Hajar Razi

Diminished response to in vivo mechanical loading in trabecular and not cortical bone in adulthood of female C57Bl/6 mice coincides with a reduction in deformation to load☆

Bone loss occurs during adulthood in both women and men and affects trabecular bone more than cortical bone. The mechanism responsible for trabecular bone loss during adulthood remains unexplained, but may be due at least in part to a reduced mechanoresponsiveness. We hypothesized that trabecular and cortical bone would respond anabolically to loading and that the bone response to mechanical loading would be reduced and the onset delayed in adult compared to postpubescent mice. We evaluated the longitudinal adaptive response of trabecular and cortical bone in postpubescent, young (10 week old) and adult (26 week old) female C57Bl/6J mice to axial tibial compression using in vivo microCT (days 0, 5, 10, and 15) and dynamic histomorphometry (day 15). Loading elicited an anabolic response in both trabecular and cortical bone in young and adult mice. As hypothesized, trabecular bone in adult mice exhibited a reduced and delayed response to loading compared to the young mice, apparent in trabecular bone volume fraction and architecture after 10 days. No difference in mechanoresponsiveness of the cortical bone was observed between young and adult mice. Finite element analysis showed that load-induced strain was reduced with age. Our results suggest that trabecular bone loss that occurs in adulthood may in part be due to a reduced mechanoresponsiveness in this tissue and/or a reduction in the induced tissue deformation which occurs during habitual loading. Therapeutic approaches that address the mechanoresponsiveness of the bone tissue may be a promising and alternate strategy to maintain trabecular bone mass during aging.

Mineralizing surface is the main target of mechanical stimulation independent of age: 3D dynamic in vivo morphometry

Mechanical loading can increase cortical bone mass by shifting the balance between bone formation and resorption towards increased formation. With advancing age resorption outpaces formation resulting in a net loss in cortical bone mass. How cortical bone (re)modeling - especially resorption - responds to mechanical loading with aging remains unclear. In this study, we investigated age-related changes in the modulation of cortical bone formation and resorption sites by mechanical loading. Using in vivo microCT we determined the kinetics of three dimensional formation and resorption parameters. To analyze age-associated adaptation, the left tibiae of young, adult and elderly female C57BL/6 mice were cyclically loaded for 2weeks. Our data showed that in the nonloaded limbs, cortical bone loss with age is the result of an imbalance of resorption to formation thickness, while the surface of resorption is comparable to formation. Loading has a much stronger effect on formation than on resorption; more specifically this effect is due to an increase in formation surface with mechanical stimulation. This is the only effect of loading which is conserved into old age. The resorption thickness is independent of loading in all age groups. Using this novel image analysis technique, we were able for the first time to quantify age-related changes in cortical (re)modeling and the adaptive capacity to mechanics. Most likely a therapy against age-related bone loss combining physical exercise and pharmaceuticals is most efficient if they each act on different parameters of the (re)modeling process. Despite some differences in skeletal aging between mice and humans, our results would suggest that physical exercise in old individuals can positively influence only the formation side of (re) modeling.

Aging Leads to a Dysregulation in Mechanically Driven Bone Formation and Resorption

Physical activity is essential to maintain skeletal mass and structure, but its effect seems to diminish with age. To test the hypothesis that bone becomes less sensitive to mechanical strain with age, we used a combined in vivo/in silico approach. We investigated how maturation and aging influence the mechanical regulation of bone formation and resorption to 2 weeks of noninvasive in vivo controlled loading in mice. Using 3D in vivo morphometrical assessment of longitudinal microcomputed tomography images, we quantified sites in the mouse tibia where bone was deposited or resorbed in response to controlled in vivo loading. We compared the (re)modeling events (formation/resorption/quiescent) to the mechanical strains induced at these sites (predicted using finite element analysis). Mice of all age groups (young, adult, and elderly) responded to loading with increased formation and decreased resorption, preferentially at high strains. Low strains were associated with no anabolic response in adult and elderly mice, whereas young animals showed a strong response. Adult animals showed a clear separation between strain ranges where formation and resorption occurred but without an intermediate quiescent “lazy zone”. This strain threshold disappeared in elderly mice, as mechanically induced (re)modeling became dysregulated, apparent in an inability to inhibit resorption or initiate formation. Contrary to what is generally believed until now, aging does not shift the mechanical threshold required to initiate formation or resorption, but rather blurs its specificity. These data suggest that pharmaceutical strategies augmenting physical exercise should consider this dysfunction in the mechanical regulation of bone (re)modeling to more effectively combat age‐related bone loss.

The influence of age on adaptive bone formation and bone resorption

Bone is a tissue with enormous adaptive capacity, balancing resorption and formation processes. It is known that mechanical loading shifts this balance towards an increased formation, leading to enhanced bone mass and mechanical performance. What is not known is how this adaptive response to mechanical loading changes with age. Using dynamic micro-tomography, we show that structural adaptive changes of trabecular bone within the tibia of living mice subjected to two weeks of in vivo cyclic loading are altered by aging. Comparisons of 10, 26 and 78 weeks old animals reveal that the adaptive capacity diminishes. Strikingly, adaptation was asymmetric in that loading increases formation more than it reduces resorption. This asymmetry further shifts the (re)modeling balance towards a net bone loss with age. Loading results in a major increase in the surface area of mineralizing bone. Interestingly, the resorption thickness is independent of loading in trabecular bone in all age groups. This data suggests that during youth, mechanical stimulation induces the recruitment of bone modeling cells whereas in old age, only bone forming cells are affected. These findings provide mechanistic insights into the processes that guide skeletal aging in mice as well as in other mammals.

Skeletal maturity leads to a reduction in the strain magnitudes induced within the bone: A murine tibia study

Bone adapts to changes in the local mechanical environment (e.g. strains) through formation and resorption processes. However, the bone adaptation response is significantly reduced with increasing age. The mechanical strains induced within the bone by external loading are determined by bone morphology and tissue material properties. Although it is known that changes in bone mass, architecture and bone tissue quality occur with age, to what extent they contribute to the altered bone adaptation response remains to be determined. This study investigated alterations in strains induced in the tibia of different aged female C57Bl/6J mice (young, 10-week-old; adult, 26-week-old; and elderly, 78-week-old) subjected to in vivo compressive loading. Using a combined in vivo/in silico approach, the strains in the bones were assessed by both strain gauging and finite element modeling experiments. In cortical bone, strain magnitudes induced at the mid-diaphysis decreased by 20% from young to adult mice and by 15% from adult to elderly mice. In the cancellous bone (at the proximal metaphysis), induced strains were 70% higher in young compared with adult and elderly mice. Taking into account previous studies showing a reduced bone adaptation response to mechanical loading in adulthood, these results suggest that the diminished adaptive response is in part due to a reduction in the strains induced within the bone.

Monitoring in vivo (re)modeling: A computational approach using 4D microCT data to quantify bone surface movements

Bone undergoes continual damage repair and structural adaptation to changing external loads with the aim of maintaining skeletal integrity throughout life. The ability to monitor bone (re)modeling would allow for a better understanding in how various pathologies and interventions affect bone turnover and subsequent bone strength. To date, however, current methods to monitor bone (re)modeling over time and in space are limited. We propose a novel method to visualize and quantify bone turnover, based on in vivo microCT imaging and a 4D computational approach. By in vivo tracking of spatially correlated formation and resorption sites over time it classifies bone restructuring into (re)modeling sequences, the spatially and temporally linked sequences of formation, resorption and quiescent periods on the bone surface. The microCT based method was validated using experimental data from an in vivo mouse tibial loading model and ex vivo data of the mouse tibia. In this application, the method allows the visualization of time-resolved cortical (re)modeling and the quantification of short-term and long-term modeling on the endocortical and periosteal surface at the mid-diaphysis of loaded and control mice tibiae. Both short-term and long-term modeling processes, independent formation and resorption events, could be monitored and modeling (spatially not correlated formation and resorption) and remodeling (resorption followed by new formation at the same site) could be distinguished on the bone surface. This novel method that combines in vivo microCT with a computational approach is a powerful tool to monitor bone turnover in animal models now and is waiting to be applied to human patients in the near future.

The Periosteal Bone Surface is Less Mechano-Responsive than the Endocortical.

Dynamic processes modify bone micro-structure to adapt to external loading and avoid mechanical failure. Age-related cortical bone loss is thought to occur because of increased endocortical resorption and reduced periosteal formation. Differences in the (re)modeling response to loading on both surfaces, however, are poorly understood. Combining in-vivo tibial loading, in-vivo micro-tomography and finite element analysis, remodeling in C57Bl/6J mice of three ages (10, 26, 78 week old) was analyzed to identify differences in mechano-responsiveness and its age-related change on the two cortical surfaces. Mechanical stimulation enhanced endocortical and periosteal formation and reduced endocortical resorption; a reduction in periosteal resorption was hardly possible since it was low, even without additional loading. Endocortically a greater mechano-responsiveness was identified, evident by a larger bone-forming surface and enhanced thickness of formed bone packets, which was not detected periosteally. Endocortical mechano-responsiveness was better conserved with age, since here adaptive response declined continuously with aging, whereas periosteally the main decay in formation response occurred already before adulthood. Higher endocortical mechano-responsiveness is not due to higher endocortical strains. Although it is clear structural adaptation varies between different bones in the skeleton, this study demonstrates that adaptation varies even at different sites within the same bone.

Shaping scaffold structures in rapid manufacturing implants: A modeling approach toward mechano-biologically optimized configurations for large bone defect†

Large segmental bone defects remain a clinical challenge. Titanium lattice-structured implants in combination with laser sintering technology promises to be an alternative to bone grafting in the treatment of critical sized bone defects. Laser sintering allows the rapid manufacturing of patient specific 3D-structured scaffolds with highly interconnected macroporous networks and tunable mechanical properties. Unknown remains to what degree the mechanical properties of these implants could be tuned, without leading to mechanical failure but still providing adequate mechanical stimuli for tissue ingrowth. The aim of this study was to evaluate various implant designs for their mechanical potential towards (a) optimized safety against stress failure and (b) optimal intrastructural straining for bone ingrowth. Finite element analyses of several lattice-structured configurations were performed. Results illustrated a strong influence of the configuration on the load carrying capacity of the constructs. The likelihood of mechanical failure was predicted to be highly dependent on structure configuration with little influence of implant porosity. Increasing porosity did not result in an increase in the implant intrastructural straining in all configurations; however, the lattice configuration was the determinant factor for implant load transfer capacity. This study provides a framework for the design of effective implants with open pore structures to ensure mechanical stability as well as promote mechanical stimulation and encourage in vivo osseointegration.

Scaffold curvature-mediated novel biomineralization process originates a continuous soft tissue-to-bone interface

A myriad of shapes are found in biological tissues, often naturally evolved to fulfill a particular function. In the field of tissue engineering, substrate geometry influences cell behavior and tissue formation in vitro, yet little is known how this translates to an in vivo scenario. Here we investigate scaffold curvature-induced tissue growth, without additional growth factors or cells, in an ovine animal model. We show that soft tissue formation follows a curvature-driven tissue growth model. The highly organized endogenous soft matrix, potentially under mechanical strain, leads to a non-standard form of biomineralization, whereby the pre-existing organic matrix is mineralized without collagen remodeling and without an intermediate cartilage ossification phase. Micro- and nanoscale characterization of the tissue microstructure using histology, backscattered electron (BSE) and second-harmonic generation (SHG) imaging and synchrotron small angle X-ray scattering (SAXS) revealed (i) continuous collagen fibers across the soft-hard tissue interface on the tip of mineralized cones, and (ii) bone remodeling by basic multicellular units (BMUs) in regions adjacent to the native cortical bone. Thus, features of soft tissue-to-bone interface resembling the insertion sites of ligaments and tendons into bone were created, using a scaffold that did not mimic the structural or biological gradients across such a complex interface at its mature state. This study provides fundamental knowledge for biomimetic scaffold design in the fields of bone regeneration and soft tissue-to-bone interface tissue engineering. STATEMENT OF SIGNIFICANCE Geometry influences cell behavior and tissue formation in vitro. However, little is known how this translates to an in vivo scenario. Here we investigate the influence of scaffold mean surface curvature on in vivo tissue growth using an ovine animal model. Based on a multiscale tissue microstructure characterization, we show a seamless integration of soft tissue into newly formed bone, resembling the insertion sites of ligaments and tendons into bone. This interface was created using a scaffold without additional growth factors or cells that did not recapitulate the structural or biological gradients across such a complex tissue interface at its mature state. These findings have important implications for biomimetic scaffold design for bone regeneration and soft tissue-to-bone interface tissue engineering.

Crack driving force in twisted plywood structures

Twisted plywood architectures can be observed in many biological materials with high fracture toughness, such as in arthropod cuticles or in lamellar bone. Main purpose of this paper is to analyze the influence of the progressive rotation of the fiber direction on the spatial variation of the crack driving force and, thus, on the fracture toughness of plywood-like structures. The theory of fiber composites is used to describe the stiffness matrix of a twisted plywood structure in a specimen-fixed coordinate system. The driving force acting on a crack propagating orthogonally to the fiber-rotation plane is studied by methods of computational mechanics, coupled with the concept of configurational forces. The analysis unfolds a spatial variation of the crack driving force with minima that are beneficial for the fracture toughness of the material. It is shown that the estimation of the crack driving force can be simplified by replacing the complicated anisotropic twisted plywood structure by an isotropic material with appropriate periodic variations of Youngs modulus, which can be constructed based either on the local stiffness or local strain energy density variations. As practical example, the concepts are discussed for a specimen with a stiffness anisotropy similar to lamellar bone. STATEMENT OF SIGNIFICANCE Twisted plywood-like structures exist in many natural fiber composites, such as bone or insect carapaces, and are known to be very fracture resistant. The crack driving force in such materials is analyzed quantitatively for the first time, using the concept of configurational forces. This tool, well established in the mechanics of materials, is introduced to the modeling of biological material systems with inhomogeneous and anisotropic material behavior. Based on this analysis, it is shown that the system can be approximated by an appropriately chosen inhomogeneous but isotropic material for the calculation of the crack driving force. The spatial variation of the crack driving force and, especially, its local minima are essential to describe the fracture properties of twisted plywood structures.