Local anisotropy in mineralized fibrocartilage and subchondral bone beneath the tendon-bone interface.

The enthesis allows the insertion of tendon into bone thanks to several remarkable strategies. This complex and clinically relevant location often features a thin layer of fibrocartilage sandwiched between tendon and bone to cope with a highly heterogeneous mechanical environment. The main purpose of this study was to investigate whether mineralized fibrocartilage and bone close to the enthesis show distinctive three-dimensional microstructural features, possibly to enable load transfer from tendon to bone. As a model, the Achilles tendon-calcaneus bone system of adult rats was investigated with histology, backscattered electron imaging and micro-computed tomography. The microstructural porosity of bone and mineralized fibrocartilage in different locations including enthesis fibrocartilage, periosteal fibrocartilage and bone away from the enthesis was characterized. We showed that calcaneus bone presents a dedicated protrusion of low porosity where the tendon inserts. A spatially resolved analysis of the trabecular network suggests that such protrusion may promote force flow from the tendon to the plantar ligament, while partially relieving the trabecular bone from such a task. Focusing on the tuberosity, highly specific microstructural aspects were highlighted. Firstly, the interface between mineralized and unmineralized fibrocartilage showed the highest roughness at the tuberosity, possibly to increase failure resistance of a region carrying large stresses. Secondly, fibrochondrocyte lacunae inside mineralized fibrocartilage, in analogy with osteocyte lacunae in bone, had a predominant alignment at the enthesis and a rather random organization away from it. Finally, the network of subchondral channels inside the tuberosity was highly anisotropic when compared to contiguous regions. This dual anisotropy of subchondral channels and cell lacunae at the insertion may reflect the alignment of the underlying collagen network. Our findings suggest that the microstructure of fibrocartilage may be linked with the loading environment. Future studies should characterize those microstructural aspects in aged and or diseased conditions to elucidate the poorly understood role of bone and fibrocartilage in enthesis-related pathologies.

The artiodactyl calcaneus as a potential 'control bone' cautions against simple interpretations of trabecular bone adaptation in the anthropoid femoral neck.

For over a century, the arched trabecular patterns of the human proximal femur have been considered to resemble tension and compression stress trajectories produced by stereotypical bending loads. This reflects conventional modeling of the human femoral neck-head region as a short cantilevered beam. Although this conception is the foundation of many biomechanical, clinical, paleontological, and comparative morphological studies of trabecular bone in various species, attempts have not been made to contrast these data to a bone that could be considered a 'control' for simple/stereotypical bending. We quantified trabecular architectural characteristics in sheep and deer calcanei as a first step in potentially establishing them as 'controls' in this context because they have arched trabecular patterns that resemble tension/compression stress trajectories, and have been shown by strain gauge measurements to be relatively simply loaded in bending. Using micro-computed tomography, calcanei from adult domesticated sheep and wild deer were analyzed where in the dorsal 'compression' and plantar 'tension' trabecular tracts they begin to separate and bending is less complex (mid-shaft), and where trabeculae extensively interconnect and loading is more complex (distal shaft). Of the eight trabecular architectural characteristics evaluated, only one (trabecular number, Tb.N) showed a probable mechanically relevant dorsal/plantar difference. However, this was paradoxically opposite in the sheep calcanei. Aside from Tb.N, the architectural characteristics showed little, if any, evidence of habitual bending. The non-uniformity of the stresses between the trabecular tracts in these bones might be reduced by load-sharing functions of their robust cortices and the nearby ligament and tendon, which might account for the similar morphologies between the tracts. These findings may help to explain why in many cases regional trabecular architectural variations seem to lack sufficient sensitivity and specificity for interpreting habitual bending in other bone regions. This cautions against simple interpretations of trabecular bone adaptation in the anthropoid femoral neck.

Microarchitecture and bone quality in the human calcaneus: local variations of fabric anisotropy.

The local variability of microarchitecture of human trabecular calcaneus bone is investigated using high-resolution micro-computed tomography (µCT) scanning. The fabric tensor is employed as the measure of the microarchitecture of the pore structure of a porous medium. It is hypothesized that a fabric tensor-dependent poroelastic ultrasound approach will more effectively predict the data variance than will porosity alone. The specific aims of the present study are as follows: (1) to quantify the morphology and local anisotropy of the calcaneus microarchitecture with respect to anatomical directions; (2) to determine the interdependence, or lack thereof, of microarchitecture parameters, fabric, and volumetric bone mineral density (vBMD); and (3) to determine the relative ability of vBMD and fabric measurements in evaluating the variance in ultrasound wave velocity measurements along orthogonal directions in the human calcaneus. Our results show that the microarchitecture in the analyzed regions of human calcanei is anisotropic, with a preferred alignment along the posterior-anterior direction. Strong correlation was found between most scalar architectural parameters and vBMD. However, no statistical correlation was found between vBMD and the fabric components, the measures of the pore microstructure orientation. Therefore, among the parameters usually considered for cancellous bone (ie, classic histomorphometric parameters such as porosity, trabecular thickness, number and separation), only fabric components explain the data variance that cannot be explained by vBMD, a global mass measurement, which lacks the sensitivity and selectivity to distinguish osteoporotic from healthy subjects because it is insensitive to directional changes in bone architecture. This study demonstrates that a multidirectional, fabric-dependent poroelastic ultrasound approach has the capability of characterizing anisotropic bone properties (bone quality) beyond bone mass, and could help to better understand anisotropic changes in bone architecture using ultrasound.

Do regional modifications in tissue mineral content and microscopic mineralization heterogeneity adapt trabecular bone tracts for habitual bending? Analysis in the context of trabecular architecture of deer calcanei.

Calcanei of mature mule deer have the largest mineral content (percent ash) difference between their dorsal 'compression' and plantar 'tension' cortices of any bone that has been studied. The opposing trabecular tracts, which are contiguous with the cortices, might also show important mineral content differences and microscopic mineralization heterogeneity (reflecting increased hemi-osteonal renewal) that optimize mechanical behaviors in tension vs. compression. Support for these hypotheses could reveal a largely unrecognized capacity for phenotypic plasticity - the adaptability of trabecular bone material as a means for differentially enhancing mechanical properties for local strain environments produced by habitual bending. Fifteen skeletally mature and 15 immature deer calcanei were cut transversely into two segments (40% and 50% shaft length), and cores were removed to determine mineral (ash) content from 'tension' and 'compression' trabecular tracts and their adjacent cortices. Seven bones/group were analyzed for differences between tracts in: first, microscopic trabecular bone packets and mineralization heterogeneity (backscattered electron imaging, BSE); and second, trabecular architecture (micro-computed tomography). Among the eight architectural characteristics evaluated [including bone volume fraction (BVF) and structural model index (SMI)]: first, only the 'tension' tract of immature bones showed significantly greater BVF and more negative SMI (i.e. increased honeycomb morphology) than the 'compression' tract of immature bones; and second, the 'compression' tracts of both groups showed significantly greater structural order/alignment than the corresponding 'tension' tracts. Although mineralization heterogeneity differed between the tracts in only the immature group, in both groups the mineral content derived from BSE images was significantly greater (P < 0.01), and bulk mineral (ash) content tended to be greater in the 'compression' tracts (immature 3.6%, P = 0.03; mature 3.1%, P = 0.09). These differences are much less than the approximately 8% greater mineral content of their 'compression' cortices (P < 0.001). Published data, suggesting that these small mineralization differences are not mechanically important in the context of conventional tests, support the probability that architectural modifications primarily adapt the tracts for local demands. However, greater hemi-osteonal packets in the tension trabecular tract of only the mature bones (P = 0.006) might have an important role, and possible synergism with mineralization and/or microarchitecture, in differential toughening at the trabeculum level for tension vs. compression strains.

The Osteoporosis Self-Assessment Tool versus alternative tests for selecting postmenopausal women for bone mineral density assessment: a comparative systematic review of accuracy.

SUMMARY: We performed a systematic review of studies comparing the Osteoporosis Self-Assessment Tool (OST) and other tests used to select women for bone mineral density (BMD) assessment. In comparative meta-analyses, we found that the accuracy of OST was similar to other tests that are based on information from the medical history. By contrast, assessment by quantitative ultrasonography at the heel was more accurate than OST in discriminating between women with high and low BMD. The methodological quality of the included studies was generally low. INTRODUCTION: Numerous tests are suggested for triaging postmenopausal women for bone mineral density (BMD) assessment by dual-energy X-ray absorptiometry. Previous studies suggest that OST, based on age and weight only, may be as accurate as more complex triage tests. We systematically compare the accuracy of OST and alternative triage tests in postmenopausal women. METHODS: We searched PubMed, Embase, Web of Science, citation lists, and conference proceedings. Our main measure of accuracy was the diagnostic odds ratio (DOR). We compared summary estimates of DOR (sDOR) for OST and alternative tests in pairwise meta-analyses by using the Moses-Littenberg approach. RESULTS: Summary estimates of DOR for OST and the clinical decision rules Simple Calculated Osteoporosis Risk Estimation (SCORE) and Osteoporosis Risk Assessment Instrument (ORAI) did not differ significantly in white women (relative sDOR: 0.57-1.17, all p >or= 0.11). By contrast, sDOR was higher for Stiffness Index assessed by calcaneal quantitative ultrasonography than for OST (relative sDOR: 1.9, p = 0.005). Studies were few in Asian and black women. Methodological quality, assessed with the Quality Assessment of Diagnostic Accuracy Studies (QUADAS) checklist, was generally low. CONCLUSIONS: In white women, the accuracy of OST and alternative clinical decision rules was similar, whereas Stiffness Index was more accurate than OST. Low study quality renders transferability to clinical settings uncertain.

Structural and biochemical analysis of the effect of immobilization followed by stretching on the calcaneal tendon of rats.

Little is known about the stretching effects on the biochemical and morphological features of tendons submitted to a long period of immobilization. Our purpose was to evaluate the response of rat tendons to stretching procedures after immobilization. The animals were separated into five experimental groups: GI--control of immobilized and euthanized animals; GII--immobilized and euthanized animals; GIII--control of immobilized animals and afterward stretched or allowed free cage activity; GIV--immobilized and stretched animals; and GV--immobilized and allowed free cage activity. Analysis in SDS-PAGE showed no remarkable differences among the groups, but a prominent collagen band was observed in GV, as compared to GIV and the control group, both in the compression and tension regions. Hydroxyproline content was highest in the compression region of GII. No differences among the groups were observed in the tension region. In regard to the concentration of noncollagenous proteins, differences were detected only in the tension region, where larger concentrations were found in the GII. When GII and GIV were compared, highest values were found in the GII. A more abundant presence of sulfated glycosaminoglycans, especially chondroitin sulfate, was detected in GIV, at the compression region of tendons. The presence of dermatan sulfate was outstanding in the compression and tension regions of the GII and GV groups. In the Ponceau SS stained sections, analyzed under polarization microscopy, GII exhibited the highest disorganization of the collagen bundles, partially recovered after stretching or with only remobilization. Our results indicate that a revision in the stretching procedures, in terms of duration and periodicity of the sessions, could benefit the efficiency of the stretching in cases of previous immobilization of tendons.

Are distributions of secondary osteon variants useful for interpreting load history in mammalian bones?

BACKGROUND/AIMS: In cortical bone, basic multicellular units (BMUs) produce secondary osteons that mediate adaptations, including variations in their population densities and cross-sectional areas. Additional important BMU-related adaptations might include atypical secondary osteon morphologies (zoned, connected, drifting, elongated, multiple canal). These variants often reflect osteonal branching that enhances toughness by increasing interfacial (cement line) complexity. If these characteristics correlate with strain mode/magnitude-related parameters of habitual loading, then BMUs might produce adaptive differences in unexpected ways. METHODS: We carried out examinations in bones loaded in habitual torsion (horse metacarpals) or bending: sheep, deer, elk, and horse calcanei, and horse radii. Atypical osteons were quantified in backscattered images from anterior, posterior, medial, and lateral cortices. Correlations were determined between atypical osteon densities, densities of all secondary osteons, and associations with habitual strain mode/magnitude or transcortical location. RESULTS: Osteon variants were not consistently associated with 'tension', 'compression', or neutral axis ('shear') regions, even when considering densities or all secondary osteons, or only osteon variants associated with relatively increased interfacial complexity. Similarly, marrow- and strain-magnitude-related associations were not consistent. CONCLUSION: These data do not support the hypothesis that spatial variations in these osteon variants are useful for inferring a habitual bending or torsional load strain history.

The effect of age on the structure and composition of rat tendon fibrocartilage.

Biochemical and morphological aspects of fibrocartilages of calcaneal and deep digital flexor tendons in rats aged 30, 180 and 730 days were analyzed. In both tendons a stronger staining with Alcian blue, indicating the presence of proteoglycans, was detected in rats of 30 and 180 days. In animals 730 days old, it was restricted to the pericellular area. Ultrastructural analysis showed a more prominent pericellular matrix in calcaneal tendon compared to the deep digital flexor tendon. The biochemical analysis showed higher levels of proteins and glycosaminoglycans in the calcaneal tendon of 30-day-old rats compared to older rats. In the deep digital flexor tendon, no significant differences were observed between ages. The small proteoglycan, fibromodulin, was detected in both tendons of all ages, but in young rats it appeared to be running as a 210 kDa component, probably due to the association with collagen chains or self-association.

Preliminary observations on the calcaneal trabecular microarchitecture of extant large-bodied hominoids.

In this pilot study, we point out potential differences between calcaneal trabecular microarchitecture in humans and nonhuman large apes, such as increased degree of anisotropy, reduced bone volume fraction, and very stereotypical orientation of the trabeculae. Even though sample size does not permit us to investigate the issue statistically, the observed differences between humans and other hominoids warrants further in-depth investigation. We also show that some measurements of the trabecular network might be dependent on sampling density, which can be difficult to deal with in the case of animals of different body masses. We also present a new visualization technique that summarizes the trabecular network orientation, and makes it more readily interpretable than the summary statistics of the underlying fabric tensor of the orientation matrix.

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.

Fractographic examination of racing greyhound central (navicular) tarsal bone failure surfaces using scanning electron microscopy.

The greyhound is a fatigue fracture model of a short distance running athlete. Greyhounds have a high incidence of central (navicular) tarsal bone (CTB) fractures, which are not associated with overt trauma. We wished to determine whether these fractures occur because of accumulation of fatigue microdamage. We hypothesized that bone from racing dogs would show site-specific microdamage accumulation, causing predisposition to structural failure. We performed a fractographic examination of failure surfaces from fractured bones using scanning electron microscopy and assessed microcracking observed at the failure surface using a visual analog scale. Branching arrays of microcracks were seen in failure surfaces of CTB and adjacent tarsal bones, suggestive of compressive fatigue failure. Branching arrays of microcracks were particularly prevalent in remodeled trabecular bone that had become compact. CTB fractures showed increased microdamage when compared with other in vivo fractures (adjacent tarsal bone and long bone fractures), and ex vivo tarsal fractures induced by monotonic loading (P < 0.02). It was concluded that greyhound racing and training often results in CTB structural failure, because of accumulation and coalescence of branching arrays of fatigue microcracks, the formation of which appears to be predisposed to adapted bone.

Biomechanical properties of human os calcanei: relationships with bone density and fractal evaluation of bone microarchitecture.

The relationship between bone strength and bone mass is well established. The link between trabecular microarchitecture and biomechanical properties has been less extensively explored. To address this question, we have tested the mechanical behaviour of calcaneus bone samples and investigated the correlations between mechanical properties on the one hand, bone density and fractal analysis of microarchitecture on the other hand. Mechanical properties of 43 human os calcanei were determined by uniaxial compression testing of samples from tuber calcanei. Ash density, bulk density and dual energy X-ray absorptiometry of the samples were measured. Fractal analysis of the trabecular bone on calcaneus radiographs was performed by two estimators derived from the fractional Brownian motion model. The mechanical properties of human os calcis were found to correlate with age and density measurements. Fractal parameters derived from the bone texture analysis showed weaker but significant correlations with bone strength. Fractal analysis of texture could account in part for the variations of bone strength, but in this study cannot explain better than density the mechanical properties of trabecular bone. Nevertheless, it provides a non-invasive means of assessing molecular bone microarchitecture.

Diffraction and interface losses in broadband ultrasound attenuation measurements of the calcaneum.

Ultrasonic investigation of bone disease most frequently involves measurements on the heel, and the parameter most often used for this is the broadband ultrasound attenuation (BUA) which is the slope of the attenuation as a function of frequency between 0.2 and 0.6 MHz. In this study, the possible losses in the BUA measurement due to diffraction and interfaces have been discussed and evaluated using existing data and a standard diffraction model. The loss due to diffraction was found to depend critically upon whether a contact or immersion technique is used. For a contact method, we estimate that the diffraction loss can be greater than 10 dB MHz-1, whereas insertion devices typically will have losses of up to 0.6 dB MHz-1, and it not always clear whether or how manufacturers have attempted to correct for this. Dispersion which is found in the os calcis has only a small effect on the diffraction loss, but it can cause a frequency dependent interface loss of about 0.2 dB MHz-1. It may be impossible to correct for these discrepancies in vivo as the necessary data for the individual components of the heel are not known. However the losses should be borne in mind as factors limiting the accuracy of the measurements and, in the case of in vitro investigations, may merit further study.

Consistency in calibrated backscattered electron images of calcified tissues and minerals analyzed in multiple imaging sessions.

Pure metal standards have been used to calibrate the operating envionment in quatitative backscattered electron (BSE) imaging of mineralized tissue, allowing comparisons to be made between various mineralization states of bone at the microscopic level. It has not previuously been documented that calibration procedures produce consistent, reliable results over multiple imaging sessions. In this study, BSE images were obtained from bones, pure metals, and a naturally occurring mineral in multiple imaging sessions over a six day period. The graylevel histogram profile (GHP) from each specimen was analyzed for changes in the shape and relative placement on the graylevel spectrum. Computer controlled calibration and a restrospective calibration method using pure aluminum and pure magnesium-aluminum-zinc demonstrated consistency between imaging sessions. Calibrated weighted mean graylevels (WMGLs) for biological meterials had an average standard deviation of 5.9 graylevels (2.4% variation) during the course of the study. WMGLs for inorganic materials had an average standard deviation of 0.9 graylevels (0.4% variation). A trend towards increased image brightness, due to specimen and/or embedding media degradation, was observed in the biological tissues. No increase in rightness was observed for the inorgtanic specimens. Kurtosis and skewness tests revealed a slight deviation from normality in all specimens, which remained consistent between multiple imaging sessions. These results demonstrate the BSE image analysis of bones and mineral can be calibrated with negligible precision error allowing comparisons between data within and between multiple imaging sessions.

[Osteoid osteoma: ultrastructural aspects]. / L'osteoma osteoide: aspetti ultrastrutturali.

The authors take into consideration the ultrastructural aspects of osteoid osteoma and discuss the cellular and subcellular features of this neoplasm. Special attention is given to functional adaptability, which is specific for this type of cell, related to the degree of development of the neoformation.

Early vascular changes in rabbit subchondral bone after repetitive impulsive loading.

The sequence of vascular and bony changes that precedes experimental osteoarthrosis was observed in rabbits. The subchondral bone underlying the weight-bearing portion of the medial tibial condyle and the talocalcaneal joint were examined after two, three, and six weeks of 50-ms and 500-ms repetitive loading at 1 Hz for 40 minutes each day. Vascular alterations were evident in the subchondral bone of the talocalcaneal joint after three weeks of a 50-ms load regime. A 10% increase in bone mass and a significant increase in the number of small diameter vessels were observed after six weeks. No changes were observed in the tibial subchondral bone, consistent with the measured attenuation of load distal to this joint. No changes were observed in either tibial or calcaneal subchondral bone in the 500-ms load group. Impulsive loading promotes early vascular changes in subchondral bone, which are developed in response to both the magnitude and the rate of loading.