Abstract. Objectives. Additive manufacturing has led to numerous innovations in orthopaedic surgery: surgical guides; surface coatings/textures; and custom implants. Most contemporary implants are made from titanium alloy (Ti-6Al-4V). Despite being widely available industrially and clinically, there is little published information on the performance of this 3D printed material for orthopaedic devices with respect to regulatory approval. The aim of this study was to document the mechanical, chemical and biological properties of selective laser sintering (SLS) manufactured specimens following medical device (TOKA®, 3D Metal Printing LTD, UK) submission and review by the UK Medicines and Healthcare Products Regulatory Agency (MHRA). Methods. All specimens were additively manufactured in Ti-6Al-4V ELI (Renishaw plc, UK). Mechanical tests were performed according to ISO6892-1, ISO9585 and ISO12107 for tensile (n=10), bending (n=3) and fatigue (n=16) respectively (University of Bath, UK). Appropriate chemical characterisation and biological tests were selected according to recommendations in ISO10993 and conducted by external laboratories (Wickham Labs, UK; Lucideon, UK; Edwards Analytical, UK) in adherence with Good Lab Practise guidelines. A toxicological review was conducted on the findings (Bibra, UK). Results. The mechanical tests demonstrated that the material performed to the specification for conventionally manufactured titanium alloy of this type (ISO5832-3). The toxicology review concluded that there were no significant concerns for the health of the patients identified in this evaluation and implantation of the TOKA® device would not result in a significant health risk to patients. Conclusions. Reflecting on our MHRA experience, additive manufacture of orthopaedic devices is still considered to be a ‘novel’ process by regulatory bodies, requiring additional safety evidence. Despite this, our findings demonstrate that there is no difference, mechanically or chemically, to the traditionally manufactured alloy material. We hope to support the widening use of 3D printed titanium alloy orthopaedic devices by publishing our route to regulatory approval. Declaration of Interest. (a) fully declare any financial or other potential conflict of interest

The current gold standard bone substitute is still autologous bone, despite the fact that its harvest demands for a second operation site, causes additional pain, discomfort, potential destruction of the grafting site, and is limited in supply. Since newly developed clinical approaches like transplantation of cells are invasive and costly, and osteoinduction by bone morphogenetic proteins is expensive and is associated with mild to severe side effects, the optimization of osteoconduction appears as promising option to realize bone substitute-based bone tissue engineering. In the 90ties of the last century, the holy grail of pore size for scaffolds in bone tissue engineering was set between 0.3 and 0.5 mm. More recent, papers from others and us indicated that the optimal microarchitecture for bone tissue engineering scaffolds in terms of pore size, constrictions, rod thickness, or rod distance is still unknown. Additive manufacturing appears as an ideal tool to study those diverse microarchitecture options since it can generate scaffolds where size and location of pores and connections between pores can be tested. For the production of our test scaffolds, we applied laser sintering of titanium and lithography-based additive manufacturing of ceramics. Histomorphometry of calvarial defects in rabbits revealed that bone formation was significantly increased by scaffolds with pore diameters in the range of 0.7–1.2 mm. Scaffolds with pores of 1.5 and 1.7 mm perform significantly worse. Therefore, pore diameters in osteoconductive bone substitutes should be 1.0–1.2 mm and thus much bigger than previously suggested. In essence, osteoconductive microarchitectures of degradable bone substitutes can be realized by lithography based additive manufacturing and this methodology appears as a promising tool for the production of personalized bone tissue engineering scaffolds to be used in cranio-maxillofacial surgery, dentistry, and orthopedics

Medial opening wedge high tibial osteotomy (MOWHTO) is the workhorse procedure for correcting varus malalignment of the knee. There have been recent developments in the synthetic options to fill the osteotomy gap. The current gold standard for filling this osteotomy gap is autologous bone graft which is associated with donor site morbidity. We would like to introduce and describe the process of utilizing the novel Osteopore® 3D printed, honeycomb structured, Polycaprolactone and β-Tricalcium Phosphate wedge for filling the gap in MOWHTO. In the advent of additive manufacturing and the quest for more biocompatible materials, the usage of the Osteopore® bone wedge in MOWHTO is a promising technique that may improve the biomechanical stability as well the healing of the osteotomy gap

Critical-sized bone defects remain challenging in the clinical setting. Autologous bone grafting remains preferred by clinicians. However, the use of autologous tissue is associated with donor-site morbidity and limited accessibility to the graft tissue. Advances in the development of synthetic bone substitutes focus on improving their osteoinductive properties. Whereas osteoinductivity has been demonstrated with ceramics, it is still a challenge in case of polymeric composites. One of the approaches to improve the regenerative properties of biomaterials, without changing their synthetic character, is the addition of inorganic ions with known osteogenic and angiogenic properties. We have previously reported that the use of a bioactive composite with high ceramic content composed of poly(ethyleneoxide terephthalate)/poly(butylene terephthalate) (1000PEOT70PBT30, PolyActive, PA) and 50% beta-tricalcium phosphate (β-TCP) with the addition of zinc in a form of a coating of the TCP particles can enhance the osteogenic differentiation of human mesenchymal stromal cells (hMSCs) (3). To further support the regenerative properties of these scaffolds, inorganic ions with known angiogenic properties, copper or cobalt, were added to the coating solution. β-TCP particles were immersed in a zinc and copper or zinc and cobalt solution with a concentration of 15 or 45 mM. 3D porous scaffolds composed of 1000PEOT70PBT30 and pure or coated β-TCP were additively manufactured by 3D fibre deposition. The osteogenic and angiogenic properties of the fabricated scaffolds were tested in vitro through culture with hMSCs and human umbilical vein endothelial cells, respectively. The materials were further evaluated through ectopic implantation in an in vivo mini-pig model. The early expression of relevant osteogenic gene markers (collagen-1, osteocalcin) of hMSCs was upregulated in the presence of lower concentration of inorganic ions. Further analysis will focus on the evaluation of ectopic bone formation and vascularisation of these scaffolds after implantation in a mini-pig ectopic intramuscular model

Developments in the field of additive manufacturing have allowed significant improvements in the design and production of scaffolds with biologically relevant features to treat bone defects. Unfortunately, the workflow to generate personalized scaffolds is source of inaccuracies leading to a poor fit between the implant and patients' bone defects. In addition, scaffolds are often brittle and fragile, uneasing their handling by surgeons, with significant risks of fracture during their insertion in the defect. Consequently, we developed organo-mineral cementitious scaffolds displaying evolutive mechanical properties which are currently being evaluated to treat maxillofacial bone deformities in veterinary clinics. Treatment of dog patients was approved by ethic and welfare committees (CERVO-2022-14-V). To date, 8 puppies with cleft palate/lip deformities received the following treatment. Two weeks prior surgery, CT-scan of patient's skull was performed to allow for surgical planning and scaffold designing. Organo-mineral printable pastes were formulated by mixing an inorganic cement precursor (α-Ca3(PO4)2) to a self-reticulating hydrogel (silanized hyaluronic acid) supplemented with a viscosifier (hydroxymethylpropylcellulose). Scaffolds were produced by robocasting of these pastes. Surgical interventions included the reconstruction of soft tissues, and the insertion of the scaffold soaked with autologous bone marrow. Bone formation was monitored 3 and 6 months after reconstruction, and a biopsy at 6 months was performed for more detailed analyses. Scaffolds displayed great handling properties and were inserted within bone defects without significant issue with a relevant bone edges/scaffold contact. Osteointegration of the scaffolds was observed after 3 months, and regeneration of the defect at 6 months seemed quite promising. Preliminary results have demonstrated a potential of the set-up strategy to treat cleft lip/palate deformities in real, spontaneous clinical setting. Translation of these innovative scaffolds to orthopedics is planned for a near future

Introduction. Kienböck's disease is generally defined as the collapse of the lunate bone, and this may lead to early wrist osteoarthritis. Replacing the collapsed lunate with an implant has regained renewed interest with the advancing technology of additive manufacturing, enabling the design of patient-specific implants. The aims of this project are (1) to determine how accurate it is to use the contralateral lunate shape as a template for patient-specific lunate implants, and (2) to study the effects of shape variations wrist kinematics using 4D-computed tomography (CT) scanning. Methods. A 3D statistical shape model (SSM) of the lunate was built based on bilateral CT scans of 54 individuals. Using SMM, shape variations of the lunate were identified and the intra- and inter-subject shape variations were compared by performing an intraclass correlation analysis. A radiolucent motor-controlled wrist-holder was designed to guide flexion/extension and radial/ulnar deviation of ex vivo wrist specimens under 4D-CT scanning. In this pilot, three shape mode variations were tested per specimen in two specimens were. After post-processing each CT, the scapholunate angle (SLA) and capitolunate angle (CLA) were measured. Results. The shape of the lunate was not symmetrical, defined as exceeding the intra-subject variation in five different shape modes. The FE tests show a generalized increase in scapholunate and capitolunate angle when using lunate implants, and comparing variation of shape modes showed that shape mode 3 has a significant effect on the measured angles (p<0.05). Discussion. The design of patient-specific lunate implants may prove to be challenging using a ‘mirror’-design as it will lead to a degree of shape asymmetry. The pilot study, to determine the effects of those shape variations on wrist kinematics suggest that the degree of shape variation observed indeed may alter the wrist kinematics, although this needs to be further investigated in study using more specimens

Introduction. The surgical treatment of critical-sized bone defects with complex three-dimensional (3D) geometries is a challenge for the treating surgeon. Additive manufacturing such as 3D printing enables the production of highly individualized bone implants meeting the shape of the patient's bone defect and including a tunable internal structure. In this study, we showcase the design process for patient-specific implants with critical-sized tibia defects. Methods. Two clinical cases of patients with critical tibia defects (size 63×20×21 mm and 50×24×17 mm) were chosen. Brainlab software was used for segmentation of CT data generating 3D models of the defects. The implant construction involves multiple stages. Initially, the outer shell is precisely defined. Subsequently, the specified volume is populated with internal structures using Voronoi, Gyroid, and NaCl crystal structures. Variation in pore size (1.6 mm and 1.0 mm) was accomplished by adjusting scaffold size and material thickness. Results. An algorithmic design process in Rhino and Grasshopper was successfully applied to generate model implants for the tibia from Ct data. By integrating a precise mesh into an outer shell, a scaffold with controlled porosity was designed. In terms of the internal design, both Voronoi and Gyroid form macroscopically homogeneous properties, while NaCl, exhibits irregularities in density and consequently, in the strength of the structure. Data implied that Voronoi and Gyroid structures adapt more precisely to complex and irregular outer shapes of the implants. Conclusion. In proof-of-principle studies customized tibia implants were successfully generated and printed as model implants based on resin. Further studies will include more patient data sets to refine the workflows and digital tools for a broader spectrum of bone defects. The algorithm-based design might offer a tremendous potential in terms of an automated design process for 3D printed implants which is essential for clinical application

Critical-sized bone defects can result from trauma, inflammation, and tumor resection. Such bone defects, often have irregular shapes, resulting in the need for new technologies to produce suitable implants. Bioprinting is an additive manufacturing method to create complex and individualised bone constructs, which can already include vital cells. In this study, we established an extrusion-based printing technology to produce osteoinductive scaffolds based on polycaprolactone (PCL) combined with calcium phosphate, which is known to induce osteogenic differentiation of stem cells. The model was created in python based on the signed distance functions. The shape of the 3D model is a ring with a diameter of 20 mm and a height of 10 mm with a spongiosa-like structure. The interconnected irregular pores have a diameter of 2 mm +/− 0.2 mm standard deviation. Extrusion-based printing was performed using the BIO X6. To produce the bioink, PCL (80 kDa) was combined with calcium phosphate nanopowder (> 150 nm particle size) under heating. After printing, 5 × 10. 6. hMSC were seeded on the construct using a rotating incubator. We were able to print a highly accurate ring construct with an interconnected pore structure. The PCL combined with calcium phosphate particles resulted in a precise printed construct, which corresponded to the 3D model. The bioink containing calcium phosphate nanoparticles had a higher printing accuracy compared to PCL alone. We found that hMSC cultured on the construct settled in close proximity to the calcium phosphate particles. The hMSC were vital for 22 days on the construct as demonstrated by life/dead staining. The extrusion printing technology enables to print a mechanically stable construct with a spongiosa-like structure. The porous PCL ring could serve as an outer matrix for implants, providing the construct the stability of natural bone. To extend this technology and to improve the implant properties, a biologised inner structure will be integrated into the scaffold in the future

The success of cementless orthopaedic implants relies on bony ingrowth and active bone remodelling. Much research effort is invested to develop implants with controllable surface roughness and internal porous architectures that encourage these biological processes. Evaluation of these implants requires long-term and costly animal studies, which do not always yield the desired outcome requiring iteration. The aim of our study is to develop a cost-effective method to prescreen design parameters prior to animal trials to streamline implant development and reduce live animal testing burden. Ex vivo porcine cancellous bone cylinders (n=6, Ø20×12mm) were extracted from porcine knee joints with a computer-numerically-controlled milling machine under sterile conditions within 4 hours of animal sacrifice. The bone discs were implanted with Ø6×12mm additive manufactured porous titanium implants and were then cultured for 21days. Half underwent static culture in medium (DMEM, 10% FBS, 1% antibiotics) at 37°C and 5% CO. 2. The rest were cultured in novel high-throughput stacked configuration in a bioreactor that simulated physiological conditions after surgery: the fluid flow and cyclic compression force were set at 10ml/min and 10–150 N (1Hz,5000 cycles/day) respectively. Stains were administered at days 7 and 14. Samples were evaluated with widefield microscopy, scanning electron microscopy (SEM) and with histology. More bone remodelling was observed on the samples cultured within the bioreactor: widefield imaging showed more remodelling at the boundaries between the implant-bone interface, while SEM revealed immature bone tissue integration within the pores of the implant. Histological analysis confirmed these results, with many more trabecular struts with new osteoid formation on the samples cultured dynamically compared to static ones. Ex vivo bone can be used to analyse new implant technologies with lower cost and ethical impact than animal trial. Physiological conditions (load and fluid flow) promoted bone ingrowth and remodelling

Although 3D-printed porous dental implants may possess improved osseointegration potential, they must exhibit appropriate fatigue strength. Finite element analysis (FEA) has the potential to predict the fatigue life of implants and accelerate their development. This work aimed at developing and validating an FEA-based tool to predict the fatigue behavior of porous dental implants. Test samples mimicking dental implants were designed as 4.5 mm-diameter cylinders with a fully porous section around bone level. Three porosity levels (50%, 60% and 70%) and two unit cell types (Schwarz Primitive (SP) and Schwarz W (SW)) were combined to generate six designs that were split between calibration (60SP, 70SP, 60SW, 70SW) and validation (50SP, 50SW) sets. Twenty-eight samples per design were additively manufactured from titanium powder (Ti6Al4V). The samples were tested under bending compression loading (ISO 14801) monotonically (N=4/design) to determine ultimate load (F. ult. ) (Instron 5866) and cyclically at six load levels between 50% and 10% of F. ult. (N=4/design/load level) (DYNA5dent). Failure force results were fitted to F/F. ult. = a(N. f. ). b. (Eq1) with N. f. being the number of cycles to failure, to identify parameters a and b. The endurance limit (F. e. ) was evaluated at N. f. = 5M cycles. Finite element models were built to predict the yield load (F. yield. ) of each design. Combining a linear correlation between FEA-based F. yield. and experimental F. ult. with equation Eq1 enabled FEA-based prediction of F. e. . For all designs, F. e. was comprised between 10% (all four samples surviving) and 15% (at least one failure) of F. ult. The FEA-based tool predicted F. e. values of 11.7% and 12.0% of F. ult. for the validation sets of 50SP and 50SW, respectively. Thus, the developed FEA-based workflow could accurately predict endurance limit for different implant designs and therefore could be used in future to aid the development of novel porous implants. Acknowledgements: This study was funded by EU's Horizon 2020 grant No. 953128 (I-SMarD). We gratefully acknowledge the expert advice of Prof. Philippe Zysset

The growing number of non-union fractures in an aging population has increased the clinical demand for tissue-engineered bone. Electrical stimulation (ES) has been described as a promising strategy for bone regeneration treatments in several clinical studies. However the underlying mechanism by which ES augments bone formation is still poorly understood and its use in bone tissue engineering (BTE) strategies is currently underexplored. Additive manufacturing (AM) technologies (Fused Deposition Modeling/3D Printing) have been widely used in BTE due to their ability to fabricate scaffolds with a high control over their structural and mechanical properties in a reproducible and scalable manner. Thus, in this work, we combined AM methods with conductive biomaterials and ES to enhance the osteogenic differentiation of human bone marrow-derived mesenchymal stem/stromal cells (hBMSCs) envisaging improved BTE strategies. First, we started by developing AM-based electro-bioreactor devices containing medical-grade electrodes (stainless steel and Ti6Al4V) to apply ES to monolayer 2D cultures and 3D cell-seeded scaffolds. Computer modeling(Finite Element Analysis-FEA) was employed to predict the magnitude/distribution of electrical fields within the ES devices and along the different conductive scaffolds. Prior to scaffold culture, 5 different ES protocols were tested in terms of their ability to promote hBMSCs proliferation and osteogenic differentiation in 2D cultures. The best performance ES protocol was then used in two different AM-based BTE strategies: 1) Two different conductive scaffolds (conductive poly lactic acid (PLA) and titanium) were seeded with hBMSCs and cultured for 21 days under osteogenic medium conditions with and without ES and their biological performance was evaluated in comparison to non-conductive standard PLA scaffolds; 2) Different PEDOT:PSS-based coating solutions were screened to obtain PEDOT:PSS/Gelatin-coated 3D polycaprolactone (PCL) scaffolds with a high(11 S.cm. -1. ) and stable electroconductivity. When cultured under ES, PEDOT:PSS/Gelatin-PCL scaffolds enhanced significantly hBMSCs osteogenic differentiation and mineralization(calcium deposition), highlighting their potential for BTE applications. Acknowledgements: Funding received from FCT through projects InSilico4OCReg (PTDC/EME-SIS/0838/2021), OptiBioScaffold (PTDC/EME-SIS/4446/2020) and BioMaterARISES (EXPL/CTM-CTM/0995/2021), and to the institutions iBB (UIDB/04565/2020), CDRSP (UIDB/04044/2020) and Associate Laboratory i4HB (LA/P/0140/2020)

Among the advanced technology developed and tested for orthopaedic surgery, the Rizzoli (IOR) has a long experience on custom-made design and implant of devices for joint and bone replacements. This follows the recent advancements in additive manufacturing, which now allows to obtain products also in metal alloy by deposition of material layer-by-layer according to a digital model. The process starts from medical image, goes through anatomical modelling, prosthesis design, prototyping, and final production in 3D printers and in case post-production. These devices have demonstrated already to be accurate enough to address properly the specific needs and conditions of the patient and of his/her physician. These guarantee also minimum removal of the tissues, partial replacements, no size related issues, minimal invasiveness, limited instrumentation. The thorough preparation of the treatment results also in a considerable shortening of the surgical and of recovery time. The necessary additional efforts and costs of custom-made implants seem to be well balanced by these advantages and savings, which shall include the lower failures and revision surgery rates. This also allows thoughtful optimization of the component-to-bone interfaces, by advanced lattice structures, with topologies mimicking the trabecular bone, possibly to promote osteointegration and to prevent infection. IOR's experience comprises all sub-disciplines and anatomical areas, here mentioned in historical order. Originally, several systems of Patient-Specific instrumentation have been exploited in total knee and total ankle replacements. A few massive osteoarticular reconstructions in the shank and foot for severe bone fractures were performed, starting from mirroring the contralateral area. Something very similar was performed also for pelvic surgery in the Oncology department, where massive skeletal reconstructions for bone tumours are necessary. To this aim, in addition to the standard anatomical modelling, prosthesis design, technical/technological refinements, and manufacturing, surgical guides for the correct execution of the osteotomies are also designed and 3D printed. Another original experience is about en-block replacement of vertebral bodies for severe bone loss, in particular for tumours. In this project, technological and biological aspects have also been addressed, to enhance osteointegration and to diminish the risk of infection. In our series there is also a case of successful custom reconstruction of the anterior chest wall. Initial experiences are in progress also for shoulder and elbow surgery, in particular for pre-op planning and surgical guide design in complex re-alignment osteotomies for severe bone deformities. Also in complex flat-foot deformities, in preparation of surgical corrections, 3D digital reconstruction and 3D printing in cheap ABS filaments have been valuable, for indication, planning of surgery and patient communication; with special materials mimicking bone strength, these 3D physical models are precious also for training and preparation of the surgery. In Paediatric surgery severe multi planar & multifocal deformities in children are addressed with personalized pre-op planning and custom cutting-guides for the necessary osteotomies, most of which require custom allografts. A number of complex hip revision surgeries have been performed, where 3D reconstruction for possible final solutions with exact implants on the remaining bone were developed. Elective surgery has been addressed as well, in particular the customization of an original total ankle replacement designed at IOR. Also a novel system with a high-tibial-osteotomy, including a custom cutting jig and the fixation plate was tested. An initial experience for the design and test of custom ankle & foot orthotics is also in progress, starting with 3D surface scanning of the shank and foot including the plantar aspect. Clearly, for achieving these results, multi-disciplinary teams have been formed, including physicians, radiologists, bioengineers and technologists, working together for the same goal

Abstract. INTRODUCTION. Polyetheretherketone (PEEK) is a high-performance thermoplastic polymer which has found increasing application in orthopaedic implant devices and has a lot of promise for ‘made-to-measure’ implants produced through additive manufacturing [1]. However, a key limitation of PEEK is that it is bioinert and there is a requirement to functionalise its surface to make the material osteoconductive to ensure a more rapid, improved and stable fixation, in vivo. One approach to solving this issue is to modify PEEK with bioactive materials, such as hydroxyapatite (HA). OBJECTIVE. To 3D PEEK/HA composite materials using a Fused Filament Fabrication (FFF) approach to enhance the properties of the PEEK matrix. METHODS. PEEK/HA composites (0–30% w/w HA/PEEK) were 3D printed using a modified Ultimaker 2+ 3D printer. The mechanical, thermal, physical, chemical and in vitro properties of the 3D printed samples were all studied as part of this work. RESULTS. The CT images of both the filament and the 3D printed samples showed that the HA material was evenly dispersed throughout the bulk all the samples. SEM/EDX measurements highlighted that HA was homogenously distributed across the surface. As the HA content of the samples increases, so does the tensile modulus, ranging from 4.2 GPa (PEEK) to 6.1 GPa (30% HA/PEEK) and are significantly higher than datasheet information of injected molded PEEK samples. All materials supported the growth of osteoblast cells on their surface. CONCLUSIONS. The results clearly show that we can successfully and easily 3D print HA/PEEK composite materials up to 30% w/w HA/PEEK. The samples produced have a homogeneous distribution of HA in both the bulk and surface of all the samples, and their mechanical performance of the PEEK is enhanced by the addition of HA

The management of maxillofacial injuries requires restoring the contours of the facial skeleton to achieve an aesthetic outcome. When fractures are simple, open reduction and rigid fixation with stock titanium osteosynthesis plates is usually sufficient. However, when the damage is more substantial (when the fracture is comminuted or in case of a bone defect) anatomical landmarks are lost and the reconstruction requires the use of titanium meshes. These meshes are usually modelled intraoperatively to restore the contours of the bone. This can be a tough and time consuming task in case of minimal invasive approach and intraoperative edema. When the injury is unilateral, printing a 3D anatomical model of the mirrored unaffected side is an easy way to accurately pre-bend the mesh preoperatively. With the emergence of “low cost” consumer 3D printers, the aim of our study was to evaluate the cost of this technique in a department of maxillofacial surgery. The first part of the study was to evaluate free software solutions available online to determine which of these could be used to create 3D virtual models from the patients' volume imaging data, mirror the model and export an STL file suitable for 3D-printing with a consumer 3D-printer. The second part was to identify the desktop 3D-printers commercially available according to the different technology used, their prices and that of consumables required. Five free software solutions were identified to create STL meshes of the patient's anatomy from thin slice CT scan DICOM data. Two more were available to repair, segment and mirror them to provide a clean STL file suitable for 3D printing with a desktop 3D printer. The prices of 2 different printers were then listed for each of the 3 additive manufacturing technologies available to date. Prices ranged from 2,299 € for the Ultimaker 2+© (Fuse Deposition Modeling, FDM), to 4,999 € for the Sintratec© printer (Selective Laser Sintering, SLS), the Formlabs 2© (stereolithography) being at an intermediate price of 3,299 €. Finally, the cost of the manufacture of a model was calculated for each of these printers. Considering a model of a supraorbital ridge printed to restore the anterior wall of the frontal sinus, the volume of the mesh is around 20 cm. 3. This represents a cost of less than 1 € with the FDM technology, 4.70 € with stereolithography and 1.50 € with the SLS printer. Since patents of additive manufacturing have become part of the public domain, the cost of 3D printing technology has fallen drastically. Desktop printers are now an investment accessible to a surgery department and the cost of the material is low. This allows the surgeons, by the mean of free software, to directly create 3D models of their patients' anatomy, mirror them if needed and manufacture a template to pre-bend titanium meshes that will be subsequently sterilized for the surgery. Having the printer in the department reduces manufacturing lead times and makes this technique possible even for urgent cases

The extracellular matrix (ECM) is the non-cellular structural support that provides cells with a network of biochemical and biomechanical factors for cellular processes. The ECM regulates cell function, differentiation and homeostasis. Here, we present a proteomics characterization of three commonly used additive manufactured polymers: polylactic acid (PLA), polyactive (PEOT/PBT) and polycaprolactone (PCL). We cultured human mesenchymal stromal cells (hMSCs) and make them undergo chondrogenic and osteogenic differentiation on 3D printed PCL, PEOT/PBT and PLA scaffolds. hMSCs were cultured in basal, chondrogenic and osteogenic media (200000 cells/scaffold) and analyzed after 35 days of culture. Differentiation was proved through biochemical assays, immunofluorescence and histology. The protein content was explored using label free liquid chromatography mass spectrometry (LC-MS), which revealed upregulated proteins and their related pathways. A higher difference was found among different media compared to the scaffold type through principal component analysis (PCA). Interestingly, in all three materials, chondrogenesis was characterized by a lower but more diverse amount of proteins. PCL induced ECM production in both differentiation media, but it led to more apoptosis and GAG degradation in the chondrogenic medium compared to the osteogenic one. During chondrogenesis in PEOT/PBT and PLA, cell differentiation resulted in the activation of stress response cascades, collagen formation and ECM remodelling. On the other hand, in osteogenesis, PCL enhanced insulin-like growth factor pathway and fibrin clot related pathways

Abstract. Objectives. Additive manufacturing (AM) enables fine control over the architecture of porous lattice structures, and the resulting mechanical performance. Orthopaedic implants may benefit from the tailored stiffness/elastic modulus of these AM biomaterials, as the stiffness can be made to closer match the properties of the replaced trabecular bone. Methods. This study used laser powder bed fusion (PBF) to create stochastic porous lattice structures in stainless steel (SS316L) and titanium alloy (Ti6Al4V), with modifications that aimed to overcome PBF manufacturing limitations of build angles. The structures were tested in uni-axial compression (n = 5) in 10 load orientations relative to the structure, including the three orthogonal axes. Results. The testing verified that no hidden peaks in elastic modulus existed in the stochastic structure. The standard deviation of the 10 elastic modulus values in the final structure decreased from 249 MPa to 101 MPa when made in SS316L and from 95.9 MPa to 52.5 MPa for Ti6Al4V, indicating the structures were more isotropic. Conclusions. These modified stochastic lattices have similar stiffness to cancellous bone and have controllable anisotropy, giving them the potential to be used within implants which match the stiffness of trabecular bone. Declaration of Interest. (b) declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported:I declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project

Abstract. Introduction. The long-term biological success of cementless orthopaedic prostheses is highly dependent on osteointegration. Pre-clinical testing of new cementless implant technology however, requires live animal testing, which has anatomical, loading, ethical and cost challenges. This proof-of-concept study aimed to develop an in vitro model to examine implant osteointegration under known loading/micromotion conditions. Methods. Fresh cancellous bone cylinders (n=8) were harvested from porcine femur and implanted with additive manufactured porous titanium implants (Ø4 × 15 mm). To simulate physiological conditions, n=3 bone cylinders were tested in a bioreactor system with a cyclic 30 µm displacement at 1Hz for 300 cycles every day for 15 days in a total of 21 days culture. The chamber was also perfused with culture medium using a peristaltic pump. Control bone cylinders were cultured under static conditions (n=5). Samples were calcein stained at day 7. Post-testing, bone cylinders were formalin fixed and bony ingrowth was measured via microscopy. Results. Viability of the freshly harvested ex vivo bone cylinders was maintained for up to 28 days. Two samples remain unanalysed due to COVID lockdown, one in each group. Similar to osteointegration seen in live animal models, evidence of bony ingrowth was seen more markedly at the bone-implant interface under dynamic conditions. This was evident by a greater intensity of calcein staining, confirming the deposition of new bone, at the bone-implant interface. In comparison, under static conditions, calcein staining was observed randomly all over the cylinder. Conclusion. This proof-of-concept study demonstrates that implant bony adaptation and ingrowth can be measured in vitro under known cyclic micromotion/loading conditions. This comparatively low cost, low ethical impact, controlled loading laboratory method has potential to accelerate the rate of implant development whilst conforming with the principles of NC3Rs. Declaration of Interest. (b) declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported:I declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research project

As compared to magnesium (Mg) and iron (Fe), solid zinc (Zn)-based absorbable implants show better degradation rates. An ideal bone substitute should provide sufficient mechanical support, but pure Zn itself is not strong enough for load-bearing medical applications. Modern processing techniques, like additive manufacturing (AM), can improve mechanical strength of Zn. To better mimic the in vivo situation in the human body, we evaluated the degradation behavior of porous Zn implants in vitro under dynamic conditions. Our study applied selective laser melting (SLM) to build topographically ordered absorbable Zn implants with superior mechanical properties. Specimens were fabricated from pure Zn powder using SLM and diamond unit cell topological design. In vitro degradation was performed under both static and dynamic conditions in a custom-built set-up under cell culture conditions (37 °C, 20% O2 and 5% CO2) for up to 28 days. Mechanical properties of the porous structures were determined according to ISO 13314: 2011 at different immersion time points. Modified ISO 10993 standards were used to evaluate biocompatibility through direct cell seeding and indirect extract-based cytotoxicity tests (MTS assay, Promega) against identically designed porous titanium (Ti-6Al-4V) specimens as reference material. Twenty-four hours after cell seeding, its efficacy was evaluated by Live-Dead staining (Abcam) and further analyzed using dual channel fluorescent optical imaging (FOI) and subsequent flow cytometric quantification. Porous Zn implants were successfully produced by means of SLM with a yield strength and Young's modulus in the range of 3.9–9.6 MPa and 265–570 MPa, respectively. Dynamic flow significantly increased the degradation rate of AM porous Zn after 28 days. Results from Zn extracts were similar to Ti-6Al-4V with >95% of cellular activity at all tested time points, confirming level 0 cytotoxicity (i.e., This study clearly shows the great potential of AM porous Zn as a bone substituting material. Moreover, we demonstrate that complex topological design permits control of mechanical properties and degradation behavior

Bioabsorbable metals hold a lot of potential as orthopaedic implant materials. Three metal families are currently being investigated: iron (Fe), magnesium (Mg) and zinc (Zn). Currently, however, biodegradation of such implants is poorly predictable. We thus used Direct Metal Printing to additively manufacture porous implants of a standardized bone-mimetic design and evaluated their mechanical properties and degradation behaviour, respectively, under in vivo-like conditions. Atomized powder was manufactured to porous implants of repetitive diamond unit cells, using a ProX DMP 320 (Layerwise, Belgium) or a custom-modified ReaLizer SLM50 metal printer. Degradation behaviour was characterized under static and dynamic conditions in a custom-built bioreactor system (37ºC, 5% CO. 2. and 20% O. 2. ) for up of 28 days. Implants were characterized by micro-CT before and after in vivo-like degradation. Mechanical characterization (according to ISO 13314: 2011) was performed on an Instron machine (10kN load cell) at different immersion times in simulated body fluid (r-SBF). Morphology and composition of degradation products were analysed (SEM, JSM-IT100, JEOL). Topographically identical titanium (Ti-6Al-4V, Ti64) specimen served as reference. Micro-CT analyses confirmed average strut sizes (420 ± 4 μm), and porosity (64%), to be close to design values. After 28 days of in vivo-like degradation, scaffolds were macroscopically covered by degradation products in an alloy-specific manner. Weight loss after cleaning also varied alloy-specifically, as did the change in pH value of the r-SBF. Corrosion time-dependent changes in Young's moduli from 1200 to 800 MPa for Mg, 1000 to 700 MPa for Zn and 48-8 MPa for iron were statistically significant. In summary, DMP allows to accurately control interconnectivity and topology of implants from all three families and micro-structured design holds potential to optimize their degradation speed. This first systematic report sheds light into how design influences degradation behaviour under in vivo-like conditions to help developing new standards for future medical device evaluation

Direct metal printed (DMP) porous iron implants possess promising mechanical and corrosion properties for various clinical application. Nevertheless, there is a requirement for better co-relation between in vitro and in vivo corrosion and biocompatibility behaviour of such biomaterials. Our present study evaluates absorption of porous iron implants under both static and dynamic conditions. Furthermore, this study characterizes their cytocompatibility using fibroblastic, osteogenic, endothelial and macrophagic cell types. In vitro degradation was performed statically and dynamically in a custom-built set-up placed under cell culture conditions (37 °C, 5% CO2 and 20% O2) for 28 days. The morphology and composition of the degradation products were analysed by scanning electron microscopy (SEM, JSM-IT100, JEOL). Iron implants before and after immersion were imaged by μCT (Quantum FX, Perkin Elmer, USA). Biocompatibility was also evaluated under static and dynamic in vitro culture conditions using L929, MG-63, HUVEC and RAW 264.7 cell lines. According to ISO 10993, cytocompatibility was evaluated directly using live/dead staining (Live and Dead Cell Assay kit, Abcam) in dual channel fluorescent optical imaging (FOI) and additionally quantified by flow cytometry. Furthermore, cytotoxicity was indirectly quantified using ISO conform extracts in proliferation assays. Strut size of DMP porous iron implants was 420 microns, with a porosity of 64% ± 0.2% as measured by micro-CT. After 28 days of physiological degradation in vitro, dynamically tested samples were covered with brownish degradation products. They revealed a 5.7- fold higher weight loss than statically tested samples, without significant changes in medium pH. Mechanical properties (E = 1600–1800 MPa) of these additively manufactured implants were still within the range of the values reported for trabecular bone, even after 28 days of biodegradation. Less than 25% cytotoxicity at 85% of the investigated time points was measured with L929 cells, while MG-63 and HUVEC cells showed 75% and 60% viability, respectively, after 24 h, with a decreasing trend with longer incubations. Cytotoxicity was analysed by two-way ANOVA and post-hoc Tukey's multiple comparisons test. Under dynamic culture conditions, live-dead staining and flow cytometric quantification showed a 2.8-fold and 5.7-fold increase in L929 and MG-63 cell survival rates, respectively, as compared to static conditions. Therefore, rationally designed and properly coated iron-based implants hold potential as a new generation of absorbable Orthopaedic implants