Alterations of bone material properties in adult patients with X-linked hypophosphatemia (XLH).

X-linked hypophosphatemia (XLH) caused by PHEX mutations results in elevated serum FGF23 levels, renal phosphate wasting and low 1,25-dihydroxyvitamin D. The glycophosphoprotein osteopontin, a potent inhibitor of mineralization normally degraded by PHEX, accumulates within the bone matrix. Conventional therapy consisting of supplementation with phosphate and vitamin D analogs is burdensome and the effects on bone material poorly characterized. We analyzed transiliac bone biopsies from four adult patients, two of them severely affected due to no diagnosis and no treatment until adulthood. We used light microscopy, qBEI and FTIRI to study histology, histomorphometry, bone mineralization density distribution, properties of the organic matrix and size of hypomineralized periosteocytic lesions. Non-treatment resulted in severe osteomalacia, twice the amount of mineralized trabecular volume, multiple osteon-like perforations, continuity of lamellae from mineralized to unmineralized areas and distinctive patches of woven bone. Periosteocytic lesions were larger than in treated patients. The latter had nearly normal osteoid thicknesses, although surface was still elevated. The median calcium content of the matrix was always within normal range, although the percentage of lowly mineralized bone areas was highly increased in non-treated patients, resulting in a marked heterogeneity in mineralization. Divalent collagen cross-links were evident independently of the mineral content of the matrix. Broad osteoid seams lacked measurable pyridinoline, a mature trivalent cross-link and exhibited considerable acidic lipid content, typically found in matrix vesicles. Based on our results, we propose a model that possibly integrates the relationship between the observed mineralization disturbances, FGF23 secretion and the known osteopontin accumulation in XLH.

Registering 2D and 3D imaging data of bone during healing.

UNLABELLED: PURPOSE/AIMS OF THE STUDY: Bone's hierarchical structure can be visualized using a variety of methods. Many techniques, such as light and electron microscopy generate two-dimensional (2D) images, while micro-computed tomography (µCT) allows a direct representation of the three-dimensional (3D) structure. In addition, different methods provide complementary structural information, such as the arrangement of organic or inorganic compounds. The overall aim of the present study is to answer bone research questions by linking information of different 2D and 3D imaging techniques. A great challenge in combining different methods arises from the fact that they usually reflect different characteristics of the real structure. MATERIALS AND METHODS: We investigated bone during healing by means of µCT and a couple of 2D methods. Backscattered electron images were used to qualitatively evaluate the tissue's calcium content and served as a position map for other experimental data. Nanoindentation and X-ray scattering experiments were performed to visualize mechanical and structural properties. RESULTS: We present an approach for the registration of 2D data in a 3D µCT reference frame, where scanning electron microscopies serve as a methodic link. Backscattered electron images are perfectly suited for registration into µCT reference frames, since both show structures based on the same physical principles. We introduce specific registration tools that have been developed to perform the registration process in a semi-automatic way. CONCLUSIONS: By applying this routine, we were able to exactly locate structural information (e.g. mineral particle properties) in the 3D bone volume. In bone healing studies this will help to better understand basic formation, remodeling and mineralization processes.

Hierarchically structured vanadium pentoxide-polymer hybrid materials.

Biomimetic composite materials consisting of vanadium pentoxide (V2O5) and a liquid crystal (LC) "gluing" polymer were manufactured exhibiting six structural levels of hierarchy, formed through LC phases. The organic matrix was a polyoxazoline with pendant cholesteryl and carboxyl units, forming a lyotropic phase with the same structural orientation extending up to hundreds of micrometers upon shearing, and binding to V2O5 via hydrogen bridges. Composites consisting of V2O5-LC polymer hybrid fibers with a pronounced layered structuring were obtained. The V2O5-LC polymer hybrid fibers consist of aligned V2O5 ribbons, composed of self-assembled V2O5 sheets, encasing a chiral nematic polymer matrix. The structures of the V2O5-LC polymer composites strongly depend on the preparation method, i.e., the phase-transfer method from aqueous to organic medium, in which the polymer forms LC phases. Notably, highly defined micro- and nanostructures were obtained when initiating the synthesis using V2O5 tactoids with preoriented nanoparticle building units, even when using isotropic V2O5 dispersions. Shear-induced hierarchical structuring of the composites was observed, as characterized from the millimeter and micrometer down to the nanometer length scales using complementary optical and electron microscopy, SAXS, µCT, and mechanical nanoindentation.

Structural and mechanical properties of the arthropod cuticle: comparison between the fang of the spider Cupiennius salei and the carapace of American lobster Homarus americanus.

Most biological materials are nanocomposites characterized by a multi-level structural hierarchy. Particularly, the arthropod cuticle is a chitin-based composite material where the mechanical properties strongly depend on both molecular chitin/protein properties, and the structural arrangement of chitin-fibrils within the protein matrix. Here materials properties and structural organization of two types of cuticle from distantly related arthropods, the wandering spider Cupiennius salei and American lobster Homarus americanus were studied using nanoindentation and X-ray diffraction. The structural analysis of the two types of cuticle including the packing and alignment of chitin-fibrils is supported by Monte Carlo simulations of the experimental X-ray data, thereby regions of parallel and rotated fibril arrangement can be clearly distinguished. The tip of the spider fang which is used to inject venom into the prey was found to be considerably harder than the lobster carapace, while its stiffness is slightly lower.

The grinding tip of the sea urchin tooth exhibits exquisite control over calcite crystal orientation and Mg distribution.

The sea urchin tooth is a remarkable grinding tool. Even though the tooth is composed almost entirely of calcite, it is used to grind holes into a rocky substrate itself often composed of calcite. Here, we use 3 complementary high-resolution tools to probe aspects of the structure of the grinding tip: X-ray photoelectron emission spectromicroscopy (X-PEEM), X-ray microdiffraction, and NanoSIMS. We confirm that the needles and plates are aligned and show here that even the high Mg polycrystalline matrix constituents are aligned with the other 2 structural elements when imaged at 20-nm resolution. Furthermore, we show that the entire tooth is composed of 2 cooriented polycrystalline blocks that differ in their orientations by only a few degrees. A unique feature of the grinding tip is that the structural elements from each coaligned block interdigitate. This interdigitation may influence the fracture process by creating a corrugated grinding surface. We also show that the overall Mg content of the tooth structural elements increases toward the grinding tip. This probably contributes to the increasing hardness of the tooth from the periphery to the tip. Clearly the formation of the tooth, and the tooth tip in particular, is amazingly well controlled. The improved understanding of these structural features could lead to the design of better mechanical grinding and cutting tools.

Evidence that treatment with risedronate in women with postmenopausal osteoporosis affects bone mineralization and bone volume.

Risedronate is used in osteoporosis treatment. Postmenopausal women enrolled in the Vertebral Efficacy with Risedronate Therapy trial received either risedronate (5 mg/day) or placebo for 3 years. Subjects received calcium and vitamin D supplementation if deficient at baseline. Lumbar spine bone mineral density (BMD) was measured at baseline and at 3 years. Quantitative back-scattered electron imaging (qBEI) was performed on paired iliac crest biopsies (risedronate, n = 18; placebo, n = 13) before and after treatment, and the mineral volume fraction in the trabecular bone was calculated. Combining dual-energy X-ray absorptiometric values with the mineral volume fraction for the same patients allowed us to calculate the relative change in trabecular bone volume with treatment. This showed that the effect on BMD was likely to be due partly to changes in matrix mineralization and partly due to changes in bone volume. After treatment, trabecular bone volume in the lumbar spine tended to increase in the risedronate group (+2.4%, nonsignificant) but there was a significant decrease (-3.7%, P < 0.05) in the placebo group. Calcium supplementation with adequate levels of vitamin D led to an approximately 3.3% increase in mineral content in the bone material independently of risedronate treatment. This increase was larger in patients with lower matrix mineralization at baseline and likely resulted from correction of calcium/vitamin D deficiency as well as from reduced bone remodeling. Combining BMD and bone mineralization density distribution data show that in postmenopausal osteoporosis 3-year treatment with risedronate preserves or may increase trabecular bone volume, unlike placebo. This analysis also allows, for the first time, separation of the contributions of bone volume and matrix mineralization to the increase in BMD.

Bone material properties in trabecular bone from human iliac crest biopsies after 3- and 5-year treatment with risedronate.

UNLABELLED: Long-term effects of risedronate on bone mineral maturity/crystallinity and collagen cross-link ratio in triple iliac crest biopsies of osteoporotic women were evaluated. In this double-blinded study, 3- and 5-year treatment with risedronate arrested the tissue aging encountered in untreated osteoporosis and in osteoporosis treated with other antiresorptives. This effect may be contributing to risedronate's antifracture efficacy. INTRODUCTION: Risedronate is widely used in the treatment of osteoporosis. It reduces bone turnover, increases BMD, and decreases fracture risk. To date, there are no data available on the long-term effects of risedronate on bone material properties in humans. MATERIALS AND METHODS: Osteoporotic women enrolled in the VERT-NA trial received either risedronate (5 mg/day, orally) or placebo for up to 5 years. All subjects received calcium. They also received vitamin D supplementation if deficient at baseline. Triple iliac crest biopsies were collected from a subset of these subjects at baseline, 3 years, and 5 years. Mineral maturity/crystallinity and collagen cross-link ratio was measured in these biopsies using Fourier transform infrared imaging. RESULTS: Patients that received placebo exhibited increased mineral maturity/crystallinity and collagen cross-link ratio after 3 and 5 years compared with baseline values. On the contrary, patients that received risedronate retained baseline values in both bone material indices throughout. A more spatially detailed analysis revealed that this was achieved mainly through beneficial effects on active bone-forming areas. Surprisingly, patients that received risedronate achieved premenopausal values at bone-forming areas in both indices after 5 years of treatment. CONCLUSION: Long-term treatment with risedronate affects bone material properties (mineral maturity/crystallinity and collagen cross-link ratio) and arrests the tissue aging apparent in untreated osteoporosis. These changes at the material level of the bone matrix may contribute to risedronate's rapid and sustained antifracture efficacy in osteoporotic patients.

Effects of 3- and 5-year treatment with risedronate on bone mineralization density distribution in triple biopsies of the iliac crest in postmenopausal women.

UNLABELLED: Long-term effects of risedronate on bone mineralization density distribution in triple transiliac crest biopsies of osteoporotic women were evaluated. In this double-blinded study, 3- and 5-year treatment with risedronate increased the degree and homogeneity of mineralization without producing hypermineralization. These changes at the material level of bone could contribute to risedronate's antifracture efficacy. INTRODUCTION: Risedronate, a nitrogen-containing bisphosphonate, is widely used in the treatment of osteoporosis. It reduces bone turnover, increases BMD, and decreases fracture risk. To date, there are no data available on the long-term effects of risedronate on bone mineralization density distribution (BMDD) in humans. MATERIALS AND METHODS: Osteoporotic women enrolled in the VERT-NA trial received either risedronate (5 mg/day, orally) or placebo for up to 5 years. All subjects received calcium and vitamin D supplementation if deficient at baseline. Triple iliac crest biopsies were collected from a subset of these subjects at baseline and 3 and 5 years. BMDD was measured in these biopsies using quantitative backscattered electron imaging, and the data were also compared with a normal reference group. RESULTS: At baseline, both risedronate and placebo groups had a lower degree and a greater heterogeneity of mineralization as well as an increase in low mineralized bone compared with the normal reference group. The degree of mineralization increased significantly in the risedronate as well as in the placebo group after 3- and 5-year treatment compared with baseline. However, the degree of mineralization did not exceed that of normal. Three-year treatment with risedronate significantly increased the homogeneity of mineralization and slightly decreased low mineralized bone compared with placebo. Surprisingly with 5-year risedronate treatment, heterogeneity of mineralization increased compared with 3-year treatment, which might indicate an increase in newly formed bone. CONCLUSIONS: Long-term treatment with risedronate affects the homogeneity and degree of mineralization without inducing hypermineralization of the bone matrix. These changes at the material level of the bone matrix may contribute to risedronate's antifracture efficacy in osteoporotic patients.