Sawa Kaneko

Effect of compressive forces on extracellular matrix in rat mandibular condylar cartilage.

To reveal the effect of compressive force on the mandibular condylar cartilage, an appliance was set on 8-week-old Wistar rats to load continuous compressive force. Immunohistochemical and histochemical analyses were performed using toluidine blue, antibodies, and probes for aggrecan, hyaluronan, type II collagen, type X collagen, and 5-bromo-2″-deoxyuridine (BrdU). Histomorphometry and statistical analyses were also performed for aggrecan and BrdU immunostaining. In toluidine blue staining, tissue metachromasia was observed in the transitional zone and the hypertrophic zone of the mandibular condylar cartilage. In histomorphometry and statistical analysis, thickness of the cartilage decreased significantly in all regions in the 3-day experimental group. However, the thickness of the cartilage in the anterior region showed recovery while it decreased continuously in the posterior region. Distributional changes of aggrecan, hyaluronan, type II collagen, and type X collagen in the experimental groups were similar to those for toluidine blue staining. The immunostained area of all these molecules decreased as a result of the decrement of the cartilage area. However, enhanced immunostaining for aggrecan in the proliferative zone was observed only in the 1-day experimental group. BrdU-positive cells, observed in the proliferating zone and the transitional zone, decreased significantly in the experimental group 3 days after force was applied. These results demonstrate that continuous compressive forces on the mandibular condylar cartilage decrease the proliferation of chondrocytes and the amount of extracellular matrices.

Effect of sympathetic nervous activity on alveolar bone loss induced by occlusal hypofunction in rats

OBJECTIVE To elucidate the effect of sympathetic nervous activity on alveolar bone loss induced by occlusal hypofunction in rat molars. DESIGN Occlusal hypofunction in the molar area was produced by attaching appliances to rat maxillary and mandibular incisors. In addition, a non-selective β-adrenergic receptor antagonist, propranolol, was administered orally to rats in drinking water to pharmacologically suppress sympathetic nervous activity. After 1 week, alveolar bones in all groups were examined by micro-CT, histomorphometry and histology to determine their trabecular bone phenotypes and histological changes. RESULTS The marrow spaces of the interradicular alveolar bone of rat mandibular first molars (M1) increased in the occlusal hypofunction group (Group H) but not in the control group (Group C), whilst these decreased in rats in the occlusal hypofunction group that were administered propranolol (Group HB). Bone volume/tissue volume (BV/TV), trabecular thickness (Tb.Th) and trabecular number (Tb.N) for interradicular alveolar bone in M1 in Group H were significantly lower than those in Group C, whereas those in Group HB remained as high as those in Group C. The number of TRAP-positive cells in Group H increased compared to that in Group C, whereas it significantly decreased in Group HB. CONCLUSIONS These results suggest that sympathetic nervous activity may influence the alveolar bone loss induced by occlusal hypofunction.

Soft diet causes greater alveolar osteopenia in the mandible than in the maxilla

OBJECTIVE To investigate changes in the bony microstructure of the upper and lower alveolar bone during masticatory loading induced by soft diet feeding in growing rats. DESIGN Three-week-old male Wistar rats were randomly divided into two groups. Rats were fed with either pellets [control group (n=6)] or a soft diet [experimental group (n=6)] for nine weeks. 3D-microstructure of the alveolar bone of the first molar region (M1) was examined by micro-CT analysis. RESULTS Micro-CT images showed increased marrow spaces of the inter-radicular alveolar bone around the rat mandibular M1 in the experimental group compared with that in the control group. The bone volume/tissue volume ratio, trabecular thickness, trabecular number, mean intercept length, trabecular width and trabecular star volume for the mandibular M1 inter-radicular alveolar bone were lower in the experimental group than in the control group. Marrow space star volume was increased in the experimental group compared with the control group. CONCLUSIONS These results suggest that alveolar osteopenia is more extensive in the mandible than the maxilla in rats that experience low masticatory loading during growth.

Effects of a mandibular lateral shift on the condyle and mandibular bone in growing rats.

OBJECTIVE To examine the effects of mandibular lateral shift on the growth and morphology of the condyle and mandibular bone in growing rats. MATERIALS AND METHODS A maxillary resin plate was designed to displace 4-week-old rat mandibles 2 mm to the left during closure. The plate was cemented, and the rats were killed after 2, 4, 8, or 12 weeks. Gross dimensions of the condylar head were measured. Radiographic films of the mandibles were exposed, and selected measurements were made. The newly formed bone in the condyle was evaluated with periodic acid and Schiffs reagent (PAS) staining. RESULTS The length of the condylar head was greater on the ipsilateral side compared with the contralateral side. The experimental rats developed an asymmetric mandible, shorter in horizontal dimension but longer in the vertical dimension on the ipsilateral side. The mandibular growth direction was also affected, with the mandible on the ipsilateral side growing in a more anterior and superior direction. The amount of newly formed bone was greater in the superior subchondral region but lower in the posterior subchondral region on the ipsilateral side compared with the contralateral side. CONCLUSIONS Mechanical stimuli delivered by a functional shift produce a series of morphological and histological responses in the condyle and lead to condylar and mandibular asymmetry in rats.

Glenoid Fossa Responses to Mandibular Lateral Shift in Growing Rats

OBJECTIVE To evaluate the morphological and histological responses of the glenoid fossa to mandibular lateral shift in growing rats. MATERIALS AND METHODS A resin plate was placed on the upper incisors of 4-week-old rats in the experimental groups to displace the mandible to the left during closure. The rats were killed after 2, 4, 8, and 12 weeks. The morphometric measurements were performed on dry skulls, and tissue blocks were processed for periodic acid and Schiffs reagent (PAS) staining to examine the new bone formation. RESULTS Gross measurements showed asymmetry in both the position and size of the fossae between the two sides after 4 weeks of lateral shift. The glenoid fossa on the ipsilateral side was repositioned relatively backward, outward and upward compared with the contralateral side and control group, whereas the fossa on the contralateral side was relocated relatively forward and downward compared with the control group. The length of the fossa was smaller on the ipsilateral side than on contralateral side and control group. At 2 weeks, the amount of newly formed bone in the posterior region of the fossa was higher in the experimental group than the control group. CONCLUSION It is suggested that the mandibular lateral shift causes asymmetry in the position and size of the glenoid fossa and that this phenomenon can be related to different bilateral directional new bone formation in the posterior region.

Functional Lateral Shift of the Mandible Effects on the Expression of ECM in Rat Temporomandibular Cartilage

OBJECTIVE To test the hypothesis that the effects of mechanical stress from a functional lateral shift of the mandible have no effect on the expression of two main condylar cartilage extracellular matrix components, type II collagen and aggrecan, in rats from early puberty to young adulthood. MATERIALS AND METHODS Functional lateral shift of the mandible was induced in experimental groups of 5-week-old male Wistar rats, using guiding appliances. The rats were sacrificed at 3, 7, 14, and 28 days post appliance attachment. The condyles were immunohistochemically evaluated for type II collagen and aggrecan (the immunoreactive areas were quantified). RESULTS As compared with the control group, on the contralateral condyles, the immunoreactivity of the experimental groups was significantly increased from 7 to 14 days. While on the ipsilateral condyles, the immunoreactive areas were significantly decreased throughout the experimental period. CONCLUSION A functional lateral shift of the mandible modulated the condylar cartilage extracellular matrix differently on each side of the condyle, which affected condylar morphology, growth, biomechanical properties, and even the susceptibility of the condylar cartilage to pathogenesis.

Influence of Occlusal Stimuli on the Remodelling of Alveolar Bone in a Rat Hypofunction-Recovery Model

Remodelling of alveolar bone is a crucial factor determining tooth movement against orthodontic forces. Occlusion is the most influential stimulus causing physiological remodelling of alveolar bone. A removable metal appliance was devised, which can sequentially induce hypofunctional occlusion at molar teeth and reestablish normal occlusion. Using this appliance, remodelling of the alveolar bone was examined by histological morphometric analyses, with particular attention to localized regions of bone resorption by identifying the receptor activator of NF-κB ligand (RANKL), a growth factor inducing osteoclastogenesis, and osteoclasts. In the occlusal hypofunction period, the number of TRAP-positive cells increased at the margin of marrow spaces, and the marrow space enlarged. RANKL-positive osteoblasts also increased, especially around the blood vessels. In bone morphometric parameters, BV/TV, Tb. Th and Tb. N decreased, and Tb. Sp and Oc. N increased. These results are consistent with active bone remodelling consisting of active resorption at the inner alveolar bone and bone formation on the outer alveolar bone surface. In the 3 days recovery group, many TRAP-positive cells were observed at the inner alveolar bone and around the bone surface. RANKL-positive osteoblasts were observed at the margin of the marrow space in alveolar bone. However, BV/TV, Tb. Th, and Tb. Sp returned to the control level in the 7 days recovery group, with decreased TRAP-positive cells and RANKL-positive osteoblasts. These findings suggest that occlusal stimuli play important roles in the maintenance of functional alveolar structure and the regulation of alveolar bone in rats.

Occlusal hypofunction causes periodontal atrophy and VEGF/VEGFR inhibition in tooth movement

OBJECTIVE To examine changes in microvasculature and the expression of vascular endothelial growth factor A (VEGF-A) and VEGF receptor 2 (VEGFR-2) in rat hypofunctional periodontal ligament (PDL) during experimental tooth movement. MATERIALS AND METHODS Twelve-week-old male Sprague-Dawley rats were divided into normal occlusion and occlusal hypofunction groups. After a 2-week bite-raising period, rat first molar was moved mesially using a 10-gf titanium-nickel alloy closed coil spring in both groups. On days 0, 1, 2, 3, and 7 after tooth movement, histologic changes were examined by micro-computed tomography and immunohistochemistry using CD31, VEGF-A, VEGFR-2, and the terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) method. RESULTS Hypofunctional molars inclined more than normal molars and did not move notably after day 1 of tooth movement. Blood vessels increased on the tension side of the PDL in normal teeth. Immunoreactivities for VEGF-A and VEGFR-2 in normal teeth were greater than those in hypofunctional teeth during tooth movement. Compressive force rapidly caused apoptosis of the PDL and vascular endothelial cells in hypofunctional teeth, but not in normal teeth. CONCLUSIONS Occlusal hypofunction induces vascular constriction through a decrease in the expression of VEGF-A and VEGFR-2, and apoptosis of the PDL and vascular cells occurs during tooth movement.

Histochemical localization of hyaluronan and versican in the rat molar dental pulp

The distribution of hyaluronan and versican in the dental pulp of the young rat was mapped histochemically. The pattern of staining showed considerable variation between different teeth and different specimens. The most common pattern was a strong reaction for hyaluronan and a weak reaction for versican in the subodontoblastic region, with the reverse deeper in the pulp. This was not an entirely consistent pattern and there was considerable regional variation in the staining intensity for both molecules. The localization of these molecules at similar sites could thus indicate related roles in the connective tissue matrix rather than any chemical bonding between them.

Masseter Muscular Weakness Affects Temporomandibular Synovitis Induced by Jaw Opening in Growing Rats

OBJECTIVE To evaluate the influence of impaired masseter function during growth on the development of temporomandibular synovitis. MATERIALS AND METHODS Sixteen 3-week-old male Wistar rats were classified into four groups. The first group served as control; and in the second group, jaw opening was forced for 3 hours when the rats were 9 weeks old. In the third and fourth groups, the masseter muscles were bilaterally resected at 3 weeks of age, and the rats in the fourth group were additionally forced to open their jaw at 9 weeks of age. All rats were sacrificed at 9 weeks. Temporomandibular joint (TMJ) tissue samples were processed for histology, and evaluated for cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expressions by immunohistochemistry to examine the inflammatory changes in the synovial membrane. RESULTS The control group showed noninflammatory changes. In the jaw-opening group, vascular dilation and weak COX-2 immunoreactivity were induced by jaw opening in the synovium. In the masseter-resection group, the masseter-resected rats exhibited moderate synovial changes while in the resection with opening group, the masseter-resected rats revealed more significant inflammatory changes including synovial hyperplasia, dilated vasculature, fibrin deposits, and intense immunoreactivity for COX-2 and iNOS, all caused by jaw opening. CONCLUSIONS These results suggest that masseter activity in the growth period is an important factor in the induction of temporomandibular synovitis.