Effect of Ovariectomy, Malnutrition and Glucocorticoid Application on Bone Properties in Sheep: A Pilot Study

The demographic changes in the human population continue to lead to an increasing incidence of osteoporosis. The main clinical symptom of osteoporosis is fracture. Fracture fixation in osteoporosis is frequently complicated by failure of fixation. There is a great need for a large-animal model of osteoporosis for controlled studies, which allows the investigation of fracture healing and fracture treatment in weak bone. Eight swiss mountain sheep, 7–9 years old, were divided into four treatment groups of two animals each. Group 1 was ovariectomized and fed a calcium/vitamin D-restricted diet (O+D). Group 2 was ovariectomized and given a daily intramuscular injection of 25 mg methylprednisolone (O+S). Group 3 was ovariectomized, fed a calcium/vitamin D-restricted diet and injected with 25 mg intramuscular methylprednisolone per day (O+D+S). Group 4 was used as an untreated, not sham operated control group. At the beginning of the study and every 2 months for 6 months the bone mineral density (BMD) was determined using quantitative computed tomography (pQCT) at the distal radius. Biopsies were taken after 6 months from vertebral bodies and femoral heads and the bone structure, i.e. trabecular thickness (Tb.Th), trabecular number (Tb.N), trabecular separation (Tb.Sp), bone surface fraction (BS/BV) and bone volume fraction (BV/TV), was determined by micro-CT. In vitro compression testing of the biopsies was performed to determine failure load and stiffness. The control group showed no changes in BMD. The greatest decrease in BMD was seen in group 3 (O+D+S), which had a decline of 58% in cancellous bone and 22% in cortical bone. In the vertebral body biopsies a prominent change in structural parameters was observed (Tb.N, –53%; Tb.Th, –63%, Tb.Sp, +150%). The changes were less pronounced in the femoral head biopsies. In the compression test the vertebral body biopsies of group 3 (O+D+S) had stiffness values 40% lower failure load 70% lower compared with the control group. The most effective method of inducing osteoporosis in sheep was found to be the combined treatment. These results need to be confirmed in a larger number of animals. Received: 4 May 2001 / Accepted: 13 December 2001     

Computed Tomography Image Analysis of the Calcaneus in Male Osteoporosis

The present study aimed to characterize bone microarchitecture assessed by computed tomography (CT) at the calcaneus in male subjects suffering from osteoporosis. Seventy-nine subjects were assessed (45 with osteoporosis and 34 control subjects matched for age). Osteoporosis was defined according to the World Health Organization classification either at the lumbar spine or at the femoral neck. Thirty-three subjects (73%) had a past history of low-energy fracture mainly represented by vertebral fractures (24/33). Nine axial sections (1 mm in width and 2 mm apart) were selected for each subject. Bone microarchitecture analysis was performed using structural (binary and skeletonized images but also skeletonization from gray levels) and fractal analyses. Bone densitometry by dual-energy X-ray absorptiometry (DXA) at the calcaneus was also performed in 73 cases. Bone mineral density (BMD) was decreased in osteoporotic patients compared with controls both at the lumbar spine and hip and also at the calcaneus (p<0.01). Also 14 microarchitectural features among 25 measured were significantly different between the two groups (p<0.01). The odds ratio for fracture per 1 control group standard deviation decrease were also significant for 13 structural features but also for BMD at the calcaneus. The odds ratios after adjustment for BMD at the calcaneus were significant for the following features (p<0.05): number of valleys, 2.8 (1.2–6.9); trabecular partition, 3.3 (1.3–7.9); apparent trabecular spacing, 1.8 (1.0–3.1); trabecular bone pattern factor, 2.2 (1.1–4.3); Euler number, 3.0 (1.1–8.7); node-to-terminus strut count, 3.3 (1.4–7.8); terminus-to-terminus strut count, 2.9 (1.2–6.9); and fractal dimension, 3.7 (1.5–9.7). Few and weak correlations were found between BMD at the calcaneus measured with DXA and features obtained from CT, suggesting that these two methods give different information about bone status. In conclusion, male osteoporosis is a disease characterized by decreased bone mass but also by microarchitectural deterioration of bone tissue which is partly independent of BMD. Received: 24 April 2001 / Accepted: 6 July 2001     

Fractal Analysis of Trabecular Bone Texture on Calcaneus Radiographs: Effects of Age, Time Since Menopause and Hormone Replacement Therapy

An analysis of trabecular bone texture based on fractal mathematics, when applied to trabecular bone images on plain radiographs, can be considered as a reflection of trabecular bone microarchitecture. It has been shown to be able to distinguish postmenopausal osteoporosis cases from controls. This cross-sectional study was carried out to investigate the influence of age, time since menopause and hormone replacement therapy (HRT) on the fractal dimension of trabecular bone texture at the calcaneus in a sample of 537 healthy women. Fractal analysis of texture was performed on calcaneus radiographs and the result expressed as the Hmean parameter (H = 2–fractal dimension). Total hip, femoral neck and lumbar spine bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry. There was a statistically significant Hmean parameter decrease with age (p<0.0001) but the degree of correlation was low (r=–0.2) compared with the correlation between age and BMD (r=–0.36 to –0.61 according to the BMD site). We found a weak but statistically significant correlation between time since menopause and Hmean (r=–0.14, p= 0.03) in the 241 postmenopausal women included in the study. Hmean was significantly lower in a group of postmenopausal women without HRT (n= 110) compared with a group of age-matched postmenopausal women with HRT (n = 110): respectively 0.683 ± 0.043 and 0.695 ± 0.038 (p= 0.03). In conclusion, this study suggests that there is a menopause- and age-related decrease in the Hmean parameter and that HRT interferes with the results of the fractal analysis of trabecular bone texture on calcaneus radiographs. Received: 2 March 2001 / Accepted: 2 October 2001     

Image Analysis of the Distal Radius Trabecular Network Using Computed Tomography

Bone texture analysis might provide information about bone structure in a noninvasive manner. In a prospective case–control cross-sectional study we investigated the value of computed tomography (CT) image analysis of the distal radius in the assessment of osteoporosis. Twenty patients suffering from postmenopausal osteoporosis were studied and compared with 21 age-matched controls. Eight slices were selected in each patient: four consecutive coronal slices and four consecutive cross-sectional slices. Bone texture analysis was performed using statistical, fractal and structural methods leading to the measurement of 32 features. Structural variables derived from histomorphometric parameters were measured after segmentation from a binary or a skeletonized image. Bone mineral density was measured by dual-energy X-ray absorptiometry both at the lumbar spine and the femoral neck. Eight of the 9 statistical features were significantly different in osteoporotic women as compared with controls (coronal slices, p < 0.05). Seven structural variables were statistically different between the two groups on coronal slices (p < 0.05): valley surface area, bone volume/tissue volume, trabecular partition, Euler’s number, trabecular bone pattern factor, node-to-node strut count and terminus-to-terminus strut count. The most significant results on coronal slices (p < 0.01) concerned 4 structural features: trabecular partition, Euler’s number, trabecular bone pattern factor and terminus-to-terminus strut count. Three features were statistically different (p < 0.01) between the two groups on cross-sectional slices (skeletonization from gray levels). A few features yielded by texture analysis were correlated with both lumbar spine and femoral neck bone mineral density, but the level of these correlations was weak (r < 0.5). In conclusion, CT image analysis of the distal radius is a useful tool for characterizing bone texture alterations in osteoporotic women. These findings are in keeping with microarchitectural osteoporosis-related changes diagnosed on bone biopsies. Received: 8 April 1998 / Accepted: 14 September 1998     

High-Resolution Computed Tomography for Architectural Characterization of Human Lumbar Cancellous Bone: Relationships with Histomorphometry and Biomechanics

The aim of the present study on human vertebral cancellous bone was to validate structural parameters measured with high-resolution (150 μm) computed tomography (HRCT) by referring to histomorphometry and to try to predict mechanical properties of bone using HRCT. Two adjacent vertical cores were removed from the central part of human L2 vertebral body taken after necropsy in 22 subjects aged 47–95 years (10 women, 12 men; mean age 79 ± 14 years). The right core was used for structural analysis performed by both HRCT and histomorphometry. Two cancellous bone specimens were extracted from the left core: a cube for HRCT and a compression test, and a cylinder for a shear test. Significant correlations were found between HRCT and histomorphometric measurements (BV/TV, trabecular thickness, separation and number, and node-strut analysis), but with higher values for most of the tomographic parameters (BV/TV and trabecular thickness determined by HRCT were overestimated by a factor 3.5 and 2.5 respectively, as compared with histomorphometry). The maximum compressive strength and Young’s modulus were highly correlated (ρ= 0.99, p<0.0005). Significant correlation was obtained between bone mineral density (determined using dual-energy X-ray absorptiometry) and the maximum compressive strength (ρ= 0.64, p= 0.002). In addition the maximum compressive strength and architectural parameters determined by HRCT or histomorphometry showed significant correlations (e.g., for HRCT, BV/TV: ρ = 0.88, p<0.0005, N.Nd/TV: ρ= 0.73, p<0.001). The shear strength was significantly correlated with BV/TV (ρ= 0.62, p= 0.002), Tb.Sp (ρ=−0.58, p= 0.004) and TSL (ρ= 0.55, p= 0.006) measured by HRCT. In conclusion, an HRCT system with 150 μm resolution is not sufficient to predict the true values of the structural parameters measured by histomorphometry, although high correlations were found between the two methods. However, we showed that a resolution of 150 μm allowed us to predict the mechanical properties of human cancellous bone. In vivo peripheral systems with such a resolution should be of interest and would deliver an acceptable radiation dose to the patient. Received: 13 October 1998 / Accepted: 16 March 1999     

In Vivo Assessment of Architecture and Micro-Finite Element Analysis Derived Indices of Mechanical Properties of Trabecular Bone in the Radius

Measurement of microstructural parameters of trabecular bone noninvasively in vivo is possible with high-resolution magnetic resonance (MR) imaging. These measurements may prove useful in the determination of bone strength and fracture risk, but must be related to other measures of bone properties. In this study in vivo MR imaging was used to derive trabecular bone structure measures and combined with micro-finite element analysis (μFE) to determine the effects of trabecular bone microarchitecture on bone mechanical properties in the distal radius. The subjects were studied in two groups: (I) postmenopausal women with normal bone mineral density (BMD) (n= 22, mean age 58 ± 7 years) and (II) postmenopausal women with spine or femur BMD −1 SD to −2.5 SD below young normal (n= 37, mean age 62 ± 11 years). MR images of the distal radius were obtained at 1.5 T, and measures such as apparent trabecular bone volume fraction (App BV/TV), spacing, number and thickness (App TbSp, TbN, TbTh) were derived in regions of interest extending from the joint line to the radial shaft. The high-resolution images were also used in a micro-finite element model to derive the directional Young’s moduli (E1, E2 and E3), shear moduli (G12, G23 and G13) and anisotropy ratios such as E1/E3. BMD at the distal radius, lumbar spine and hip were assessed using dual-energy X-ray absorptiometry (DXA). Bone formation was assessed by serum osteocalcin and bone resorption by serum type I collagen C-terminal telopeptide breakdown products (serum CTX) and urinary CTX biochemical markers. The trabecular architecture displayed considerable anisotropy. Measures of BMD such as the ultradistal radial BMD were lower in the osteopenic group (p<0.01). Biochemical markers between the two groups were comparable in value and showed no significant difference between the two groups. App BV/TV, TbTh and TbN were higher, and App TbSp lower, in the normal group than the osteopenic group. All three directional measures of elastic and shear moduli were lower in the osteopenic group compared with the normal group. Anisotropy of trabecular bone microarchitecture, as measured by the ratios of the mean intercept length (MIL) values (MIL1/MIL3, etc.), and the anisotropy in elastic modulus (E1/E3, etc.), were greater in the osteopenic group compared with the normal group. The correlations between the measures of architecture and moduli are higher than those between elastic moduli and BMD. Stepwise multiple regression analysis showed that while App BV/TV is highly correlated with the mechanical properties, additional structural measures do contribute to the improved prediction of the mechanical measures. This study demonstrates the feasibility and potential of using MR imaging with μFE modeling in vivo in the study of osteoporosis. Received: 13 December 2000 / Accepted: 30 May 2001     

New Model-Independent Measures of Trabecular Bone Structure Applied to In Vivo High-Resolution MR Images

Complementing measurements of bone mass with measurements of the architectural status of trabecular bone is expected to improve predictions of fracture risk in osteoporotic patients and improve the assessment of response to drug therapy. With high-resolution MRI the trabecular network can be imaged with 156×156×500 mm³ voxels, sufficient to depict individual trabeculae, albeit with inaccurate thickness. In this work, distance transformation techniques were applied to the three-dimensional image of the distal radius of postmenopausal patients. Structural indices such as trabecular number (app.Tb.N), thickness (app.Tb.Th) and separation (app.Tb.Sp) were determined without model assumptions. A new metric index, the apparent intra-individual distribution of separations (app.Tb.Sp.SD), is introduced. The reproducibility of the MR procedure and structure assessment was determined on volunteers, and the coefficient of variation was found to be 2.7–4.6% for the mean values of structural indices and 7.7% for app.Tb.Sp.SD. The distance transformation methods were then applied to two groups of patients: one of postmenopausal women without vertebral fracture and one of postmenopausal women with at least one vertebral fracture. It was found that app.Tb.Sp.SD discriminates fracture subjects from non-fracture patients as well as dual-energy X-ray absorptiometry (DXA) measurements of the radius and the spine, but not as well as DXA of the hip. Using receiver operating characteristic analysis, the area under the curve (AUC) values were 0.67 for app.Tb.Sp.SD, 0.72 for DXA radius, 0.67 for DXA spine and 0.81 for DXA of the hip. A combination of MR indices reached an AUC of 0.75. Age-adjusted odds ratio ranged from 1.85 to 2.03 for app.Tb.N, app.Tb.Sp and app.Tb.Sp.SD (p<0.003). We conclude that in vivo high-resolution MRI not only has the potential of imaging trabecular bone, but in combination with novel metrics may offer new insight into the structural changes occurring in postmenopausal women. Received: 7 November 2000 / Accepted: 20 August 2001     

Image-Based Assessment of Spinal Trabecular Bone Structure from High-Resolution CT Images

The goal of this study was to assess whether a high-resolution CT measure of trabecular bone structure can enhance the discrimination between subjects with or without a vertebral fracture and having overall low hip or spine bone mineral density (BMD) by dual-energy X-ray absorptiometry (DXA). Sixty-one women with low BMD by DXA (T-score <–2.5 at hip or spine) were examined. Twenty women had sustained a vertebral fracture. Quantitative CT (QCT) BMD and high-resolution CT spinal scans were performed on a whole-body CT scanner. For the high-resolution images (0.31 mm pixel, 1.5 mm thick slice), trabecular bone was segmented from marrow using an adaptive threshold, region growth and skeletonization step. From the processed image we measured the apparent trabecular bone fraction (BV/TV), apparent trabecular thickness (I.Th) and apparent trabecular spacing (I.Sp). We also assessed the connectivity of the marrow space using region growing to derive a mean (H_A) and maximum (H_M) hole size. Despite the fact that the study population was preselected to have a low BMD by DXA, QCT BMD was highly associated with (p <0.005) with fracture status. All structural parameters were correlated (r ~ 0.64 to 0.79) with BMD with p <0.003 and showed significant differences between the fracture and non-fracture group. However, except for H_A, this difference did not remain significant after adjustment for BMD. When BMD and then H_A was entered into a paired linear regression model to predict fracture outcome, H_A contributed with p= 0.03 and BMD with p= 0.86. ROC analysis was applied and showed that H_A, BMD, I.Th and I.Sp discriminated the two groups with areas of 0.76, 0.75, 0.71 and 0.68, respectively. These findings suggest that an assessment of vertebral trabecular structure from high-resolution CT images is useful in discriminating subjects with vertebral fractures and potentially useful for predicting future fractures. Received: 10 October 1997 / Accepted: 4 December 1997     

Bone Changes due to Glucocorticoid Application in an Ovariectomized Animal Model for Fracture Treatment in Osteoporosis

In a pilot experiment comparing four different modalities for inducing osteoporosis in the sheep, a combination of ovariectomy, calcium/vitamin D-restricted diet and steroid administration was found to generate the highest decrease in bone mineral density (BMD). The aim of the present study was to quantify the outcome of this triple treatment in an animal model of osteoporosis in terms of alteration in bone mass, bone structure and bone mechanics. A total of 32 sheep were divided into two equal groups. Group 1 (age 3–5 years) was used as a normal control. Group 2 (age 7–9 years) was ovariectomized, fed a calcium/vitamin D-restricted diet and injected with methylprednisolone (MP) over 7 months (22 weeks MP solution, 6 weeks MP suspension). The BMD at the distal radius and tibia was determined preoperatively and at repeated intervals bilaterally using quantitative computed tomography. Steroid blood levels were determined 4 and 24 h after selected injections. BMD was measured at L3 and L4 after 7 months. Biopsies were taken from iliac crests, vertebral bodies and femoral heads, and bone structure parameters investigated by three-dimensional micro-CT. Compressive mechanical properties of cancellous bone were determined from biopsies of vertebral bodies and femoral heads. After 7 months of osteoporosis induction the BMD of cancellous bone decreased 36 ± 3% in the radius and 39 ± 4% in the tibia. Steroid blood levels 24 h after injection of MP suspension were significantly higher than after injection of MP solution. Changes in structural parameters of cancellous bone from the iliac crest, lumbar spine and femoral head in group 2 indicated osteoporosis-associated changes. In group 2 there was a significant reduction in BMD of the lumbar spine and a significant reduction in stiffness and failure load in compression testing of biopsies of lumbar vertebrae. In sheep, changes in the structural parameters of bone such as trabecular number and separation during osteoporosis induction are comparable to the human situation. The sheep model presented seems to meet the criteria for an osteoporosis model for fracture treatment with respect to mechanical and morphometric bone properties. Received: 4 May 2001 / Accepted: 6 December 2001     

Relationship Between Structural Parameters, Bone Mineral Density and Fracture Load in Lumbar Vertebrae, Based on High-Resolution Computed Tomography, Quantitative Computed Tomography and Compression Tests

Different noninvasive techniques for the assessment of the individual fracture risk in osteoporosis are introduced, and the relation between structural properties of high-resolution computed tomography (HR-CT) images of vertebral bodies, their bone mineral density (BMD) and the fracture load is analyzed. In 24 unfractured lumbar vertebrae with different degrees of demineralization from six specimens, the trabecular and cortical BMD was determined using quantitative CT. A lateral X-ray image revealed the number of fractures in the entire spine. A structural analysis of spongy and cortical bone was performed based on the HR-CT images. In the spongiosa, the fractal dimension was calculated as a function of the threshold value. In the cortical shell, the maximum number of clusters of low BMD was determined at varying threshold values. After the CT measurements the vertebrae were excised and compressed until fractured. On the basis of the spongiosa BMD and the number of fractures, 3 cases were found to be severely osteoporotic; the other 3 cases showed osteopenia. The average fracture loads were determined as 3533 N for the non-osteoporotic cases (range 2602–5802 N) and 1725 N for the osteoporotic cases (range 1311–2490 N). The parameters were determined as follows: average spongiosa BMD 115.2 mg/ml (101.8–135.3 mg/ml) for the non-osteoporotic cases, 46.2 mg/ml (34.8–57.6 mg/ml) for the osteoporotic cases; average cortical BMD 285.1 mg/ml (216.4–361.9 mg/ml) for the non-osteoporotic cases, 136.1 mg/ml (142.5–215.2 mg/ml) for the osteoporotic cases; spongiosa structure: average 0.5 (range 0.32–0.75) for the non-osteoporotic cases, average 1.05 (range 0.87–1.24) for the osteoporotic cases; cortical structure: average 81 (range 55–104) for the non-osteoporotic cases), average 136 (range 102–159) for the osteoporotic cases. Single parameters (BMD and structure) and weighted sums of these parameters were correlated with the fracture load, resulting in correlation coefficients of r _sBMD= 0.82 (spongiosa BMD), r _cBMD= 0.82 (cortical BMD), r _sStr=–0.75 (spongiosa structure) and r _cStr=–0.86 (cortical structure). The weighted sum of cortical and spongiosa BMD resulted in r _BMD= 0.86, of cortical and spongiosa structure in r _Str=–0.86. A weighted combination of all four parameters correlates with the fracture load at r ₄= 0.89, all correlations being statistically significant (p<0.0001). The four individual parameters show only a slight overlap between non-osteoporotic and osteoporotic subjects. The high correlation of the cortical BMD and the structural parameter in cortical bone indicates the important contribution of the cortical shell to vertebral stability. A weighted sum of multiple parameters results in a higher correlation with the fracture load and does not show an overlap between the two groups. It is best suited to estimate the individual fracture risk. The presented methods are generally applicable in vivo; and allow an improvement of the diagnosis of osteoporosis compared with the measurement of the BMD alone. Received: 7 November 1997 / Accepted: 28 September 1998     

Biomechanics of Bone: Determinants of Skeletal Fragility and Bone Quality

Bone fragility can be defined by biomechanical parameters, including ultimate force (a measure of strength), ultimate displacement (reciprocal of brittleness) and work to failure (energy absorption). Bone fragility is influenced by bone size, shape, architecture and tissue ‘quality’. Many osteoporosis treatments build bone mass but also change tissue quality. Antiresorptive therapies, such as bisphosphonates, substantially reduce bone turnover, impairing microdamage repair and causing increased bone mineralization, which can increase the brittleness of bone. Anabolic therapies, such as parathyroid hormone (PTH-(1–84)) or teriparatide (PTH-(1–34)), increase bone turnover and porosity, which offset some of the positive effects on bone strength. Osteoporosis therapies may also affect bone architecture by causing the redistribution of bone structure. Restructuring of bone during treatment may change bone fragility, even in the absence of drug effects on bone mineral density (BMD). This effect may explain why some drugs can affect fracture incidence disproportionately to changes in BMD. For instance, in a recent clinical trial, PTH-(1–34) therapy caused a dose-related increase in spinal BMD without any dose-dependent effect on the observed decrease in spinal fracture incidence. This apparent disassociation between spinal BMD and bone fragility is probably due to effects of PTH-(1–34) on bone architecture within vertebral bodies. While it has been shown that BMD is highly heritable, bone mineral distribution and architecture are also under strong genetic influence. Recent findings suggest that different genes regulate trabecular and cortical structures within lumbar vertebrae, producing a wide range of bone architectural designs. These findings suggest that there is no single optimal bone architecture; instead many different architectural solutions produce adequate bone strength. Received: 14 June 2001 / Accepted: 3 September 2001     

High Resolution Magnetic Resonance Imaging of the Calcaneus: Age-Related Changes in Trabecular Structure and Comparison with Dual X-Ray Absorptiometry Measurements

A high-resolution magnetic resonance imaging (MRI) protocol, together with specialized image processing techniques, was applied to the quantitative measurement of age-related changes in calcaneal trabecular structure. The reproducibility of the technique was assessed and the annual rates of change for several trabecular structure parameters were measured. The MR-derived trabecular parameters were compared with calcaneal bone mineral density (BMD), measured by dual X-ray absorptiometry (DXA) in the same subjects. Sagittal MR images were acquired at 1.5 T in 23 healthy women (mean age: 49.3 ± 16.6 [SD]), using a three-dimensional gradient echo sequence. Image analysis procedures included internal gray-scale calibration, bone and marrow segmentation, and run-length methods. Three trabecular structure parameters, apparent bone volume (ABV/TV), intercept thickness (I.Th), and intercept separation (I.Sp) were calculated from the MR images. The short- and long-term precision errors (mean %CV) of these measured parameters were in the ranges 1–2% and 3–6%, respectively. Linear regression of the trabecular structure parameters vs. age showed significant correlation: ABV/TV (r ²= 33.7%, P < 0.0037), I.Th (r ²= 26.6%, P < 0.0118), I.Sp (r ²= 28.9%, P < 0.0081). These trends with age were also expressed as annual rates of change: ABV/TV (− 0.52%/year), I.Th (−0.33%/year), and I.Sp (0.59%/year). Linear regression analysis also showed significant correlation between the MR-derived trabecular structure parameters and calcaneal BMD values. Although a larger group of subjects is needed to better define the age-related changes in trabecular structure parameters and their relation to BMD, these preliminary results demonstrate that high-resolution MRI may potentially be useful for the quantitative assessment of trabecular structure. Received: 11 March 1996 / Accepted: 9 July 1996     

Effect of Collagen and Mineral Content on the High-Frequency Ultrasonic Properties of Human Cancellous Bone

The technology surrounding ultrasonic bone assessment is evolving rapidly as investigators explore the utility of new ultrasonic parameters and different ultrasonic frequencies. This study had three main goals. The first was to perform in vitro measurements of the speed of sound (SOS) and normalized broadband ultrasonic attenuation (nBUA) in specimens of normal human cancellous bone using a 2.25 MHz broadband measurement system. The second was to explore the utility of a backscatter-based parameter called apparent integrated backscatter (AIB). The third goal was to investigate the roles that collagen and mineral content play in affecting each of these three ultrasonic parameters. This was accomplished by chemically treating the specimens to remove one or the other of these two important constituents of bone. Our results showed that in most cases SOS and nBUA correlated well (p<0.05) with bone mineral density (BMD) as measured by quantitative computed tomography (QCT). In contrast, AIB did not correlate strongly with BMD. When the specimens were demineralized, decreases were produced in SOS (19–39%) and nBUA (44–58%). Changes produced in AIB were not significant except along the superoinferior direction, in which a 12% decrease was measured. When the specimens were decollagenized, decreases were produced in SOS (10–12%). In contrast, increases were produced in both nBUA (35–77%) and AIB (14–15%). From this study we conclude that high-frequency ultrasonic measurements may yield useful information about the content and organization of both collagen and mineral in cancellous bone. Received: 30 November 2000 / Accepted: 22 June 2001     

Graphic Trace Analysis of Ultrasound at the Phalanges May Differentiate Between Subjects with Primary Hyperparathyroidism and with Osteoporosis: A Pilot Study

Bone loss characterizes both primary hyperparathyroidism (PHPT) and osteoporosis (OP) but with a different histologic pattern, and this could partially explain the different fracture incidence in these two populations. Quantitative ultrasound (QUS), influenced by bone structural parameters other than bone mineral density (BMD), could evidence these differences, opening new perspectives in the evaluation of patients with metabolic bone diseases. The aim of the present study was to investigate the usefulness of QUS graphic trace parameters, assessed at the phalanx, in discriminating between PHPT bone disease and osteoporosis. We studied 34 patients with PHPT (mean age 59.7 ± 12.7 years), 35 patients with OP (mean age 60.6 ± 7.1 years) and 34 healthy subjects as controls (mean age 59.1 ± 9.4 years). In all subjects QUS measurements were performed at the phalanx with a Bone Profiler (IGEA, Italy), obtaining the amplitude-dependent speed of sound (AD-SoS), fast wave amplitude (FWA), signal dynamic (SDy), bone transmission time (BTT) and ultrasound bone profile index (UBPI). Moreover, serum calcium, phosphorus, parathyroid hormone (PTH), bone isoenzyme of alkaline phosphatase (B-ALP) and ionized calcium were measured in all subjects in the morning under fasting conditions. In PHPT patients BTT was correlated with PTH, ionized calcium and B-ALP levels (r=–0.47, –0.57 and –0.44, respectively; p <0.01), whereas FWA, SDy and UBPI correlated only with B-ALP (r=–0.43, –0.46 and –0.50, respectively; p <0.01). Moreover, FWA, SDY and UBPI were significantly (p<0.01) lower and BTT significantly (p<0.001) higher in OP than in PHPT patients. UBPI, BTT, FWA and the BTT/FWA ratio, but not SDy, were able to discriminate between the two groups (area under the curve =0.66, 0.69, 0.67 and 0.81, respectively).  Our findings show that ultrasound signal parameters are differently influenced by bone changes characterizing primary hyperparathyroidism or osteoporosis. This suggests that the QUS signal could be a useful instrument in discriminating and studying some of the bone alterations typical of metabolic bone diseases. Received: 15 February 2001 / Accepted: 27 August 2001     

Cut-off Values Determined for Vertebral Fracture by Peripheral Quantitative Computed Tomography in Japanese Women

In spite of the benefits of bone mass measurement by dual-energy X-ray absorptiometry (DXA), the use of DXA has limitations. It is unable to assess a true bone density, and cannot discriminate between the trabecular and cortical bone compartments. Ultradistal radius bone density was measured using peripheral quantitative computed tomography (pQCT) to determine reference values for total bone density (BD), trabecular bone density (TBD), polar strength strain index (pSSI), total bone mineral content (BC), trabecular bone mineral content (TBC), cortical bone density (CBD), cortical bone mineral content (CBC) and polar cross-sectional moment of inertia (pCSMI) in the Japanese female population, and to ascertain the cut-off values of the measured indicators that could most efficiently discriminate osteoporotic subjects with vertebral fractures. A total of 5266 healthy Japanese women aged 20–89 years were included in this study to determine Japanese reference values. Additionally, 621 who had undergone radiographic examination of the thoracic and lumbar spine at the time of pQCT measurement were selected to determine the cut-off values of BD, TBD, pSSI and other indicators for vertebral fractures. All the healthy subjects were divided into 5 year age groups. The BD showed nonsignificant changes from the 20–24 year age group to the 45–49 year age group, and fell significantly thereafter. The TBD maintained a plateau until the 40–44 year group, which corresponds to the young adult mean (YAM) values of the lumbar spine, femoral neck and radius BMDs measured using DXA. The TBD decreased significantly thereafter. The pSSI did not change significantly from the 20–24 year age group to the 45–49 year age group, and decreased slightly in the 50–54 year age group and markedly after 55–59 years. The cut-off values for the discrimination of vertebral fractures were obtained by the calculation of sensitivities, specificities and the area under the curves obtained using age-adjusted receiver operating characteristics (ROC) analysis. Odds ratios and 95% confidence limits (CL) were calculated using age-adjusted logistic analysis. The cut-off values for vertebral fractures, the area under the ROC curves (AUC) and odds ratios were 270.1 mg/cm³ (−2.2 SD, 66.6% of YAM), 0.689 ± 0.025, 2.10 (1.63, 2.70) for BD, 104.8 mg/cm³ (−2.2 SD, 53.5% of YAM), 0.699 ± 0.023, 2.17 (1.69, 2.77) for TBD and 192.8 mm³ (−1.9 SD, 59.8% of YAM), 0.631 ± 0.028, 1.72 (1.34, 2.21) for pSSI, respectively. These findings suggest that ultradistal radius BMD measured using pQCT can be used to discriminate women with vertebral fractures. Received: 3 August 2000 / Accepted: 5 March 2001     

The Effects of Pregnancy and Lactation on Bone Mineral Density

We performed a prospective study of bone mineral density (BMD) in 38 women during their first full-term pregnancy until 12 months postpartum. BMD measurements at lumbar spine [L2–L4 (LS)] and forearm [distal 33% (RD) and ultradistal (RUD) region of the radius] were made within 3 months before conception, after delivery, and at 6 and 12 months postpartum. In mid-pregnancy the DXA examination was carried out only at the forearm. Patients were grouped according to duration of lactation as group I, II or III (0–1, 1–6, 6–12 months respectively). During pregnancy there was a significant difference between baseline and delivery (p< 0.001) in the LS, RUD and RD BMD values. In group I there was no statistically significant difference in LS BMD between visits following pregnancy. The RUD BMD loss was recovered by 6 months postpartum (PP6). Group II showed continuous bone loss from delivery until PP6 at LS and RUD. In group III the LS BMD loss continued throughout the lactation period. The RUD BMD dropped (4.9%) until PP6 then increased by 3.0% as measured at 12 months postpartum (PP12). There was no significant change in RD BMD in any of three groups during lactation. At LS bone loss between delivery and PP12 correlated well with the duration of lactation (r=−0.727; p<0.001). We suggest that calcium needed for fetal skeletal growth during pregnancy was gained from maternal trabecular and cortical sites and that calcium needed for infant growth during lactation was drawn mainly from the maternal trabecular skeleton in our patients. The effect of pregnancy and lactation on the maternal bone mass was spontaneously compensated after weaning. Received: 13 July 2000 / Accepted: 19 April 2001     

Combined Calcium and Vitamin D3 Supplementation in Elderly Women: Confirmation of Reversal of Secondary Hyperparathyroidism and Hip Fracture Risk: The Decalyos II Study

Vitamin D insufficiency and low calcium intake contribute to increase parathyroid function and bone fragility in elderly people. Calcium and vitamin D supplements can reverse secondary hyperparathyroidism thus preventing hip fractures, as proved by Decalyos I. Decalyos II is a 2-year, multicenter, randomized, double-masked, placebo-controlled confirmatory study. The intention-to-treat population consisted of 583 ambulatory institutionalized women (mean age 85.2 years, SD = 7.1) randomized to the calcium–vitamin D₃ fixed combination group (n= 199); the calcium plus vitamin D₃ separate combination group (n= 190) and the placebo group (n= 194). Fixed and separate combination groups received the same daily amount of calcium (1200 mg) and vitamin D₃ (800 IU), which had similar pharmacodynamic effects. Both types of calcium-vitamin D₃ regimens increased serum 25-hydroxyvitamin D and decreased serum intact parathyroid hormone to a similar extent, with levels returning within the normal range after 6 months. In a subgroup of 114 patients, femoral neck bone mineral density (BMD) decreased in the placebo group (mean =–2.36% per year, SD = 4.92), while remaining unchanged in women treated with calcium-vitamin D₃ (mean = 0.29% per year, SD = 8.63). The difference between the two groups was 2.65% (95% CI =–0.44, 5.75%) with a trend in favor of the active treatment group. No significant difference between groups was found for changes in distal radius BMD and quantitative ultrasonic parameters at the os calcis. The relative risk (RR) of HF in the placebo group compared with the active treatment group was 1.69 (95% CI = 0.96, 3.0), which is similar to that found in Decalyos I (RR = 1.7; 95% CI = 1.0, 2.8). Thus, these data are in agreement with those of Decalyos I and indicate that calcium and vitamin D₃ in combination reverse senile secondary hyperparathyroidism and reduce both hip bone loss and the risk of hip fracture in elderly institutionalized women. Received: 23 March 2001 / Accepted: 28 October 2001     

Relationship Between Plain Radiographic Patterns and Three- dimensional Trabecular Architecture in The Human Calcaneus

The purpose of this study was to determine the relationship between three-dimensional (3D) trabecular structure and two-dimensional plain radiographic patterns. An in vitro cylinder of human calcaneal trabecular bone was three-dimensionally imaged by micro-CT using synchrotron radiation, at 33.4 μm resolution. The original 3D image was processed using 14 distinct sequences of morphologic operations, i.e., of dilations and erosions, to obtain a total of 15 3D models or images of calcaneal trabecular bone. These 15 models had distinct densities (volume fractions) and architectures. The 3D structure of each calcaneal model was assessed using mean intercept length (fabric), by averaging individual fabric measurements associated with each medial-lateral image slice, and determining the relative anisotropy, R_3D, of the structure. A summated pattern or plain radiograph was also computed from the 3D image data for each calcaneal model. Each summated pattern was then locally thresholded, and the resulting two-dimensional (2D) binary image analyzed using the same fabric analysis as used for the 3D data. The anisotropy of the 2D summated pattern was denoted by R_x-ray. The volume fractions of the 15 models ranged from 0.08 to 0.19 with a mean of 0.14. The medial-lateral anisotropies, R_3D, ranged from 1.38 to 2.54 with a mean of 1.88. The anisotropy of the 2D summated patterns, R_x-ray, ranged from 1.35 to 2.18 with a mean of 1.71. The linear correlation of the 3D trabecular architecture, R_3D, with the radiographic trabecular architecture, R_x-ray, was 0.99 (p<0.0001). This study shows that the plain radiograph contains architectural information directly related to the underlying 3D structure. A well-controlled sequential reproducible plain radiograph may prove useful for monitoring changes in trabecular architecture in vivo and in identifying those individuals at increased risk of osteoporotic fracture. Received: 9 December 1997 / Accepted: 3 September 1998     

Bone Quality Parameters of the Distal Radius as Assessed by pQCT in Normal and Fractured Women

The aim of this study was to test the ability of some indicators of different aspects of bone quality (assessed by peripheral quantitative computed tomography in the distal radius) to discriminate between fractured and nonfractured individuals. The study compared 214 women aged 45–85 years, free of any bone-affecting treatment, of whom 107 had suffered a Colles” fracture in the previous 6 months and 107 did not. The determinations included bone tissue or mineral “mass” indicators (trabecular, cortical and total volumetric mineral content, cortical bone area); bone “density” estimates (trabecular, cortical and total volumetric mineral density), and the Cartesian (rectangular) and polar moments of inertia as influences of cross-sectional architecture on resistance to bending and torsional loads, respectively.  The influences of body height, weight and age on the tomographic indicators were minimized by adjusting the data according to the partial coefficients of multiple stepwise regressions. The adjusted values of all the indicators were lower in fractured than in nonfractured groups. The prevalence of fractures was directly related to the actual values of the indicators, rather than the age or body habitus of the individuals. The significance of these differences between the assessed indicators decreased in the following order: trabecular “mass” > trabecular “density” > cortical or total “mass” > cortical architecture > total or cortical “density” indicators. Within the same type of bone, the tissue or mineral “mass” indicators performed better than the “density” indicators. The cortical bone density did not give useful information, probably because of technical difficulties. Odds-ratios and receiver-operating characteristic (ROC) analyses confirmed those features. The selected “cut-off” values of the indicators as determined by the ROC curves (very close to those determined by the inflexion points of the logistic reression curves) may indicate reference limits to detect persons at risk of fracture according to the type of information provided by each variable. These results show that these tomographic indicators discriminate well between fractured and nonfractured individuals, and should be suitable to assess how total, cortical and trabecular bone strength in the distal radius could affect different kinds of strength regardless of the age or body habitus of the individual. Their ability to estimate fracture risk from different biomechanical points of view should be assessed by adequately designed, prospective studies. Received: June 2000 / Accepted: January 2001     

Digital X-ray Radiogrammetry Predicts Hip,Wrist and Vertebral Fracture Risk in Elderly Women: A Prospective Analysis from the Study of Osteoporotic Fractures

Digital X-ray radiogrammetry (DXR) is a technique that uses automated image analysis of standard hand radiographs to estimate bone mineral density (DXR-BMD). Previous studies have shown that DXR-BMD measurements have high precision, are strongly correlated with forearm BMD and are lower in individuals with prevalent fractures. To determine whether DXR-BMD measurements predict wrist, hip and vertebral fracture risk we conducted a case–cohort study within a prospective study of 9704 community-dwelling elderly women (the Study of Osteoporotic Fractures). We compared DXR-BMD, and BMD of the radius (proximal and distal), calcaneus, femoral neck and posteroanterior lumbar spine in women who subsequently suffered a wrist (n= 192), hip (n= 195), or vertebral fracture (n= 193) with randomly selected controls from the same cohort (n= 392–398). DXR-BMD was estimated from hand radiographs acquired at the baseline visit. The radiographs were digitized and the Pronosco X-posure System was used to compute DXR-BMD from the second through fourth metacarpals. Wrist fractures were confirmed by radiographic reports and hip fractures were confirmed by radiographs. Vertebral fractures were defined using morphometric analysis of lateral spine radiographs acquired at baseline and an average of 3.7 years later. Age-adjusted odds ratio (OR, vertebral fracture) or relative hazard (RH, wrist and hip fracture) for a 1 SD decrease in BMD were computed. All BMD measurements were similar for prediction of wrist (RH = 1.5–2.1) and vertebral fracture (OR = 1.8–2.5). Femoral neck BMD best predicted hip fracture (RH = 3.0), while the relative hazards for all other BMD measurements were similar (RH = 1.5–1.9). These prospective data indicate that DXR-BMD performs as well as other peripheral BMD measurements for prediction of wrist, hip and vertebral fractures. Therefore, DXR-BMD may be useful for prediction of fracture risk in clinical settings where hip BMD is not available. Received: 27 April 2001 / Accepted: 10 October 2001