Dimorphic Mechanisms of Fragility in Diabetes Mellitus: the Role of Reduced Collagen Fibril Deformation.

Diabetes mellitus (DM) is an emerging metabolic disease, and the management of diabetic bone disease poses a serious challenge worldwide. Understanding the underlying mechanisms leading to high fracture risk in DM is hence of particular interest and urgently needed to allow for diagnosis and treatment optimization. In a case-control postmortem study, the whole 12th thoracic vertebra and cortical bone from the mid-diaphysis of the femur from male individuals with type 1 diabetes mellitus (T1DM) (n = 6; 61.3 ± 14.6 years), type 2 diabetes mellitus (T2DM) (n = 11; 74.3 ± 7.9 years), and nondiabetic controls (n = 18; 69.3 ± 11.5) were analyzed with clinical and ex situ imaging techniques to explore various bone quality indices. Cortical collagen fibril deformation was measured in a synchrotron setup to assess changes at the nanoscale during tensile testing until failure. In addition, matrix composition was analyzed including determination of cross-linking and non-crosslinking advanced glycation end-products like pentosidine and carboxymethyl-lysine. In T1DM, lower fibril deformation was accompanied by lower mineralization and more mature crystalline apatite. In T2DM, lower fibril deformation concurred with a lower elastic modulus and tendency to higher accumulation of non-crosslinking advanced glycation end-products. The observed lower collagen fibril deformation in diabetic bone may be linked to altered patterns mineral characteristics in T1DM and higher advanced glycation end-product accumulation in T2DM. © 2022 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Impact of test environment on the fracture resistance of cortical bone.

Water is a crucial component of bone, affecting the interplay of collagen and minerals and contributing to bone's high strength and ductility. Dehydration has been shown to significantly effect osseous mechanical properties; however, studies comparing the effects of various dehydrating environments on fracture toughness of bone are scarce. Accordingly, the crack resistance curve (R-curve) behavior of human and sheep cortical bone was characterized in a bio-bath, in ambient pressure air, and in scanning electron microscopes (SEMs) under three different environmental conditions (water vapor pressure, air pressure, and high-vacuum). The aim of this work was to better understand the impact of test environment on both intrinsic and extrinsic toughening and hence crack initiation toughness, K0 and crack growth resistance, dK/dΔa. Results show significantly lower K0 values for samples that were tested inside SEMs combined with pronounced extrinsic toughening through microcracking and crack path deflections out of the mode I plane. Importantly, all three SEM test environments gave similar results, and thus it does not matter which type of SEM is used. Ex situ testing of hydrated samples revealed similar K0 for both environments but elevated crack growth resistance for testing in ambient air relative to the bio-bath. Our data reveals the experimental difficulties to directly observe microscale crack propagation in cortical bone that resembles the in vivo situation. Ex situ testing immersed in Hanks' Balanced Salt Solution (HBSS) with subsequent crack path analysis, while tedious, is thought to presents the most realistic picture of the in vivo structure-fracture property relations in biological tissue.

Effect of variations in tissue-level ductility on human vertebral strength.

Although the ductility of bone tissue is a unique element of bone quality and varies with age and across the population, the extent to which and mechanisms by which typical population-variations in tissue-level ductility can alter whole-bone strength remains unclear. To provide insight, we conducted a finite element analysis parameter study of whole-vertebral (monotonic) compressive strength on six human L1 vertebrae. Each model was generated from micro-CT scans, capturing the trabecular micro-architecture in detail, and included a non-linear constitutive model for the bone tissue that allowed for plastic yielding, different strengths in tension and compression, large deformations, and, uniquely, localized damage once a specified limit in tissue-level ultimate strain was exceeded. Those strain limits were based on reported (mean ± SD) values from cadaver experiments (8.8 ± 3.7% strain for trabecular tissue and 2.2 ± 0.9% for cortical tissue). In the parameter study, the strain limits were varied by ±1 SD from their mean values, for a combination of nine analyses per specimen; bounding values of zero and unlimited post-yield strain were also modeled. The main outcomes from the finite element analysis were the vertebral compressive strength and the amount of failed (yielded or damaged) tissue at the overall structure-level failure. Compared to a reference case of using the mean values of ultimate strain, we found that varying both trabecular and cortical tissue ultimate strains by ±1 SD changed the computed vertebral strength by (mean ± SD) ±6.9 ± 1.1% on average. Mechanistically, that modest effect arose because the proportion of yielded tissue (without damage) was 0.9 ± 0.3% of all the bone tissue across the nine cases and the proportion of damaged tissue (i.e. tissue exceeding the prescribed tissue-level ultimate strain) was 0.2 ± 0.1%. If the types of variations in tissue-level ductility investigated here accurately represent real typical variations in the population, the consistency of our results across specimens and the modest effect size together suggest that typical variations in tissue-level ductility only have a modest impact on vertebral compressive strength, in large part because so few trabeculae are damaged at the load capacity of the bone.

Multiscale bone quality analysis in osteoarthritic knee joints reveal a role of the mechanosensory osteocyte network in osteophytes.

Osteophytes - bony outgrowths on joint structures - are found in healthy individuals but are specifically present in late osteoarthritis (OA). Osteophyte development and function is not well understood, yet biomechanical stimuli are thought to be critical. Bone adapts to mechanical forces via the cellular network of osteocytes. The involvement of osteocytes in osteophyte formation and maturation has not been unravelled. Forty-three osteophytes from tibias of 23 OA patients (65 ± 9 years) were analysed. The trabecular bone structure of osteophytes presented with fewer trabeculae of lower bone mineral density compared to subchondral bone. We identified 40% early stage and 60% late stage osteophytes that significantly differed in their trabecular bone characteristics. Osteophyte bone revealed a higher number of osteocytes and a lower number of empty osteocyte lacunae per bone area than the subchondral bone. We found that OA osteophytes consist of younger bone material comprised of woven and lamellar bone with the capacity to develop into a late stage osteophyte potentially via the involvement of the osteocyte network. Our analysis of OA osteophytes implies a transition from woven to lamellar bone as in physiological bone growth within a pathological joint. Therefore, osteophyte development and growth present a valuable research subject when aiming to investigate the osteogenic signalling cascade.

Bisphosphonate treatment changes regional distribution of trabecular microstructure in human lumbar vertebrae.

BACKGROUND: In osteoporosis patients, antiresorptive treatments such as alendronate reduce the resorption of trabecular bone and thus minimize vertebral fracture risk. However, fracture risk reduction efficacy of antiresorptive drugs varies between skeletal sites and is highest for vertebral bone. In human vertebrae, cancellous bone is distributed heterogeneously between regions. This microstructural heterogeneity is changing with patient age and is likely to play a major role in vertebral failure mechanisms and fracture susceptibility. Whether antiresorptive treatment affects the heterogeneity of vertebral microstructure in osteoporosis has not been unraveled. METHODS: Our aim was to assess whether antiresorptive treatment would have a region-dependent influence on vertebral trabecular bone. Therefore, we used high-resolution peripheral quantitative computed tomography (HR-pQCT), microcomputed tomography (microCT) and uniaxial compression testing to determine the structure and mechanical properties of trabecular bone cores from anterior and posterior regions of 22 lumbar vertebrae from elderly osteoporotic women. We analyzed age-matched ex vivo bone samples from bisphosphonate-treated female osteoporosis patients (age: 82 ±â€¯7y, bisphosphonate treatment period: 4 ±â€¯2 years) along treatment-naïve female controls (82 ±â€¯7y). RESULTS: MicroCT analysis showed a significantly lower bone volume fraction (p = 0.006) and lower trabecular number (p = 0.003) for the anterior bone cores compared to posterior bone cores in the treatment-naïve group. The bisphosphonate-treated group had a more homogeneous bone volume distribution and did not show significant regional differences in bone volume, it however also displayed significantly different trabecular numbers (p = 0.016). In bone cores of the bisphosphonate-treated group, trabeculae were thicker in comparison to treatment-naïve controls (p = 0.011). Differences in bone volume further resulted in different maximum forces during compression testing between the samples. In addition, the percental difference between BV/TVµCT in anterior and posterior bone cores was lower in bisphosphonate-treated vertebrae when vertebrae with directly adjacent fractures (n = 3) were excluded. CONCLUSION: In conclusion, regional trabecular bone microstructure in lumbar vertebrae of bisphosphonate-treated women was more homogeneous compared to treatment-naïve controls. Bisphosphonate treatment, which specifically targets resorption surfaces common in anterior vertebral bone, might have resulted in a region-specific preservation of vertebral microstructure and loading capacity. This could have positive implications for the reduction of wedge fracture risk and add to the explanation of the higher efficacy of fracture risk reduction in vertebrae in comparison to other fracture regions.

Subregional areal bone mineral density (aBMD) is a better predictor of heterogeneity in trabecular microstructure of vertebrae in young and aged women than subregional trabecular bone score (TBS).

BACKGROUND: Currently, bone densitometry fails to identify nearly half of those elderly patients at immediate fracture risk. To improve clinical assessment of vertebral fracture risk, we aimed to determine how the DXA-based 2D parameter Trabecular Bone Score (TBS) relates to subregional variability in 3D trabecular microstructure in young and elderly women compared to aBMD. METHODS: T12 vertebrae from 29 women (11 young: 32 ±â€¯6 years, 18 aged: 71 ±â€¯5 years) were DXA-scanned ex vivo in anterior-posterior (AP) and lateral projection providing vertebral aBMD and TBS. Additionally, aBMD and TBS were measured for three horizontal (superior, mid-horizontal, inferior) and three vertical subregions (anterior, mid-vertical, posterior) and related to 3D microstructure indices, i.e. bone volume per tissue volume (BV/TV), trabecular number and thickness (Tb.N, Tb.Th), based on HRpQCT. RESULTS: Subregional high-resolution tomography showed significant differences in trabecular parameters for both age groups: In horizontal subregions, BV/TV was lowest superiorly, Tb.Th was highest mid-horizontally, and Tb.N was lowest mid-horizontally and highest inferiorly. Correspondingly, aBMD varied between horizontal subregions, with differences depending on projection direction. TBS varied only in lateral projections of the aged group, with lower values for the mid-horizontal subregion. In vertical subregions, BV/TV, Tb.N, and aBMD were highest posteriorly for both groups. TBS did not differ between vertical subregions. Regression analysis showed aBMD as a predictor explained more of the variance in subregional 3D microstructure compared to TBS. Stepwise multi-regression analysis revealed only three combinations of subregion, projection, and group where aBMD and TBS were both significant predictors. CONCLUSIONS: Subregional aBMD reflects variations in trabecular bone microstructure better than subregional TBS for trisected regions. Specifically, lateral aBMD identifies microstructural heterogeneities independent of age and may improve prediction of vertebral strength and susceptibility to specific fracture types.

Publisher Correction: Calcium and vitamin-D deficiency marginally impairs fracture healing but aggravates posttraumatic bone loss in osteoporotic mice.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Inter-site variability of the osteocyte lacunar network in the cortical bone underpins fracture susceptibility of the superolateral femoral neck.

BACKGROUND: The osteocytic lacunar network is considered to be an integral player in the regulation of bone homeostasis, and reduction in osteocytes is associated with reduced bone strength. Here, we analyzed site-specific patterns in osteocyte characteristics and matrix composition in the cortical compartment of the femoral neck to reveal the structural basis of its fragility. METHODS: Cross-sections of the human femoral neck - one of the most common fracture sites - were acquired from 12 female cadavers (age 34-86 years) and analyzed with backscattered scanning electron microscopy and high-resolution micro-computed tomography (µ-CT). The 2D/3D density and size of the osteocyte lacunae as well as bone mineral density distribution (BMDD) were measured in two regions subject to different biomechanical loads in vivo: the inferomedial (medial) region (habitually highly loaded in compression) and the superolateral (lateral) region (lower habitual loading intensity). Using quantitative polarized light microscopy, collagen fiber orientation was quantified in these two regions, accordingly. RESULTS: In 2D measurements, the inferomedial region displayed lower mineralization heterogeneity, 19% higher osteocyte lacunar density (p = 0.005), but equal lacunar size compared to the superolateral region. 3D measurements confirmed a significantly higher osteocyte lacunar density in the inferomedial region (p = 0.015). Osteocyte lacunar density decreased in aged individuals, and inter-site differences were reduced. Site-specific osteocyte characteristics were not accompanied by changes in collagen fiber orientation. CONCLUSIONS: Since osteocyte characteristics may provide valuable insights into bone mechanical competence, the variations in osteocyte properties might reflect the increased fracture susceptibility of the superolateral neck.

Increased mechanical loading through controlled swimming exercise induces bone formation and mineralization in adult zebrafish.

Exercise promotes gain in bone mass through adaptive responses of the vertebrate skeleton. This mechanism counteracts age- and disease-related skeletal degradation, but remains to be fully understood. In life sciences, zebrafish emerged as a vertebrate model that can provide new insights into the complex mechanisms governing bone quality. To test the hypothesis that musculoskeletal exercise induces bone adaptation in adult zebrafish and to characterize bone reorganization, animals were subjected to increased physical exercise for four weeks in a swim tunnel experiment. Cellular, structural and compositional changes of loaded vertebrae were quantified using integrated high-resolution analyses. Exercise triggered rapid bone adaptation with substantial increases in bone-forming osteoblasts, bone volume and mineralization. Clearly, modeling processes in zebrafish bone resemble processes in human bone. This study highlights how exercise experiments in adult zebrafish foster in-depth insight into aging-related bone diseases and can thus catalyze the search for appropriate prevention and new treatment options.

Bone tissue aging affects mineralization of cement lines.

Cement lines are known as thin peripheral boundaries of the osteons. With a thickness below 5 µm their composition of inorganic and organic compounds has been a matter of debate. Here, we hypothesized that cement lines become hypermineralized and their degree of mineralization is not constant but related to the tissue age of the osteon. Therefore, we analyzed the calcium content of osteons and their corresponding cement lines in a range of different tissue ages reflected by osteonal mineralization levels in femoral cortical bone of both postmenopausal women with osteoporosis and bisphosphonate-treated cases. Quantitative backscattered electron imaging (qBEI) showed that cement lines are hypermineralized entities with consistently higher calcium content than their corresponding osteons (mean calcium content: 29.46 ±â€¯0.80 vs. 26.62 ±â€¯1.11 wt%; p < 0.001). Micro-Raman spectroscopy complemented the qBEI data by showing a significantly higher phosphate/amide I ratio in the cement lines compared to the osteonal bone (8.78 ±â€¯0.66 vs. 6.33 ±â€¯0.58, p < 0.001), which was both due to an increased phosphate peak and a reduced amide I peak in cement lines. A clear positive correlation of cement line mineralization and the mineralization of the osteon was observed (r = 0.839, p = 0.003). However, the magnitude of the difference between cement line and osteonal calcium content decreased with increased osteonal calcium content (r = -0.709, p < 0.001), suggesting diverging mineralization dynamics in these osseous entities. The number of mineralized osteocyte lacunae per osteon bone area correlated positively with both osteonal and cement line calcium content (p < 0.01). The degree of mineralization of cement lines may represent another tissue-age related phenomenon, given that it strongly relates to the osteonal mineralization level. Understanding of the cement lines' mineralization and their changes in aging and disease states is important for predicting crack propagation pathways and fracture resistance mechanisms in human cortical bone.

Calcium and vitamin-D deficiency marginally impairs fracture healing but aggravates posttraumatic bone loss in osteoporotic mice.

Calcium and vitamin-D (Ca/VitD) deficiency is a major risk factor for osteoporosis. It may also contribute to the compromised bone healing frequently observed in osteoporotic patients, since calcium is essential for fracture-callus mineralization. Additionally, clinical data suggest systemic bone loss following fracture, which may aggravate osteoporosis and thus increase the risk for fragility fractures in osteoporotic patients further. However, the role of Ca/VitD in fracture healing and posttraumatic bone turnover has to date been poorly investigated. Here, we studied bone regeneration and posttraumatic bone turnover in C57BL/6 J mice with ovariectomy-induced osteoporosis. Mice were fed a standard or a Ca/VitD-deficient diet. Notably, fracture healing was only marginally disturbed in Ca/VitD-deficient mice. However, deficient mice displayed significantly increased serum parathyroid hormone levels and osteoclast activity, as well as reduced bone mass in the intact skeleton post-fracture, suggesting considerably enhanced calcium mobilization from the intact skeleton during bone regeneration. Ca/VitD supplementation initiated post-fracture prevented posttraumatic bone loss by reducing bone resorption and furthermore improved bone repair. These results imply that adequate Ca/VitD supply post-fracture is essential to provide sufficient calcium for callus-mineralization in order to prevent posttraumatic bone loss and to reduce the risk for secondary fractures in osteoporotic patients with Ca/VitD deficiency.

The Formation of Calcified Nanospherites during Micropetrosis Represents a Unique Mineralization Mechanism in Aged Human Bone.

Osteocytes-the central regulators of bone remodeling-are enclosed in a network of microcavities (lacunae) and nanocanals (canaliculi) pervading the mineralized bone. In a hitherto obscure process related to aging and disease, local plugs in the lacuno-canalicular network disrupt cellular communication and impede bone homeostasis. By utilizing a suite of high-resolution imaging and physics-based techniques, it is shown here that the local plugs develop by accumulation and fusion of calcified nanospherites in lacunae and canaliculi (micropetrosis). Two distinctive nanospherites phenotypes are found to originate from different osteocytic elements. A substantial deviation in the spherites' composition in comparison to mineralized bone further suggests a mineralization process unlike regular bone mineralization. Clearly, mineralization of osteocyte lacunae qualifies as a strong marker for degrading bone material quality in skeletal aging. The understanding of micropetrosis may guide future therapeutics toward preserving osteocyte viability to maintain mechanical competence and fracture resistance of bone in elderly individuals.

Magnetic Particle / Magnetic Resonance Imaging: In-Vitro MPI-Guided Real Time Catheter Tracking and 4D Angioplasty Using a Road Map and Blood Pool Tracer Approach.

PURPOSE: In-vitro evaluation of the feasibility of 4D real time tracking of endovascular devices and stenosis treatment with a magnetic particle imaging (MPI) / magnetic resonance imaging (MRI) road map approach and an MPI-guided approach using a blood pool tracer. MATERIALS AND METHODS: A guide wire and angioplasty-catheter were labeled with a thin layer of magnetic lacquer. For real time MPI a custom made software framework was developed. A stenotic vessel phantom filled with saline or superparamagnetic iron oxide nanoparticles (MM4) was equipped with bimodal fiducial markers for co-registration in preclinical 7T MRI and MPI. In-vitro angioplasty was performed inflating the balloon with saline or MM4. MPI data were acquired using a field of view of 37.3×37.3×18.6 mm3 and a frame rate of 46 volumes/sec. Analysis of the magnetic lacquer-marks on the devices were performed with electron microscopy, atomic absorption spectrometry and micro-computed tomography. RESULTS: Magnetic marks allowed for MPI/MRI guidance of interventional devices. Bimodal fiducial markers enable MPI/MRI image fusion for MRI based roadmapping. MRI roadmapping and the blood pool tracer approach facilitate MPI real time monitoring of in-vitro angioplasty. Successful angioplasty was verified with MPI and MRI. Magnetic marks consist of micrometer sized ferromagnetic plates mainly composed of iron and iron oxide. CONCLUSIONS: 4D real time MP imaging, tracking and guiding of endovascular instruments and in-vitro angioplasty is feasible. In addition to an approach that requires a blood pool tracer, MRI based roadmapping might emerge as a promising tool for radiation free 4D MPI-guided interventions.

Hypochlorhydria-induced calcium malabsorption does not affect fracture healing but increases post-traumatic bone loss in the intact skeleton.

Efficient calcium absorption is essential for skeletal health. Patients with impaired gastric acidification display low bone mass and increased fracture risk because calcium absorption is dependent on gastric pH. We investigated fracture healing and post-traumatic bone turnover in mice deficient in Cckbr, encoding a gastrin receptor that affects acid secretion by parietal cells. Cckbr-/- mice display hypochlorhydria, calcium malabsorption, and osteopenia. Cckbr-/- and wildtype (WT) mice received a femur osteotomy and were fed either a standard or calcium-enriched diet. Healed and intact bones were assessed by biomechanical testing, histomorphometry, micro-computed tomography, and quantitative backscattering. Parathyroid hormone (PTH) serum levels were determined by enzyme-linked immunosorbent assay. Fracture healing was unaffected in Cckbr-/- mice. However, Cckbr-/- mice displayed increased calcium mobilization from the intact skeleton during bone healing, confirmed by significantly elevated PTH levels and osteoclast numbers compared to WT mice. Calcium supplementation significantly reduced secondary hyperparathyroidism and bone resorption in the intact skeleton in both genotypes, but more efficiently in WT mice. Furthermore, calcium administration improved bone healing in WT mice, indicated by significantly increased mechanical properties and bone mineral density of the fracture callus, whereas it had no significant effect in Cckbr-/- mice. Therefore, under conditions of hypochlorhydria-induced calcium malabsorption, calcium, which is essential for callus mineralization, appears to be increasingly mobilized from the intact skeleton in favor of fracture healing. Calcium supplementation during fracture healing prevented systemic calcium mobilization, thereby maintaining bone mass and improving fracture healing in healthy individuals whereas the effect was limited by gastric hypochlorhydria. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:1914-1921, 2016.

Addition of a Fluoride-containing Radiopacifier Improves Micromechanical and Biological Characteristics of Modified Calcium Silicate Cements.

INTRODUCTION: Calcium silicate cements (CSCs) with the addition of nanohydroxyapatite and calcium carbonate play a critical role in dental applications. To further improve their properties, particularly radiopacity and biointeractivity, the fluoride-containing radiopacifier ytterbium trifluoride (YbF3) was added to their composition, and biological and mechanical characteristics were evaluated. METHODS: YbF3 was added to 3 different CSCs: cement I (CSC + calcium carbonate), cement II (CSC + nanohydroxyapatite), and Portland cement. Material characterization encompassed measurements of pH, calcium, ytterbium, and fluoride ion release; radiopacity; setting time; porosity; microindentation properties; wettability; and Fourier transform infrared spectroscopic, x-ray diffraction, and scanning electron microscopic analyses. Osteoblast- and osteoclast-like cells were grown on the materials' surface to evaluate their adherence. RESULTS: The addition of calcium carbonate, nanohydroxyapatite, and 30 wt% of YbF3 improved radiopacity and the setting time of experimental cements. The pH values did not differ among the groups. The greatest ytterbium and fluoride releases occurred in the Portland cement + YbF3 group. Combined x-ray diffraction and Fourier transform infrared spectroscopic analysis showed the presence of calcium hydroxide and calcium silicate hydrates. In addition, the presence of calcium ytterbium fluoride and ytterbium oxide proved that YbF3 reacted with cement compounds. Wettability of cement I + YbF3 was superior to other formulations, but its porosity and microindentation properties were weaker than in the Portland cement + YbF3 mixture. Cement II + YbF3 presented micromechanical indentation and porosity characteristics similar to the Portland-based cement formulation. Osteoclast- and osteoblast-like cells adhered to the cements' surfaces without alteration of the cell structural integrity. CONCLUSIONS: YbF3-containing CSCs with nanostructured hydroxyapatite and calcium carbonate are well suited for dental application.

Multi-level characterization of human femoral cortices and their underlying osteocyte network reveal trends in quality of young, aged, osteoporotic and antiresorptive-treated bone.

Characterization of bone's hierarchical structure in aging, disease and treatment conditions is imperative to understand the architectural and compositional modifications to the material and its mechanical integrity. Here, cortical bone sections from 30 female proximal femurs - a frequent fracture site - were rigorously assessed to characterize the osteocyte lacunar network, osteon density and patterns of bone matrix mineralization by backscatter-electron imaging and Fourier-transform infrared spectroscopy in relation to mechanical properties obtained by reference-point indentation. We show that young, healthy bone revealed the highest resistance to mechanical loading (indentation) along with higher mineralization and preserved osteocyte-lacunar characteristics. In contrast, aging and osteoporosis significantly alter bone material properties, where impairment of the osteocyte-lacunar network was evident through accumulation of hypermineralized osteocyte lacunae with aging and even more in osteoporosis, highlighting increased osteocyte apoptosis and reduced mechanical competence. But antiresorptive treatment led to fewer mineralized lacunae and fewer but larger osteons signifying rejuvenated bone. In summary, multiple structural and compositional changes to the bone material were identified leading to decay or maintenance of bone quality in disease, health and treatment conditions. Clearly, antiresorptive treatment reflected favorable effects on the multifunctional osteocytic cells that are a prerequisite for bone's structural, metabolic and mechanosensory integrity.