High-Scale 3D-Bioprinting Platform for the Automated Production of Vascularized Organs-on-a-Chip.

3D bioprinting possesses the potential to revolutionize contemporary methodologies for fabricating tissue models employed in pharmaceutical research and experimental investigations. This is enhanced by combining bioprinting with advanced organs-on-a-chip (OOCs), which includes a complex arrangement of multiple cell types representing organ-specific cells, connective tissue, and vasculature. However, both OOCs and bioprinting so far demand a high degree of manual intervention, thereby impeding efficiency and inhibiting scalability to meet technological requirements. Through the combination of drop-on-demand bioprinting with robotic handling of microfluidic chips, a print procedure is achieved that is proficient in managing three distinct tissue models on a chip within only a minute, as well as capable of consecutively processing numerous OOCs without manual intervention. This process rests upon the development of a post-printing sealable microfluidic chip, that is compatible with different types of 3D-bioprinters and easily connected to a perfusion system. The capabilities of the automized bioprint process are showcased through the creation of a multicellular and vascularized liver carcinoma model on the chip. The process achieves full vascularization and stable microvascular network formation over 14 days of culture time, with pronounced spheroidal cell growth and albumin secretion of HepG2 serving as a representative cell model.

Sensing Levofloxacin with an RNA Aptamer as a Bioreceptor.

To combat the growing threat of antibiotic resistance, environmental testing for antibiotic contamination is gaining an increasing role. This study aims to develop an easy-to-use assay for the detection of the fluoroquinolone antibiotic levofloxacin. Levofloxacin is used in human and veterinary medicine and has been detected in wastewater and river water. An RNA aptamer against levofloxacin was selected using RNA Capture-SELEX. The 73 nt long aptamer folds into three stems with a central three-way junction. It binds levofloxacin with a Kd of 6 µM and discriminates the closely related compound ciprofloxacin. Furthermore, the selection process was analyzed using a next-generation sequencing approach to better understand the sequence evolution throughout the selection. The aptamer was used as a bioreceptor for the development of a lateral flow assay. The biosensor exploited the innate characteristic of RNA Capture-SELEX to select aptamers that displace a complementary DNA oligonucleotide upon ligand binding. The lateral flow assay achieved a limit of visual detection of 100 µM. While the sensitivity of this assay constrains its immediate use in environmental testing, the present study can serve as a template for the selection of RNA aptamer-based biosensors.

A new 3D-printed polylactic acid-bioglass composite for bone tissue engineering induces angiogenesis in vitro and in ovo.

Large bone defects such as those that occur after trauma or resections due to cancer still are a challenge for surgeons. Main challenge in this area is to find a suitable alternative to the gold-standard therapy, which is highly risky, and a promising option is to use biomaterials manufactured by 3D printing. In former studies, we demonstrated that the combination of polylactic acid (PLA) and bioglass (BG) resulted in a stable 3D-printable material, and porous and finely structured scaffolds were printed. These scaffolds exhibited osteogenic and anti-inflammatory properties. This 3D-printed material fulfills most of the requirements described in the diamond concept of bone healing. However, the question remains as to whether it also meets the requirements concerning angiogenesis. Therefore, the aim of this study was to analyze the effects of the 3D-printed PLA-BG composite material on angiogenesis. In vitro analyses with human umbilical vein endothelial cells (HUVECs) showed a positive effect of increasing BG content on viability and gene expression of endothelial markers. This positive effect was confirmed by an enhanced vascular formation analyzed by Matrigel assay and chicken chorioallantoic membrane (CAM) assay. In this work, we demonstrated the angiogenic efficiency of a 3D-printed PLA-BG composite material. Recalling the osteogenic potential of this material demonstrated in former work, we manufactured a mechanically stable, 3D-printable, osteogenic and angiogenic material, which could be used for bone tissue engineering.

Influence of the physico-chemical bioink composition on the printability and cell biological properties in 3D-bioprinting of a liver tumor cell line.

The selection of a suitable matrix material is crucial for the development of functional, biomimetic tissue and organ models. When these tissue models are fabricated with 3D-bioprinting technology, the requirements do not only include the biological functionality and physico-chemical properties, but also the printability. In our work, we therefore present a detailed study of seven different bioinks with the focus on a functional liver carcinoma model. Agarose, gelatin, collagen and their blends were selected as materials based on their benefits for 3D cell culture and Drop-on-Demand (DoD) bioprinting. The formulations were characterized for their mechanical (G' of 10-350 Pa) and rheological (viscosity 2-200 Pa*s) properties as well as albumin diffusivity (8-50 µm2/s). The cellular behavior was exemplarily shown for HepG2 cells by monitoring viability, proliferation and morphology over 14 days, while the printability on a microvalve DoD printer was evaluated by drop volume monitoring in flight (100-250 nl), camera imaging of the wetting behavior and microscopy of the effective drop diameter (700 µm and more). We did not observe negative effects on cell viability or proliferation, which is due to the very low shear stresses inside the nozzle (200-500 Pa). With our method, we could identify the strengths and weaknesses of each material, resulting in a material portfolio. By specifically selecting certain materials or blends, cell migration and possible interaction with other cells can be directed as indicated by the results of our cellular experiments.

In vitro Evaluation of a 20% Bioglass-Containing 3D printable PLA Composite for Bone Tissue Engineering.

Three-dimensional (3D) printing is considered a key technology in the production of customized scaffolds for bone tissue engineering. In a previous work, we developed a 3D printable, osteoconductive, hierarchical organized scaffold system. The scaffold material should be osteoinductive. Polylactic acid (PLA) (polymer)/Bioglass (BG) (mineral/ion source) composite materials are promising. Previous studies of PLA/BG composites never exceed BG fractions of 10%, as increase of bioactive BG component negatively affects the printability of the composite material. Here, we test a novel, 3D printable PLA/BG composite with BG fractions up to 20% for its biological activity in vitro. PLA/BG filaments suitable for microstructure 3D printing were spun and the effect of different BG contents (5%, 10%, and 20%) in this material on mesenchymal stem cell (MSC) activity was tested in vitro. Our results showed that all tested composites are biocompatible. MSC cell adherence and metabolic activity increase with increasing BG content. The presence of BG component in scaffold has only slight effect on osteogenic gene expression, but it has significant suppressive effect on the expression of inflammatory genes in MSC. In addition, the material did not provoke any significant inflammatory response in whole-blood stimulation assay. The results show that by increasing the BG content, the bioactivity can be further enhanced.

Vascularization in Bioartificial Parenchymal Tissue: Bioink and Bioprinting Strategies.

Among advanced therapy medicinal products, tissue-engineered products have the potential to address the current critical shortage of donor organs and provide future alternative options in organ replacement therapy. The clinically available tissue-engineered products comprise bradytrophic tissue such as skin, cornea, and cartilage. A sufficient macro- and microvascular network to support the viability and function of effector cells has been identified as one of the main challenges in developing bioartificial parenchymal tissue. Three-dimensional bioprinting is an emerging technology that might overcome this challenge by precise spatial bioink deposition for the generation of a predefined architecture. Bioinks are printing substrates that may contain cells, matrix compounds, and signaling molecules within support materials such as hydrogels. Bioinks can provide cues to promote vascularization, including proangiogenic signaling molecules and cocultured cells. Both of these strategies are reported to enhance vascularization. We review pre-, intra-, and postprinting strategies such as bioink composition, bioprinting platforms, and material deposition strategies for building vascularized tissue. In addition, bioconvergence approaches such as computer simulation and artificial intelligence can support current experimental designs. Imaging-derived vascular trees can serve as blueprints. While acknowledging that a lack of structured evidence inhibits further meta-analysis, this review discusses an end-to-end process for the fabrication of vascularized, parenchymal tissue.

Recent trends in bioartificial muscle engineering and their applications in cultured meat, biorobotic systems and biohybrid implants.

Recent advances in tissue engineering and biofabrication technology have yielded a plethora of biological tissues. Among these, engineering of bioartificial muscle stands out for its exceptional versatility and its wide range of applications. From the food industry to the technology sector and medicine, the development of this tissue has the potential to affect many different industries at once. However, to date, the biofabrication of cultured meat, biorobotic systems, and bioartificial muscle implants are still considered in isolation by individual peer groups. To establish common ground and share advances, this review outlines application-specific requirements for muscle tissue generation and provides a comprehensive overview of commonly used biofabrication strategies and current application trends. By solving the individual challenges and merging various expertise, synergetic leaps of innovation that inspire each other can be expected in all three industries in the future.

Manufacturing, characterization, and degradation of a poly(lactic acid) warp-knitted spacer fabric scaffold as a candidate for tissue engineering applications.

Three-dimensional bioabsorbable textiles represent a novel technology for the manufacturing of tissue engineering scaffolds. In the present study, 3D bioabsorbable poly(lactic acid) (PLA) spacer fabric scaffolds are fabricated by warp-knitting and their potential for tissue engineering is explored in vitro. Changes in physical properties and mechanical performance with different heat setting treatments are assessed. To characterize the microenvironment experienced by cells in the scaffolds, yarn properties are investigated prior to, and during, hydrolytic degradation. The differences in yarn morphology, thermal properties, infrared spectra, and mechanical properties are investigated and monitored during temperature accelerated in vitro degradation tests in phosphate buffered saline (PBS) solution at 58 °C and pH 7.4 for 55 days. Yarn and textile cytocompatibility are tested to assess the effect of materials employed, manufacturing conditions, post processing and sterilization on cell viability, together with the cytocompatibility of the textile degradation products. Results show that the heat setting process can be used to modify scaffold properties, such as thickness, porosity, pore size and stiffness within the range useful for tissue regeneration. Scaffold degradation rate in physiological conditions is estimated by comparing yarn degradation data with PLA degradation data from literature. This will potentially allow the prediction of scaffold mechanical stability in the long term and thus its suitability for the remodelling of different tissues. Mouse calvaria preosteoblast MC3T3-E1 cells attachment and proliferation are observed on the scaffold over 12 days of in vitro culture by 4',6-diamidino-2-phenylindole (DAPI) fluorescent staining and DNA quantification. The present work shows the potential of spacer fabric scaffolds as a versatile and scalable scaffold fabrication technique, having the ability to create a microenvironment with appropriate physical, mechanical, and degradation properties for 3D tissue engineering. The high control and tunability of spacer fabric properties makes it a promising candidate for the regeneration of different tissues in patient-specific applications.

Scalable Biofabrication: A Perspective on the Current State and Future Potentials of Process Automation in 3D-Bioprinting Applications.

Biofabrication, specifically 3D-Bioprinting, has the potential to disruptively impact a wide range of future technological developments to improve human well-being. Organs-on-Chips could enable animal-free and individualized drug development, printed organs may help to overcome non-treatable diseases as well as deficiencies in donor organs and cultured meat may solve a worldwide environmental threat in factory farming. A high degree of manual labor in the laboratory in combination with little trained personnel leads to high costs and is along with strict regulations currently often a hindrance to the commercialization of technologies that have already been well researched. This paper therefore illustrates current developments in process automation in 3D-Bioprinting and provides a perspective on how the use of proven and new automation solutions can help to overcome regulatory and technological hurdles to achieve an economically scalable production.

3D-Printed PLA-Bioglass Scaffolds with Controllable Calcium Release and MSC Adhesion for Bone Tissue Engineering.

Large bone defects are commonly treated by replacement with auto- and allografts, which have substantial drawbacks including limited supply, donor site morbidity, and possible tissue rejection. This study aimed to improve bone defect treatment using a custom-made filament for tissue engineering scaffolds. The filament consists of biodegradable polylactide acid (PLA) and a varying amount (up to 20%) of osteoconductive S53P4 bioglass. By employing an innovative, additive manufacturing technique, scaffolds with optimized physico-mechanical and biological properties were produced. The scaffolds feature adjustable macro- and microporosity (200-2000 µm) with adaptable mechanical properties (83-135 MPa). Additionally, controllable calcium release kinetics (0-0.25 nMol/µL after 24 h), tunable mesenchymal stem cell (MSC) adhesion potential (after 24 h by a factor of 14), and proliferation (after 168 h by a factor of 18) were attained. Microgrooves resulting from the 3D-printing process on the surface act as a nucleus for cell aggregation, thus being a potential cell niche for spheroid formation or possible cell guidance. The scaffold design with its adjustable biomechanics and the bioglass with its antimicrobial properties are of particular importance for the preclinical translation of the results. This study comprehensibly demonstrates the potential of a 3D-printed bioglass composite scaffold for the treatment of critical-sized bone defects.

Fabrication of biomimetic networks using viscous fingering in flexographic printing.

Mammalian tissue comprises a plethora of hierarchically organized channel networks that serve as routes for the exchange of liquids, nutrients, bio-chemical cues or electrical signals, such as blood vessels, nerve fibers, or lymphatic conduits. Despite differences in function and size, the networks exhibit a similar, highly branched morphology with dendritic extensions. Mimicking such hierarchical networks represents a milestone in the biofabrication of tissues and organs. Work to date has focused primarily on the replication of the vasculature. Despite initial progress, reproducing such structures across scales and increasing biofabrication efficiency remain a challenge. In this work, we present a new biofabrication method that takes advantage of the viscous fingering phenomenon. Using flexographic printing, highly branched, inter-connective channel structures with stochastic, biomimetic distribution and dendritic extensions can be fabricated with unprecedented efficiency. Using gelatin (5%-35%) as resolvable sacrificial material, the feasability of the proposed method is demonstrated on the example of a vascular network. By selectively adjusting the printing velocity (0.2-1.5 m s-1), the anilox roller dip volume (4.5-24 ml m-2) as well as the shear viscosity of the printing material used (10-900 mPas), the width of the structures produced (30-400 µm) as well as their distance (200-600 µm) can be specifically determined. In addition to the flexible morphology, the high scalability (2500-25 000 mm2) and speed (1.5 m s-1) of the biofabrication process represents an important unique selling point. Printing parameters and hydrogel formulations are investigated and tuned towards a process window for controlled fabrication of channels that mimic the morphology of small blood vessels and capillaries. Subsequently, the resolvable structures were casted in a hydrogel matrix enabling bulk environments with integrated channels. The perfusability of the branched, inter-connective structures was successfully demonstrated. The fabricated networks hold great potential to enable nutrient supply in thick vascularized tissues or perfused organ-on-a-chip systems. In the future, the concept can be further optimized and expanded towards large-scale and cost-efficient biofabrication of vascular, lymphatic or neural networks for tissue engineering and regenerative medicine.

Toward 3D-bioprinting of an endocrine pancreas: A building-block concept for bioartificial insulin-secreting tissue.

Three-dimensional bioprinting of an endocrine pancreas is a promising future curative treatment for patients with insulin secretion deficiency. In this study, we present an end-to-end concept from the molecular to the macroscopic level. Building-blocks for a hybrid scaffold device of hydrogel and functionalized polycaprolactone were manufactured by 3D-(bio)printing. Pseudoislet formation from INS-1 cells after bioprinting resulted in a viable and proliferative experimental model. Transcriptomics showed an upregulation of proliferative and ß-cell-specific signaling cascades, downregulation of apoptotic pathways, overexpression of extracellular matrix proteins, and VEGF induced by pseudoislet formation and 3D-culture. Co-culture with endothelial cells created a natural cellular niche with enhanced insulin secretion after glucose stimulation. Survival and function of pseudoislets after explantation and extensive scaffold vascularization of both hydrogel and heparinized polycaprolactone were demonstrated in vivo. Computer simulations of oxygen, glucose and insulin flows were used to evaluate scaffold architectures and Langerhans islets at a future perivascular transplantation site.

Targeted Printing of Cells: Evaluation of ADA-PEG Bioinks for Drop on Demand Approaches.

A novel approach, in the context of bioprinting, is the targeted printing of a defined number of cells at desired positions in predefined locations, which thereby opens up new perspectives for life science engineering. One major challenge in this application is to realize the targeted printing of cells onto a gel substrate with high cell survival rates in advanced bioinks. For this purpose, different alginate-dialdehyde-polyethylene glycol (ADA-PEG) inks with different PEG modifications and chain lengths (1-8 kDa) were characterized to evaluate their application as bioinks for drop on demand (DoD) printing. The biochemical properties of the inks, printing process, NIH/3T3 fibroblast cell distribution within a droplet and shear forces during printing were analyzed. Finally, different hydrogels were evaluated as a printing substrate. By analysing different PEG chain lengths with covalently crosslinked and non-crosslinked ADA-PEG inks, it was shown that the influence of Schiff's bases on the viscosity of the corresponding materials is very low. Furthermore, it was shown that longer polymer chains resulted in less stable hydrogels, leading to fast degradation rates. Several bioinks highly exhibit biocompatibility, while the calculated nozzle shear stress increased from approx. 1.3 and 2.3 kPa. Moreover, we determined the number of cells for printed droplets depending on the initial cell concentration, which is crucially needed for targeted cell printing approaches.

Investigation and comparison of resin materials in transparent DLP-printing for application in cell culture and organs-on-a-chip.

Organs-on-a-Chip (OOCs) have recently led to major discoveries and a better understanding of 3D cell organization, cell-cell interactions and tissue response to drugs and biological cues. However, their complexity and variability are still limited by the available fabrication technology. Transparent, cytocompatible and high-resolution 3D-printing could overcome these limitations, offering a flexible and low-cost alternative to soft lithography. Many advances have been made in stereolithography printing regarding resin formulation and the general printing process, but a systematic analysis of the printing process steps, employed resins and post-treatment procedures with a strong focus on the requirements in OOCs is missing. To fill this gap, this work provides an in-depth analysis of three different resin systems in comparison to polystyrene (PS) and poly(dimethylsiloxane) (PDMS), which can be considered the gold-standards in cell culture and microfluidics. The resins were characterized with respect to transparency, cytocompatibility and print resolution. These properties are not only governed by the resin composition, but additionally by the post-treatment procedure. The investigation of the mechanical (elastic modulus ∼2.2 GPa) and wetting properties (∼60° native / 20° plasma treated) showed a behavior very similar to PS. In addition, the absorbance of small molecules was two orders of magnitude lower in the applied resins (diffusion constant ∼0.01 µm2 s-1) than for PDMS (2.5 µm2 s-1), demonstrating the intrinsic suitability of these materials for OOCs. Raman spectroscopy and UV/VIS spectrophotometry revealed that post-treatment increased monomer conversion up to 2 times and removed photo initiator residues, leading to an increased transparency of up to 50% and up to 10-times higher cell viability. High magnification fluorescence imaging of HUVECs and L929 cells cultivated on printed dishes shows the high optical qualities of prints fabricated by the Digital Light Processing (DLP) printer. Finally, components of microfluidic chips such as high-aspect ratio pillars and holes with a diameter of 50 µm were printed. Concluding, the suitability of DLP-printing for OOCs was demonstrated by filling a printed chip with a cell-hydrogel mixture using a microvalve bioprinter, followed by the successful cultivation under perfusion. Our results highlight that DLP-printing has matured into a robust fabrication technology ready for application in extensive and versatile OOC research.

Scaffolds for Cultured Meat on the Basis of Polysaccharide Hydrogels Enriched with Plant-Based Proteins.

The world population is growing and alternative ways of satisfying the increasing demand for meat are being explored, such as using animal cells for the fabrication of cultured meat. Edible biomaterials are required as supporting structures. Hence, we chose agarose, gellan and a xanthan-locust bean gum blend (XLB) as support materials with pea and soy protein additives and analyzed them regarding material properties and biocompatibility. We successfully built stable hydrogels containing up to 1% pea or soy protein. Higher amounts of protein resulted in poor handling properties and unstable gels. The gelation temperature range for agarose and gellan blends is between 23-30 °C, but for XLB blends it is above 55 °C. A change in viscosity and a decrease in the swelling behavior was observed in the polysaccharide-protein gels compared to the pure polysaccharide gels. None of the leachates of the investigated materials had cytotoxic effects on the myoblast cell line C2C12. All polysaccharide-protein blends evaluated turned out as potential candidates for cultured meat. For cell-laden gels, the gellan blends were the most suitable in terms of processing and uniform distribution of cells, followed by agarose blends, whereas no stable cell-laden gels could be formed with XLB blends.

Biosynthetic, biomimetic, and self-assembled vascularized Organ-on-a-Chip systems.

Organ-on-a-Chip (OOC) devices have seen major advances in the last years with respect to biological complexity, physiological composition and biomedical relevance. In this context, integration of vasculature has proven to be a crucial element for long-term culture of thick tissue samples as well as for realistic pharmacokinetic, toxicity and metabolic modelling. With the emergence of digital production technologies and the reinvention of existing tools, a multitude of design approaches for guided angio- and vasculogenesis is available today. The underlying production methods can be categorized into biosynthetic, biomimetic and self-assembled vasculature formation. The diversity and importance of production approaches, vascularization strategies as well as biomaterials and cell sourcing are illustrated in this work. A comprehensive technological review with a strong focus on the challenge of producing physiologically relevant vascular structures is given. Finally, the remaining obstacles and opportunities in the development of vascularized Organ-on-a-Chip platforms for advancing drug development and predictive disease modelling are noted.

Warp-Knitted Spacer Fabrics: A Versatile Platform to Generate Fiber-Reinforced Hydrogels for 3D Tissue Engineering.

Mesenchymal stem cells (MSCs) possess huge potential for regenerative medicine. For tissue engineering approaches, scaffolds and hydrogels are routinely used as extracellular matrix (ECM) carriers. The present study investigated the feasibility of using textile-reinforced hydrogels with adjustable porosity and elasticity as a versatile platform for soft tissue engineering. A warp-knitted poly (ethylene terephthalate) (PET) scaffold was developed and characterized with respect to morphology, porosity, and mechanics. The textile carrier was infiltrated with hydrogels and cells resulting in a fiber-reinforced matrix with adjustable biological as well as mechanical cues. Finally, the potential of this platform technology for regenerative medicine was tested on the example of fat tissue engineering. MSCs were seeded on the construct and exposed to adipogenic differentiation medium. Cell invasion was detected by two-photon microscopy, proliferation was measured by the PrestoBlue assay. Successful adipogenesis was demonstrated using Oil Red O staining as well as measurement of secreted adipokines. In conclusion, the given microenvironment featured optimal mechanical as well as biological properties for proliferation and differentiation of MSCs. Besides fat tissue, the textile-reinforced hydrogel system with adjustable mechanics could be a promising platform for future fabrication of versatile soft tissues, such as cartilage, tendon, or muscle.

Matrix-specific mechanism of Fe ion release from laser-generated 3D-printable nanoparticle-polymer composites and their protein adsorption properties.

Nanocomposites have been widely applied in medical device fabrication and tissue-engineering applications. In this context, the release of metal ions as well as protein adsorption capacity are hypothesized to be two key processes directing nanocomposite-cell interactions. The objective of this study is to understand the polymer-matrix effects on ion release kinetics and their relations with protein adsorption. Laser ablation in macromolecule solutions was employed for synthesizing Au and Fe nanoparticle-loaded nanocomposites based on thermoplastic polyurethane (TPU) and alginate. Confocal microscopy revealed a three-dimensional homogeneous dispersion of laser-generated nanoparticles in the polymer. The physicochemical properties revealed a pronounced dependence upon embedding of Fe and Au nanoparticles in both polymer matrices. Interestingly, the total Fe ion concentration released from alginate gels under static conditions decreased with increasing mass loadings, a phenomenon only found in the Fe-alginate system and not in the Cu/Zn-alginate and Fe-TPU control system (where the effects were proportioonal to the nanoparticle load). A detailed mechanistic examination of iron the ion release process revealed that it is probably not the redox potential of metals and diffusion of metal ions alone, but also the solubility of nano-metal oxides and affinity of metal ions for alginate that lead to the special release behaviors of iron ions from alginate gels. The amount of adsorbed bovine serum albumin (BSA) and collagen I on the surface of both the alginate and TPU composites was significantly increased in contrast to the unloaded control polymers and could be correlated with the concentration of released Fe ions and the porosity of composites, but was independent of the global surface charge. Interestingly, these effects were already highly pronounced at minute loadings with Fe nanoparticles down to 200 ppm. Moreover, the laser-generated Fe or Au nanoparticle-loaded alginate composites were shown to be a suitable bioink for 3D printing. These findings are potentially relevant for ion-sensitive bio-responses in cell differentiation, endothelisation, vascularisation, or wound healing.

Bioprinting Cell- and Spheroid-Laden Protein-Engineered Hydrogels as Tissue-on-Chip Platforms.

Human tissues, both in health and disease, are exquisitely organized into complex three-dimensional architectures that inform tissue function. In biomedical research, specifically in drug discovery and personalized medicine, novel human-based three-dimensional (3D) models are needed to provide information with higher predictive value compared to state-of-the-art two-dimensional (2D) preclinical models. However, current in vitro models remain inadequate to recapitulate the complex and heterogenous architectures that underlie biology. Therefore, it would be beneficial to develop novel models that could capture both the 3D heterogeneity of tissue (e.g., through 3D bioprinting) and integrate vascularization that is necessary for tissue viability (e.g., through culture in tissue-on-chips). In this proof-of-concept study, we use elastin-like protein (ELP) engineered hydrogels as bioinks for constructing such tissue models, which can be directly dispensed onto endothelialized on-chip platforms. We show that this bioprinting process is compatible with both single cell suspensions of neural progenitor cells (NPCs) and spheroid aggregates of breast cancer cells. After bioprinting, both cell types remain viable in incubation for up to 14 days. These results demonstrate a first step toward combining ELP engineered hydrogels with 3D bioprinting technologies and on-chip platforms comprising vascular-like channels for establishing functional tissue models.

3D-Printing of Hierarchically Designed and Osteoconductive Bone Tissue Engineering Scaffolds.

In Bone Tissue Engineering (BTE), autologous bone-regenerative cells are combined with a scaffold for large bone defect treatment (LBDT). Microporous, polylactic acid (PLA) scaffolds showed good healing results in small animals. However, transfer to large animal models is not easily achieved simply by upscaling the design. Increasing diffusion distances have a negative impact on cell survival and nutrition supply, leading to cell death and ultimately implant failure. Here, a novel scaffold architecture was designed to meet all requirements for an advanced bone substitute. Biofunctional, porous subunits in a load-bearing, compression-resistant frame structure characterize this approach. An open, macro- and microporous internal architecture (100 µm-2 mm pores) optimizes conditions for oxygen and nutrient supply to the implant's inner areas by diffusion. A prototype was 3D-printed applying Fused Filament Fabrication using PLA. After incubation with Saos-2 (Sarcoma osteogenic) cells for 14 days, cell morphology, cell distribution, cell survival (fluorescence microscopy and LDH-based cytotoxicity assay), metabolic activity (MTT test), and osteogenic gene expression were determined. The adherent cells showed colonization properties, proliferation potential, and osteogenic differentiation. The innovative design, with its porous structure, is a promising matrix for cell settlement and proliferation. The modular design allows easy upscaling and offers a solution for LBDT.