Spatial and temporal distribution of CD44 and osteopontin in fracture callus.
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The multifunctional adhesion molecule CD44 is a major cell-surface receptor for hyaluronic acid (HUA). Recent data suggest that it may also bind the ubiquitous bone-matrix protein, osteopontin (OPN). Because OPN has been shown to be a potentially important protein in bone remodelling, we investigated the hypothesis that OPN interactions with the CD44 receptor on bone cells participate in the regulation of the healing of fractures. We examined the spatial and temporal patterns of expression of OPN and CD44 in healing fractures of rat femora by in situ hybridisation and immunohistochemistry. We also localised HUA in the fracture callus using biotinylated HUA-binding protein. OPN was expressed in remodelling areas of the hard callus and was found in osteocytes, osteoclasts and osteoprogenitor cells, but not in cuboidal osteoblasts which were otherwise shown to express osteocalcin. The OPN signal in osteocytes was not uniformly distributed, but was restricted to specific regions near sites where OPN mRNA-positive osteoclasts were attached to bone surfaces. In the remodelling callus, intense immunostaining for CD44 was detected in osteocyte lacunae, along canaliculi, and on the basolateral plasma membrane of osteoclasts, but not in the cuboidal osteoblasts. HUA staining was detected in fibrous tissues but little was observed in areas of hard callus where bone remodelling was progressing. Our findings suggest that OPN, rather than HUA, is the major ligand for CD44 on bone cells in the remodelling phase of healing of fractures. They also raise the possibility that such interactions may be involved in the communication of osteocytes with each other and with osteoclasts on bone surfaces. The interactions between CD44 and OPN may have important clinical implications in the repair of skeletal tissues. (+info)
Down-regulation of human osteoblast PTH/PTHrP receptor mRNA in end-stage renal failure.
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BACKGROUND: Resistance to the action of parathyroid hormone (PTH) has been demonstrated in end-stage renal failure and is considered to be important in the pathogenesis of secondary hyperparathyroidism. The mechanism of resistance is unknown. However, altered regulation of cellular PTH/PTH-related protein (PTH/PTHrP) receptor (PTH1R) has been assumed to be important. METHODS: We have used in situ hybridization to examine PTH1R mRNA expression by osteoblasts in human bone and have compared the expression in high- and low-turnover renal bone disease, high-turnover nonrenal bone disease (healing fracture callus and Pagetic bone), and normal bone. Bone biopsies were formalin fixed, ethylenediaminetetraacetic acid decalcified, and paraffin wax embedded. A 1.8 kb PTH1R cDNA probe, labeled with 35S, was used, and the hybridization signal was revealed by autoradiography. The density of signal over osteoblasts was quantitated using a semiautomated Leica image analysis software package. RESULTS: The mean density of PTH1R mRNA signal over osteoblasts in renal high-turnover bone was only 36% of that found in nonrenal high-turnover bone (P < 0.05) and 51% of that found in normal bone (P < 0.05). Osteoblast PTH1R mRNA signal in adynamic bone from individuals with diabetes mellitus was 28% of normal bone (P < 0.05) and 54% of that found in renal high-turnover bone (P < 0.05). CONCLUSIONS: These results demonstrate a down-regulation of osteoblast PTH1R mRNA in end-stage renal failure in comparison to normal and high-turnover bone from otherwise healthy individuals, and provide an insight into the mechanisms of "skeletal resistance" to the actions of PTH. (+info)
Low-intensity pulsed ultrasound accelerates bone maturation in distraction osteogenesis in rabbits.
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We investigated the effects of low-intensity pulsed ultrasound on distraction osteogenesis in a rabbit model. Callotasis of the right tibia was performed in 70 male Japanese white rabbits using mini-external fixators. In the first part of the study in 64 animals using normal distraction (waiting period seven days; distraction rate 0.5 mm/12 hours; distraction period ten days), we evaluated the distraction site by radiography, measurement of the bone mineral density (BMD), mechanical testing, and histology. In the second part in six rabbits using fast distraction (waiting period 0 days; distraction rate 1.5 mm/12 hours; distraction period seven days) the site was evaluated radiologically. Half of the animals (35) had received ultrasound to their right leg (30 mW/cm2) for 20 minutes daily after ceasing distraction (ultrasound group), while rigid fixation only was maintained in the other half (control group). With normal distraction, the hard callus area, as shown by radiography, the BMD, and the findings on mechanical testing, were significantly greater in those receiving ultrasound than in the control group. Histological analysis showed no tissue damage attributable to exposure to ultrasound. With fast distraction, immature bone regeneration was observed radiologically in the control group, while bone maturation was achieved in the ultrasound group. We conclude that ultrasound can accelerate bone maturation in distraction osteogenesis in rabbits, even in states of poor callotasis. (+info)
A new look at osteogenesis imperfecta. A clinical, radiological and biochemical study of forty-two patients.
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In a clinical, radiological and biochemical study of forty-two patients from Oxford with osteogenesis imperfecta, it was found that patients could be divided simply into mild, moderate and severe groups according to deformity of long bones. In the severe group (seventeen patients) a family history of affected members was uncommon and fractures began earlier and were more frequent than in the mild group (twenty-two patients); sixteen patients in the severe group had scoliosis and eleven had white sclerae; no patients in the mild group had white sclerae or scoliosis. Radiological examination of the femur showed only minor modelling defects in patients in the mild group, whereas in the severe group five distinct appearances of bone (thin, thick, cystic and buttressed bones, and those with hyperplastic callus) were seen. The polymeric (structural) collagen from skin was unstable to depolymerisation in patients in the severe group, but normal in amount, whereas the reverse was found in the mild group. This division according to long bone deformity may provide, a basis for future research more useful than previous classifications. (+info)
Stress-relaxation plates and the remodeling of callus and cortex under the plate in rabbits.
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OBJECTIVE: To study the influence of a stress-relaxation plate on the remodeling of callus and cortex under the plate. METHODS: The bilateral tibia diaphysis of New Zealand rabbit were osteotomized and fixed with stress-relaxation plate (SRP) and rigid plate (RP), respectively. Polarized light microscopy and transmission electron microscopy (TEM) were used to study the remodeling of callus and the cortex under the plate from 4 to 24 weeks postoperatively. RESULTS: Polarized light microscopy: the structural changes of callus and cortex beneath the plate are similar in the SRP and RP groups at the early postoperative stage, manifesting an alignment disorder of collagen fibers with a weak birefringence in the callus and absorption cavities in the cortex under the plate. After the twelfth postoperative week, the SRP group showed callus starting to transform to lamellar bone and absorption cavities in the cortex under the plate becoming smaller. By contrast in the RP group the absorption cavities in the callus and cortex under the plate became larger and the whole layer of cortex was cancellated. TEM: the active osteoclasts appeared in both SRP and RP groups in the period from 4 to 8 weeks postoperatively. In the SRP group, many functionally active osteoblasts could be seen on the surface of the bone, while in the RP group, the osteoblasts were not very active. By 24 weeks postoperatively, the shape of osteocytes were normal but the number of the osteoclasts were small in the SRP group. In the RP group, the osteoclasts became more active and osteocytic osteolysis was manifested. CONCLUSIONS: Fixation with SRP not only enhanced callus remodeling, but also abated the degree of osteoporosis in the cortex under the plate. This approach may lead to an improved osteosynthetic apparatus. (+info)
Interobserver and intraobserver variation in the assessment of the healing of tibial fractures after intramedullary fixation.
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The reliability of the radiological assessment of the healing of tibial fractures remains undetermined. We examined the inter- and intraobserver agreement of the healing of such fractures among four orthopaedic trauma surgeons who, on two separate occasions eight weeks apart, independently assessed the radiographs of 30 patients with fractures of the tibial shaft which had been treated by intramedullary fixation. The radiographs were selected from a database to represent fractures at various stages of healing. For each radiograph, the surgeon scored the degree of union, quantified the number of cortices bridged by callus or with a visible fracture line, described the extent and quality of the callus, and provided an overall rating of healing. The interobserver chance-corrected agreement using a quadratically weighted kappa (kappa) statistic in which values of 0.61 to 0.80 represented substantial agreement were as follows: radiological union scale (kappa= 0.60); number of cortices bridged by callus (kappa = 0.75); number of cortices with a visible fracture line (kappa= 0.70); the extent of the callus (kappa = 0.57); and general impression of fracture healing (kappa = 0.67). The intraobserver agreement of the overall impression of healing (kappa = 0.89) and the number of cortices bridged by callus (kappa = 0.82) or with a visible fracture line (kappa = 0.83) was almost perfect. There are no validated scales which allow surgeons to grade fracture healing radiologically. Among those examined, the number of cortices bridged by bone appears to be a reliable, and easily measured radiological variable to assess the healing of fractures after intramedullary fixation. (+info)
Diminished callus size and cartilage synthesis in alpha 1 beta 1 integrin-deficient mice during bone fracture healing.
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Integrins mediate cell adhesion to extracellular matrix components. Integrin alpha 1 beta 1 is a collagen receptor expressed on many mesenchymal cells, but mice deficient in alpha 1 integrin (alpha1-KO) have no gross structural defects. Here, the regeneration of a fractured long bone was studied in alpha1-KO mice. These mice developed significantly less callus tissue than the wild-type (WT) mice, and safranin staining revealed a defect in cartilage formation. The mRNA levels of nine extracellular matrix genes in calluses were evaluated by Northern blotting. During the first 9 days the mRNA levels of cartilage-related genes, including type II collagen, type IX collagen, and type X collagen, were lower in alpha1-KO mice than in WT mice, consistent with the reduced synthesis of cartilaginous matrix appreciated in tissue sections. Histological observations also suggested a diminished number of chondrocytes in the alpha 1-KO callus. Proliferating cell nuclear antigen staining revealed a reduction of mesenchymal progenitors at the callus site. Although, the number of mesenchymal stem cells (MSCs) obtained from WT and alpha 1-KO whole marrow was equal, in cell culture the proliferation rate of the MSCs of alpha 1-KO mice was slower, recapitulating the in vivo observation of reduced callus cell proliferation. The results demonstrate the importance of proper collagen-integrin interaction in fracture healing and suggest that alpha1 integrin plays an essential role in the regulation of MSC proliferation and cartilage production. (+info)
Ultrasound stimulation of maxillofacial bone healing.
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A substantial part of the maxillofacial surgery practice deals with maxillofacial bone healing. In the past decades, low-intensity ultrasound treatment has been shown to reduce the healing time of fresh fractures of the extremities up to 38%, and to heal delayed and non-unions up to 90% and 83%, respectively. Based on the assumption that the process of bone healing in the bones of the extremities and maxillofacial skeleton is essentially the same, the potential of ultrasound to stimulate maxillofacial bone healing was investigated. Although limited evidence is available to support the susceptibility of maxillofacial bone to the ultrasound signal, ultrasound may be of value in the treatment of delayed unions, in callus maturation after distraction, and in the treatment of osteoradionecrosis. (+info)