While conventionally manufactured metallic biomaterials can hardly meet all the requirements for bone implants including complex geometry, exact dimensions, adequate biodegradability, bone-matching mechanical properties, and biological function, two additional tools have recently appeared in the arsenal of biomaterials scientists which promise to deliver the desired combination of properties. First, the unique mechanical, electrical, and biological properties of graphene (Gr) and its derivatives (GDs), e.g., a Young's modulus up to 1 TPa, can be utilized to create metal matrix composites in which GDs of varied contents (typically not more than 2 wt%), sizes (lateral sizes from a few nanometers to several micrometers), surface areas (up to the theoretical value of 2630 m²/g), and layer numbers (typically up to 10) are embedded in the biodegradable metal matrix, thereby endowing the composite implants with extraordinary properties. Second, the distinct advantages of additive manufacturing (AM) make it possible for GD-containing composite materials to precisely mimic the complex shapes and structures of bones at multiple length scales. Here, a comprehensive review of the recent advances in the development of GD-containing biodegradable metal matrix composites (GBMMCs), ranging from composite fabrication, including composite powder preparation, and AM processes, to the evaluation of AM composites in terms of their mechanical and biological properties, is presented. Furthermore, the constraints in processing composite powders, the advantages and disadvantages of applicable AM techniques, and the mechanisms of mechanical reinforcement, biodegradation modulation, osteogenesis improvement, and cytotoxicity/antibacterial balance are critically analyzed. Thereafter, the foreseeable challenges faced in the development of the next-generation of bone implants based on GBMMCs are presented and some future directions of research are identified.

Additively manufactured (AM) iron (Fe)-based scaffolds have been developed as promising biodegradable bone-substituting biomaterials. Multi-material extrusion-based 3D printing has recently yielded Fe-manganese (Mn) alloy-based scaffolds that can resolve ferromagnetism and cytotoxicity associated with Fe-based biomaterials. Herein, we, for the first time, present the findings from in vivo study on extrusion-based AM FeMn-akermanite (Ak) scaffolds for critical-size bone defect repair. The scaffolds comprised Fe, 35 wt% Mn, and 20 or 30 vol% Ak, with microporous struts and 61–63 % porosity. Both scaffolds exhibited mechanical properties within the range of trabecular bone and provided suitable sites for Ca/P deposition during in vitro biodegradation. In vitro cell cultures demonstrated favorable cell responses without negating the osteogenic potential of cells. An in vivo study was conducted in a murine semi-orthotopic subcutaneous model. With this model, 4 bovine bone plugs were implanted subcutaneously with critical-size defects created at their cores. Scaffolds were placed into these critical-size defects to assess biodegradation and bone formation. After 16 weeks, the volume of scaffolds decreased by 6–8 %. The FeMn-20Ak scaffolds retained their yield strength and elastic modulus during the 16 weeks in vivo, whereas the mechanical integrity of FeMn-30Ak scaffolds deteriorated after mechanical push-out tests. Excellent osseointegration of both scaffold groups was apparent. 3D reconstruction of CT images revealed that FeMn-30Ak scaffolds had more newly formed tissue in the macro-pores than FeMn-20Ak. Altogether, our findings demonstrate the potential of AM FeMn-Ak scaffolds as biodegradable bone substitutes, encouraging further in vivo research in a large animal model.

Medical devices contribute to the carbon footprint generated by the healthcare sector. The development of implants and biomaterials using recycled waste materials promotes sustainable advances in tissue engineering. Additively manufactured (AM) bone-substituting biomaterials with multifunctional properties, e.g., biodegradability, antibacterial and osteogenic potential, can contribute to sustainable healthcare. Biodegradable biomaterials eliminate secondary surgeries to remove implants, reduce post-surgical complications, and enhance patient recovery, thus decreasing the energy usage and waste associated with medical treatments. Herein, we present porous iron (Fe) scaffolds incorporating 20 vol% waste-derived eggshell particles for bone substitution. The Fe-eggshell scaffolds were fabricated using direct ink writing (DIW) technique and underwent post-AM heat treatment. During sintering, the eggshell's main component – CaCO₃, transformed into CaO. Atomic diffusion between α-Fe and CaO phases resulted in the formation of Ca₂Fe₂O₅ phase at the interface. The scaffolds were 70 % porous and displayed a biodegradation rate of 0.11 mm/year. The mechanical properties were comparable to trabecular bone and the scaffolds endured 3 million loading cycles at 0.7σ_y in r-SBF. The scaffolds showed apatite-forming ability, evidenced by the formation of (carbonaceous) hydroxyapatite, which are conducive to preosteoblast adhesion, proliferation, and differentiation. RT-qPCR analysis confirmed the osteogenic potential of the specimens as evidenced by the upregulated expression of osteopontin and osteocalcin as compared to Ti6Al4V controls. Furthermore, the scaffolds exhibited bactericidal activity (>3.9-log CFU reduction) against methicillin-sensitive and multidrug-resistant strains of Staphylococcus aureus and delayed their biofilm formation. Our research showcases the exceptional multifunctionality of DIW Fe-eggshell composite scaffolds for the sustainable development of orthopedic biomaterials. Statement of significance: We aim to improve the biofunctionalities and sustainability of biodegradable bone substitutes, by developing the extrusion-based 3D printed porous Fe composite scaffolds containing eggshell-derived CaO bioceramics. Our results demonstrated that Fe-eggshell scaffolds exhibited hydroxyapatite-forming ability in simulated body fluid, having mechanical properties in the range of trabecular bone even after 4 weeks biodegradation, supported the proliferation of preosteoblasts and upregulated the expression of osteogenic genes. Moreover, the scaffolds were bactericidal against methicillin-sensitive and multi-drug resistant strains Staphylococcus aureus and delayed their biofilm formation.

Additive manufacturing (AM), also known as 3D printing, is gaining the attention of various industries as a viable alternative to conventional manufacturing, empowering design freedom, novel architectures, composition control, and sustainability. Meanwhile, metal matrix composites (MMCs) are being investigated for orthopedic implant applications due to their flexibility to achieve excellent strength, corrosion resistance, and bioactivity. Combining these two research fronts by utilizing AM for manufacturing multi-functional MMC bone scaffolds, having specific structures and compositions, has led to the recent development of a new generation of biomaterials with enhanced material properties not achievable with monolithic counterparts. Aimed at understanding the status of the research on the topic and identifying the remaining challenges, this review article discusses the utilization of AM for realizing the design vision of different MMC scaffolds, focusing on the synergistic combination of mechanical and biological characteristics, such as enhanced biodegradability, strength, and osteogenic properties. It starts by discussing the requirements for orthopedic implants and different AM techniques utilized thus far for manufacturing them, especially MMC orthopedic implants. Then, it delves into different MMCs, including Ti-, Mg-, and Fe-matrix composites that have been 3D printed into bone-substituting scaffolds and discusses their recent progress and specific characteristics. Finally, we identify the knowledge gaps and potential directions for developing MMCs further toward clinically viable, advanced orthopedic implants.

The development of high-fidelity three-dimensional (3D) tissue models can minimize the need for animal models in clinical medicine and drug development. However, physical limitations regarding the distances within which diffusion processes are effective impose limitations on the size of such constructs. That is because larger-size constructs experience necrosis, especially in their centers, due to the cells residing deep inside such constructs not receiving enough oxygen and nutrients. This hampers the sustained in vitro growth of the tissues which is required for achieving functional microtissues. To address this challenge, we used three types of 3D printing technologies to create perfusable networks at different length scales and integrate them into such constructs. Toward this aim, networks incorporating porous conduits with increasingly complex configurations were designed and fabricated using fused deposition modeling, stereolithography, and two-photon polymerization while optimizing the printing conditions for each of these technologies. Furthermore, following network embedding in hydrogels, contrast agent-enhanced micro-computed tomography and confocal fluorescence microscopy were employed to characterize one of the essential network functionalities, namely the diffusion function. The investigations revealed the effects of various design parameters on the diffusion behavior of the porous conduits over 24 h. We found that the number of pores exerts the most significant influence on the diffusion behavior of the contrast agent, followed by variations in the pore size and hydrogel concentration. The analytical approach and the findings of this study establish a solid base for a new technological platform to fabricate perfusable multiscale 3D porous networks with complex designs while enabling the customization of diffusion characteristics to meet specific requirements for sustained in vitro tissue growth. Statement of significance: This study addresses an essential limitation of current 3D tissue engineering, namely, sustaining tissue viability in larger constructs through optimized nutrient and oxygen delivery. By utilizing advanced 3D printing techniques this research proposes the fabrication of perfusable, multiscale and customizable networks that enhance diffusion and enable cell access to essential nutrients throughout the construct. The findings highlighted the role of network characteristics on the diffusion of a model compound within a hydrogel matrix. This work represents a promising technological platform for creating advanced in vitro 3D tissue models that can reduce the use of animal models in research involving tissue regeneration, disease models and drug development.

Tissue engineering approaches for cartilage tissue regeneration are expanding to include the complex features of the tissue, such as the biological and mechanical gradients. Many of these approaches are, however, based on the use of multiple biomaterials or concentrations, and crosslinking methods that make it difficult to integrate and control the properties of the resulting scaffolds. In this study, a 3D bioprinted scaffold with a stiffness gradient was fabricated by using a single biomaterial type and concentration combined with a directional ionic crosslinking method. The scaffolds revealed a gradient in stiffness from 39.8 ± 6.6 kPa at the top to 60.6 ± 10.9 kPa at the bottom of the scaffolds. Live/dead analysis of human chondrocytes embedded in the scaffolds showed no negative effects of the stiffness gradient on cell viability over 28 days. The induced stiffness gradient led to a gradient in cell density and sulfated glycosaminoglycan deposition in the bioprinted tissue constructs with enhanced values in the softer top region of the scaffolds as compared to the stiffer bottom part. This study showed a novel method to generate scaffolds with stiffness gradients from a single biomaterial and indicates that such scaffolds could be used to spatially regulate the behavior of chondrocytes and the associated deposition of the cartilage matrix.

Magnetic particles (MPs), due to their unique physical and chemical properties, have emerged as promising tools in bone tissue engineering. Their incorporation into scaffolds or uptake by bone cells, combined with exposure to external magnetic fields, has been shown in various studies to enhance cell adhesion, proliferation, and osteogenic differentiation. In this review, the state-of-the-art is presented on the synthesis processes of magnetized cells (MCs) and magnetized scaffolds (MSs), as well as the biological and mechanical effects of scaffold-free MCs, cell-seeded MSs, and MC-seeded MSs under externally applied magnetic fields on bone tissue engineering. Furthermore, the specific applications of these systems is highlighted, such as non-contact mechanical stimulation, and discuss their application to advance bone tissue engineering strategies.

OBJECTIVE: The treatment of mature biofilm in implant-associated infections (IAI) has become increasingly challenging, mainly due to the rise of antibiotic-resistant bacteria. While many antibacterial biomaterials harness their functionality through their surface properties, alternating magnetic field (AMF)-induced hyperthermia offers an approach from a fundamentally different angle. METHOD: To summarize and compare the practice of assessing AMF-induced hyperthermia in vitro and in vivo as treatment for implant-associated infections and the efficacy of this therapy, a literature search was conducted and 18 articles were selected based on relevance. RESULTS AND CONCLUSION: The studies have demonstrated that AMF-induced hyperthermia can effectively eliminate biofilms as a standalone treatment or in combination with antimicrobials. Although thermal tissue damage is an inherent concern, it can be controlled and reduced by implementing short intermittent heating patterns around 65-75ºC while still preserving antibacterial efficacy. However, clear guidelines for evaluating safety, particularly regarding thermal injury, are still lacking and should be a key focus of future work.

Osteoimmunomodulation (OIM) is emerging as a key biofunctionality of orthopedic implants. Biomaterial surface geometries can modulate the interactions between immune cells and osteoprogenitors at the bone-implant interface, positively affecting osteogenic differentiation and implant osseointegration. This review highlights the recent advancements in geometry-induced OIM (G-OIM) across multiple length scales (nano to mesoscale, including multiscale topographies and 3D scaffolds), identifying relations between specific geometries and subsequent mechanisms of OIM, as provided by the coculture model used. Our review reveals surface geometries with OIM potential at each length scale. These effects can be mediated by both M1 and M2 macrophages, wherein the pathway depends on the shape and length scale of the geometrical cues provided (e.g., integrin-mediated mechanotransduction for nanoscale topographies and macrophage contact inhibition for micropatterns). Most studies assess G-OIM predominantly based on geometry-induced macrophage polarization and its paracrine effect on osteoprogenitors. However, few studies utilizing direct coculture reveal the key role of the direct interplay between macrophages, osteoprogenitors, and biomaterial for OIM. The novel field of G-OIM is advancing at a high pace. It could benefit from improved, clinically relevant coculture models involving human-derived cells and technological developments in biomaterial design and fabrication. Such advances could establish (G-)OIM as a transformative approach for regenerative immunoengineering of orthopedic implants. Statement of significance: Osteoimmunomodulation, the ability of biomaterials to modulate the interactions between immune cells and skeletal cells to enhance osteogenesis, is increasingly recognized as a crucial biofunctionality for orthopedic biomaterials. Various biomaterial surface geometries can be used to target osteoimmune pathways. Given the complexity of these interactions, suitable coculture models are essential for studying the underlying cellular mechanisms. This review reveals the state-of-the-art results on geometry-induced osteoimmunomodulation. Not only does this review discuss approaches that have been taken thus far in terms of biomaterial geometry design at various length scales, but it also highlights the role of the coculture model in osteoimmunomodulation and the importance of advances in these in vitro models to improve the translation of research to clinical practice.

Titanium surfaces featuring high-aspect ratio (HAR) nanopillars can have antimicrobial and osteogenic properties. Nevertheless, the impact of these surfaces on immune cells and their potential for immunomodulation remain unclear. In this study, the effects of HAR titanium nanopillars produced by dry-etching (DETi) on the response of unstimulated (M0) and pro-inflammatory (M1) murine macrophages (J774A.1) have been explored. The findings revealed changes in the morphology and crystallinity of the DETi nanopillars along their height. After 48 h of culture, both M0 and M1 stimulated macrophages displayed a more elongated morphology, a smoother cell surface, and shorter cellular protrusions on the more hydrophilic and rough DETi surfaces. Furthermore, DETi surfaces induced polarization of M0 cells towards M2 phenotypes, whereas M1 stimulated cells showed M2-like elongated morphologies while maintaining a stronger pro-inflammatory response to DETi surfaces relative to the glass control. The findings indicate that the DETi surfaces can induce morphological changes in macrophages and specific immunomodulatory effects depending on their initial phenotype, highlighting the potential of such biomaterials to incorporate an immunomodulatory biofunctionality next to the osteogenic and bactericidal ones.

Two-photon polymerization (2PP) is an additive manufacturing technology capable of producing polymeric 3D nano- to mesoscale structures with design flexibility and sub-micron resolution. This study investigates the influence of 2PP printing parameters on the morphology and mechanical properties of solid and porous microstructures fabricated from three commercial resins: IP-Q, IP-S, and IP-polydimethylsiloxane (IP-PDMS). To evaluate micromechanical behavior, micropillar compression tests are conducted using IP-Q, which has not been extensively characterized. Porous structures retained 80–85% of the stiffness of solids for IP-Q and IP-S, and 50% for IP-PDMS. Fourier transform infrared spectroscopy showed degrees of conversion of 38% for IP-Q and 61% for IP-S and IP-PDMS. The optimal printing parameters for IP-Q micropillars were a laser power of 50 mW, slicing distance (s) of 1.2 μm, and hatching distance (h) of 1 μm. These settings correspond to a peak laser intensity of 1.58 × 10⁻¹¹ W cm⁻², a focal spot diameter (dxy) of 3.17 μm, a Rayleigh length (zR) of 10.13 μm, and a voxel overlap (δ) of 0.6. These conditions yielded a Young's modulus of 3.7 GPa and yield strength of 75.21 MPa. Overall, the findings emphasize the challenges of parameter optimization when introducing porosity and comparing materials. The results provide a systematic framework for tailoring 2PP processing to guide biomedical microdevice design.

High-performance soft–hard interfaces are inherently difficult to fabricate due to the dissimilar mechanical properties of both materials, especially when connecting extremely soft biomaterials, such as hydrogels, to much harder biomaterials, such as rigid polymers. Nevertheless, there is significant clinical demand for synthetic soft–hard interfaces. Here, soft–hard interface geometries are proposed, designed with the aid of computational analyses and fabricated as 3D-printed hydrogel-to-polylactide (PLA) structures. Two primary interlocking geometries (i.e., anti-trapezoidal (AT) and double-hook (DH)) are used to study the envelope of 2.5D geometric interlocking designs, fabricated through hybrid 3D printing, combining pneumatic extrusion with fused deposition modeling. Finite-element analysis, uniaxial tensile tests, and digital image correlation (DIC) are used to characterize the geometries and identify parameters that significantly influence their mechanical performance. These findings reveal significant differences between geometric designs, where DH geometries performed significantly better than AT geometries, exhibiting a 190% increase in the maximum force, F_max, and a 340% increase in the fracture toughness, W. Compared to the control groups (i.e., flat, inset, and 90° interfaces), F_max and W values increased by 500%–990% and 350%–1200%, respectively. The findings of this study can serve as a guideline for the design and fabrication of efficient soft–hard interfaces with performances close to predicted values.

The Poisson's ratio and elastic modulus are two parameters determining the elastic behavior of biomaterials. While the effects of elastic modulus on the cell response is widely studied, very little is known regarding the effects of the Poisson's ratio. The micro-architecture of meta-biomaterials determines not only the Poisson's ratio but also several other parameters that also influence cell response, such as porosity, pore size, and effective elastic modulus. It is, therefore, very challenging to isolate the effects of the Poisson's ratio from those of other micro-architectural parameters. Here, we computationally design meta-biomaterials with controlled Poisson's ratios, ranging between -0.74 and +0.74, while maintaining consistent porosity, pore size, and effective elastic modulus. The 3D meta-biomaterials were additively manufactured at the micro-scale using two-photon polymerization (2PP), and were mechanically evaluated at the meso‑scale. The response of murine preosteoblasts to these meta-biomaterials was then studied using in vitro cell culture models. Meta-biomaterials with positive Poisson's ratios resulted in higher metabolic activity than those with negative values. The cells could attach and infiltrate all meta-biomaterials from the bottom to the top, fully covering the scaffolds after 17 days of culture. Interestingly, the meta-biomaterials exhibited different cell-induced deformations (e.g., shrinkage or local bending) as observed via scanning electron microscopy. The outcomes of osteogenic differentiation (i.e., Runx2 immunofluorescent staining) and matrix mineralization (i.e., Alizarin red staining) assays indicated the significant potential impact of these meta-biomaterials in the field of bone tissue engineering, paving the way for the development of advanced bone meta-implants. Statement of significance: We studied the influence of Poisson's ratio on bone cell response in meta-biomaterials. While elastic modulus effects are well-studied, the impact of Poisson's ratio, especially negative values found in architected biomaterials, remains largely unexplored. The complexity arises from intertwined micro-architectural parameters, such as porosity and elastic modulus, making it challenging to isolate the Poisson's ratio. To overcome this limitation, this study employed rational computational design to create meta-biomaterials with controlled Poisson's ratios, alongside consistent effective elastic modulus, porosity, and pore size. The study reveals that two-photon polymerized 3D meta-biomaterials with positive Poisson's ratios displayed higher metabolic activity, while all the developed meta-biomaterials supported osteogenic differentiation of preosteoblasts as well as matrix mineralization. The outcomes pave the way for the development of advanced 3D bone tissue models and meta-implants.

The currently available treatments for inner ear disorders often involve systemic drug administration, leading to suboptimal drug concentrations and side effects. Cochlear implants offer a potential solution by providing localized and sustained drug delivery to the cochlea. While the mechanical characterization of both the implants and their constituent material is crucial to ensure functional performance and structural integrity during implantation, this aspect has been mostly overlooked. This study proposes a novel methodology for the mechanical characterization of our recently developed cochlear implant design, namely, rectangular and cylindrical, fabricated using two-photon polymerization (2 PP) with a novel photosensitive resin (IP-Q™). We used in silico computational models and ex silico experiments to study the mechanics of our newly designed implants when subjected to torsion mimicking the foreseeable implantation procedure. Torsion testing on the actual-sized implants was not feasible due to their small size (0.6 × 0.6 × 2.4 mm³). Therefore, scaled-up rectangular cochlear implants (5 × 5 × 20 mm³, 10 × 10 × 40 mm³, and 20 × 20 × 80 mm³) were fabricated using stereolithography and subjected to torsion testing. Finite element analysis (FEA) accurately represented the linear behavior observed in the torsion experiments. We then used the validated Finite element analysis models to study the mechanical behavior of real-sized implants fabricated from the IP-Q resin. Mechanical characterization of both implant designs, with different inner porous structures (pore size: 20 μm and 60 μm) and a hollow version, revealed that the cylindrical implants exhibited approximately three times higher stiffness and mechanical strength as compared to the rectangular ones. The influence of the pore sizes on the mechanical behavior of these implant designs was found to be small. Based on these findings, the cylindrical design, regardless of the pore size, is recommended for further research and development efforts.

The existing 3D printing methods exhibit certain fabrication-dependent limitations for printing curved constructs that are relevant for many tissues. Four-dimensional (4D) printing is an emerging technology that is expected to revolutionize the field of tissue engineering and regenerative medicine (TERM). 4D printing is based on 3D printing, featuring the introduction of time as the fourth dimension, in which there is a transition from a 3D printed scaffold to a new, distinct, and stable state, upon the application of one or more stimuli. Here, we present an overview of the current developments of the 4D printing technology for TERM, with a focus on approaches to achieve temporal changes of the shape of the printed constructs that would enable biofabrication of highly complex structures. To this aim, the printing methods, types of stimuli, shape-shifting mechanisms, and cell-incorporation strategies are critically reviewed. Furthermore, the challenges of this very recent biofabrication technology as well as the future research directions are discussed. Our findings show that the most common printing methods so far are stereolithography (SLA) and extrusion bioprinting, followed by fused deposition modelling, while the shape-shifting mechanisms used for TERM applications are shape-memory and differential swelling for 4D printing and 4D bioprinting, respectively. For shape-memory mechanism, there is a high prevalence of synthetic materials, such as polylactic acid (PLA), poly(glycerol dodecanoate) acrylate (PGDA), or polyurethanes. On the other hand, different acrylate combinations of alginate, hyaluronan, or gelatin have been used for differential swelling-based 4D transformations. TERM applications include bone, vascular, and cardiac tissues as the main target of the 4D (bio)printing technology. The field has great potential for further development by considering the combination of multiple stimuli, the use of a wider range of 4D techniques, and the implementation of computational-assisted strategies.

This study proposes a new concept for an on-demand drug releasing device intended for integration into additively manufactured (i.e., 3D printed) orthopedic implants. The system comprises a surface with conduits connected to a subsurface reservoir used for storage and on-demand release of antimicrobial agents, covered with a cap that prevents the antibacterial agents from being released until alternating magnetic field (AMF) raises the temperature of the cap, thus, releasing the stored drug. To demonstrate this concept, Ti6Al4V specimens are directly 3D printed using selective laser melting and their surface, reservoirs, and drug releasing properties are characterized. A new synthetic antimicrobial peptide, SAAP-148, is thereafter tested for its cytotoxic, osteogenic, and immunomodulatory effects at concentrations relevant for its minimal bactericidal concentration (MBC) and is compared with its natural analogue, LL-37. The results showed that AMF successfully activated the release from the 3D printed loaded samples. Both peptides demonstrated to be non-cytotoxic within the MBC levels for macrophages and preosteoblasts and did not influence their osteoimmunomodulatory behavior. The findings of this study indicate that the proposed concept is technically feasible and has the potential to be used for the development of bone implants with on-demand delivery systems to fight IAI without systemic or continuous local release of antibiotics.

The antibacterial biofunctionality of bone implants is essential for the prevention and treatment of implant-associated infections (IAI). In vitro co-culture models are utilized to assess this and study bacteria-host cell interactions at the implant interface, aiding our understanding of biomaterial and the immune response against IAI without impeding the peri-implant bone tissue regeneration. This paper reviews existing co-culture models together with their characteristics, results, and clinical relevance. A total of 36 studies were found involving in vitro co-culture models between bacteria and osteogenic or immune cells at the interface with orthopedic antibacterial biomaterials. Most studies (∼67%) involved co-culture models of osteogenic cells and bacteria (osteo-bac), while 33% were co-culture models of immune cells and bacterial cells (im-bac). All models involve direct co-culture of two different cell types. The cell seeding sequence (simultaneous, bacteria-first, and cell-first) was used to mimic clinically relevant conditions and showed the greatest effect on the outcome for both types of co-culture models. The im-bac models are considered more relevant for early peri-implant infections, whereas the osteo-bac models suit late infections. The limitations of the current models and future directions to develop more relevant co-culture models to address specific research questions are also discussed.

Macrophage responses following the implantation of orthopaedic implants are essential for successful implant integration in the body, partly through intimate crosstalk with human marrow stromal cells (hMSCs) in the process of new bone formation. Additive manufacturing (AM) and plasma electrolytic oxidation (PEO) in the presence of silver nanoparticles (AgNPs) are promising techniques to achieve multifunctional titanium implants. Their osteoimmunomodulatory properties are, however, not yet fully investigated. Here, we studied the effects of implants with AgNPs on human macrophages and the crosstalk between hMSCs and human macrophages when co-cultured in vitro with biofunctionalised AM Ti6Al4V implants. A concentration of 0.3 g/L AgNPs in the PEO electrolyte was found to be optimal for both macrophage viability and inhibition of bacteria growth. These specimens also caused a decrease of the macrophage tissue repair related factor C-C Motif Chemokine Ligand 18 (CCL18). Nevertheless, co-cultured hMSCs could osteogenically differentiate without any adverse effects caused by the presence of macrophages that were previously exposed to the PEO (±AgNPs) surfaces. Further evaluation of these promising implants in a bony in vivo environment with and without infection is highly recommended to prove their potential for clinical use.

Individual cells and multicellular systems respond to cell-scale curvatures in their environments, guiding migration, orientation, and tissue formation. However, it remains largely unclear how cells collectively explore and pattern complex landscapes with curvature gradients across the Euclidean and non-Euclidean spectra. Here, we show that mathematically designed substrates with controlled curvature variations induce multicellular spatiotemporal organization of preosteoblasts. We quantify curvature-induced patterning and find that cells generally prefer regions with at least one negative principal curvature. However, we also show that the developing tissue can eventually cover unfavorably curved territories, can bridge large portions of the substrates, and is often characterized by collectively aligned stress fibers. We demonstrate that this is partly regulated by cellular contractility and extracellular matrix development, underscoring the mechanical nature of curvature guidance. Our findings offer a geometric perspective on cell-environment interactions that could be harnessed in tissue engineering and regenerative medicine applications.

Hearing loss is a highly prevalent multifactorial disorder affecting 20% of the global population. Current treatments using the systemic administration of drugs are therapeutically ineffective due to the anatomy of the cochlea and the existing blood–labyrinth barrier. Local drug delivery systems can ensure therapeutic drug concentrations locally while preventing adverse effects caused by high dosages of systemically administered drugs. Here, we aimed to design, fabricate, and characterize a local drug delivery system for the human cochlea. The design was relevant to the size of the human ear, included two different shapes, and incorporated two different microporous structures acting as reservoirs for drug loading and release. The four cochlear implant designs were printed using the two-photon polymerization (2PP) technique and the IP-Q photoresist. The optimized 2PP process enabled the fabrication of the cochlear implants with great reproducibility and shape fidelity. Rectangular and cylindrical implants featuring cylindrical and tapered tips, respectively, were successfully printed. Their outer dimensions were 0.6 × 0.6 × 2.4 mm3 (L × W × H). They incorporated internal porous networks that were printed with high accuracy, yielding pore sizes of 17.88 ± 0.95 μm and 58.15 ± 1.62 μm for the designed values of 20 μm and 60 μm, respectively. The average surface roughness was 1.67 ± 0.24 μm, and the water contact angle was 72.3 ± 3.0°. A high degree of polymerization (~90%) of the IP-Q was identified after printing, and the printed material was cytocompatible with murine macrophages. The cochlear implants designed and 3D printed in this study, featuring relevant sizes for the human ear and tunable internal microporosity, represent a novel approach for personalized treatment of hearing loss through local drug delivery.