed-coil springs for 28 days, 10-, 25-, 50-, and 100-g mesial force was applied to the maxillary left first molars. Micro-CT was taken in the same rat at 0, 1, 2, 3, 10, 14, and 28 days. The width of the PDL was measured in the pressure and tension sides from 0 to 3 days. Angular and linear measurements were used to evaluate molar position at day 0, 10, 14, and 28. The finite element model (FEM) was constructed to evaluate the initial stress distribution, molar displacement, and center of rotation of the molar. RESULTS The initial evaluation of PDL width showed no statistical differences among different force magnitudes. Tooth movement was registered 1 hour after force application and gradually increased with time. From day 10, greater tooth movement was observed when 10 g of force was applied. The FEM showed that the center of rotation in the molar is located in the center of five roots at the apical third of the molar roots. CONCLUSION The rats molar movement mainly consists of mesial tipping, extrusion of distal roots, intrusion of mesial root, palatal inclination, and mesial rotation. Although the initial tooth movement after the application of different force magnitudes until day 3 was not remarkably different, 10 g of force produced more tooth movement compared with heavier forces at day 28.

Interleukin-4 directly inhibits tumor necrosis factor-α-mediated osteoclast formation in mouse bone marrow macrophages

Recently it has been found that osteoclast differentiation is induced by tumor necrosis factor (TNF)-alpha. Interleukin (IL)-4 was reported to suppress osteoclast differentiation and bone resorption. However, no study has investigated the effect of IL-4 on TNF-alpha-induced osteoclast formation. In this study, we investigated whether IL-4 inhibits TNF-alpha-mediated osteoclast formation in mouse bone marrow derived macrophages (BMM). First, IL-4 suppresses RANKL-induced osteoclast formation and bone resorption. Next, when BMM were cultured with TNF-alpha, osteoclast-like cells were formed. When they were cultured with both TNF-alpha and IL-4, osteoclast formation and bone resorption was suppressed by IL-4 in a dose-dependent manner. It has been recently found that TNF-alpha and RANKL synergistically promote osteoclastogenesis. Finally, we investigated whether IL-4 had the ability to inhibit synergistic TNF-alpha and RANKL-induced osteoclastogenesis, with the result that it effectively inhibited the synergistic osteoclast formation in a dose-dependent manner. We conclude that IL-4 can strongly inhibit osteoclast formation that is related to both physiological bone resorption induced by RANKL and pathological bone resorption induced by TNF-alpha.