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Age-related changes in bone microstructure can inform our understanding the biology of both extant and fossil birds, but to date, histological work in birds, and particularly work using high-resolution 3D imaging, has largely been restricted to limited growth stages. We used minimally destructive synchrotron radiation-based X-ray computed tomography to visualise and measure key morphological and histological traits in 3D across development in the domestic duck and ring-necked pheasant. We use these measurements to build on the database of key reference material for interpreting bone histology. We found that growth patterns differed between the two species, with the ducks showing rapid growth in their lower limbs and early lower limb maturation, while pheasants grew more slowly, reflecting their later age at maturity. In the pheasant, both walking and flight occur early and their upper and lower limbs grew at similar rates. In the duck, flight and wing development are delayed until the bird is almost at full body mass. Through juvenile development, the second moment of area for the duck wing was low but increased rapidly towards the age of flight, at which point it became significantly greater than that of the lower limb, or the pheasant. On a microstructural level, both cortical porosity and canal diameter were related to cortical bone deposition rate. In terms of orientation, vascular canals in the bone cortex were more laminar in the humerus and femur compared with the tibiotarsus, and laminarity increased through juvenile development in the humerus, but not the tibiotarsus, possibly reflecting torsional vs compressive loading. These age-related changes in cortical bone vascular microstructure of the domestic duck and pheasant will help understanding the biology of both extant and fossil birds, including age estimation, growth rate and growth patterns, and limb function., (© 2022 The Authors. Journal of Anatomy published by John Wiley & Sons Ltd on behalf of Anatomical Society.)

Morphological variation in the hominoid capitate has been linked to differences in habitual locomotor activity due to its importance in movement and load transfer at the midcarpal joint proximally and carpometacarpal joints distally. Although the shape of bones and their articulations are linked to joint mobility, the internal structure of bones has been shown experimentally to reflect, at least in part, the loading direction and magnitude experienced by the bone. To date, it is uncertain whether locomotor differences among hominoids are reflected in the bone microarchitecture of the capitate. Here, we apply a whole-bone methodology to quantify the cortical and trabecular architecture (separately and combined) of the capitate across bipedal (modern Homo sapiens), knuckle-walking (Pan paniscus, Pan troglodytes, Gorilla sp.), and suspensory (Pongo sp.) hominoids (n = 69). It is hypothesized that variation in bone microarchitecture will differentiate these locomotor groups, reflecting differences in habitual postures and presumed loading force and direction. Additionally, it is hypothesized that trabecular and cortical architecture in the proximal and distal regions, as a result of being part of mechanically divergent joints proximally and distally, will differ across these portions of the capitate. Results indicate that the capitate of knuckle-walking and suspensory hominoids is differentiated from bipedal Homo primarily by significantly thicker distal cortical bone. Knuckle-walking taxa are further differentiated from suspensory and bipedal taxa by more isotropic trabeculae in the proximal capitate. An allometric analysis indicates that size is not a significant determinate of bone variation across hominoids, although sexual dimorphism may influence some parameters within Gorilla. Results suggest that internal trabecular and cortical bone is subjected to different forces and functional adaptation responses across the capitate (and possibly other short bones). Additionally, while separating trabecular and cortical bone is normal protocol of current whole-bone methodologies, this study shows that when applied to carpals, removing or studying the cortical bone separately potentially obfuscates functionally relevant signals in bone structure., (© 2021 The Authors. Journal of Anatomy published by John Wiley & Sons Ltd on behalf of Anatomical Society.)

The human clavicle (i.e. collarbone) is an unusual long bone due to its signature S-shaped curve and variability in macrostructure observed between individuals. Because of the complex nature of how the upper limb moves, as well as due to its complex musculoskeletal arrangement, the biomechanics, in particular the mechanical loadings, of the clavicle are not fully understood. Given that bone remodeling can be influenced by bone stress, the histologic organization of Haversian bone offers a hypothesis of responses to force distributions experienced across a bone. Furthermore, circularly polarized light microscopy can be used to determine the orientation of collagen fibers, providing additional information on how bone matrix might organize to adapt to direction of external loads. We examined Haversian density and collagen fiber orientation, along with cross-sectional geometry, to test whether the clavicle midshaft shows unique adaptation to atypical load-bearing when compared with the sternal (medial) and acromial (lateral) shaft regions. Because fractures are most common at the midshaft, we predicted that the cortical bone structure would show both disparities in Haversian remodeling and nonrandomly oriented collagen fibers in the midshaft compared with the sternal and acromial regions. Human clavicles (n = 16) were sampled via thin-sections at the sternal, middle, and acromial ends of the shaft, and paired sample t-tests were employed to evaluate within-individual differences in microstructural or geometric properties. We found that Haversian remodeling is slightly but significantly reduced in the middle of the bone. Analysis of collagen fiber orientation indicated nonrandom fiber orientations that are overbuilt for tensile loads or torsion but are poorly optimized for compressive loads throughout the clavicle. Geometric properties of percent bone area, polar second moment of area, and shape (I max /I min ) confirmed the conclusions drawn by existing research on clavicle macrostructure. Our results highlight that mediolateral shape changes might be accompanied by slight changes in Haversian density, but bone matrix organization is predominantly adapted to resisting tensile strains or torsion throughout and may be a major factor in the risk of fracture when experiencing atypical compression., (© 2019 Anatomical Society.)

Here we present detailed regional bone thickness and cross-sectional measurements from full adult ribs using high resolution CT scans processed with a cortical bone mapping technique. Sixth ribs from 33 subjects ranging from 24 to 99 years of age were used to produce average cortical bone thickness maps and to provide average ± 1SD corridors for expected cross-section properties (cross-sectional areas and inertial moments) as a function of rib length. Results obtained from CT data were validated at specific rib locations using direct measurements from cut sections. Individual thickness measurements from CT had an accuracy (mean error) and precision (SD error) of -0.013 ± 0.167 mm (R 2 coefficient of determination of 0.84). CT-based measurement errors for rib cross-sectional geometry were -0.1 ± 13.1% (cortical bone cross-sectional area) and 4.7 ± 1.8% (total cross-sectional area). Rib cortical bone thickness maps show the expected regional variation across a typical rib's surface. The local mid-rib maxima in cortical thickness along the pleural rib aspect ranged from range 0.9 to 2.6 mm across the study population with an average map maximum of 1.4 mm. Along the cutaneous aspect, rib cortical bone thickness ranged from 0.7 to 1.9 mm with an average map thickness of 0.9 mm. Average cross-sectional properties show a steady reduction in total cortical bone area from 10% along the rib's length through to the sternal end, whereas overall cross-sectional area remains relatively constant along the majority of the rib's length before rising steeply towards the sternal end. On average, male ribs contained more cortical bone within a given cross-section than was seen for female ribs. Importantly, however, this difference was driven by male ribs having larger overall cross-sectional areas, rather than by sex differences in the bone thickness observed at specific local cortex sites. The cortical bone thickness results here can be used directly to improve the accuracy of current human body and rib models. Furthermore, the measurement corridors obtained from adult subjects across a wide age range can be used to validate future measurements from more widely available image sources such as clinical CT where gold standard reference measures (e.g. such as direct measurements obtained from cut sections) are otherwise unobtainable., (© 2019 Anatomical Society.)

Bone remodeling, one of the main processes that regulate bone microstructure, consists of bone resorption followed by the deposition of secondary bone at the same location. Remodeling intensity varies among taxa, but a characteristically compact cortex is ubiquitous in the long bones of mature terrestrial mammals. A previous analysis found that cortical bone in a few 'tree sloth' (Bradypus and Choloepus) specimens is heavily remodeled and characterized by numerous immature secondary osteons, suggesting that these animals were remodeling their bones at high rate until late in their ontogeny. This study aims at testing if this remodeling is generally present in 'tree sloths', using a quantitative analysis of the humeral cortical compactness (CC) among xenarthrans. The results of the investigation of humeral diaphyseal cross-sections of 26 specimens belonging to 10 xenarthran species including specimens from both extinct and extant species indicate that in 'tree sloths' the CC is significantly lower than in the other sampled xenarthrans. No significant difference was found between the CC of the two genera of 'tree sloths'. Our results are consistent with the hypothesis that the cortical bone of 'tree sloths' in general undergoes intense and balanced remodeling that is maintained until late (possibly throughout) in their ontogeny. In the light of xenarthran phylogeny, low CC represents another convergence between the long-separated 'tree sloth' lineages. Although the exact structural and/or functional demands that are associated with this trait are hitherto unknown, several hypotheses are suggested here, including a relationship to their relatively low metabolism and to the mechanical demands imposed upon the bones by the suspensory posture and locomotion, which was independently acquired by the two genera of 'tree sloths'., (© 2018 Anatomical Society.)

Vascular canals in cortical bone during growth and development typically show an anisotropic pattern with canals falling into three main categories: circumferential, radial, and longitudinal. Two major hypotheses attempt to explain the preferred orientations in bone: that vascular canal orientation is optimized to resist a predominant strain direction from functional loading, or that it reflects growth requirements and velocity. We use a controlled growth experiment in broiler chickens to investigate the effect of growth rate on vascular canal orientation. Using feed restriction we set up a fast growing control group and a slow growing restricted group. We compared the microstructure in the humerus and the femur at 42 days of age using synchrotron micro-computed tomography (micro-CT), a three-dimensional (3D) method that visualizes the full canal network. We measured the 3D orientation of each canal in the whole cross-section of the bone cortex using a set of custom ImageJ scripts. Using these orientations we compute laminar, radial, and longitudinal indices that measure the proportion of circumferential, radial, and longitudinal canals, by unit of length, in the cortex. Following previous studies we hypothesized that vascular canal orientation is related to growth, with radial canals linked to a faster growth rate and related to functional loading through a high laminar index in flight bones which reflects torsional loading resulting from active flight. The control group had final body weights that were nearly twice the final weights of the restricted group and higher absolute growth rates. We found consistent patterns in the comparison between the humerus and the femur in both groups, with the humerus having higher laminar and longitudinal indices, and a lower radial index than the femur. The control group had higher radial indices and lower laminar and longitudinal indices in both the humerus and the femur than the restricted group. The higher radial indices in our control group point to a link between radial canals and faster growth, and between laminar canals and slower growth, while the higher laminar indices in the humerus point to a link between circumferential canals and torsional loading. Overall, our results indicate that the orientation of the cortical canal network in a bone is the consequence of a complex interaction between the growth rate of that bone and functional loading environment., (© 2018 The Authors. Journal of Anatomy published by John Wiley & Sons Ltd on behalf of Anatomical Society.)

Cortical bone porosity and specifically the orientation of vascular canals is an area of growing interest in biomedical research and comparative/paleontological anatomy. The potential to explain microstructural adaptation is of great interest. However, the determinants of the development of canal orientation remain unclear. Previous studies of birds have shown higher proportions of circumferential canals (called laminarity) in flight bones than in hindlimb bones, and interpreted this as a sign that circumferential canals are a feature for resistance to the torsional loading created by flight. We defined the laminarity index as the percentage of circumferential canal length out of the total canal length. In this study we examined the vascular canal network in the humerus and femur of a sample of 31 bird and 24 bat species using synchrotron micro-computed tomography (micro-CT) to look for a connection between canal orientation and functional loading. The use of micro-CT provides a full three-dimensional (3D) map of the vascular canal network and provides measurements of the 3D orientation of each canal in the whole cross-section of the bone cortex. We measured several cross-sectional geometric parameters and strength indices including principal and polar area moments of inertia, principal and polar section moduli, circularity, buckling ratio, and a weighted cortical thickness index. We found that bat cortices are relatively thicker and poorly vascularized, whereas those of birds are thinner and more highly vascularized, and that according to our cross-sectional geometric parameters, bird bones have a greater resistance to torsional stress than the bats; in particular, the humerus in birds is more adapted to resist torsional stresses than the femur. Our results show that birds have a significantly (P = 0.031) higher laminarity index than bats, with birds having a mean laminarity index of 0.183 in the humerus and 0.232 in the femur, and bats having a mean laminarity index of 0.118 in the humerus and 0.119 in the femur. Counter to our expectation, the birds had a significantly higher laminarity index in the femur than in the humerus (P = 0.035). To evaluate whether this discrepancy was a consequence of methodology we conducted a comparison between our 3D method and an analogue to two-dimensional (2D) histological measurements. This comparison revealed that 2D methods significantly underestimate (P < 0.001) the amount of longitudinal canals by an average of 20% and significantly overestimate (P < 0.001) the laminarity index by an average of 7.7%, systematically mis-estimating indices of vascular canal orientations. In comparison with our 3D results, our approximated 2D measurement had the same results for comparisons between the birds and bats but found significant differences only in the longitudinal index between the humerus and the femur for both groups. The differences between our 3D and pseudo-2D results indicate that differences between our findings and the literature may be partially based in methodology. Overall, our results do not support the hypothesis that the bones of flight are more laminar, suggesting a complex relation between functional loading and microstructural adaptation., (© 2018 The Authors. Journal of Anatomy published by John Wiley & Sons Ltd on behalf of Anatomical Society.)

The histomorphometric properties of the subtrochanteric femoral region have rarely been investigated. The aim of this study was to investigate the age-associated variations and regional differences of histomorphometric and osteocytic properties in the cortical bone of the subtrochanteric femoral shaft, and the association between osteocytic and histological cortical bone parameters. Undecalcified histological sections of the subtrochanteric femoral shaft were obtained from cadavers (n = 20, aged 18-82 years, males). They were cut and stained using modified Masson-Goldner stain. Histomorphometric parameters of cortical bone were analysed with ×50 and ×100 magnification after identifying cortical bone boundaries using our previously validated method. Within cortical bone areas, only complete osteons with typical concentric lamellae and cement line were selected and measured. Osteocytic parameters of cortical bone were analyzed under phase contrast microscopy and epifluorescence within microscopic fields (0.55 mm 2 for each). The cortical widths of the medial and lateral quadrants were significantly higher than other quadrants (P < 0.01). Osteonal area per cortical bone area was lower and cortical porosities were higher in the posterior quadrant than in the other quadrants (P < 0.05). Osteocyte lacunar number per cortical bone area was found higher in the young subjects (≤ 50 years) than in the older ones (> 50 years) both before and after adjustments for body height and weight (P < 0.05). Moreover, significant but low correlations were found between the cortical bone and osteocytic parameters (0.20 ≤ R 2  ≤ 0.35, P < 0.05). It can be concluded that in healthy males, the cortical histomorphometric parameters differ between the anatomical regions of the subtrochanteric femoral shaft, and are correlated with the osteocytic parameters from the same site. These findings may be of use when discussing mechanisms that predispose patients to decreasing bone strength., (© 2017 Anatomical Society.)