No influence of alimentary zinc on the healing of calvarial defects filled with osteopromotive substances in rats.

Zinc has been demonstrated to play an important role in bone metabolism and is required for normal growth. However, no studies have investigated the influence of zinc on calvarial bone healing in aged or adult rats. The aim of the study was to evaluate whether alimentary zinc supplementation and depletion affect bone healing of calvarial defects implanted with osteopromotive substances in adult rats. Two 5 mm full thickness critical size bone defects were trephined in the central part of each parietal bone of 60 six-month-old male Wistar rats. The bone defects were filled with demineralized bone matrix (DBM), autogenous bone chips, or were left as unfilled controls. The rats were divided into three groups of 20 rats each and received a semi-synthetic diet containing 20, 60, or 120 mg zinc/kg. After 4 months, the biomechanical integrity of the healing defects was evaluated by a punch out test and the healed defects were examined with histomorphometry. Statistical analysis of the data was carried out by two-way analysis of variance and Wilcoxon's non-parametric signed rank test. Biomechanical testing revealed that the maximum load was significantly higher in DBM-filled defects than in those filled with autogenous bone, and that the defects filled with autogenous bone were stronger than the unfilled controls. The biomechanical findings indicated that the alimentary zinc content did not influence the healing of calvarial defects. No significant difference in maximum load could be established between the three diet groups for any of the filling materials, whereas the highest zinc supplement resulted in an increase in the relative extension on mineralizing surfaces in the control group. Thus, healing of adult rat calvarial defects is not influenced by alimentary zinc supplementation or depletion. Defects filled with DBM were significantly stronger and exhibited significantly more new bone formation than defects filled with autogenous bone or unfilled controls.

Biomechanical evaluation of rat skull defects, 1, 3, and 6 months after implantation with osteopromotive substances.

PURPOSE: To compare the mechanical strength of surgically created and healed rat calvarial defects having been filled with three different osteopromotive substances: hydroxyapatite, intramembraneous demineralised bone matrix (DBM), and autogenous bone chips. MATERIAL: Sixty adult male Wistar rats were divided into three groups of 20 animals, each group representing healing times of one, three, or six months. METHODS: Identical 5mm bilateral critical size defects were trephined into the parietal bones and hydroxyapatite, DBM, or autogenous bone chips were implanted into the defects, or left as unfilled controls. The repaired defects were evaluated biomechanically using a modified punch out test 1, 3, or 6 months postoperatively. RESULTS: The maximum load carried in the DBM group was significantly higher than in the bone chips, hydroxyapatite, and control groups after one month of healing. The mean bone strength did not increase significantly after the first month of healing in any of the groups. The DBM group showed a significantly higher load to failure than the other groups at all three observation periods. The mean maximum load to failure of the bone chips and hydroxyapatite groups was slightly higher than in the control group; however the difference was not statistically significant. None of the control experimental bone defects demonstrated any bone formation. CONCLUSION: Defects filled with hydroxyapatite and bone chips were not significantly stronger than unfilled controls, whereas defects filled with DBM were significantly stronger than all other defects after 1, 3, and 6 months of healing.

An expression relating breaking stress and density of trabecular bone.

Bone mineral density (BMD) is the principal diagnostic tool used in clinical settings to diagnose and monitor osteoporosis. Experimental studies on ex vivo bone samples from multiple skeletal locations have been used to propose that their breaking stress bears a power-law relationship to volumetric BMD, with a location-dependent index. We argue that a power-law cannot represent effects of trabecular removal, which is one of the leading causes of reduction in bone strength. A new expression, proposed on the basis of theoretical and numerical analysis of a mathematical model, is tested using previously published data on bone samples from iliac crest and vertebral body. It represents the experimental biomechanical data at least as well as the power-law, and provides means for extrapolating results from small biopsy samples to an entire bone. In addition, changes caused by trabecular thinning and anisotropy can be modeled by the expression.

Mice deficient in 11beta-hydroxysteroid dehydrogenase type 1 lack bone marrow adipocytes, but maintain normal bone formation.

Glucocorticoids (GCs) exert potent, but poorly characterized, effects on the skeleton. The cellular activity of GCs is regulated at a prereceptor level by 11beta-hydroxysteroid dehydrogenases (11betaHSDs). The type 1 isoform, which predominates in bone, functions as a reductase in intact cells and regenerates active cortisol (corticosterone) from circulating inert 11-keto forms. The aim of the present study was to investigate the role of this intracrine activation of GCs on normal bone physiology in vivo using mice deficient in 11betaHSD1 (HSD1(-/-)). The HSD1(-/-) mice exhibited no significant changes in cortical or trabecular bone mass compared with wild-type (Wt) mice. Aged HSD1(-/-) mice showed age-related bone loss similar to that observed in Wt mice. Histomorphometric analysis showed similar bone formation and bone resorption parameters in HSD1(-/-) and Wt mice. However, examination of bone marrow composition revealed a total absence of marrow adipocytes in HSD1(-/-) mice. Cells from Wt and HSD1(-/-) mice exhibited similar growth rates as well as similar levels of production of osteoblastic markers. The adipocyte-forming capacity of in vitro cultured bone marrow stromal cells and trabecular osteoblasts was similar in HSD1(-/-) and Wt mice. In conclusion, our results suggest that 11betaHSD1 amplification of intracellular GC actions in mice may be required for bone marrow adipocyte formation, but not for bone formation. The clinical relevance of this observation remains to be determined.

Comparison of feline nasal cavity dimensions measured by acoustic rhinometry and nasal casts.

BACKGROUND: The goal of this study was to evaluate the relationship between feline nasal cavity geometry determined in vivo by acoustic rhinometry (AR(in vivo)) and by nasal cavity casts. Cast cross-sectional areas were measured by acoustic rhinometry (AR(cast)), a fluid-displacement method (FDM), and slicing. A volume comparison between AR(in vivo) and AR(cast) was studied in cats with varying degrees of nasal obstruction after application of phenylpropanolamine, saline, or compound 48/80. METHOD: After measurements of AR(in vivo), impression material was injected into the nasal cavity to produce casts. Subsequently, the cross-sectional areas of the nasal impressions were measured by AR(cast) and FDM using ethanol. All casts were weighed to determine exact volume. Six casts also were sliced into segments of equal thickness for determination of cross-sectional area. RESULTS: Cast volume determined by AR(cast) was consistent with results obtained using FDM and weight. Volumes of the first 3 cm determined by AR(in vivo) ranged between 78 +/- 9% of cast volumes determined by AR(cast) for decongested cavities and 16 +/- 15% for congested cavities. CONCLUSION: AR(in vivo) does not reflect cast geometry, probably because of (1) underestimation by AR because of methodological problems caused by the cavity geometry, (2) deformation of compliant structures within the nasal passageways resulting from the casting procedure, and/or (3) the casting material reaches parts of the nasal cavity not accessible to sound, e.g., sinuses or recesses. Nevertheless, this study does not preclude the use of AR as a sensitive method suited to evaluate relative changes in nasal volume caused by experimental challenges of the nasal mucosa. Compared with AR(in vivo), casts still may be of use but it is less sensitive to measure relative changes after experimental challenge.

Age-related changes in cortical bone content of insulin-like growth factor binding protein (IGFBP)-3, IGFBP-5, osteoprotegerin, and calcium in postmenopausal osteoporosis: a cross-sectional study.

Serum GH and IGF-I levels decline with increasing age, whereas osteoprotegerin (OPG) increases. IGFs as well as OPG are present in bone matrix and mediate the effects of many upstream hormones (e.g. estrogen). To evaluate whether changes in these proteins may to some extent explain the decrease in bone mass in postmenopausal or senile osteoporosis, we measured bone contents of IGF-I, IGF-II, IGF binding protein (IGFBP)-3, IGFBP-5, and OPG in combined extracts obtained after EDTA and guanidine hydrochloride extraction in 60 postmenopausal women aged 47-74 (mean, 63) yr with a previous distal forearm fracture and a hip or spine Z-score less than 0. We found age-related increases in IGFBP-3 (r = 0.35; P < 0.01), IGFBP-5 (r = 0.59; P < 0.001), and OPG (r = 0.36; P < 0.01) in cortical bone, significantly inversely correlated with femoral neck and lumbar spine BMD. A correlation between age and OPG was also detected in trabecular bone (r = 0.27; P < 0.05). A pronounced age-related decrease in cortical calcium contents (r = -0.60; P < 0.001), positively correlated with femoral neck and lumbar spine BMD, was also found. No age-related changes were detected for IGF-I or IGF-II. The present study demonstrates age-related changes in cortical bone contents of IGFBPs, calcium, and OPG, possibly related to the pathophysiology of postmenopausal osteoporosis. As for OPG, our findings probably represent compensatory responses to increased osteoclastic resorption.

The structural and hormonal basis of sex differences in peak appendicular bone strength in rats.

To identify the structural and hormonal basis for the lower incidence of fractures in males than females, sex differences in femoral mid-shaft geometry and breaking strength were studied in growth hormone (GH)-replete and -deficient male and female rats. Sexual dimorphism appeared during growth. Cortical thickening occurred almost entirely by acquisition of bone on the outer (periosteal) surface in males and mainly on the inner (endocortical) surface in females. By 8 months of age, males had 22% greater bone width and 33% greater breaking strength than females. Gonadectomy (Gx) at 6 weeks reduced sex differences in bone width to 7% and strength to 21% by halving periosteal bone formation in males and doubling it in females. Gx had no net effect on the endocortical surface in males but abolished endocortical bone acquisition in females. GH deficiency halved periosteal bone formation and had no net effect on the endocortical surface in males, but abolished bone acquisition on both surfaces in females, leaving males with 17% greater bone width and 44% greater breaking strength than females. Sex hormone deficiency produces greater bone fragility in males than females by removing a stimulator of periosteal growth in males and removing an inhibitor of periosteal growth in females. GH deficiency produces less bone fragility in males than females because males retain androgen-dependent periosteal bone formation while bone acquisition on both surfaces is abolished in females. Thus, periosteal growth is independently and additively stimulated by androgens and GH in males, inhibited by estrogen, and stimulated by GH in females. The hormonal regulation of bone surfaces establishes the amount and spatial distribution of bone and so the sexual dimorphism in its strength.

Polarized light microscopic analyses of collagen fibers in the rat incisor periodontal ligament in relation to areas, regions, and ages.

We prepared decalcified sagittal sections (20 microm thick) from the incisal, middle, and basal regions of the mandibular incisor of male Wistar rats aged 2, 6, 12, and 24 months, and examined the sections using polarized light microscopy. Most of the birefringent fibers appeared to run obliquely across the periodontal ligament. Birefringent fibers running parallel to the long axis of the incisor were also found in the intermediate area of the ligament. Similar fiber architecture was observed in all four age groups. Quantitative analysis showed that the retardation values of collagen were higher in the bone- and tooth-related areas and lower in the intermediate area of the ligament. The values for the bone- and tooth-related areas increased from the basal toward the incisal regions in all four age groups. Age-related changes in the retardation values were found only in the incisal region of the incisor. In the incisal region, the values for the bone- and tooth-related areas increased markedly from 2-24 months of age, whereas those for the intermediate area increased slightly but significantly with age. Our findings indicate that the degrees of molecular organization and alignment of collagen fibers in the bone- and tooth-related areas of the ligament are higher than those in the intermediate area and increase near the incisal region and with age. It is also suggested that the collagen fibers in the intermediate area remain immature along the long axis of the incisor throughout the life span of the animal.

Prevention of bone loss in ovariectomized rats by combined treatment with risedronate and 1alpha,25-dihydroxyvitamin D3.

Bisphosphonates inhibit bone loss through inhibition of osteoclast-mediated bone resorption. At low doses, vitamin D metabolites can prevent bone loss in models of osteopenia in rats by an antiresorptive effect, while at high doses they also stimulate osteoblast activity and show an anabolic effect. Therefore, combined therapy with bisphosphonates and vitamin D analogs might be expected to be more effective than either treatment alone. It was the aim of this study to compare the efficacy of risedronate and of the naturally occurring vitamin D hormone 1alpha,25-dihydroxyvitamin D3 (calcitriol), alone and in combination, for the prevention of ovariectomy-induced bone loss in rats. One hundred ten female 4-month-old Sprague-Dawley rats were used for this experiment. Ninety rats were bilaterally ovariectomized (OVX), 10 rats were sham-operated (SHAM), and 10 rats were killed at the time of surgery as a baseline control. Groups of rats (10 rats/group) received vehicle or daily doses of 0.1 mg or 0.5 mg of risedronate or 0.05 microg or 0.1 microg of calcitriol/kg body weight, alone and in combination. Both compounds were administered orally via gavage, commencing on the day after surgery. Although estrogen deficiency-induced bone loss was prevented by individual prophylactic administration of risedronate or calcitriol, OVX rats treated with a combination of risedronate and calcitriol had higher bone mineral density (BMD), cancellous bone area (B.Ar), and bone strength in long bones and vertebrae compared with rats receiving risedronate alone. Furthermore, calcitriol enhanced the suppressive effects of risedronate on osteoclast number and partially counteracted the suppressive effects of risedronate on bone formation and histomorphometric indices of osteoblast team performance. Risedronate did not reduce the anabolic effect of calcitriol, and at the high dose it normalized hypercalcemia in calcitriol-treated OVX rats. Therefore, this study in OVX rats suggests that combined therapy with bisphosphonates and vitamin D analogs may offer advantages over the treatment with bisphosphonates or vitamin D analogs alone.

Combination of topological parameters and bone volume fraction better predicts the mechanical properties of trabecular bone.

Trabecular bone structure may complement bone volume/total volume fraction (BV/TV) in the prediction of the mechanical properties. Nonetheless, the direct in vivo use of information pertaining to trabecular bone structure necessitates some predictive analytical model linking structure measures to mechanical properties. In this context, the purpose of this study was to combine BV/TV and topological parameters so as to better estimate the mechanical properties of trabecular bone. Thirteen trabecular bone mid-sagittal sections were imaged by magnetic resonance (MR) imaging at the resolution of 117 x 117x 300 microm(3). Topological parameters were evaluated in applying the 3D-line skeleton graph analysis (LSGA) technique to the binary MR images. The same images were used to estimate the elastic moduli by finite element analysis (FEA). In addition to the mid-sagittal section, two cylindrical samples were cored from each vertebra along vertical and horizontal directions. Monotonic compression tests were applied to these samples to measure both vertical and horizontal ultimate stresses. BV/TV was found as a strong predictor of the mechanical properties, accounting for 89-94% of the variability of the elastic moduli and for 69-86% of the variability of the ultimate stresses. Topological parameters and BV/TV were combined following two analytical formulations, based on: (1) the normalization of the topological parameters; and on (2) an exponential fit-model. The normalized parameters accounted for 96-98% of the variability of the elastic moduli, and the exponential model accounted for 80-95% of the variability of the ultimate stresses. Such formulations could potentially be used to increase the prediction of the mechanical properties of trabecular bone.