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Remodeling of bone and bones: effects of altered mechanical stress on the regeneration of transplanted bones.

We divided 116 rats weighing 50 gm into four groups with tails either left in situ or transplanted as follows: straight in situ: untreated controls; bent in situ: five caudal vertebrae (CV) in the loop; straight transplants: three CV skinned and transplanted autologously; and bent transplants: five CV skinned, bent to form a loop, and transplanted autologously. Tails were radiographed weekly up to 6 weeks and at 12 weeks, and microradiographic and histological studies were undertaken on selected specimens. At 12 weeks the bones in the apex of the loop of tails left in situ appeared bent with a straight-to-convex shaft on the outer side and a thicker, more concave one on the inner side. In the transplanted bent segments the bone shaft died and initially the reverse occurred: the outer shaft thickened and the inner resorbed completely. A new concave inner diaphysis then formed so that the bones in both instances were essentially similar in final shape. In the bent transplants the surviving osteogenic tissues regenerated and, adapting to the altered forces, formed a new bone shaft. This involved a change in the direction, amount, and nature of endochondral, periosteal, and regenerative growth and subsequent remodeling of bone. The results support previous observations that, within limits, the strain in the osteogenic envelope is an important factor in adaptation of bones to changing stress and that, where the envelope is deficient, the surviving tissues have the capacity to regenerate and repair defects in the bone so that it best resists the changing stresses applied to it.

Animals↗

Why does bone matrix contain non-collagenous proteins? The possible roles of osteocalcin, osteonectin, osteopontin and bone sialoprotein in bone mineralisation and resorption.

Four major non-collagenous bone proteins were localised by single and double immuno- histochemistry during de novo mineralisation and bone resorption. Both osteopontin and bone sialoprotein were localised ahead of the mineralisation front, suggesting that both proteins are necessary for the initiation of bone mineralisation. This supports previous suggestions that bone sialoprotein acts as a crystal nucleator. The role of osteopontin is less certain, but might be related to ensuring that only the right type of crystal is formed. Osteocalcin and osteonectin were not present in areas of first crystal formation, but were present in the fully mineralised matrix. Their role may be to control the size and speed of crystal formation. Osteopontin, bone sialoproteins and osteocalcin (but not osteonectin) were also present at reversal lines. Interpreting this localisation together with information from the literature, the following functions are suggested during resorption: Osteocalcin may act as a chemoattractant for osteoclasts, while both osteopontin and bone sialoprotein may facilitate the binding of osteoclasts via the arg-gly-asp motif.

Animals↗

Disruption of four-and-a-half LIM 2 decreases bone mineral content and bone mineral density in femur and tibia bones of female mice.

Four-and-a-half LIM 2 (FHL2) is a member of a family of LIM domain proteins which mediate protein-protein interactions. FHL2 acts as a coactivator and binds to important regulators of bone formation such as insulin-like growth factor binding protein (IGFBP)-5, androgen receptor, and beta-catenin. We hypothesized that FHL2 is an important regulator of bone formation. We evaluated growth and skeletal parameters in FHL2 knockout (KO) and wild-type (WT) mice at 4, 8, and 12 weeks of age. At 4 weeks of age, lack of FHL2 reduced femur, tibia, and total bone mineral content (BMC) and body weight in all mice. A gender-by-treatment interaction (P <or= 0.05) was observed for several parameters due to a greater reduction in females. Specifically, femur BMC was reduced 11-27% at 8 and 12 weeks of age and BMD was reduced 7-13% at all ages in female KO mice (P < 0.05). A similar reduction was observed in the tibias at 8 weeks of age. A 6% reduction (P = 0.07) in femur cortical thickness was observed at 12 weeks of age in female KO mice. Interestingly, a gender-specific reduction in IGFBP-5 expression was observed in the femurs of female KO mice. During differentiation of bone marrow stromal cells into osteoblasts, expression of osteocalcin, alkaline phosphatase, and bone sialoprotein was reduced 47-96% in FHL2 KO cells (P < 0.001). In conclusion, FHL2 is an important regulator of peak bone mass, lack of FHL2 produces gender- and site-specific effects on bone accretion and IGFBP-5 expression, and FHL2 is important for optimal osteoblast differentiation in vitro.

Animals↗

The biphasic effect of triiodothyronine compared to bone resorbing effect of PTH on bone modelling of mouse long bone in vitro.

To examine the effects of T3 on fetal long bone modelling the radii and ulnae of 16 day old fetal mice were grown in vitro for two days. Their growth, mineralization, and resorption were assessed by measuring diaphyseal length, calcium and phosphorus content, hydroxyproline content, and the release of incorporated 45Ca. The effects of T3 were compared to the effects of 1-34 PTH, a known resorbing agent, on the same system. Devitalized bones were used as a control. The results showed that T3 had a biphasic effect. At high concentrations (10(-5) M-10(-6) M) T3 inhibited the growth of the bones as indicated by their diaphyseal length and hydroxyproline content. Calcium and phosphorus content were significantly decreased while 45Ca release was increased. Similar effects were also found after the addition of 1-34 PTH to the media. However, T3, at lower concentrations (10(-7) M-10(-9) M), stimulated the growth and calcification of the bones as indicated by an increase in diaphyseal length and the hydroxyproline, calcium, and phosphorus content. 45Ca release was significantly decreased at these concentrations. Neither T3 nor 1-34 PTH affected devitalized bones in the same system. The results suggest that at physiological concentrations, T3 has a direct, anabolic effect on bone, which may explain its major role in the growth process of various species. At high doses, however, T3 stimulates bone resorption in a way similar to PTH.

Animals↗

Induction of new bone by ceramic bovine bone with recombinant human bone morphogenetic protein 2 and transforming growth factor beta.

Bone morphogenetic protein (BMP) has the ability to induce ectopic bone, while the action of transforming growth factor beta (TGF beta) is to stimulate proliferation of osteoblasts and chondrocytes as well as the production of extracellular matrix. The aim of the present study was to study their synergistic actions in bone formation. Three kinds of complexes, recombinant human BMP2 (rhBMP2), TGF beta and rhBMP2/TGF beta in ceramic bovine bone (CBB), were made and then implanted into the thigh muscle pouches of mice. The histological reactions of the implanted areas were studied at intervals of 3, 5, 7, 14 and 21 days. The results showed that, except for the implants with TGF beta alone, both rhBMP2 and rhBMP2/TGF beta implants exhibited new ectopic bone formation. The morphometric study revealed that the quantity of newly formed bone induced by rhBMP2/TGF beta was obviously greater than by rhBMP2 alone. These results indicate that TGF beta in combination with BMP may enhance formation of ectopic bone.

Animals↗

Osteoporosis and bone functional adaptation: mechanobiological regulation of bone architecture in growing and adult bone, a review.

During life, bone is continually optimized for its load-bearing role by a process of functionally adaptive (re)modelling. This process, which is more active in growing bone, is dominated by high-magnitude, high-rate strains, presented in an unusual distribution. Adaptation occurs at an organ level, involving changes in whole bone architecture and bone mass. The repetitive coordinated bone loading associated with habitual activity may have little role in the preservation of bone mass, and may even reduce the osteogenic potential of an otherwise highly osteogenic stimulus. Cells of the osteocyte/osteoblast network are best placed to appreciate mechanical strain. Among the strain-related responses they show, is a reduced rate of apoptosis. This may serve to regulate and target osteoclast activity. A more complete understanding of the stimuli and pathways involved in both the physiology and pathology of this structural homeostatic mechanism will allow the design of more appropriate exercise regimens and targeted pharmacological interventions to limit morbidity and mortality by reducing bone fragility.

Adaptation, Physiological↗

Repair of segmental bone defects using bioactive bone cement: comparison with PMMA bone cement.

We developed a bioactive bone cement (BABC) that consists of apatite and wollastonite containing glass ceramic (AW-GC) powder and bisphenol-A-glycidyl dimethacrylate (Bis-GMA) based resin. In the present study, the effectiveness of the BABC for repair of segmental bone defects under load-bearing conditions was examined using a rabbit tibia model. Polymethylmethacrylate (PMMA) bone cement was used as a control. A 15-mm length of bone was resected from the middle of the shaft of the tibia, and the tibia was fixed by two Kirschner wires. The defects were replaced by cement. Each cement was used in 12 rabbits; six rabbits were sacrificed at 12 and 25 weeks after surgery, and the tibia containing the bone cement was excised and tension tested. At both the intervals studied, the failure loads of the BABC were significantly higher than those of the PMMA cement. The BABC was in direct contact with bone, whereas soft tissue was observed between the cement and bone in all PMMA cement specimens. Results indicated that the BABC was useful as a bone substitute under load-bearing conditions.

Animals↗

Bone mineral density in children and adolescents with juvenile diabetes: selective measurement of bone mineral density of trabecular and cortical bone using peripheral quantitative computed tomography.

Bone mineral density (BMD) was studied in 21 children and adolescents with type I diabetes and in age- and sex-matched healthy controls. BMD was selectively measured in trabecular and total bone using peripheral quantitative computed tomography (pQCT). Cortical bone density was calculated. There was a decrease of trabecular bone density (-18.9%, p < 0.01), total bone density (-9.0%, NS) and cortical bone density (-5.1%, NS) in diabetes. Trabecular bone density was inversely correlated with the duration of diabetes and the concentration of glycosylated hemoglobin (HbA1) (r = -0.48, p = 0.027 and r = -0.63, p = 0.002, respectively). Total BMD correlated inversely with HbA1 (r = -0.52, p = 0.017). pQCT allows the selective measurement of metabolically active trabecular bone where changes of mineralization first occur. We conclude that pQCT is a useful method for investigating BMD in diabetes.

Adolescent↗

Bone-demineralized bone-bone graft for ligament reconstruction in rats.

Bone-tendon-bone (B-T-B) type grafts were prepared for ligament reconstruction by harvesting the radius from rats, wrapping both ends of each bone with a laboratory film leaving only the central 10 mm exposed, and demineralizing the central part by immersing the bone in 1.75% HCl solution. In the grafts prepared, the central part of the bone became semi-translucent and flexible while the ends remained as hard bone tissue, thus forming a B-T-B type graft. The tensile strength of the grafts was greater than that of the medial collateral ligaments of the rats and about the same as of their anterior cruciate ligaments. No inflammation or other adverse reaction was noticed in experimental subcutaneous transplantation and the grafts showed excellent biocompatibility. In experimental ligament reconstruction, the test animals did not show any impairment on gait. There was invasion of fibroblasts into the graft at 4 weeks, and the fibroblasts were found through the whole graft and what looked like ligament tissue could be seen macroscopically at 8 weeks. Besides, bone tissue had infiltrated into the inside cavity of the non-demineralized part of the graft and proliferation, resulting in good bone union. The results obtained suggest that the grafts prepared in this study have a sufficient potential as B-T-B type grafts for ligament reconstruction.

Animals↗

Immunohistology of bone proteins, bone quality, and bone turnover.

The role played by noncollagenous proteins in bone metabolism, originally conceived as mainly related to the promotion and regulation of matrix mineralization, probably involves a number of regulatory functions. Local composition of the bone matrix may affect cell responses to regulatory agents in different ways, which include binding of growth factors within the pericellular environment, and integrins on the cell surface. The focus of studies on the expression and localization of bone proteins in intact tissues should therefore move towards a detailed dissection of heterogeneity of matrix locales throughout bone development, growth, and postnatal remodeling. So far, such studies have provided valuable information on the diverse phenotypic profiles of maturational stages of bone cells, and more recently, on actual differences in secretory output of cells otherwise comprised in the 'mature' osteoblastic compartment. Still, a main gap in current information is represented by the paucity of data on bone protein expression and deposition during post-natal remodeling and its diseases. Filling this gap will require a more focused interest in post-natal (trabecular) bone, and the development of appropriate protocols of tissue preparation (noncollagenous) proteins in bone physiology.

Bone Matrix↗

Commencing, continuing and stopping brisk walking: effects on bone mineral density, quantitative ultrasound of bone and markers of bone metabolism in postmenopausal women.

Regular walking is associated with reduced risk of fracture and, in our recent randomized trial, reduced calcaneal bone loss relative to controls. The present follow-up study compared the effects on dual-energy X-ray absorptiometry, ultrasound and biochemical indices of bone density and metabolism of (i) taking up (ii) continuing with and (iii) ceasing brisk walking for exercise. Subjects were 68 postmenopausal women aged 60-70 years. Twenty previously sedentary women remained sedentary (Sed/Sed) whilst 17 took up brisk walking (Sed/Walk). Fifteen women who had been walking regularly for 1 year returned to their former sedentary lifestyle (Walk/Sed), whilst 16 continued brisk walking over a second year (Walk/Walk). Bone mineral density (BMD), broadband ultrasonic attenuation (BUA), and biochemical markers of bone formation (serum osteocalcin, C-terminal propeptide of type I collagen and bone alkaline phosphatase) and resorption (urinary deoxypyridinoline) were assessed at baseline and 12 months. Women in the Sed/Walk and Walk/Walk groups completed a mean (SEM) of 16.9 (0.7) and 20.8 (1.2) min of brisk walking per day, respectively. Changes in BMD did not differ significantly between groups. Calcaneal BMD decreased significantly in Walk/Sed women [by 2.7 (1.4)%; p = 0.01] whilst changes in other groups were not significant. Calcaneal BUA increased significantly (p = 0.02) in Sed/Walk women [by 7.4 (3.3)%] relative to other groups. Urinary deoxypyridinoline increased over the year in the Sed/Sed group but there were no significant changes in biochemical markers in other groups. Women taking up brisk walking for exercise showed no change in BMD but a significant increase in calcaneal BUA. There was no significant effect on BMD or BUA of continuing brisk walking but calcaneal BMD declined on ceasing brisk walking. Bone resorption increased in sedentary women but not exercisers, suggesting the effect on exercise on bone in postmenopausal women could be through amelioration of this increased turnover.

Absorptiometry, Photon↗

Direct action of 1,25-dihydroxyvitamin D on bone: VDRKO bone shows excessive bone formation in normal mineral condition.

In the present study, the direct role of Vitamin D in bone metabolism was investigated. Vitamin D has been suggested to be an important hormone for bone metabolism, but there has been little evidence that Vitamin D actively participates in this process. Here, we show the direct action of Vitamin D by transplanting the bone of the Vitamin D receptor null mutant mice (VDR-/-) to the wild-type mouse. This procedure allowed us to investigate the changes in the bone without VDR in the normal humoral environment. Unexpectedly, the volume and the density of the VDR-/- bone transplanted to the wild-type mouse were significantly increased compared with the control (wild-type bone transplanted to the wild-type mouse). We show that Vitamin D has key roles in bone metabolism negatively.

Alkaline Phosphatase↗

Decreased trabecular bone mineral density in children with idiopathic short stature: normalization of bone density and increased bone turnover after 1 year of growth hormone treatment.

Patients with growth hormone (GH) deficiency have impaired bone mineral metabolism; treatment with GH leads to an improvement in their bone mineral density (BMD). The effect of GH on the BMD of children with idiopathic short stature is unknown. We studied 14 short, slowly growing, otherwise healthy, prepubertal children without GH deficiency (7 girls and 7 boys) with a chronological age of 10.9 +/- 1.4 years, bone age of 8.8 +/- 1.5 years, and height of 127.8 +/- 8.5 cm (height SD score of -2.2 +/- 0.5). Growth velocity increased from 3.9 +/- 1.1 cm/y to 8.0 +/- 1.9 cm/y, and height SD score improved from -2.2 +/- 0.5 to -1.8 +/- 0.5 after 12 months of GH treatment (P <.007 and P <.001, respectively). Baseline lumbar spine BMD was decreased when compared with that of a control group of children matched for bone age and height (0.645 +/- 0.09 g/cm(2) vs 0.730 +/- 0.08 g/cm(2); P <.003). Lumbar spine BMD increased in subjects with ISS after 1 year of GH treatment from 0.645 +/- 0.09 g/cm(2) to 0.808 +/- 0.04 g/cm(2) (P <.05), reaching levels similar to those of control subjects, followed up without therapy (0.808 +/- 0.04 g/cm(2) vs 0.760 +/- 0.08 g/cm(2)); lumbar spine BMD increased 25.3% in the subjects with ISS and 4.1% in the control subjects. Femoral neck BMD did not change during treatment. Serum concentrations of the carboxy-terminal propeptide of type 1 collagen increased from 231.6 +/- 65.5 microg/L to 351.6 +/- 87.2 microg/L, and levels of the carboxy-terminal cross-linked telopeptide of type 1 collagen increased from 9.9 +/- 5.9 microg/L to 13.9 +/- 2.4 microg/L. Children with ISS have decreased lumbar spine BMD, which increases with GH therapy, reaching levels similar to those of control subjects. Bone turnover increased as indicated by a rise in bone formation and bone resorption markers.

Biomarkers↗

Transforming growth factor beta (TGF-beta) levels in the conditioned media of human bone cells: relationship to donor age, bone volume, and concentration of TGF-beta in human bone matrix in vivo.

Transforming growth factor-beta (TGF-beta) is thought to play an important role in human bone remodeling. In the present study, we examined constitutive differences in TGF-beta levels in primary bone cell cultures from the iliac crest of 112 women, aged 28-79 years. TGF-beta1 was the major TGF-beta isoform in the conditioned media, as determined by neutralizing TGF-beta activity with specific antibodies against TGF-beta1-3 in the mink lung cell bioassay, and by enzyme-linked immunoassay (ELISA). TGF-beta1 levels in the conditioned media did not change with donor age. There was a lack of association between TGF-beta levels in vitro and the concentration of matrix-associated TGF-beta in vivo. TGF-beta1 levels failed to be associated with the local trabecular bone volume in the complete study population (r = +0.15, p = 0.16, n = 89). A significant association between TGF-beta1 levels and bone volume was present in premenopausal women (r = +0.39, p = 0.02, n = 33), but was largely accounted for by the two samples with the highest TGF-beta concentrations. In conclusion, our data suggest that TGF-beta1 is the major TGF-beta isoform produced by human bone cells in vitro, and that the constitutive secretion of TGF-beta by bone cells does not change with age. Whether constitutive differences in TGF-beta secretion may be a determinant of human bone mass remains to be clarified in further studies.

Adult↗