The mechanoresponse of bone is closely related to the osteocyte lacunocanalicular network architecture.

Organisms rely on mechanosensing mechanisms to adapt to changes in their mechanical environment. Fluid-filled network structures not only ensure efficient transport but can also be employed for mechanosensation. The lacunocanalicular network (LCN) is a fluid-filled network structure, which pervades our bones and accommodates a cell network of osteocytes. For the mechanism of mechanosensation, it was hypothesized that load-induced fluid flow results in forces that can be sensed by the cells. We use a controlled in vivo loading experiment on murine tibiae to test this hypothesis, whereby the mechanoresponse was quantified experimentally by in vivo micro-computed tomography (µCT) in terms of formed and resorbed bone volume. By imaging the LCN using confocal microscopy in bone volumes covering the entire cross-section of mouse tibiae and by calculating the fluid flow in the three-dimensional (3D) network, we could perform a direct comparison between predictions based on fluid flow velocity and the experimentally measured mechanoresponse. While local strain distributions estimated by finite-element analysis incorrectly predicts preferred bone formation on the periosteal surface, we demonstrate that additional consideration of the LCN architecture not only corrects this erroneous bias in the prediction but also explains observed differences in the mechanosensitivity between the three investigated mice. We also identified the presence of vascular channels as an important mechanism to locally reduce fluid flow. Flow velocities increased for a convergent network structure where all of the flow is channeled into fewer canaliculi. We conclude that, besides mechanical loading, LCN architecture should be considered as a key determinant of bone adaptation.

Microstructural heterogeneity of the collagenous network in the loaded and unloaded periodontal ligament and its biomechanical implications.

The periodontal ligament (PDL) is a highly heterogeneous fibrous connective tissue and plays a critical role in distributing occlusal forces and regulating tissue remodeling. Its mechanical properties are largely determined by the extracellular matrix, comprising a collagenous fiber network interacting with the capillary system as well as interstitial fluid containing proteoglycans. While the phase-contrast micro-CT technique has portrayed the 3D microscopic heterogeneity of PDL, the topological parameters of its network, which is crucial to understanding the multiscale constitutive behavior of this tissue, has not been characterized quantitatively. This study aimed to provide new understanding of such microscopic heterogeneity of the PDL with quantifications at both tissue and collagen network levels in a spatial manner, by combining phase-contrast micro-CT imaging and a purpose-built image processing algorithm for fiber analysis. Both variations within a PDL and among the PDL with different shapes, i.e. round-shaped and kidney-shaped PDLs, are described in terms of tissue thickness, fiber distribution, local fiber densities, and fiber orientation (namely azimuthal and elevation angles). Furthermore, the tissue and collagen fiber network responses to mechanical loading were evaluated in a similar manner. A 3D helical alignment pattern was observed in the fiber network, which appears to regulate and adapt a screw-like tooth motion under occlusion. The microstructural heterogeneity quantified here allows development of sample-specific constitutive models to characterize the PDL's functional and pathological loading responses, thereby providing a new multiscale framework for advancing our knowledge of this complex limited mobility soft-hard tissue interface.

The effect of aging on the nanostructure of murine alveolar bone and dentin.

INTRODUCTION: Alveolar bone, dentin, and cementum provide a striking example of structurally different collagen-based mineralized tissues separated only by periodontal ligament. While alveolar bone is strongly remodeled, this does not hold for dentin and cementum. However, additional dentin can be deposited on the inner surface of the pulp chamber also in older age. By investigating alveolar bone and molar of mice, the aim of our study is to detect changes in the mineral nanostructure with aging. MATERIALS AND METHODS: Buccal-lingual sections of the mandible and first molar from C57BL/6 mice of three different age groups (young 5 weeks, adult 22 weeks and old 23 months) were characterized using synchrotron small and wide-angle X-ray scattering. Local average thickness and length of the apatite particles were mapped with several line scans covering the alveolar bone and the tooth. RESULTS: In alveolar bone, a spatial gradient was seen to develop with age with the thickest and longest particles in the distal part of the bone. The mineral particles in dentin were found to be become thicker, but then decrease of average length from adult to old animals. The mineral particle characteristics of dentin close to the pulp chamber were not only different to the rest of the tooth, but also when comparing the different age groups and even between individual animals in the same age group. CONCLUSIONS: These results indicated that mineral particle characteristics were found to evolve differently between molar and alveolar bone as a function of age.

Heterogeneity of the osteocyte lacuno-canalicular network architecture and material characteristics across different tissue types in healing bone.

Various tissue types, including fibrous connective tissue, bone marrow, cartilage, woven and lamellar bone, coexist in healing bone. Similar to most bone tissue type, healing bone contains a lacuno-canalicular network (LCN) housing osteocytes. These cells are known to orchestrate bone remodeling in healthy bone by sensing mechanical strains and translating them into biochemical signals. The structure of the LCN is hypothesized to influence mineralization processes. Hence, the aim of the present study was to visualize and match spatial variations in the LCN topology with mineral characteristics, within and at the interfaces of the different tissue types that comprise healing bone. We applied a correlative multi-method approach to visualize the LCN architecture and quantify mineral particle size and orientation within healing femoral bone in a mouse osteotomy model (26 weeks old C57BL/6 mice). This approach revealed structural differences across several length scales during endochondral ossification within the following regions: calcified cartilage, bony callus, cortical bone and a transition zone between the cortical and callus region analyzed 21 days after the osteotomy. In this transition zone, we observed a continuous convergence of mineral characteristics and osteocyte lacunae shape as well as discontinuities in the lacunae volume and LCN connectivity. The bony callus exhibits a 34% higher lacunae number density and 40% larger lacunar volume compared to cortical bone. The presented correlations between LCN architecture and mineral characteristics improves our understanding of how bone develops during healing and may indicate a contribution of osteocytes to bone (re)modeling.

Towards a Connectomic Description of the Osteocyte Lacunocanalicular Network in Bone.

PURPOSE OF REVIEW: Osteocytes are the most abundant bone cells. They are completely encased in mineralized tissue, sitting inside lacunae that are connected by a multitude of canaliculi. In recent years, the osteocyte network has been shown to fulfill endocrine functions and to communicate with a number of other organs. This review addresses emerging knowledge on the connectome of the lacunocanalicular network in different types of bone tissue. RECENT FINDINGS: Recent advances in three-dimensional imaging technology started to reveal parameters that are well known from general theory to characterize the function of networks, such as network density, degree of nodes, or shortest path length through the network. The connectome of the lacunocanalicular network differs in some aspects between lamellar and woven bone and seems to change with age. More research is needed to relate network structure to function, such as intercellular transport or communication and its role in mechanosensation, as well as to understand the effect of diseases.

Late stages of mineralization and their signature on the bone mineral density distribution.

PURPOSE: Experimental measurements of bone mineral density distributions (BMDDs) enable a determination of secondary mineralization kinetics in bone, but the maximum degree of mineralization and how this maximum is approached remain uncertain. We thus test computationally different hypotheses on late stages of bone mineralization by simulating BMDDs in low-turnover conditions. MATERIALS AND METHODS: An established computational model of the BMDD that accounts for mineralization and remodeling processes was extended to limit mineralization to various maximum calcium capacities of bone. Simulated BMDDs obtained by reducing turnover rate from the reference trabecular BMDD under different assumptions on late stage mineralization kinetics were compared with experimental BMDDs of low-turnover bone. RESULTS: Simulations show that an abrupt stopping of mineralization near a maximum calcium capacity induces a pile-up of minerals in the BMDD statistics that is not observed experimentally. With a smooth decrease of mineralization rate, imposing low maximum calcium capacities helps to match peak location and width of simulated low-turnover BMDDs with peak location and width of experimental BMDDs, but results in a distinctive asymmetric peak shape. No tuning of turnover rate and maximum calcium capacity was able to explain the differences found in experimental BMDDs between trabecular bone (high turnover) and femoral cortical bone (low turnover). CONCLUSIONS: Secondary mineralization in human bone does not stop abruptly, but continues slowly up to a calcium content greater than 30 wt% Ca. The similar mineral heterogeneity seen in trabecular and femoral cortical bones at different peak locations was unexplained by the turnover differences tested.

Coalignment of osteocyte canaliculi and collagen fibers in human osteonal bone.

During bone formation osteocytes get connected with each other via a dense network of canaliculi within the mineralized bone matrix. Important functions attributed to the osteocyte network include the control of bone remodeling and a contribution to mineral homeostasis. To detect structural clues of the formation and functionality of the network, this study analyzes the structure and orientation of the osteocyte lacuno-canalicular network (OLCN), specifically in relation to the concentric bone lamellae within human osteons. The network structure within 49 osteons from four samples of cortical bone from the femoral midshaft of middle-aged healthy women was determined by a combination of rhodamine staining and confocal laser scanning microscopy followed by computational image analysis. A quantitative evaluation showed that 64±1% of the canalicular length has an angle smaller than 30° to the direction towards the osteon center, while the lateral network - defined by an orientation angle larger than 60° - comprises 16±1%. With the same spatial periodicity as the bone lamellae, both radial and lateral network show variations in the network density and order. However, only the preferred orientation of the lateral network twists when crossing a lamella. This twist agrees with the preferred orientation of the fibrous collagen matrix. The chirality of the twist was found to be individual-specific. The coalignment between network and matrix extends to the orientation of the elongated osteocyte lacunae. The intimate link between OLCN and collagen matrix implies an interplay between osteocyte processes and the arrangement of the surrounding collagen fibers during osteoid formation.

The nanostructure of murine alveolar bone and its changes due to type 2 diabetes.

Alveolar bone - the bony ridge containing the tooth sockets - stands out by its remodeling activity where bone is being formed and resorbed at a much higher rate than in any other bony tissue. Teeth that are anchored in the jaw through the periodontal ligament exert very large localized loads during mastication that could lead to a unique adaptation of the collagen/mineral structure in the bone. Our aim was to characterize the nanostructure of alveolar bone and to determine the influence of diabetes on structural characteristics of the mineralized matrix. Using small- and wide-angle X-ray scattering (SAXS/WAXS), we studied a spontaneous diabetic mouse model (KK+) and its corresponding healthy controls (KK-) (n=6) to determine the size and mutual alignment of the mineral nanoparticles embedded in the collagen matrix. On cross-sections (buccal-lingual) of the first molar multiple line scans with a spatial resolution of 30µm were performed on each sample, from the lingual to the buccal side of the mandible. Mineral particle thickness and length are decreasing towards the tooth in both buccal and lingual sides of alveolar bone. While mineral particles are well aligned with the long axis of the tooth on the buccal side, they are in a quarter of the measurements oriented along two preferred directions on the lingual side. These nanostructural differences can be interpreted as the result of an asymmetric loading during mastication, leading to a tilting of the tooth in its socket. In diabetic mice particle thicknesses are smaller compared to control animals.

Anisotropy in bone demineralization revealed by polarized far-IR spectroscopy.

Bone material is composed of an organic matrix of collagen fibers and apatite nanoparticles. Previously, vibrational spectroscopy techniques such as infrared (IR) and Raman spectroscopy have proved to be particularly useful for characterizing the two constituent organic and inorganic phases of bone. In this work, we tested the potential use of high intensity synchrotron-based far-IR radiation (50-500 cm(-1)) to gain new insights into structure and chemical composition of bovine fibrolamellar bone. The results from our study can be summarized in the following four points: (I) compared to far-IR spectra obtained from synthetic hydroxyapatite powder, those from fibrolamellar bone showed similar peak positions, but very different peak widths; (II) during stepwise demineralization of the bone samples, there was no significant change neither to far-IR peak width nor position, demonstrating that mineral dissolution occurred in a uniform manner; (III) application of external loading on fully demineralized bone had no significant effect on the obtained spectra, while dehydration of samples resulted in clear differences. (IV) using linear dichroism, we showed that the anisotropic structure of fibrolamellar bone is also reflected in anisotropic far-IR absorbance properties of both the organic and inorganic phases. Far-IR spectroscopy thus provides a novel way to functionally characterize bone structure and chemistry, and with further technological improvements, has the potential to become a useful clinical diagnostic tool to better assess quality of collagen-based tissues.

Spatial variations in the osteocyte lacuno-canalicular network density and analysis of the connectomic parameters.

Osteocyte lacuno-canalicular network (LCN) is comprised of micrometre-sized pores and submicrometric wide channels in bone. Accumulating evidence suggests multiple functions of this network in material transportation, mechanobiological signalling, mineral homeostasis and bone remodelling. Combining rhodamine staining and confocal laser scanning microscopy, the longitudinal cross-sections of six mouse tibiae were imaged, and the connectome of the network was quantified with a focus on the spatial heterogeneities of network density, connectivity and length of canaliculi. In-vivo loading and double calcein labelling on these tibiae allowed differentiating the newly formed bone from the pre-existing regions. The canalicular density of the murine cortical bone varied between 0.174 and 0.243 µm/µm3, and therefore is three times larger than the corresponding value for human femoral midshaft osteons. The spatial heterogeneity of the network was found distinctly more pronounced across the cortex than along the cortex. We found that in regions with a dense network, the LCN conserves its largely tree-like character, but increases the density by including shorter canaliculi. The current study on healthy mice should serve as a motivating starting point to study the connectome of genetically modified mice, including models of bone diseases and of reduced mechanoresponse.

Logistics of Bone Mineralization in the Chick Embryo Studied by 3D Cryo FIB-SEM Imaging.

During skeletal development, bone growth and mineralization require transport of substantial amounts of calcium, while maintaining very low concentration. How an organism overcomes this major logistical challenge remains mostly unexplained. To shed some light on the dynamics of this process, cryogenic focused ion beam-scanning electron microscopy (cryo-FIB/SEM) is used to image forming bone tissue at day 13 of a chick embryo femur. Both cells and matrix in 3D are visualized and observed as calcium-rich intracellular vesicular structures. Counting the number of these vesicles per unit volume and measuring their calcium content based on the electron back-scattering signal, the intracellular velocity at which these vesicles need to travel to transport all the calcium required for the mineral deposited in one day within the collagenous tissue can be estimated. This velocity at 0.27 µm s-1 is estimated, which is too large for a diffusion process and rather suggests active transport through the cellular network. It is concluded that calcium logistics is hierarchical and based on several transport mechanisms: first through the vasculature using calcium-binding proteins and the blood flow, then active transport over tens of micrometers through the network of osteoblasts and osteocytes, and finally diffusive transport over the last one or two microns.

Subcanalicular Nanochannel Volume Is Inversely Correlated With Calcium Content in Human Cortical Bone.

The spatial distribution of mineralization density is an important signature of bone growth and remodeling processes, and its alterations are often related to disease. The extracellular matrix of some vertebrate mineralized tissues is known to be perfused by a lacunocanalicular network (LCN), a fluid-filled unmineralized structure that harbors osteocytes and their fine processes and transports extracellular fluid and its constituents. The current report provides evidence for structural and compositional heterogeneity at an even smaller, subcanalicular scale. The work reveals an extensive unmineralized three-dimensional (3D) network of nanochannels (~30 nm in diameter) penetrating the mineralized extracellular matrix of human femoral cortical bone and encompassing a greater volume fraction and surface area than these same parameters of the canaliculi comprising the LCN. The present study combines high-resolution focused ion beam-scanning electron microscopy (FIB-SEM) to investigate bone ultrastructure in 3D with quantitative backscattered electron imaging (qBEI) to estimate local bone mineral content. The presence of nanochannels has been found to impact qBEI measurements fundamentally, such that volume percentage (vol%) of nanochannels correlates inversely with weight percentage (wt%) of calcium. This mathematical relationship between nanochannel vol% and calcium wt% suggests that the nanochannels could potentially provide space for ion and small molecule transport throughout the bone matrix. Collectively, these data propose a reinterpretation of qBEI measurements, accounting for nanochannel presence in human bone tissue in addition to collagen and mineral. Further, the results yield insight into bone mineralization processes at the nanometer scale and present the possibility for a potential role of the nanochannel system in permitting ion and small molecule diffusion throughout the extracellular matrix. Such a possible function could thereby lead to the sequestration or occlusion of the ions and small molecules within the extracellular matrix. © 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).

Structural and functional heterogeneity of mineralized fibrocartilage at the Achilles tendon-bone insertion.

A demanding task of the musculoskeletal system is the attachment of tendon to bone at entheses. This region often presents a thin layer of fibrocartilage (FC), mineralized close to the bone and unmineralized close to the tendon. Mineralized FC deserves increased attention, owing to its crucial anchoring task and involvement in enthesis pathologies. Here, we analyzed mineralized FC and subchondral bone at the Achilles tendon-bone insertion of rats. This location features enthesis FC anchoring tendon to bone and sustaining tensile loads, and periosteal FC facilitating bone-tendon sliding with accompanying compressive and shear forces. Using a correlative multimodal investigation, we evaluated potential specificities in mineral content, fiber organization and mechanical properties of enthesis and periosteal FC. Both tissues had a lower degree of mineralization than subchondral bone, yet used the available mineral very efficiently: for the same local mineral content, they had higher stiffness and hardness than bone. We found that enthesis FC was characterized by highly aligned mineralized collagen fibers even far away from the attachment region, whereas periosteal FC had a rich variety of fiber arrangements. Except for an initial steep spatial gradient between unmineralized and mineralized FC, local mechanical properties were surprisingly uniform inside enthesis FC while a modulation in stiffness, independent from mineral content, was observed in periosteal FC. We interpreted these different structure-property relationships as a demonstration of the high versatility of FC, providing high strength at the insertion (to resist tensile loading) and a gradual compliance at the periosteal surface (to resist contact stresses). STATEMENT OF SIGNIFICANCE: Mineralized fibrocartilage (FC) at entheses facilitates the integration of tendon in bone, two strongly dissimilar tissues. We focus on the structure-function relationships of two types of mineralized FC, enthesis and periosteal, which have clearly distinct mechanical demands. By investigating them with multiple high-resolution methods in a correlative manner, we demonstrate differences in fiber architecture and mechanical properties between the two tissues, indicative of their mechanical roles. Our results are relevant both from a medical viewpoint, targeting a clinically relevant location, as well as from a material science perspective, identifying FC as high-performance versatile composite.

3D Interrelationship between Osteocyte Network and Forming Mineral during Human Bone Remodeling.

During bone remodeling, osteoblasts are known to deposit unmineralized collagenous tissue (osteoid), which mineralizes after some time lag. Some of the osteoblasts differentiate into osteocytes, forming a cell network within the lacunocanalicular network (LCN) of bone. To get more insight into the potential role of osteocytes in the mineralization process of osteoid, sites of bone formation are three-dimensionally imaged in nine forming human osteons using focused ion beam-scanning electron microscopy (FIB-SEM). In agreement with previous observations, the mineral concentration is found to gradually increase from the central Haversian canal toward pre-existing mineralized bone. Most interestingly, a similar feature is discovered on a length scale more than 100-times smaller, whereby mineral concentration increases from the LCN, leaving around the canaliculi a zone virtually free of mineral, the size of which decreases with progressing mineralization. This suggests that the LCN controls mineral formation but not just by diffusion of mineralization precursors, which would lead to a continuous decrease of mineral concentration from the LCN. The observation is, however, compatible with the codiffusion and reaction of precursors and inhibitors from the LCN into the bone matrix.

Tomographic X-ray scattering based on invariant reconstruction: analysis of the 3D nanostructure of bovine bone.

Small-angle X-ray scattering (SAXS) is an effective characterization technique for multi-phase nanocomposites. The structural complexity and heterogeneity of biological materials require the development of new techniques for the 3D characterization of their hierarchical structures. Emerging SAXS tomographic methods allow reconstruction of the 3D scattering pattern in each voxel but are costly in terms of synchrotron measurement time and computer time. To address this problem, an approach has been developed based on the reconstruction of SAXS invariants to allow for fast 3D characterization of nanostructured inhomogeneous materials. SAXS invariants are scalars replacing the 3D scattering patterns in each voxel, thus simplifying the 6D reconstruction problem to several 3D ones. Standard procedures for tomographic reconstruction can be directly adapted for this problem. The procedure is demonstrated by determining the distribution of the nanometric bone mineral particle thickness (T parameter) throughout a macroscopic 3D volume of bovine cortical bone. The T parameter maps display spatial patterns of particle thickness in fibrolamellar bone units. Spatial correlation between the mineral nano-structure and microscopic features reveals that the mineral particles are particularly thin in the vicinity of vascular channels.

Mineral density differences between femoral cortical bone and trabecular bone are not explained by turnover rate alone.

Bone mineral density distributions (BMDDs) are a measurable property of bone tissues that depends strongly on bone remodelling and mineralisation processes. These processes can vary significantly in health and disease and across skeletal sites, so there is high interest in analysing these processes from experimental BMDDs. Here, we propose a rigorous hypothesis-testing approach based on a mathematical model of mineral heterogeneity in bone due to remodelling and mineralisation, to help explain differences observed between the BMDD of human femoral cortical bone and the BMDD of human trabecular bone. Recent BMDD measurements show that femoral cortical bone possesses a higher bone mineral density, but a similar mineral heterogeneity around the mean compared to trabecular bone. By combining this data with the mathematical model, we are able to test whether this difference in BMDD can be explained by (i) differences in turnover rate; (ii) differences in osteoclast resorption behaviour; and (iii) differences in mineralisation kinetics between the two bone types. We find that accounting only for differences in turnover rate is inconsistent with the fact that both BMDDs have a similar spread around the mean, and that accounting for differences in osteoclast resorption behaviour leads to biologically inconsistent bone remodelling patterns. We conclude that the kinetics of mineral accumulation in bone matrix must therefore be different in femoral cortical bone and trabecular bone. Although both cortical and trabecular bone are made up of lamellar bone, the different mineralisation kinetics in the two types of bone point towards more profound structural differences than usually assumed.

Network architecture strongly influences the fluid flow pattern through the lacunocanalicular network in human osteons.

A popular hypothesis explains the mechanosensitivity of bone due to osteocytes sensing the load-induced flow of interstitial fluid squeezed through the lacunocanalicular network (LCN). However, the way in which the intricate structure of the LCN influences fluid flow through the network is largely unexplored. We therefore aimed to quantify fluid flow through real LCNs from human osteons using a combination of experimental and computational techniques. Bone samples were stained with rhodamine to image the LCN with 3D confocal microscopy. Image analysis was then performed to convert image stacks into mathematical network structures, in order to estimate the intrinsic permeability of the osteons as well as the load-induced fluid flow using hydraulic circuit theory. Fluid flow was studied in both ordinary osteons with a rather homogeneous LCN as well as a frequent subtype of osteons-so-called osteon-in-osteons-which are characterized by a ring-like zone of low network connectivity between the inner and the outer parts of these osteons. We analyzed 8 ordinary osteons and 9 osteon-in-osteons from the femur midshaft of a 57-year-old woman without any known disease. While the intrinsic permeability was 2.7 times smaller in osteon-in-osteons compared to ordinary osteons, the load-induced fluid velocity was 2.3 times higher. This increased fluid velocity in osteon-in-osteons can be explained by the longer path length, needed to cross the osteon from the cement line to the Haversian canal, including more fluid-filled lacunae and canaliculi. This explanation was corroborated by the observation that a purely structural parameter-the mean path length to the Haversian canal-is an excellent predictor for the average fluid flow velocity. We conclude that osteon-in-osteons may be particularly significant contributors to the mechanosensitivity of cortical bone, due to the higher fluid flow in this type of osteons.

Properties and role of interfaces in multimaterial 3D printed composites.

In polyjet printing photopolymer droplets are deposited on a build tray, leveled off by a roller and cured by UV light. This technique is attractive to fabricate heterogeneous architectures combining compliant and stiff constituents. Considering the layer-by-layer nature, interfaces between different photopolymers can be formed either before or after UV curing. We analyzed the properties of interfaces in 3D printed composites combining experiments with computer simulations. To investigate photopolymer blending, we characterized the mechanical properties of the so-called digital materials, obtained by mixing compliant and stiff voxels according to different volume fractions. We then used nanoindentation to measure the spatial variation in mechanical properties across bimaterial interfaces at the micrometer level. Finally, to characterize the impact of finite-size interfaces, we fabricated and tested composites having compliant and stiff layers alternating along different directions. We found that interfaces formed by deposition after curing were sharp whereas those formed before curing showed blending of the two materials over a length scale bigger than individual droplet size. We found structural and functional differences of the layered composites depending on the printing orientation and corresponding interface characteristics, which influenced deformation mechanisms. With the wide dissemination of 3D printing techniques, our results should be considered in the development of architectured materials with tailored interfaces between building blocks.

Newly formed and remodeled human bone exhibits differences in the mineralization process.

During human skeletal growth, bone is formed via different processes. Two of them are: new bone formation by depositing bone at the periosteal (outer) surface and bone remodeling corresponding to a local renewal of tissue. Since in remodeling formation is preceded by resorption, we hypothesize that modeling and remodeling could require radically different transport paths for ionic precursors of mineralization. While remodeling may recycle locally resorbed mineral, modeling implies the transport over large distances to the site of bone apposition. Therefore, we searched for potential differences of size, arrangement and chemical composition of mineral particles just below surfaces of modeling and remodeling sites in femur midshaft cross-sections from healthy children. These bone sites were mapped using scanning synchrotron X-ray scattering, Raman microspectroscopy, energy dispersive X-ray analysis and quantitative backscattered electron microscopy. The results show clear differences in mineral particle size and composition between the sites, which cannot be explained by a change in the rate of mineral apposition or accumulation. At periosteal modeling sites, mineral crystals are distinctly larger, display higher crystallinity and exhibit a lower calcium to phosphorus ratio and elevated Na and Mg content. The latter may originate from Mg used for phase stabilization of mineral precursors and therefore indicate different time periods for mineral transport. We conclude that the mineralization process is distinctively different between modeling and remodeling sites due to varying requirements for the transport distance and, therefore, the stability of non-crystalline ionic precursors, resulting in distinct compositions of the deposited mineral phase. STATEMENT OF SIGNIFICANCE: In growing children new bone is formed either due to apposition of bone tissue e.g. at the outer ridge of long bones to allow growth in thickness (bone modeling), or in cavities inside the mineralized matrix when replacing tissue (bone remodeling). We demonstrate that mineral crystal shape and composition are not the same between these two sites, which is indicative of differences in mineralization precursors. We suggest that this may be due to a longer mineral transport distance to sites of new bone formation as compared to remodeling where mineral can be locally recycled.

Mechanoregulation of Bone Remodeling and Healing as Inspiration for Self-Repair in Materials.

The material bone has attracted the attention of material scientists due to its fracture resistance and ability to self-repair. A mechanoregulated exchange of damaged bone using newly synthesized material avoids the accumulation of fatigue damage. This remodeling process is also the basis for structural adaptation to common loading conditions, thereby reducing the probability of material failure. In the case of fracture, an initial step of tissue formation is followed by a mechanobiological controlled restoration of the pre-fracture state. The present perspective focuses on these mechanobiological aspects of bone remodeling and healing. Specifically, the role of the control function is considered, which describes mechanoregulation as a link between mechanical stimulation and the local response of the material through changes in structure or material properties. Mechanical forces propagate over large distances leading to a complex non-local feedback between mechanical stimulation and material response. To better understand such phenomena, computer models are often employed. As expected from control theory, negative and positive feedback loops lead to entirely different time evolutions, corresponding to stable and unstable states of the material system. After some background information about bone remodeling and healing, we describe a few representative models, the corresponding control functions, and their consequences. The results are then discussed with respect to the potential design of synthetic materials with specific self-repair properties.