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International audience; Photopolymer-based additive manufacturing has received increasing attention in the field of acoustics over the past decade, specifically towards the design of tissue-mimicking phantoms and passive components for ultrasound imaging and therapy. While these applications rely on an accurate characterization of the longitudinal bulk properties of the materials, emerging applications involving periodic micro-architectured media also require the knowledge of the transverse bulk properties to achieve the desired acoustic behavior. However, a robust knowledge of these properties is still lacking for such attenuating materials. Here, we report on the longitudinal and transverse bulk properties, i.e., frequency-dependent phase velocities and attenuations, of photopolymer materials, which were characterized in the MHz regime using a double through-transmission method in oblique incidence. Samples were fabricated using two different printing technologies (stereolithography and polyjet) to assess the impact of two important factors of the manufacturing process: curing and material mixing. Overall, the experimentally observed dispersion and attenuation could be satisfactorily modeled using a power law attenuation to identify a reduced number of intrinsic ultrasound parameters. As a result, these parameters, and especially those reflecting transverse bulk properties, were shown to be very sensitive to slight variations of the manufacturing process.

The hardest and toughest tissues formed by living organisms are organic-mineral composites termed biominerals 1,2. When they are crystalline, their mesostructure includes the nano- and micro-scale crystallite size, shape, arrangement, and orientation. Mesostructures vary enormously across marine CaCO3 biominerals (aragonite, vaterite, calcite) because they result from divergent evolution: biominerals were formed long after organisms diverged from one another 3,4. Despite such diversity, CaCO3 marine biominerals share a convergent character: adjacent crystals are similarly oriented 5-32. The reason for such convergence is unclear. Here, we show with quantitative, precise measurements at the nanoscale that the slight misorientation is consistently between 1°-40° in diverse biominerals. Can this slight misorientation confer a desirable materials property and therefore an evolutionary advantage to the forming organisms? We test and confirm this hypothesis with nanoindentation in diverse biominerals, geologic aragonite, and in abiotic, slightly misoriented, synthetic spherulites. Molecular dynamics (MD) simulations of bicrystals reveal that aragonite, vaterite, calcite, exhibit toughness peaks when they are misoriented by 10°, 20°, 30°, respectively, demonstrating that slight misorientation alone increases crack deflection and therefore fracture toughness. Slight misorientation, along with other previously known and co-existing toughening mechanisms, was selected repeatedly and convergently, during the course of evolution, to postpone fracture and thus provide organisms with competitive advantage. We anticipate slight misorientation-toughening to be a starting point for more sophisticated materials synthesis and additive manufacturing in many fields. Compared to previously known toughening mechanisms, in fact, the advantages of slight misorientation are that it can and does occur in synthetic materials, it requires one material only and no specific top-down architecture, it is easily achieved by self-assembly of organic molecules (e.g. aspirin, chocolate), polymers, metals, and ceramics 29 well beyond biominerals.

International audience; Functional grading is a distinctive feature adopted by nature to improve the transition between tissues that present a strong mismatch in mechanical properties, a relevant example being the tendon-to-bone attachment. Recent progress in multi-material additive manufacturing now allows for the design and fabrication of bioinspired functionally graded soft-to-hard composites. Nevertheless, this emerging technology depends on several design variables, including both material and mechanistic ingredients, that are likely to affect the mechanical performance of such composites. In this paper, a model-based approach is developed to describe the interaction of ultrasound waves with homogeneous and heterogeneous additively manufactured samples, which respectively display a variation either of the material ingredients (e.g., ratio of the elementary constituents) or of their spatial arrangement (e.g., functional gradients, damage). Measurements are performed using longitudinal bulk waves, which are launched and detected using a linear transducer array. First, model is calibrated by exploiting the signals measured on the homogeneous samples, which allow identifying relationships between the model parameters and the material composition. Second, the model is validated by comparing the signals measured on the heterogeneous samples with those predicted numerically. Overall, the reported results pave the way for characterizing and optimizing multi-material systems that display complex bioinspired features.

A contrast in Poisson ratio is a possible strategy to enhance the stiffness of composite structures. In solid materials Poisson ratio is hardly tailorable unless cellular architectures are considered. Here, we first investigated the effect of a single re-entrant inclusion acting as a defect into a regular (non-re-entrant) honeycomb lattice. Building on this, we generated regular patterns of re-entrant inclusions into a regular hexagonal cellular matrix and we characterized the apparent stiffness and Poisson ratio of the obtained structures. We also explored the role of the intrinsic material properties of the inclusion as well as of its closest environment on the interplay between the deformations of different phases in the lattice. Our main finding is that a small fraction of re-entrant inclusions (around 12%) is sufficient to generate a substantial augmentation in stiffness (300%) at constant overall relative density and without inducing strong anisotropy. Eventually, we fabricated by 3D polyjet printing bi-material composite architectures to demonstrate the superior mechanical behavior obtained exploiting the Poisson effect.

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.

Functional gradients are material transitions that are found in nature and are known to result in materials with superior properties and multiple functionalities. The emerging multi-material 3D printing (=additive manufacturing, AM) techniques provide a powerful tool for the design and fabrication of bioinspired functionally graded materials (FGMs). In particular, the spatial distribution of materials can be controlled at the level of individual volumetric pixels (voxels i.e., cubes with side lengths of 20–40 μm), thereby ensuring accuracy, reliability, and reproducibility of the obtained properties and allowing for systematic studies of how various design variables affect the deformation and fracture behaviors of FGMs. Here, we designed, 3D printed, and mechanically tested tensile and notched FGMs specimens with step-wise (i.e., 5-, 10-, and 15-steps) and continuous (sigmoid and linear) gradients. The deformation and fracture mechanisms of these FGM composites were studied using digital image correlation, digital microscopy, and scanning electron microscopy. We further characterized the chemical composition and local mechanical properties of FGM composites using XPS and nanoindentation measurements, respectively. Tensile test specimens with a continuous gradient (i.e., linear) exhibited much higher Young's moduli (≈3-folds) and ultimate strengths (≈2-folds) but lower elongations (≈2-folds drop) as compared to those with stepwise gradients (i.e., 5-steps). Similarly, notched specimens with linear gradients exhibited 2-folds higher values of the stiffness and fracture stress, but 1.5-folds lower fracture strains as compared to those with 5-steps gradients. Although we found non-uniform highly concentrated strain distributions in all specimens, FGMs with linear gradients showed a smoother strain distribution and smaller crack blunting zones as compared to those with stepwise gradients. Our results imply that for stiffness and strength linear-gradient perform better than abrupt hard-soft-hard specimens.

Anchorage of orthopedic implants depends on the interfacial bonding between the implant and the host bone as well as on the mass and microstructure of peri-implant bone, with all these factors being continuously regulated by the biological process of bone (re)modeling. In osteoporotic bone implant integration may be jeopardized not only by lower peri-implant bone quality but also by reduced intrinsic regeneration ability. The first aim of this review is to provide a critical overview of the influence of osteoporosis on bone regeneration post-implantation. Mechanical stimulation can trigger bone formation and inhibit bone resorption; thus, judicious administration of mechanical loading can be used as an effective non-pharmacological treatment to enhance implant anchorage. Our second aim is to report recent achievements on the application of external mechanical stimulation to improve the quantity of peri-implant bone. The review focuses on peri-implant bone changes in osteoporotic conditions and following mechanical loading, prevalently using small animals and in vivo monitoring approaches. We intend to demonstrate the necessity to reveal new biological information on peri-implant bone mechanobiology to better target implant anchorage and fracture fixation in osteoporotic conditions. This article is protected by copyright. All rights reserved

Biomedical Engineering / Biomedizinische Technik, 58 (Supplement 1), ISSN:0013-5585, ISSN:1862-278X

Bone is a complex material which exhibits several hierarchical levels of structural organization. At the submicron-scale, the local tissue porosity gives rise to discontinuities in the bone matrix which have been shown to influence damage behavior. Computational tools to model the damage behavior of bone at different length scales are mostly based on finite element (FE) analysis, with a range of algorithms developed for this purpose. Although the local mechanical behavior of bone tissue is influenced by microstructural features such as bone canals and osteocyte lacunae, they are often not considered in FE damage models due to the high computational cost required to simulate across several length scales, i.e., from the loads applied at the organ level down to the stresses and strains around bone canals and osteocyte lacunae. Hence, the aim of the current study was twofold: First, a multilevel FE framework was developed to compute, starting from the loads applied at the whole bone scale, the local mechanical forces acting at the micrometer and submicrometer level. Second, three simple microdamage simulation procedures based on element removal were developed and applied to bone samples at the submicrometer-scale, where cortical microporosity is included. The present microdamage algorithm produced a qualitatively analogous behavior to previous experimental tests based on stepwise mechanical compression combined with in situ synchrotron radiation computed tomography. Our results demonstrate the feasibility of simulating microdamage at a physiologically relevant scale using an image-based meshing technique and multilevel FE analysis; this allows relating microdamage behavior to intracortical bone microstructure.

Although mechanical stimulation is considered a promising approach to accelerate implant integration, our understanding of load-driven bone formation and resorption around implants is still limited. This lack of knowledge may delay the development of effective loading protocols to prevent implant loosening, especially in osteoporosis. In healthy bone, formation and resorption are mechanoregulated processes. In the intricate context of peri-implant bone regeneration, it is not clear whether bone (re)modelling can still be load-driven. Here, we investigated the mechanical control of peri-implant bone (re)modelling with a well-controlled mechanobiological experiment. We applied cyclic mechanical loading after implant insertion in tail vertebrae of oestrogen depleted mice and we monitored peri-implant bone response by in vivo micro-CT. Experimental data were combined with micro-finite element simulations to estimate local tissue strains in (re)modelling locations. We demonstrated that a substantial increase in bone mass around the implant could be obtained by loading the entire bone. This augmentation could be attributed to a large reduction in bone resorption rather than to an increase in bone formation. We also showed that following implantation, mechanical regulation of bone (re)modelling was transiently lost. Our findings should help to clarify the role of mechanical stimulation on the maintenance of peri-implant bone mass.

High dietary protein imposes a metabolic acid load requiring excretion and buffering by the kidney. Impaired acid excretion in CKD, with potential metabolic acidosis, may contribute to the progression of CKD. Here, we investigated the renal adaptive response of acid excretory pathways in mice to high-protein diets containing normal or low amounts of acid-producing sulfur amino acids (SAA) and examined how this adaption requires the RhCG ammonia transporter. Diets rich in SAA stimulated expression of enzymes and transporters involved in mediating NH4 (+) reabsorption in the thick ascending limb of the loop of Henle. The SAA-rich diet increased diuresis paralleled by downregulation of aquaporin-2 (AQP2) water channels. The absence of Rhcg transiently reduced NH4 (+) excretion, stimulated the ammoniagenic pathway more strongly, and further enhanced diuresis by exacerbating the downregulation of the Na(+)/K(+)/2Cl(-) cotransporter (NKCC2) and AQP2, with less phosphorylation of AQP2 at serine 256. The high protein acid load affected bone turnover, as indicated by higher Ca(2+) and deoxypyridinoline excretion, phenomena exaggerated in the absence of Rhcg. In animals receiving a high-protein diet with low SAA content, the kidney excreted alkaline urine, with low levels of NH4 (+) and no change in bone metabolism. Thus, the acid load associated with high-protein diets causes a concerted response of various nephron segments to excrete acid, mostly in the form of NH4 (+), that requires Rhcg. Furthermore, bone metabolism is altered by a high-protein acidogenic diet, presumably to buffer the acid load.

The determining factors for the fixation of uncemented screws in bone are the bone-implant interface and the peri-implant bone. The goal of this work was to explore the role of the peri-implant bone architecture on the mechanics of the bone-implant system. In particular, the specific aims of the study were to investigate: (i) the impact of the different architectural parameters, (ii) the effects of disorder, and (iii) the deformations in the peri-implant region. A three-dimensional beam lattice model to describe trabecular bone was developed. Various microstructural features of the lattice were varied in a systematic way. Implant pull-out tests were simulated, and the stiffness and strength of the bone-implant system were computed. The results indicated that the strongest decrease in pull-out strength was obtained by trabecular thinning, whereas pull-out stiffness was mostly affected by trabecular removal. These findings could be explained by investigating the peri-implant deformation field. For small implant displacements, a large amount of trabeculae in the peri-implant region were involved in the load transfer from implant to bone. Therefore, trabecular removal in this region had a strong negative effect on pull-out stiffness. Conversely, at higher displacements, deformations mainly localized in the trabeculae in contact with the implant; hence, thinning those trabeculae produced the strongest decrease in the strength of the system. Although idealized, the current approach is helpful for a mechanical understanding of the role played by peri-implant bone.

Stem cell-mediated gene therapy for fracture repair, utilizes genetically engineered mesenchymal stem cells (MSCs) for the induction of bone growth and is considered a promising approach in skeletal tissue regeneration. Previous studies have shown that murine nonunion fractures can be repaired by implanting MSCs over-expressing recombinant human bone morphogenetic protein-2 (rhBMP-2). Nanoindentation studies of bone tissue induced by MSCs in a radius fracture site indicated similar elastic modulus compared to intact murine bone, eight weeks post treatment. In the present study we sought to investigate temporal changes in microarchitecture and biomechanical properties of repaired murine radius bones, following the implantation of MSCs. High resolution micro computed tomography (Micro-CT) was performed 10 and 35 weeks post MSC implantation, followed by micro finite element (Micro-FE) analysis. The results have shown that the regenerated bone tissue remodels over time, as indicated by a significant decrease in bone volume, total volume and connectivity density combined with an increase in mineral density. In addition, the axial stiffness of limbs repaired with MSCs was 2 to 1.5 times higher compared to the contralateral intact limbs, at 10 and 35 weeks post treatment. These results could be attributed to the fusion that occurred between in the ulna and radius bones. In conclusion, although MSCs induce bone formation, which exceeds the fracture site, significant remodeling of the repair callus occurs over time. In addition, limbs treated with an MSC graft demonstrated superior biomechanical properties, which could indicate the clinical benefit of future MSC application in nonunion fracture repair.

The heterogeneous distribution of mineral content in trabecular bone reflects the continuous renewal of bone material in bone remodeling and the subsequent increase in mineral content in the newly formed bone packets. The bone mineralization density distribution (BMDD) is typically used to describe this nonuniform mineral content of the bone matrix. Our mathematical model describes changes of the BMDD of trabecular bone as a function of bone resorption and deposition rates and the mineralization kinetics in a newly formed bone packet. Input parameters used in the simulations were taken from experimental studies. The simulations of the time evolution of the BMDD after increase in bone turnover (perimenopausal period) resulted in a shift of the BMDD toward lower values of the mineral content. Transiently, there was a broadening of the BMDD configuration partly showing two peaks, which points to a strongly heterogeneous distribution of the mineral. Conversely, when the remodeling rate was reduced (antiresorptive therapy), the BMDD shifted toward higher values of the mineral content. There was a transient narrowing of the distribution before broadening again to reach the new steady state. Results from this latter simulation are in good agreement with measurements of the BMDD of patients after 3 and 5 yr of treatment with risedronate. Based on available experimental data on bone remodeling, this model gives reliable predictions of changes in BMDD, an important factor of bone material quality. With the availability of medications with a known effect on bone turnover, this knowledge opens the possibility for therapeutic manipulation of the BMDD.

Although it is beyond doubt that mechanical stimulation is crucial to maintain bone mass, its role in preserving bone architecture is much less clear. Commonly, it is assumed that mechanics helps to conserve the trabecular network since an “accidental” thinning of a trabecula due to a resorption event would result in a local increase of load, thereby activating bone deposition there. However, considering that the thin trabecula is part of a network, it is not evident that load concentration happens locally on the weakened trabecula. The aim of this work was to clarify whether mechanical load has a protective role for preserving the trabecular network during remodeling. Trabecular bone is made dynamic by a remodeling algorithm, which results in a thickening/thinning of trabeculae with high/low strain energy density. Our simulations show that larger deviations from a regular cubic lattice result in a greater loss of trabeculae. Around lost trabeculae, the remaining trabeculae are on average thinner. More generally, thin trabeculae are more likely to have thin trabeculae in their neighborhood. The plausible consideration that a thin trabecula concentrates a higher amount of strain energy within itself is therefore only true when considering a single isolated trabecula. Mechano-regulated remodeling within a network-like architecture leads to local concentrations of thin trabeculae., Biomechanics and Modeling in Mechanobiology, 14 (4), ISSN:1617-7959, ISSN:1617-7940

Perfusion bioreactors are known to exert shear stresses on cultured cells, leading to cell differentiation and enhanced extracellular matrix deposition on scaffolds. The influence of the scaffold’s porous microstructure is investigated for a polycaprolactone (PCL) scaffold with a regular microarchitecture and a silk fibroin (SF) scaffold with an irregular network of interconnected pores. Their complex 3D geometries are imaged by micro-computed tomography and used in direct pore-level simulations of the entire scaffold–bioreactor system to numerically solve the governing mass and momentum conservation equations for fluid flow through porous media. The velocity field and wall shear stress distribution are determined for both scaffolds. The PCL scaffold exhibited an asymmetric distribution with peak and plateau, while the SF scaffold exhibited a homogenous distribution and conditioned the flow more efficiently than the PCL scaffold. The methodology guides the design and optimization of the scaffold geometry.

This study sought to determine whether deficient Igf1 expression in osteocytes would affect loading-induced osteogenic response. Tibias of osteocyte Igf1 conditional knockout (KO) mice (generated by cross-breeding Igf1 floxed mice with Dmp1- Cre transgenic mice) and wild-type (WT) littermates were subjected to four-point bending for 2 wk. Microcomputed tomography confirmed that the size of tibias of conditional mutants was smaller. Loading with an equivalent loading strain increased periosteal woven bone and endosteal lamellar bone formation in WT mice but not in conditional KO mice. Consistent with the lack of an osteogenic response, the loading failed to upregulate expression of early mechanoresponsive genes ( Igf1, Cox-2, c-fos) or osteogenic genes ( Cbfa-1, and osteocalcin) in conditional KO bones. The lack of osteogenic response was not due to reduced osteocyte density or insufficient loading strain. Deficient osteocyte Igf1 expression reduced the loading-induced upregulation of expression of canonical Wnt signaling genes ( Wnt10b, Lrp5, Dkk1, sFrp2). The loading also reduced (by 40%) Sost expression in WT mice, but the loading not only did not reduce but upregulated (∼1.5-fold) Sost expression in conditional KO mice. Conditional disruption of Igf1 in osteocytes also abolished the loading-induced increase in the bone β-catenin protein level. These findings suggest an impaired response in the loading-induced upregulation of the Wnt signaling in conditional KO mice. In summary, conditional disruption of Igf1 in osteocytes abolished the loading-induced activation of the Wnt signaling and the corresponding osteogenic response. In conclusion, osteocyte-derived IGF-I plays a key determining role in bone mechanosensitivity.

Alpha-actinin is a cytoskeleton-binding protein involved in the assembly and regulation of the actin filaments. In this work molecular dynamics method was applied to investigate the mechanical behaviour of the human skeletal muscle alpha-actinin. Five configurations were unfolded at an elongation speed of 0.1 nm/ps in order to investigate the conformational changes occurring during the extension process. Moreover, a sensitivity analysis at different velocities was performed for one of the R2-R3 spectrin-like repeat configuration extracted in order to evaluate the effect of the pulling speed on the mechanical behaviour of the molecule. Two different behaviours were recognized with respect to the pulling speed. In particular, at speed higher than 0.025 nm/ps a continuous rearrangement without evident force peaks was obtained, on the contrary at lower speed evident peaks in the range 500-750 pN were detected. R3 repeat resulted more stable than R2 during mechanical unfolding, due to the lower hydrophobic surface available to the solvent. The characterization of the R2-R3 units can be useful for the development of cytoskeleton network models based on stiffness values obtained by analyses performed at the molecular level.

Many plant tissues and organs are capable of moving due to changes in the humidity of the environment, such as the opening of the seed capsule of the ice plant and the opening of the pine cone. These are fascinating examples for the materials engineer, as these tissues are non-living and move solely through the differential swelling of anisotropic tissues and in principle may serve as examples for the bio-inspired design of artificial actuators. In this paper, we model the microstructure of the wild wheat awn (Triticum turgidum ssp. dicoccoides) by finite elements, especially focusing on the specific microscopic features of the active part of the awn. Based on earlier experimental findings, cell walls are modeled as multilayered cylindrical tubes with alternating cellulose fiber orientation in successive layers. It is shown that swelling upon hydration of this system leads to the formation of gaps between the layers, which could act as valves, thus enabling the entry of water into the cell wall. This supports the hypothesis that this plywood-like arrangement of cellulose fibrils enhances the effect of ambient humidity by accelerated water or vapor diffusion along the gaps. The finite element model shows that a certain distribution of axially and tangentially oriented fibers is necessary to generate sufficient tensile stresses within the cell wall to open nanometer-sized gaps between cell wall layers.

It is widely accepted that osteocytes transduce mechanical signals in bone. To better understand the micromechanical response of the osteocyte lacunae to imposed macroscopic strains finite element (FE) models were introduced to predict how mechanical loads act on the lacuno canalicular network (LCN). Yet these FE analyses were based on idealized LCN models. Recent progress in imaging allows quantitative assessment of bone microstructure in 3D down to the level of individual osteocyte lacunae and its canaliculi. Our goal was to study the influence of the microstructure on the mechanical response to imposed macroscopic strains by combining true 3D LCN data with image based FE modeling.