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Ekubaru Y., Gokcekaya O., Ishimoto T., et al. Excellent strength–ductility balance of Sc-Zr-modified Al–Mg alloy by tuning bimodal microstructure via hatch spacing in laser powder bed fusion. Materials and Design, 221, 110976. https://doi.org/10.1016/j.matdes.2022.110976., The bimodal microstructure, which comprises ultrafine grains (UFGs) forming along the melt pool boundary and relatively coarse grains inside the melt pool, is a characteristic of the Sc-Zr-modified Al–Mg-based alloy (Scalmalloy) microstructure manufactured using laser powder bed fusion (LPBF). Focusing on this microstructural feature, we investigated the improvement in the mechanical properties of LPBF-fabricated Scalmalloy by tailoring the volume fraction of UFGs. Our approach was to decrease the laser hatch spacing (d) from 0.1 to 0.04 mm, while the volume fraction of UFGs increased from 34.6 ± 0.6 % (d = 0.1 mm) to 59.5 ± 0.5 % (d = 0.06 mm). The tensile yield stress increased from 296 ± 9 (d = 0.1 mm) to 380 ± 6 MPa (d = 0.06 mm), while maintaining a large elongation (14.8 % ± 1.2 %). The yield stress and elongation were superior to those of the cast counterparts by 2.9 and 4.0 times, respectively. In the sample with d = 0.04 mm, pores formed owing to excessive thermal energy input. Additionally, we investigated multiple strengthening mechanisms of the as-fabricated alloy. This is the first study to improve the mechanical properties of LPBF-fabricated Scalmalloy by optimizing the track-to-track interval and tuning the UFG fraction.

Ishimoto T., Kobayashi Y., Takahata M., et al. Outstanding in vivo mechanical integrity of additively manufactured spinal cages with a novel “honeycomb tree structure” design via guiding bone matrix orientation. Spine Journal, 22, 10, 1742. https://doi.org/10.1016/j.spinee.2022.05.006., BACKGROUND CONTEXT: Therapeutic devices for spinal disorders, such as spinal fusion cages, must be able to facilitate the maintenance and rapid recovery of spinal function. Therefore, it would be advantageous that future spinal fusion cages facilitate rapid recovery of spinal function without secondary surgery to harvest autologous bone. PURPOSE: This study investigated a novel spinal cage configuration that achieves in vivo mechanical integrity as a devise/bone complex by inducing bone that mimicked the sound trabecular bone, hierarchically and anisotropically structured trabeculae strengthened with a preferentially oriented extracellular matrix. STUDY DESIGN/SETTINGS: In vivo animal study. METHODS: A cage possessing an anisotropic through-pore with a grooved substrate, that we termed “honeycomb tree structure,” was designed for guiding bone matrix orientation; it was manufactured using a laser beam powder bed fusion method through an additive manufacturing processes. The newly designed cages were implanted into sheep vertebral bodies for 8 and 16 weeks. An autologous bone was not installed in the newly designed cage. A pull-out test was performed to evaluate the mechanical integrity of the cage/bone interface. Additionally, the preferential orientation of bone matrix consisting of collagen and apatite was determined. RESULTS: The cage/host bone interface strength assessed by the maximum pull-out load for the novel cage without an autologous bone graft (3360±411 N) was significantly higher than that for the conventional cage using autologous bone (903±188 N) after only 8 weeks post-implantation. CONCLUSIONS: These results highlight the potential of this novel cage to achieve functional fusion between the cage and host bone. Our study provides insight into the design of highly functional spinal devices based on the anisotropic nature of bone. CLINICAL SIGNIFICANCE: The sheep spine is similar to the human spine in its stress condition and trabecular bone architecture

Hamai R., Sakai S., Shiwaku Y., et al. Octacalcium phosphate crystals including a higher density dislocation improve its materials osteogenecity. Applied Materials Today, 26, 101279. https://doi.org/10.1016/j.apmt.2021.101279., Herein, we show that the enhanced osteogenecity of octacalcium phosphate (OCP) biomaterial, recently identified as an important element in hybrid organic–inorganic nanocomposites involved in the initial hydroxyapatite crystal expansion in mammal bones, results from an enhanced chemical property, stemming from the presence of lattice strain and dislocations. Two types of OCPs were synthesized by wet-chemical processing in the presence (c-OCP) and absence (w-OCP) of gelatin, respectively, and subjected to structural, chemical, and biological analyses. High-resolution transmission electron microscopy (HRTEM) and fast Fourier transform (FFT) analyses revealed that c-OCP crystals contained approximately six times higher edge dislocations with Burgers vectors perpendicular to a-axis than that in the case of w-OCP. The dissolution of c-OCP crystal in tris-HCl buffer occurred toward the long axis of the crystal, most likely, toward the lattice strain along the c-axis direction, while w-OCP crystal dissolved toward the a-axis direction. The study suggested that the increment of internal energy by the higher dislocation density contributed promoting c-OCP dissolution and hydrolysis through decreasing the activation energy. c-OCP crystal displayed enhanced in vitro mesenchymal stem 2D cell and 3D spheroid differentiation, in vivo bone formation, and apatite crystallographic orientation in critical-sized rat calvarial defect model as compared to w-OCP crystal, at the same time, converting to apatite structure earlier than w-OCP. The present study demonstrates that dislocation-related dissolution along with enhanced conversion of OCP is a determinant in bone induction, which may be relevant to normal bone development utilizing OCP biomaterials.

Nagase T., Todai M., Wang P., et al. Design and development of (Ti, Zr, Hf)-Al based medium entropy alloys and high entropy alloys. Materials Chemistry and Physics, 276, 125409. https://doi.org/10.1016/j.matchemphys.2021.125409., The design and development of TiZrHfAl medium entropy alloy (MEA), and the TiZrHfAlNb0.2 and TiZrHfAlV0.2 high entropy alloys (HEAs) is described. The combination of 4th subgroup elements (Ti, Zr, and Hf) with Al is discussed based on the periodic table and taxonomy of HEAs. The alloys were designed using alloy parameters for HEAs, predicted ground state diagrams from the Materials Project, and the calculation of phase diagrams (CALPHAD). Rapid solidification was effective to suppress the formation of intermetallic compounds, resulting in BCC/B2 phase formation. Significant differences in the constituent phases and Vickers hardness between ingots and melt-spun ribbons were found among the TiZrHfAl MEA, TiZrHfAlNb0.2, and TiZrHfAlV0.2 HEAs.

Gokcekaya O., Ergun C., Webster T.J., et al. Influence of precursor deficiency sites for borate incorporation on the structural and biological properties of boronated hydroxyapatite. Ceramics International; https://doi.org/https://doi.org/10.1016/j.ceramint.2022.10.232., The biological properties of hydroxyapatite (HA) are significantly influenced by its compositional characteristics especially doping elements and/or Ca/P ratio, which can be altered by precursor chemistry. In this study, a group of boronated (B-incorporated) hydroxyapatite (BHA) was synthesized using a precipitation method by setting the Ca/P ratio to the stoichiometric value of HA (1.67), while altering the precursor chemistry by adjusting either (Ca + B)/P (Ca-deficient precursor, BC) or Ca/(P + B) (P-deficient precursor, BP). After heat-treatment, the partial decomposition of the BC was observed, forming tricalcium phosphate as the byproduct, however, the BP showed phase stability at all temperatures. The B-ionic species in the form of (BO₂)⁻ and (BO₃)³⁻ were incorporated into the HA structure at the (PO₄)³⁻ and (OH)⁻ positions, respectively. The incorporation of the B species also facilitated the incorporation of (CO₃)²⁻ groups specifically in the BPs. This is the first finding on BHA reporting that preferential (CO₃)²⁻ incorporation depends on the precursor chemistry used. As a result, osteoblast adhesion was superior on the BPs compared to pure HA owing to the carbonated structure, increasing cell spreading area. As such, this in vitro study highlighted that the present P-deficient precursor approach for synthesizing BHA improved biocompatibility properties and should, thus, be further considered for the next-generation of improved orthopedic applications.

Naito K., Kuwahara Y., Yamamoto H., et al. Improvement of acid resistance of Zn-doped dentin by newly generated chemical bonds. Materials and Design, 215, 110412. https://doi.org/10.1016/j.matdes.2022.110412., Dental caries, the world's most prevalent infectious disease, is caused by the diffusion of hydroxyl ions into tooth structures. To prevent dental caries, the application of fluoride (F) and zinc (Zn) ions to teeth surfaces are potential effective measures. In this study, The ionic influence, especially the chemical bond of F and Zn, on the acid resistance of dentin were investigated by particle induced X-ray / gamma-ray emission, X-ray diffraction, X-ray photoelectron spectroscopy and X-ray absorption spectroscopy. The results showed Zn was distributed in the limited surface layer of dentin without altering its crystal structure. From the Zn K edge extended X-ray absorption fine structure, Zn incorporated into dentin was surrounded by oxygen and demonstrated four-fold coordination. The bond length and chemical state of Zn–O in Zn doped dentin suggested newly generated Zn–O covalent bond, which may improve acid resistance of dentin. This study showed that the atomic and molecular structures, such as the molecular distances and chemical state, influenced acid resistance of teeth, emphasizing the validity of chemical state analysis for understanding properties in biomaterials.

Iijima Y., Nagase T., Matsugaki A., et al. Design and development of Ti–Zr–Hf–Nb–Ta–Mo high-entropy alloys for metallic biomaterials. Materials and Design, 202, 109548. https://doi.org/10.1016/j.matdes.2021.109548., Applying empirical alloy parameters (including Mo equivalent), the predicted ground state diagram, and thermodynamic calculations, noble nonequiatomic Ti–Zr–Hf–Nb–Ta–Mo high-entropy alloys for metallic biomaterials (BioHEAs) were designed and newly developed. It is found that the Moeq and valence electron concentration (VEC) parameters are useful for alloy design involving BCC structure formation in bio medium-entropy alloys and BioHEAs. Finally, we find a Ti28.33Zr28.33Hf28.33Nb6.74Ta6.74Mo1.55 (at.%) BioHEA that exhibits biocompatibility comparable to that of CP–Ti, higher mechanical strength than CP–Ti, and an appreciable room-temperature tensile ductility. The current findings pave the way for new Ti–Zr–Hf–Nb–Ta–Mo BioHEAs development and are applicable for another BioHEA alloys system.

Okada R., Yamato K., Kawakami M., et al. Low magnetic field promotes recombinant human BMP-2-induced bone formation and influences orientation of trabeculae and bone marrow-derived stromal cells. Bone Reports, 14, 100757. https://doi.org/10.1016/j.bonr.2021.100757., Effects of high magnetic fields [MFs, ≥ 1 T (T)] on osteoblastic differentiation and the orientation of cells or matrix proteins have been reported. However, the effect of low MFs (< 1 T) on the orientation of bone formation is not well known. This study was performed to verify the effects of low MFs on osteoblastic differentiation, bone formation, and orientation of both cells and newly formed bone. An apparatus was prepared with two magnets (190 mT) aligned in parallel to generate a parallel MF. In vitro, bone marrow-derived stromal cells of rats were used to assess the effects of low MFs on cell orientation, osteoblastic differentiation, and mineralization. A bone morphogenetic protein (BMP)-2-induced ectopic bone model was used to elucidate the effect of low MFs on microstructural indices, trabecula orientation, and the apatite c-axis orientation of newly formed bone. Low MFs resulted in an increased ratio of cells oriented perpendicular to the direction of the MF and promoted osteoblastic differentiation in vitro. Moreover, in vivo analysis demonstrated that low MFs promoted bone formation and changed the orientation of trabeculae and apatite crystal in a direction perpendicular to the MF. These changes led to an increase in the mechanical strength of rhBMP-2-induced bone. These results suggest that the application of low MFs has potential to facilitate the regeneration of bone with sufficient mechanical strength by controlling the orientation of newly formed bone.

Tsutsumi Y., Ishimoto T., Oishi T., et al. Crystallographic texture- and grain boundary density-independent improvement of corrosion resistance in austenitic 316L stainless steel fabricated via laser powder bed fusion. Additive Manufacturing, 45, 102066. https://doi.org/10.1016/j.addma.2021.102066., Improvement of corrosion resistance of austenitic 316L stainless steel via laser powder bed fusion (LPBF) is currently a prominent research topic; however, the effects of crystallographic texture and the related grain boundary density on the corrosion resistance of LPBF-fabricated parts have not been elucidated. For biomedical applications, crystallographic texture control from a single crystalline-like to randomly oriented polycrystalline microstructure is highly attractive for optimizing the mechanical properties (particularly the Young's modulus) of implants. An investigation of the impacts of crystallographic planes and grain boundaries exposed to the biological environment on corrosion behavior is necessary. 316L stainless steels with different crystallographic textures and grain boundary densities were successfully fabricated via LPBF. The corrosion resistances of the LPBF-fabricated specimens were comprehensively assessed by anodic polarization, dissolution, and crevice corrosion repassivation tests. The LPBF-fabricated specimens showed extremely high pitting potentials in the physiological saline compared with the commercially available counterparts, and importantly, excellent pitting corrosion resistance was observed irrespective of the crystallographic planes and grain boundary density exposed. Moreover, the LPBF-fabricated specimens did not show metastable pitting corrosion even in an accelerated test using an acid solution. The repassivation behavior of the specimens was not affected by LPBF. Such a drastic improvement in the corrosion resistances of the LPBF-fabricated specimens might be attributed to suppression of inclusion coarsening owing to the rapid cooling rate during solidification in LPBF. By using LPBF, the desired crystallographic texture can be introduced based on the desired mechanical properties without concern for corrosiveness.

Matsuzaka T., Matsugaki A., Nakano T.. Control of osteoblast arrangement by osteocyte mechanoresponse through prostaglandin E2 signaling under oscillatory fluid flow stimuli. Biomaterials, 279, 121203. https://doi.org/10.1016/j.biomaterials.2021.121203., Anisotropic collagen/apatite microstructure is a prominent determinant of bone tissue functionalization; in particular, bone matrix modulates its anisotropic microstructure depending on the surrounding mechanical condition. Although mechanotransduction in bones is governed by osteocyte function, the precise mechanisms linking mechanical stimuli and anisotropic formation of collagen/apatite microstructure are poorly understood. Here we developed a novel anisotropic mechano-coculture system which enables the understanding of the biological mechanisms regulating the oriented bone matrix formation, which is constructed by aligned osteoblasts. The developed model provides bone-mimetic coculture platform that enables simultaneous control of mechanical condition and osteoblast-osteocyte communication with an anisotropic culture scaffold. The engineered coculture device helps in understanding the relationship between osteocyte mechanoresponses and osteoblast arrangement, which is a significant contributor to anisotropic organization of bone tissue. Our study showed that osteocyte responses to oscillatory flow stimuli regulated osteoblast arrangement through soluble molecular interactions. Importantly, we found that prostaglandin E2 is a novel determinant for oriented collagen/apatite organization of bone matrix, through controlling osteoblast arrangement.

Ishimoto T., Hagihara K., Hisamoto K., et al. Stability of crystallographic texture in laser powder bed fusion: Understanding the competition of crystal growth using a single crystalline seed. Additive Manufacturing, 43, 102004. https://doi.org/10.1016/j.addma.2021.102004., In metal additive manufacturing, crystallographic orientation control is a promising method for tailoring the functions of metallic parts. However, despite its importance in the fabrication of texture-controlled functional parts, the stability of the crystallographic texture is not widely discussed. Herein, the crystallographic texture stability under laser powder bed fusion was investigated. Two methodologies were employed. One is that a laser scanning strategy was alternately changed for a specific number of layers. The other is a “seeding” experiment in which single-crystalline substrates with controlled crystallographic orientations in the building (z-) direction and the xy-plane (perpendicular to the building direction) were used as the starting substrate. The transient zone width, where the crystallographic orientation was inherited from the layer beneath, was analyzed to evaluate the texture stability. The crystallographic direction of the seed within the xy-plane, rather than the building direction, determined the transient zone width, i.e., the texture stability. In particular, the texture in the newly deposited portion was stable when the laser scanning direction matched the <100> orientation in the underneath layer, otherwise the crystal orientation switched rapidly, such that the <100> orientation was parallel to the scanning direction. Interestingly, the crystallographic orientation along the building direction in the underneath layer hardly impacted the stability of the texture. Therefore, for the first time, it has been clarified that the <100> orientation in the scanning direction, rather than the building direction, was preferentially stabilized, whereas the orientation in the other directions secondary stabilized.

Matsugaki A., Yamazaki D., Nakano T.. Selective patterning of netrin-1 as a novel guiding cue for anisotropic dendrogenesis in osteocytes. Materials Science and Engineering C, 108, 110391. https://doi.org/10.1016/j.msec.2019.110391., Although protein patterning approaches have found widespread applications in tuning surface characteristics of biomaterials, selective control of growth in cell body and dendrites utilizing such platforms remains difficult. The functional roles assumed by cell body and dendrites in a physiological milieu have extremely high specificity. In particular, osteocytes embedded inside the mineralized bone matrix are interconnected via dendritic cell processes characterized by an anisotropic arrangement of the lacunar-canalicular system, where the fluid-flow inside the canaliculi system regulates the mechanoresponsive functionalization of bone. Control of cellular networks connected by dendritic cell processes is, therefore, imperative for constructing artificially controlled bone-mimetic structures and as an extension, for gaining insights into the molecular mechanisms underlying dendrogenesis inside the mineralized bone matrix. Here, we report an innovative strategy to induce controlled elongation of cell body or dendritic process structures in selective directions by using the inkjet printing technique. Artificial runways employing netrin-1, inspired by neural architecture, were utilized to trigger controlled elongation in the osteocyte dendritic processes in desired directions. This is the first report, to the best of our knowledge, demonstrating that anisotropic dendrogenesis of osteocytes can be controlled with selective patterning of extracellular proteins, specifically via the axon guidance ligand netrin-1.

Gokcekaya O., Hayashi N., Ishimoto T., et al. Crystallographic orientation control of pure chromium via laser powder bed fusion and improved high temperature oxidation resistance. Additive Manufacturing, 36, 101624. https://doi.org/10.1016/j.addma.2020.101624., This is the first comprehensive study on the development of a cubic crystallographic texture in pure chromium (Cr) manufactured using laser powder bed fusion (LPBF) with different laser energy densities to alter its microstructure and high-temperature oxidation behavior. An increase in the laser energy density led to the formation of a strong crystallographic texture, which was preferentially oriented in the (100) plane, and there were microstructural improvements in the pure Cr. The grain size of the (100)-oriented Cr was larger than that of the randomly oriented Cr. In addition, the high-angle grain boundary and coincident site lattice (CSL) boundary characteristics were altered. The (100)-oriented Cr exhibited a decrease in the oxide thickness that was due to the decrease in the grain boundary density with a larger grain size and an increase in the CSL boundary ratio. In contrast, the Cr with a random texture showed higher oxidation kinetics and spallation of the oxide layer. The oxidation kinetics of the pure Cr manufactured using LPBF obeyed the parabolic rate law. However, the crystal orientation affected the oxidation of the Cr as the (100)-oriented pure Cr displayed a lower parabolic rate constant, indicating that the (100)-oriented Cr was oxidation-resistant. This is the first report to demonstrate the cubic crystallographic texture formation and the improvement of high-temperature oxidation resistance in Cr manufactured using LPBF.

Nagase T., Iijima Y., Matsugaki A., et al. Design and fabrication of Ti–Zr-Hf-Cr-Mo and Ti–Zr-Hf-Co-Cr-Mo high-entropy alloys as metallic biomaterials. Materials Science and Engineering C, 107, 110322. https://doi.org/10.1016/j.msec.2019.110322., Novel TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo high-entropy alloys for metallic biomaterials (bio-HEAs) were developed based on the combination of Ti–Nb–Ta–Zr–Mo alloy system and Co–Cr–Mo alloy system as commercially-used metallic biomaterials. Ti–Zr-Hf-Cr-Mo and Ti–Zr-Hf-Co-Cr-Mo bio-HEAs were designed using (a) a tree-like diagram for alloy development, (b) empirical alloy parameters for solid-solution-phase formation, and (c) thermodynamic calculations focused on solidification. The newly-developed bio-HEAs overcomes the limitation of classical metallic biomaterials by the improvement of (i) mechanical hardness and (ii) biocompatibility all together. The TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo bio-HEAs showed superior biocompatibility comparable to that of commercial-purity Ti. The superior biocompatibility, high mechanical hardness and low liquidus temperature for the material processing in TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo bio-HEAs compared with the Ti–Nb–Ta–Zr–Mo bio-HEAs gave the authenticity of the application of bio-HEAs for orthopedic implants with multiple functions.

Nakanishi Y., Matsugaki A., Kawahara K., et al. Unique arrangement of bone matrix orthogonal to osteoblast alignment controlled by Tspan11-mediated focal adhesion assembly. Biomaterials, 209, 103. https://doi.org/10.1016/j.biomaterials.2019.04.016., During tissue construction, cells coordinate extracellular matrix (ECM)assembly depending on the cellular arrangement. The traditional understanding of the relationship between the ECM and cells is limited to the orientation-matched interaction between them. Indeed, it is commonly accepted that the bone matrix (collagen/apatite)is formed along osteoblast orientation. Nonetheless, our recent findings are contrary to the above theory; osteoblasts on nanogrooves organize formation of the bone matrix perpendicular to cell orientation. However, the precise molecular mechanisms underlying the orthogonal organization of bone matrix are still unknown. Here, we show that mature fibrillar focal adhesions (FAs)facilitate the perpendicular arrangement between cells and bone matrix. The osteoblasts aligned along nanogrooves expressed highly mature fibrillar FAs mediated by integrin clustering. Microarray analysis revealed that Tspan11, a member of the transmembrane tetraspanin protein family, was upregulated in cells on the nanogrooved surface compared with that in cells on isotropic, flat, or rough surfaces. Tspan11 silencing significantly disrupted osteoblast alignment and further construction of aligned bone matrix orthogonal to cell orientation. Our results demonstrate that the unique bone matrix formation orthogonal to cell alignment is facilitated by FA maturation. To the best of our knowledge, this report is the first to show that FA assembly mediated by Tspan11 determines the direction of bone matrix organization.

Wang P., Todai M., Nakano T.. Beta titanium single crystal with bone-like elastic modulus and large crystallographic elastic anisotropy. Journal of Alloys and Compounds, 782, 667. https://doi.org/10.1016/j.jallcom.2018.12.236., To develop single crystalline beta titanium implant as new hard tissue replacements for suppressing the stress shielding, we design a Ti-26.6Nb-6.7Al alloy (at. %) single crystal that exhibits large crystallographic elastic anisotropy and low Young's modulus. The anisotropy factor, A, reaches 3.42 that is the highest among all the reported values. The Young's modulus along <100> direction, E100, is only 36 GPa that is similar to the Young's modulus of cortical bone. These results prove our proposed design strategy and provide a new path to design beta titanium single crystal with bone-like elastic modulus for implant to minimize stress shielding.

Ishimoto T., Suetoshi R., Cretin D., et al. Quantitative ultrasound (QUS) axial transmission method reflects anisotropy in micro-arrangement of apatite crystallites in human long bones: A study with 3-MHz-frequency ultrasound. Bone, 127, 82. https://doi.org/10.1016/j.bone.2019.05.034., Anisotropic arrangement of apatite crystallites, i.e., preferential orientation of the apatite c-axis, is known to be an important bone quality parameter that governs the mechanical properties. However, noninvasive evaluation of apatite orientation has not been achieved to date. The present paper reports the potential of quantitative ultrasound (QUS) for noninvasive evaluation of the degree of apatite orientation in human bone for the first time. A novel QUS instrument for implementation of the axial transmission (AT) method is developed, so as to achieve precise measurement of the speed of sound (SOS) in the cortex (cSOS) of human long bone. The advantages of our QUS instrument are the following: (i) it is equipped with a cortical bone surface-morphology detection system to correct the ultrasound transmission distance, which should be necessary for AT measurement of long bone covered by soft tissue of non-uniform thickness; and (ii) ultrasound with a relatively high frequency of 3 MHz is employed, enabling thickness-independent cSOS measurement even for the thin cortex by preventing guide wave generation. The reliability of the proposed AT measurement system is confirmed through comparison with the well-established direct transmission (DT) method. The cSOS in human long bone is found to exhibit considerable direction-dependent anisotropy; the axial cSOS (3870 ± 66 m/s) is the highest, followed by the tangential (3411 ± 94 m/s) and radial (3320 ± 85 m/s) cSOSs. The degree of apatite orientation exhibits the same order, despite the unchanged bone mineral density. Multiple regression analysis reveals that the cSOS of human long bone strongly reflects the apatite orientation. The cSOS determined by the AT method is positively correlated with that determined by the DT method and sensitively reflects the apatite orientation variation, indicating the validity of the AT instrument developed in this study. Our instrument will be beneficial for noninvasive evaluation of the

Hagihara K., Yamasaki M., Kawamura Y., et al. Strengthening of Mg-based long-period stacking ordered (LPSO) phase with deformation kink bands. Materials Science and Engineering A, 763, 138163. https://doi.org/10.1016/j.msea.2019.138163., The mechanical properties of the Mg-based LPSO-phase are expected to be strongly affected by the microstructure due to its anisotropic crystal structure. However, the fine details have not been sufficiently understood yet. This study first clarified the detailed microstructural factors that govern the strength of the LPSO-phase by examining alloys with microstructures that were significantly varied via directional solidification and extrusion processes. Refining the microstructure is significantly effective for strengthening LPSO-phase alloys. The yield stress of LPSO-phase alloys with random texture was previously reported to be increased by reducing the “length” of plate-like LPSO-phase grains. In addition, it was found in this study that the formation stress in the deformation kink band, which is a unique deformation mode in an LPSO-phase alloy, can be increased by decreasing the “thickness” of the grains. Furthermore, the study used directionally solidified crystals provided direct evidence that the introduction of the deformation kink band effectively increases the yield stress and work-hardening rate of alloys by hindering the motion of basal dislocations. This “kink-band strengthening” was found to have considerable temperature dependence. The strengthening is significant at or below 200 °C, but the effect gradually decreases above 300 °C and is accompanied by the operation of non-basal slip. The results quantitatively clarified that kink-band strengthening is one predominant reason why the LPSO-phase extruded alloy exhibits an unusually high yield stress at any loading orientation.

Nagase T., Todai M., Hori T., et al. Microstructure of equiatomic and non-equiatomic Ti-Nb-Ta-Zr-Mo high-entropy alloys for metallic biomaterials. Journal of Alloys and Compounds, 753, 412. https://doi.org/10.1016/j.jallcom.2018.04.082., The microstructures of equiatomic TiNbTaZrMo (Ti20Nb20Ta20Zr20Mo20—subscript numerals denote at.%) and non-equiatomic Ti2.6NbTaZrMo (Ti39.4Nb15.2Ta15.2Zr15.2Mo15.2) high-entropy alloys (HEAs) were investigated for use in metallic biomaterials, and discussed based on their thermodynamics. Equiaxial dendrite structures were observed in the as-cast specimens. Ta, Nb, and Mo were abundant in the main dendrite phase with a body centered cubic (bcc) structure, while Ti and Zr showed a tendency to be abundant in the inter-dendrite region with a bcc structure. The distribution of the constituent elements can be explained through the distribution coefficients during solidification estimated using thermodynamic calculations. The thermodynamic calculations focusing on the solidification process were effective not only for the evaluation of the solidification microstructure, but also for the design of Ti-Nb-Ta-Zr-Mo-based HEAs. The non-equiatomic Ti2.6NbTaZrMo HEA (Ti39.4Nb15.2Ta15.2Zr15.2Mo15.2) was designed based on thermodynamic calculations and the solidification microstructure was studied.

Hagihara K., Li Z., Yamasaki M., et al. Strain-rate dependence of deformation behavior of LPSO-phases. Materials Letters 214, 119 (2018); https://doi.org/https://doi.org/10.1016/j.matlet.2017.11.117., This is the first report clarifying the influence of the strain rate on the deformation behavior of Mg-based long-period stacking ordered (LPSO) phases with 14H, 18R, and 10H structures. The flow stress by basal slip showed a weakly positive or negligible strain-rate dependence, while the flow stress accompanied by the formation of deformation kink bands showed a unique negative strain-rate dependence. These results give the first experimental evidence on the recent proposal that Zn and Y atoms segregate at the kink band boundaries and hinder their migration, from the viewpoint of the mechanical properties.

Ishimoto T., Yamada K., Takahashi H., et al. Trabecular health of vertebrae based on anisotropy in trabecular architecture and collagen/apatite micro-arrangement after implantation of intervertebral fusion cages in the sheep spine. Bone, 108, 25. https://doi.org/10.1016/j.bone.2017.12.012., Healthy trabecular bone shows highly anisotropic trabecular architecture and the preferential orientation of collagen and apatite inside a trabecula, both of which are predominantly directed along the cephalocaudal axis. This makes trabecular bone stiff in the principally loaded direction (cephalocaudal axis). However, changes in these anisotropic trabecular characteristics after the insertion of implant devices remain unclear. We defined the trabecular architectural anisotropy and the preferential orientation of collagen and apatite as parameters of trabecular bone health. In the present study, we analyzed these parameters after the implantation of two types of intervertebral fusion cages, open and closed box-type cages, into sheep spines for 2 and 4 months. Alteration and evolution of trabecular health around and inside the cages depended on the cage type and implantation duration. At the boundary region, the values of trabecular architectural anisotropy and apatite orientation for the closed-type cages were similar to those for isotropic conditions. In contrast, significantly larger anisotropy was found for open-type cages, indicating that the open-type cage tended to maintain trabecular anisotropy. Inside the open-type cage, trabecular architectural anisotropy and apatite orientation significantly increased with time after implantation. Assessing trabecular anisotropy might be useful for the evaluation of trabecular health and the validation and refinement of implant designs.

Hagihara K., Li Z., Yamasaki M., et al. Strain-rate dependence of deformation behavior of LPSO-phases. Materials Letters 214, 119 (2018); https://doi.org/https://doi.org/10.1016/j.matlet.2017.11.117., This is the first report clarifying the influence of the strain rate on the deformation behavior of Mg-based long-period stacking ordered (LPSO) phases with 14H, 18R, and 10H structures. The flow stress by basal slip showed a weakly positive or negligible strain-rate dependence, while the flow stress accompanied by the formation of deformation kink bands showed a unique negative strain-rate dependence. These results give the first experimental evidence on the recent proposal that Zn and Y atoms segregate at the kink band boundaries and hinder their migration, from the viewpoint of the mechanical properties.

Gokcekaya O., Ergun C., Webster T.J., et al. Influence of precursor deficiency sites for borate incorporation on the structural and biological properties of boronated hydroxyapatite. Ceramics International; https://doi.org/https://doi.org/10.1016/j.ceramint.2022.10.232., The biological properties of hydroxyapatite (HA) are significantly influenced by its compositional characteristics especially doping elements and/or Ca/P ratio, which can be altered by precursor chemistry. In this study, a group of boronated (B-incorporated) hydroxyapatite (BHA) was synthesized using a precipitation method by setting the Ca/P ratio to the stoichiometric value of HA (1.67), while altering the precursor chemistry by adjusting either (Ca + B)/P (Ca-deficient precursor, BC) or Ca/(P + B) (P-deficient precursor, BP). After heat-treatment, the partial decomposition of the BC was observed, forming tricalcium phosphate as the byproduct, however, the BP showed phase stability at all temperatures. The B-ionic species in the form of (BO₂)⁻ and (BO₃)³⁻ were incorporated into the HA structure at the (PO₄)³⁻ and (OH)⁻ positions, respectively. The incorporation of the B species also facilitated the incorporation of (CO₃)²⁻ groups specifically in the BPs. This is the first finding on BHA reporting that preferential (CO₃)²⁻ incorporation depends on the precursor chemistry used. As a result, osteoblast adhesion was superior on the BPs compared to pure HA owing to the carbonated structure, increasing cell spreading area. As such, this in vitro study highlighted that the present P-deficient precursor approach for synthesizing BHA improved biocompatibility properties and should, thus, be further considered for the next-generation of improved orthopedic applications.

Hagihara K., Li Z., Yamasaki M., et al. Strain-rate dependence of deformation behavior of LPSO-phases. Materials Letters 214, 119 (2018); https://doi.org/https://doi.org/10.1016/j.matlet.2017.11.117., This is the first report clarifying the influence of the strain rate on the deformation behavior of Mg-based long-period stacking ordered (LPSO) phases with 14H, 18R, and 10H structures. The flow stress by basal slip showed a weakly positive or negligible strain-rate dependence, while the flow stress accompanied by the formation of deformation kink bands showed a unique negative strain-rate dependence. These results give the first experimental evidence on the recent proposal that Zn and Y atoms segregate at the kink band boundaries and hinder their migration, from the viewpoint of the mechanical properties.

Gokcekaya O., Ergun C., Webster T.J., et al. Influence of precursor deficiency sites for borate incorporation on the structural and biological properties of boronated hydroxyapatite. Ceramics International; https://doi.org/https://doi.org/10.1016/j.ceramint.2022.10.232., The biological properties of hydroxyapatite (HA) are significantly influenced by its compositional characteristics especially doping elements and/or Ca/P ratio, which can be altered by precursor chemistry. In this study, a group of boronated (B-incorporated) hydroxyapatite (BHA) was synthesized using a precipitation method by setting the Ca/P ratio to the stoichiometric value of HA (1.67), while altering the precursor chemistry by adjusting either (Ca + B)/P (Ca-deficient precursor, BC) or Ca/(P + B) (P-deficient precursor, BP). After heat-treatment, the partial decomposition of the BC was observed, forming tricalcium phosphate as the byproduct, however, the BP showed phase stability at all temperatures. The B-ionic species in the form of (BO₂)⁻ and (BO₃)³⁻ were incorporated into the HA structure at the (PO₄)³⁻ and (OH)⁻ positions, respectively. The incorporation of the B species also facilitated the incorporation of (CO₃)²⁻ groups specifically in the BPs. This is the first finding on BHA reporting that preferential (CO₃)²⁻ incorporation depends on the precursor chemistry used. As a result, osteoblast adhesion was superior on the BPs compared to pure HA owing to the carbonated structure, increasing cell spreading area. As such, this in vitro study highlighted that the present P-deficient precursor approach for synthesizing BHA improved biocompatibility properties and should, thus, be further considered for the next-generation of improved orthopedic applications.

Naito K., Kuwahara Y., Yamamoto H., et al. Improvement of acid resistance of Zn-doped dentin by newly generated chemical bonds. Materials and Design, 215, 110412. https://doi.org/10.1016/j.matdes.2022.110412., Dental caries, the world's most prevalent infectious disease, is caused by the diffusion of hydroxyl ions into tooth structures. To prevent dental caries, the application of fluoride (F) and zinc (Zn) ions to teeth surfaces are potential effective measures. In this study, The ionic influence, especially the chemical bond of F and Zn, on the acid resistance of dentin were investigated by particle induced X-ray / gamma-ray emission, X-ray diffraction, X-ray photoelectron spectroscopy and X-ray absorption spectroscopy. The results showed Zn was distributed in the limited surface layer of dentin without altering its crystal structure. From the Zn K edge extended X-ray absorption fine structure, Zn incorporated into dentin was surrounded by oxygen and demonstrated four-fold coordination. The bond length and chemical state of Zn–O in Zn doped dentin suggested newly generated Zn–O covalent bond, which may improve acid resistance of dentin. This study showed that the atomic and molecular structures, such as the molecular distances and chemical state, influenced acid resistance of teeth, emphasizing the validity of chemical state analysis for understanding properties in biomaterials.

Nagase T., Todai M., Hori T., et al. Microstructure of equiatomic and non-equiatomic Ti-Nb-Ta-Zr-Mo high-entropy alloys for metallic biomaterials. Journal of Alloys and Compounds, 753, 412. https://doi.org/10.1016/j.jallcom.2018.04.082., The microstructures of equiatomic TiNbTaZrMo (Ti20Nb20Ta20Zr20Mo20—subscript numerals denote at.%) and non-equiatomic Ti2.6NbTaZrMo (Ti39.4Nb15.2Ta15.2Zr15.2Mo15.2) high-entropy alloys (HEAs) were investigated for use in metallic biomaterials, and discussed based on their thermodynamics. Equiaxial dendrite structures were observed in the as-cast specimens. Ta, Nb, and Mo were abundant in the main dendrite phase with a body centered cubic (bcc) structure, while Ti and Zr showed a tendency to be abundant in the inter-dendrite region with a bcc structure. The distribution of the constituent elements can be explained through the distribution coefficients during solidification estimated using thermodynamic calculations. The thermodynamic calculations focusing on the solidification process were effective not only for the evaluation of the solidification microstructure, but also for the design of Ti-Nb-Ta-Zr-Mo-based HEAs. The non-equiatomic Ti2.6NbTaZrMo HEA (Ti39.4Nb15.2Ta15.2Zr15.2Mo15.2) was designed based on thermodynamic calculations and the solidification microstructure was studied.

Ishimoto T., Suetoshi R., Cretin D., et al. Quantitative ultrasound (QUS) axial transmission method reflects anisotropy in micro-arrangement of apatite crystallites in human long bones: A study with 3-MHz-frequency ultrasound. Bone, 127, 82. https://doi.org/10.1016/j.bone.2019.05.034., Anisotropic arrangement of apatite crystallites, i.e., preferential orientation of the apatite c-axis, is known to be an important bone quality parameter that governs the mechanical properties. However, noninvasive evaluation of apatite orientation has not been achieved to date. The present paper reports the potential of quantitative ultrasound (QUS) for noninvasive evaluation of the degree of apatite orientation in human bone for the first time. A novel QUS instrument for implementation of the axial transmission (AT) method is developed, so as to achieve precise measurement of the speed of sound (SOS) in the cortex (cSOS) of human long bone. The advantages of our QUS instrument are the following: (i) it is equipped with a cortical bone surface-morphology detection system to correct the ultrasound transmission distance, which should be necessary for AT measurement of long bone covered by soft tissue of non-uniform thickness; and (ii) ultrasound with a relatively high frequency of 3 MHz is employed, enabling thickness-independent cSOS measurement even for the thin cortex by preventing guide wave generation. The reliability of the proposed AT measurement system is confirmed through comparison with the well-established direct transmission (DT) method. The cSOS in human long bone is found to exhibit considerable direction-dependent anisotropy; the axial cSOS (3870 ± 66 m/s) is the highest, followed by the tangential (3411 ± 94 m/s) and radial (3320 ± 85 m/s) cSOSs. The degree of apatite orientation exhibits the same order, despite the unchanged bone mineral density. Multiple regression analysis reveals that the cSOS of human long bone strongly reflects the apatite orientation. The cSOS determined by the AT method is positively correlated with that determined by the DT method and sensitively reflects the apatite orientation variation, indicating the validity of the AT instrument developed in this study. Our instrument will be beneficial for noninvasive evaluation of the

Hagihara K., Yamasaki M., Kawamura Y., et al. Strengthening of Mg-based long-period stacking ordered (LPSO) phase with deformation kink bands. Materials Science and Engineering A, 763, 138163. https://doi.org/10.1016/j.msea.2019.138163., The mechanical properties of the Mg-based LPSO-phase are expected to be strongly affected by the microstructure due to its anisotropic crystal structure. However, the fine details have not been sufficiently understood yet. This study first clarified the detailed microstructural factors that govern the strength of the LPSO-phase by examining alloys with microstructures that were significantly varied via directional solidification and extrusion processes. Refining the microstructure is significantly effective for strengthening LPSO-phase alloys. The yield stress of LPSO-phase alloys with random texture was previously reported to be increased by reducing the “length” of plate-like LPSO-phase grains. In addition, it was found in this study that the formation stress in the deformation kink band, which is a unique deformation mode in an LPSO-phase alloy, can be increased by decreasing the “thickness” of the grains. Furthermore, the study used directionally solidified crystals provided direct evidence that the introduction of the deformation kink band effectively increases the yield stress and work-hardening rate of alloys by hindering the motion of basal dislocations. This “kink-band strengthening” was found to have considerable temperature dependence. The strengthening is significant at or below 200 °C, but the effect gradually decreases above 300 °C and is accompanied by the operation of non-basal slip. The results quantitatively clarified that kink-band strengthening is one predominant reason why the LPSO-phase extruded alloy exhibits an unusually high yield stress at any loading orientation.

Matsugaki A., Yamazaki D., Nakano T.. Selective patterning of netrin-1 as a novel guiding cue for anisotropic dendrogenesis in osteocytes. Materials Science and Engineering C, 108, 110391. https://doi.org/10.1016/j.msec.2019.110391., Although protein patterning approaches have found widespread applications in tuning surface characteristics of biomaterials, selective control of growth in cell body and dendrites utilizing such platforms remains difficult. The functional roles assumed by cell body and dendrites in a physiological milieu have extremely high specificity. In particular, osteocytes embedded inside the mineralized bone matrix are interconnected via dendritic cell processes characterized by an anisotropic arrangement of the lacunar-canalicular system, where the fluid-flow inside the canaliculi system regulates the mechanoresponsive functionalization of bone. Control of cellular networks connected by dendritic cell processes is, therefore, imperative for constructing artificially controlled bone-mimetic structures and as an extension, for gaining insights into the molecular mechanisms underlying dendrogenesis inside the mineralized bone matrix. Here, we report an innovative strategy to induce controlled elongation of cell body or dendritic process structures in selective directions by using the inkjet printing technique. Artificial runways employing netrin-1, inspired by neural architecture, were utilized to trigger controlled elongation in the osteocyte dendritic processes in desired directions. This is the first report, to the best of our knowledge, demonstrating that anisotropic dendrogenesis of osteocytes can be controlled with selective patterning of extracellular proteins, specifically via the axon guidance ligand netrin-1.

Gokcekaya O., Hayashi N., Ishimoto T., et al. Crystallographic orientation control of pure chromium via laser powder bed fusion and improved high temperature oxidation resistance. Additive Manufacturing, 36, 101624. https://doi.org/10.1016/j.addma.2020.101624., This is the first comprehensive study on the development of a cubic crystallographic texture in pure chromium (Cr) manufactured using laser powder bed fusion (LPBF) with different laser energy densities to alter its microstructure and high-temperature oxidation behavior. An increase in the laser energy density led to the formation of a strong crystallographic texture, which was preferentially oriented in the (100) plane, and there were microstructural improvements in the pure Cr. The grain size of the (100)-oriented Cr was larger than that of the randomly oriented Cr. In addition, the high-angle grain boundary and coincident site lattice (CSL) boundary characteristics were altered. The (100)-oriented Cr exhibited a decrease in the oxide thickness that was due to the decrease in the grain boundary density with a larger grain size and an increase in the CSL boundary ratio. In contrast, the Cr with a random texture showed higher oxidation kinetics and spallation of the oxide layer. The oxidation kinetics of the pure Cr manufactured using LPBF obeyed the parabolic rate law. However, the crystal orientation affected the oxidation of the Cr as the (100)-oriented pure Cr displayed a lower parabolic rate constant, indicating that the (100)-oriented Cr was oxidation-resistant. This is the first report to demonstrate the cubic crystallographic texture formation and the improvement of high-temperature oxidation resistance in Cr manufactured using LPBF.

Matsuzaka T., Matsugaki A., Nakano T.. Control of osteoblast arrangement by osteocyte mechanoresponse through prostaglandin E2 signaling under oscillatory fluid flow stimuli. Biomaterials, 279, 121203. https://doi.org/10.1016/j.biomaterials.2021.121203., Anisotropic collagen/apatite microstructure is a prominent determinant of bone tissue functionalization; in particular, bone matrix modulates its anisotropic microstructure depending on the surrounding mechanical condition. Although mechanotransduction in bones is governed by osteocyte function, the precise mechanisms linking mechanical stimuli and anisotropic formation of collagen/apatite microstructure are poorly understood. Here we developed a novel anisotropic mechano-coculture system which enables the understanding of the biological mechanisms regulating the oriented bone matrix formation, which is constructed by aligned osteoblasts. The developed model provides bone-mimetic coculture platform that enables simultaneous control of mechanical condition and osteoblast-osteocyte communication with an anisotropic culture scaffold. The engineered coculture device helps in understanding the relationship between osteocyte mechanoresponses and osteoblast arrangement, which is a significant contributor to anisotropic organization of bone tissue. Our study showed that osteocyte responses to oscillatory flow stimuli regulated osteoblast arrangement through soluble molecular interactions. Importantly, we found that prostaglandin E2 is a novel determinant for oriented collagen/apatite organization of bone matrix, through controlling osteoblast arrangement.

Ishimoto T., Hagihara K., Hisamoto K., et al. Stability of crystallographic texture in laser powder bed fusion: Understanding the competition of crystal growth using a single crystalline seed. Additive Manufacturing, 43, 102004. https://doi.org/10.1016/j.addma.2021.102004., In metal additive manufacturing, crystallographic orientation control is a promising method for tailoring the functions of metallic parts. However, despite its importance in the fabrication of texture-controlled functional parts, the stability of the crystallographic texture is not widely discussed. Herein, the crystallographic texture stability under laser powder bed fusion was investigated. Two methodologies were employed. One is that a laser scanning strategy was alternately changed for a specific number of layers. The other is a “seeding” experiment in which single-crystalline substrates with controlled crystallographic orientations in the building (z-) direction and the xy-plane (perpendicular to the building direction) were used as the starting substrate. The transient zone width, where the crystallographic orientation was inherited from the layer beneath, was analyzed to evaluate the texture stability. The crystallographic direction of the seed within the xy-plane, rather than the building direction, determined the transient zone width, i.e., the texture stability. In particular, the texture in the newly deposited portion was stable when the laser scanning direction matched the <100> orientation in the underneath layer, otherwise the crystal orientation switched rapidly, such that the <100> orientation was parallel to the scanning direction. Interestingly, the crystallographic orientation along the building direction in the underneath layer hardly impacted the stability of the texture. Therefore, for the first time, it has been clarified that the <100> orientation in the scanning direction, rather than the building direction, was preferentially stabilized, whereas the orientation in the other directions secondary stabilized.

Tsutsumi Y., Ishimoto T., Oishi T., et al. Crystallographic texture- and grain boundary density-independent improvement of corrosion resistance in austenitic 316L stainless steel fabricated via laser powder bed fusion. Additive Manufacturing, 45, 102066. https://doi.org/10.1016/j.addma.2021.102066., Improvement of corrosion resistance of austenitic 316L stainless steel via laser powder bed fusion (LPBF) is currently a prominent research topic; however, the effects of crystallographic texture and the related grain boundary density on the corrosion resistance of LPBF-fabricated parts have not been elucidated. For biomedical applications, crystallographic texture control from a single crystalline-like to randomly oriented polycrystalline microstructure is highly attractive for optimizing the mechanical properties (particularly the Young's modulus) of implants. An investigation of the impacts of crystallographic planes and grain boundaries exposed to the biological environment on corrosion behavior is necessary. 316L stainless steels with different crystallographic textures and grain boundary densities were successfully fabricated via LPBF. The corrosion resistances of the LPBF-fabricated specimens were comprehensively assessed by anodic polarization, dissolution, and crevice corrosion repassivation tests. The LPBF-fabricated specimens showed extremely high pitting potentials in the physiological saline compared with the commercially available counterparts, and importantly, excellent pitting corrosion resistance was observed irrespective of the crystallographic planes and grain boundary density exposed. Moreover, the LPBF-fabricated specimens did not show metastable pitting corrosion even in an accelerated test using an acid solution. The repassivation behavior of the specimens was not affected by LPBF. Such a drastic improvement in the corrosion resistances of the LPBF-fabricated specimens might be attributed to suppression of inclusion coarsening owing to the rapid cooling rate during solidification in LPBF. By using LPBF, the desired crystallographic texture can be introduced based on the desired mechanical properties without concern for corrosiveness.

Ishimoto T., Yamada K., Takahashi H., et al. Trabecular health of vertebrae based on anisotropy in trabecular architecture and collagen/apatite micro-arrangement after implantation of intervertebral fusion cages in the sheep spine. Bone, 108, 25. https://doi.org/10.1016/j.bone.2017.12.012., Healthy trabecular bone shows highly anisotropic trabecular architecture and the preferential orientation of collagen and apatite inside a trabecula, both of which are predominantly directed along the cephalocaudal axis. This makes trabecular bone stiff in the principally loaded direction (cephalocaudal axis). However, changes in these anisotropic trabecular characteristics after the insertion of implant devices remain unclear. We defined the trabecular architectural anisotropy and the preferential orientation of collagen and apatite as parameters of trabecular bone health. In the present study, we analyzed these parameters after the implantation of two types of intervertebral fusion cages, open and closed box-type cages, into sheep spines for 2 and 4 months. Alteration and evolution of trabecular health around and inside the cages depended on the cage type and implantation duration. At the boundary region, the values of trabecular architectural anisotropy and apatite orientation for the closed-type cages were similar to those for isotropic conditions. In contrast, significantly larger anisotropy was found for open-type cages, indicating that the open-type cage tended to maintain trabecular anisotropy. Inside the open-type cage, trabecular architectural anisotropy and apatite orientation significantly increased with time after implantation. Assessing trabecular anisotropy might be useful for the evaluation of trabecular health and the validation and refinement of implant designs.

Okada R., Yamato K., Kawakami M., et al. Low magnetic field promotes recombinant human BMP-2-induced bone formation and influences orientation of trabeculae and bone marrow-derived stromal cells. Bone Reports, 14, 100757. https://doi.org/10.1016/j.bonr.2021.100757., Effects of high magnetic fields [MFs, ≥ 1 T (T)] on osteoblastic differentiation and the orientation of cells or matrix proteins have been reported. However, the effect of low MFs (< 1 T) on the orientation of bone formation is not well known. This study was performed to verify the effects of low MFs on osteoblastic differentiation, bone formation, and orientation of both cells and newly formed bone. An apparatus was prepared with two magnets (190 mT) aligned in parallel to generate a parallel MF. In vitro, bone marrow-derived stromal cells of rats were used to assess the effects of low MFs on cell orientation, osteoblastic differentiation, and mineralization. A bone morphogenetic protein (BMP)-2-induced ectopic bone model was used to elucidate the effect of low MFs on microstructural indices, trabecula orientation, and the apatite c-axis orientation of newly formed bone. Low MFs resulted in an increased ratio of cells oriented perpendicular to the direction of the MF and promoted osteoblastic differentiation in vitro. Moreover, in vivo analysis demonstrated that low MFs promoted bone formation and changed the orientation of trabeculae and apatite crystal in a direction perpendicular to the MF. These changes led to an increase in the mechanical strength of rhBMP-2-induced bone. These results suggest that the application of low MFs has potential to facilitate the regeneration of bone with sufficient mechanical strength by controlling the orientation of newly formed bone.

Iijima Y., Nagase T., Matsugaki A., et al. Design and development of Ti–Zr–Hf–Nb–Ta–Mo high-entropy alloys for metallic biomaterials. Materials and Design, 202, 109548. https://doi.org/10.1016/j.matdes.2021.109548., Applying empirical alloy parameters (including Mo equivalent), the predicted ground state diagram, and thermodynamic calculations, noble nonequiatomic Ti–Zr–Hf–Nb–Ta–Mo high-entropy alloys for metallic biomaterials (BioHEAs) were designed and newly developed. It is found that the Moeq and valence electron concentration (VEC) parameters are useful for alloy design involving BCC structure formation in bio medium-entropy alloys and BioHEAs. Finally, we find a Ti28.33Zr28.33Hf28.33Nb6.74Ta6.74Mo1.55 (at.%) BioHEA that exhibits biocompatibility comparable to that of CP–Ti, higher mechanical strength than CP–Ti, and an appreciable room-temperature tensile ductility. The current findings pave the way for new Ti–Zr–Hf–Nb–Ta–Mo BioHEAs development and are applicable for another BioHEA alloys system.

Ishimoto T., Kobayashi Y., Takahata M., et al. Outstanding in vivo mechanical integrity of additively manufactured spinal cages with a novel “honeycomb tree structure” design via guiding bone matrix orientation. Spine Journal, 22, 10, 1742. https://doi.org/10.1016/j.spinee.2022.05.006., BACKGROUND CONTEXT: Therapeutic devices for spinal disorders, such as spinal fusion cages, must be able to facilitate the maintenance and rapid recovery of spinal function. Therefore, it would be advantageous that future spinal fusion cages facilitate rapid recovery of spinal function without secondary surgery to harvest autologous bone. PURPOSE: This study investigated a novel spinal cage configuration that achieves in vivo mechanical integrity as a devise/bone complex by inducing bone that mimicked the sound trabecular bone, hierarchically and anisotropically structured trabeculae strengthened with a preferentially oriented extracellular matrix. STUDY DESIGN/SETTINGS: In vivo animal study. METHODS: A cage possessing an anisotropic through-pore with a grooved substrate, that we termed “honeycomb tree structure,” was designed for guiding bone matrix orientation; it was manufactured using a laser beam powder bed fusion method through an additive manufacturing processes. The newly designed cages were implanted into sheep vertebral bodies for 8 and 16 weeks. An autologous bone was not installed in the newly designed cage. A pull-out test was performed to evaluate the mechanical integrity of the cage/bone interface. Additionally, the preferential orientation of bone matrix consisting of collagen and apatite was determined. RESULTS: The cage/host bone interface strength assessed by the maximum pull-out load for the novel cage without an autologous bone graft (3360±411 N) was significantly higher than that for the conventional cage using autologous bone (903±188 N) after only 8 weeks post-implantation. CONCLUSIONS: These results highlight the potential of this novel cage to achieve functional fusion between the cage and host bone. Our study provides insight into the design of highly functional spinal devices based on the anisotropic nature of bone. CLINICAL SIGNIFICANCE: The sheep spine is similar to the human spine in its stress condition and trabecular bone architecture

Hamai R., Sakai S., Shiwaku Y., et al. Octacalcium phosphate crystals including a higher density dislocation improve its materials osteogenecity. Applied Materials Today, 26, 101279. https://doi.org/10.1016/j.apmt.2021.101279., Herein, we show that the enhanced osteogenecity of octacalcium phosphate (OCP) biomaterial, recently identified as an important element in hybrid organic–inorganic nanocomposites involved in the initial hydroxyapatite crystal expansion in mammal bones, results from an enhanced chemical property, stemming from the presence of lattice strain and dislocations. Two types of OCPs were synthesized by wet-chemical processing in the presence (c-OCP) and absence (w-OCP) of gelatin, respectively, and subjected to structural, chemical, and biological analyses. High-resolution transmission electron microscopy (HRTEM) and fast Fourier transform (FFT) analyses revealed that c-OCP crystals contained approximately six times higher edge dislocations with Burgers vectors perpendicular to a-axis than that in the case of w-OCP. The dissolution of c-OCP crystal in tris-HCl buffer occurred toward the long axis of the crystal, most likely, toward the lattice strain along the c-axis direction, while w-OCP crystal dissolved toward the a-axis direction. The study suggested that the increment of internal energy by the higher dislocation density contributed promoting c-OCP dissolution and hydrolysis through decreasing the activation energy. c-OCP crystal displayed enhanced in vitro mesenchymal stem 2D cell and 3D spheroid differentiation, in vivo bone formation, and apatite crystallographic orientation in critical-sized rat calvarial defect model as compared to w-OCP crystal, at the same time, converting to apatite structure earlier than w-OCP. The present study demonstrates that dislocation-related dissolution along with enhanced conversion of OCP is a determinant in bone induction, which may be relevant to normal bone development utilizing OCP biomaterials.

Ekubaru Y., Gokcekaya O., Ishimoto T., et al. Excellent strength–ductility balance of Sc-Zr-modified Al–Mg alloy by tuning bimodal microstructure via hatch spacing in laser powder bed fusion. Materials and Design, 221, 110976. https://doi.org/10.1016/j.matdes.2022.110976., The bimodal microstructure, which comprises ultrafine grains (UFGs) forming along the melt pool boundary and relatively coarse grains inside the melt pool, is a characteristic of the Sc-Zr-modified Al–Mg-based alloy (Scalmalloy) microstructure manufactured using laser powder bed fusion (LPBF). Focusing on this microstructural feature, we investigated the improvement in the mechanical properties of LPBF-fabricated Scalmalloy by tailoring the volume fraction of UFGs. Our approach was to decrease the laser hatch spacing (d) from 0.1 to 0.04 mm, while the volume fraction of UFGs increased from 34.6 ± 0.6 % (d = 0.1 mm) to 59.5 ± 0.5 % (d = 0.06 mm). The tensile yield stress increased from 296 ± 9 (d = 0.1 mm) to 380 ± 6 MPa (d = 0.06 mm), while maintaining a large elongation (14.8 % ± 1.2 %). The yield stress and elongation were superior to those of the cast counterparts by 2.9 and 4.0 times, respectively. In the sample with d = 0.04 mm, pores formed owing to excessive thermal energy input. Additionally, we investigated multiple strengthening mechanisms of the as-fabricated alloy. This is the first study to improve the mechanical properties of LPBF-fabricated Scalmalloy by optimizing the track-to-track interval and tuning the UFG fraction.

Nagase T., Iijima Y., Matsugaki A., et al. Design and fabrication of Ti–Zr-Hf-Cr-Mo and Ti–Zr-Hf-Co-Cr-Mo high-entropy alloys as metallic biomaterials. Materials Science and Engineering C, 107, 110322. https://doi.org/10.1016/j.msec.2019.110322., Novel TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo high-entropy alloys for metallic biomaterials (bio-HEAs) were developed based on the combination of Ti–Nb–Ta–Zr–Mo alloy system and Co–Cr–Mo alloy system as commercially-used metallic biomaterials. Ti–Zr-Hf-Cr-Mo and Ti–Zr-Hf-Co-Cr-Mo bio-HEAs were designed using (a) a tree-like diagram for alloy development, (b) empirical alloy parameters for solid-solution-phase formation, and (c) thermodynamic calculations focused on solidification. The newly-developed bio-HEAs overcomes the limitation of classical metallic biomaterials by the improvement of (i) mechanical hardness and (ii) biocompatibility all together. The TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo bio-HEAs showed superior biocompatibility comparable to that of commercial-purity Ti. The superior biocompatibility, high mechanical hardness and low liquidus temperature for the material processing in TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo bio-HEAs compared with the Ti–Nb–Ta–Zr–Mo bio-HEAs gave the authenticity of the application of bio-HEAs for orthopedic implants with multiple functions.

Nagase T., Todai M., Wang P., et al. Design and development of (Ti, Zr, Hf)-Al based medium entropy alloys and high entropy alloys. Materials Chemistry and Physics, 276, 125409. https://doi.org/10.1016/j.matchemphys.2021.125409., The design and development of TiZrHfAl medium entropy alloy (MEA), and the TiZrHfAlNb0.2 and TiZrHfAlV0.2 high entropy alloys (HEAs) is described. The combination of 4th subgroup elements (Ti, Zr, and Hf) with Al is discussed based on the periodic table and taxonomy of HEAs. The alloys were designed using alloy parameters for HEAs, predicted ground state diagrams from the Materials Project, and the calculation of phase diagrams (CALPHAD). Rapid solidification was effective to suppress the formation of intermetallic compounds, resulting in BCC/B2 phase formation. Significant differences in the constituent phases and Vickers hardness between ingots and melt-spun ribbons were found among the TiZrHfAl MEA, TiZrHfAlNb0.2, and TiZrHfAlV0.2 HEAs.

Wang P., Todai M., Nakano T.. Beta titanium single crystal with bone-like elastic modulus and large crystallographic elastic anisotropy. Journal of Alloys and Compounds, 782, 667. https://doi.org/10.1016/j.jallcom.2018.12.236., To develop single crystalline beta titanium implant as new hard tissue replacements for suppressing the stress shielding, we design a Ti-26.6Nb-6.7Al alloy (at. %) single crystal that exhibits large crystallographic elastic anisotropy and low Young's modulus. The anisotropy factor, A, reaches 3.42 that is the highest among all the reported values. The Young's modulus along <100> direction, E100, is only 36 GPa that is similar to the Young's modulus of cortical bone. These results prove our proposed design strategy and provide a new path to design beta titanium single crystal with bone-like elastic modulus for implant to minimize stress shielding.

Nakanishi Y., Matsugaki A., Kawahara K., et al. Unique arrangement of bone matrix orthogonal to osteoblast alignment controlled by Tspan11-mediated focal adhesion assembly. Biomaterials, 209, 103. https://doi.org/10.1016/j.biomaterials.2019.04.016., During tissue construction, cells coordinate extracellular matrix (ECM)assembly depending on the cellular arrangement. The traditional understanding of the relationship between the ECM and cells is limited to the orientation-matched interaction between them. Indeed, it is commonly accepted that the bone matrix (collagen/apatite)is formed along osteoblast orientation. Nonetheless, our recent findings are contrary to the above theory; osteoblasts on nanogrooves organize formation of the bone matrix perpendicular to cell orientation. However, the precise molecular mechanisms underlying the orthogonal organization of bone matrix are still unknown. Here, we show that mature fibrillar focal adhesions (FAs)facilitate the perpendicular arrangement between cells and bone matrix. The osteoblasts aligned along nanogrooves expressed highly mature fibrillar FAs mediated by integrin clustering. Microarray analysis revealed that Tspan11, a member of the transmembrane tetraspanin protein family, was upregulated in cells on the nanogrooved surface compared with that in cells on isotropic, flat, or rough surfaces. Tspan11 silencing significantly disrupted osteoblast alignment and further construction of aligned bone matrix orthogonal to cell orientation. Our results demonstrate that the unique bone matrix formation orthogonal to cell alignment is facilitated by FA maturation. To the best of our knowledge, this report is the first to show that FA assembly mediated by Tspan11 determines the direction of bone matrix organization.

Ishimoto T., Hagihara K., Hisamoto K., et al. Stability of crystallographic texture in laser powder bed fusion: Understanding the competition of crystal growth using a single crystalline seed. Additive Manufacturing, 43, 102004. https://doi.org/10.1016/j.addma.2021.102004., In metal additive manufacturing, crystallographic orientation control is a promising method for tailoring the functions of metallic parts. However, despite its importance in the fabrication of texture-controlled functional parts, the stability of the crystallographic texture is not widely discussed. Herein, the crystallographic texture stability under laser powder bed fusion was investigated. Two methodologies were employed. One is that a laser scanning strategy was alternately changed for a specific number of layers. The other is a “seeding” experiment in which single-crystalline substrates with controlled crystallographic orientations in the building (z-) direction and the xy-plane (perpendicular to the building direction) were used as the starting substrate. The transient zone width, where the crystallographic orientation was inherited from the layer beneath, was analyzed to evaluate the texture stability. The crystallographic direction of the seed within the xy-plane, rather than the building direction, determined the transient zone width, i.e., the texture stability. In particular, the texture in the newly deposited portion was stable when the laser scanning direction matched the <100> orientation in the underneath layer, otherwise the crystal orientation switched rapidly, such that the <100> orientation was parallel to the scanning direction. Interestingly, the crystallographic orientation along the building direction in the underneath layer hardly impacted the stability of the texture. Therefore, for the first time, it has been clarified that the <100> orientation in the scanning direction, rather than the building direction, was preferentially stabilized, whereas the orientation in the other directions secondary stabilized.

Matsugaki A., Yamazaki D., Nakano T.. Selective patterning of netrin-1 as a novel guiding cue for anisotropic dendrogenesis in osteocytes. Materials Science and Engineering C, 108, 110391. https://doi.org/10.1016/j.msec.2019.110391., Although protein patterning approaches have found widespread applications in tuning surface characteristics of biomaterials, selective control of growth in cell body and dendrites utilizing such platforms remains difficult. The functional roles assumed by cell body and dendrites in a physiological milieu have extremely high specificity. In particular, osteocytes embedded inside the mineralized bone matrix are interconnected via dendritic cell processes characterized by an anisotropic arrangement of the lacunar-canalicular system, where the fluid-flow inside the canaliculi system regulates the mechanoresponsive functionalization of bone. Control of cellular networks connected by dendritic cell processes is, therefore, imperative for constructing artificially controlled bone-mimetic structures and as an extension, for gaining insights into the molecular mechanisms underlying dendrogenesis inside the mineralized bone matrix. Here, we report an innovative strategy to induce controlled elongation of cell body or dendritic process structures in selective directions by using the inkjet printing technique. Artificial runways employing netrin-1, inspired by neural architecture, were utilized to trigger controlled elongation in the osteocyte dendritic processes in desired directions. This is the first report, to the best of our knowledge, demonstrating that anisotropic dendrogenesis of osteocytes can be controlled with selective patterning of extracellular proteins, specifically via the axon guidance ligand netrin-1.

Nagase T., Iijima Y., Matsugaki A., et al. Design and fabrication of Ti–Zr-Hf-Cr-Mo and Ti–Zr-Hf-Co-Cr-Mo high-entropy alloys as metallic biomaterials. Materials Science and Engineering C, 107, 110322. https://doi.org/10.1016/j.msec.2019.110322., Novel TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo high-entropy alloys for metallic biomaterials (bio-HEAs) were developed based on the combination of Ti–Nb–Ta–Zr–Mo alloy system and Co–Cr–Mo alloy system as commercially-used metallic biomaterials. Ti–Zr-Hf-Cr-Mo and Ti–Zr-Hf-Co-Cr-Mo bio-HEAs were designed using (a) a tree-like diagram for alloy development, (b) empirical alloy parameters for solid-solution-phase formation, and (c) thermodynamic calculations focused on solidification. The newly-developed bio-HEAs overcomes the limitation of classical metallic biomaterials by the improvement of (i) mechanical hardness and (ii) biocompatibility all together. The TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo bio-HEAs showed superior biocompatibility comparable to that of commercial-purity Ti. The superior biocompatibility, high mechanical hardness and low liquidus temperature for the material processing in TiZrHfCr0.2Mo and TiZrHfCo0.07Cr0.07Mo bio-HEAs compared with the Ti–Nb–Ta–Zr–Mo bio-HEAs gave the authenticity of the application of bio-HEAs for orthopedic implants with multiple functions.