Biofabrication Method Research Articles

3D in vitro modeling of the exocrine pancreatic unit using tomographic volumetric bioprinting

Pancreatic ductal adenocarcinoma (PDAC) is the most common type of pancreatic cancer, a leading cause of cancer-related deaths globally. Initial lesions of PDAC develop within the exocrine pancreas' functional units, with tumor progression driven by interactions between PDAC and stromal cells. Effective therapies require anatomically and functionally relevantin vitrohuman models of the pancreatic cancer microenvironment. We employed tomographic volumetric bioprinting, a novel biofabrication method, to create human fibroblast-laden constructs mimicking the tubuloacinar structures of the exocrine pancreas. Human pancreatic ductal epithelial (HPDE) cells overexpressing the KRAS oncogene (HPDE-KRAS) were seeded in the multiacinar cavity to replicate pathological tissue. HPDE cell growth and organization within the structure were assessed, demonstrating the formation of a thin epithelium covering the acini inner surfaces. Immunofluorescence assays showed significantly higher alpha smooth muscle actin (α-SMA) vs. F-actin expression in fibroblasts co-cultured with cancerous versus wild-type HPDE cells. Additionally,α-SMA expression increased over time and was higher in fibroblasts closer to HPDE cells. Elevated interleukin (IL)-6 levels were quantified in supernatants from co-cultures of stromal and HPDE-KRAS cells. These findings align with inflamed tumor-associated myofibroblast behavior, serving as relevant biomarkers to monitor early disease progression and target drug efficacy. To our knowledge, this is the first demonstration of a 3D bioprinted model of exocrine pancreas that recapitulates its true 3-dimensional microanatomy and shows tumor triggered inflammation.

Evaluation of anti-vibriocidal, antioxidant properties and cytotoxicity of bio fabricated/green synthesized silver nanoparticles using Euphorbia hitra L. leaf extract

The increasing focus on combating antimicrobial resistance (AMR) in aquaculture sector is driving innovations and better improvements to save billions of dollars globally. Two gram- negative bacteria which cause vibriosis, Vibrio parahaemolytics and Vibrio harveyi, are now resistant to multiple antibiotics. Green technology has evolved to produce novel antimicrobials with reduced potential for AMR and sustainable synthesis. The present study utilized a one-pot biofabrication method to synthesize silver nanoparticles (AgNPs) using Euphorbia hitra leaf aqueous extract (EHLAE) through a bottom-up approach. The qualitative phytochemical examination of EHLAE confirmed the existence of diverse components including steroids, carbohydrates, alkaloids, flavonoids, tannins, terpenoids, saponins, glycosides, and phenols. These compounds have crucial role as reducing and capping AgNPs. The obtained gAgNPs were subjected to evaluate the physico-chemical properties such as Surface Plasmon Resonance (SPR), functional groups, and crystalline structure, were verified by UV-Vis spectroscopy, Fourier Transform Infrared Spectroscopy (FTIR), and X-ray diffraction (XRD). The morphology, elemental composition, and size distribution of the AgNPs synthesized using a green method were confirmed using advanced techniques including Field Emission Scanning Electron Microscope (FESEM) with Energy Dispersive X-ray spectroscopy (EDX), High-Resolution Transmission Electron Microscope (HRTEM), and Dynamic Light Scattering (DLS). The silver nanoparticles (AgNPs) shown potent anti-vibriocidal activity against both Vibrio parahaemolyticus and Vibrio harveyi and the Minimum Inhibitory Concentration (MIC) for both bacteria were consistently 31.25 µg mL−1. The in vitro antioxidant tests showed effective results and the found IC50 values are 306 µg mL−1 for DPPH and 6.46 µg mL−1 for FRAP assays. Furthermore, the assessment of in vitro cytotoxicity on Vero cell lines revealed minimal toxicity, as evidenced by an IC50 value of 191.51 µg mL−1. The results reveals that the green synthesized AgNPs using Euphorbia hitra leaf have the potential to be used as effective alternatives for therapeutic purposes in combating the increasing problem of antimicrobial resistance in the aquaculture industry.

Chaos-Assisted Production of Micro-Architected Spheres (CAPAS).

Hydrogel droplets with inner compartments are valuable in various fields, including tissue engineering. A droplet-based biofabrication method is presented for the chaos-assisted production of architected spheres (CAPAS) for the rapid generation of multilayered hydrogel spheres (ranging from 0.6 to 3.5mm in diameter) at high-throughput rates (up to 2000 spheres per min). This method is based on the use of chaotic advection generated by a Kenics static mixer (KSM) nozzle. The configuration of the KSM (i.e., the number of mixing elements) determines the number of compartments within the sphere. Sphere size is adjusted by flow rate, printhead outlet diameter, polymer concentration (sodium alginate or gelatin-methacryloyl (GelMA)), and crosslinking bath composition. This versatile system operates in dripping and jetting modes, preserving multilayered architecture in both modes. Proof-of-concept experiments with breast cancer (MDA-MB-231), human dermal fibroblast (HDF), and murine myoblast (C2C12) lines show over 80% cell viability immediately post-fabrication, maintained over extended culture (14 or 30 days). CAPAS is used to create a breast cancer model with cancer-tissue-like and healthy-tissue-like micro-niches to test paclitaxel doses. It is envisioned that CAPAS will enable high-throughput fabrication of hydrogel spheres for tissue engineering, chemical engineering, and material sciences applications.

Bioactive gelatin-sheets as novel biopapers to support prevascularization organized by laser-assisted bioprinting for bone tissue engineering

Despite significant advances in the management of patients with oral cancer, maxillofacial reconstruction after ablative surgery remains a clinical challenge. In bone tissue engineering, biofabrication strategies have been proposed as promising alternatives to solve issues associated with current therapies and to produce bone substitutes that mimic both the structure and function of native bone. Among them, laser-assisted bioprinting (LAB) has emerged as a relevant biofabrication method to print living cells and biomaterials with micrometric resolution onto a receiving substrate, also called ‘biopaper’. Recent studies have demonstrated the benefits of prevascularization using LAB to promote vascularization and bone regeneration, but mechanical and biological optimization of the biopaper are needed. The aim of this study was to apply gelatin-sheet fabrication process to the development of a novel biopaper able to support prevascularization organized by LAB for bone tissue engineering applications. Gelatin-based sheets incorporating bioactive glasses (BGs) were produced using various freezing methods and crosslinking (CL) parameters. The different formulations were characterized in terms of microstructural, physical, mechanical, and biological properties in monoculture and coculture. Based on multi-criteria analysis, a rank scoring method was used to identify the most relevant formulations. The selected biopaper underwent additional characterization regarding its ability to support mineralization and vasculogenesis, its bioactivity potential and in vivo degradability. The biopaper ‘Gel5wt% BG1wt%—slow freezing—CL160 °C 24 h′ was selected as the best candidate, due to its suitable properties including high porosity (91.69 ± 1.55%), swelling ratio (91.61 ± 0.60%), Young modulus (3.97 × 104 ± 0.97 × 104 Pa) but also its great cytocompatibility, osteogenesis and bioactivity properties. The preorganization of human umbilical vein endothelial cell using LAB onto this new biopaper led to the formation of microvascular networks. This biopaper was also shown to be compatible with 3D-molding and 3D-stacking strategies. This work allowed the development of a novel biopaper adapted to LAB with great potential for vascularized bone biofabrication.

Influence of Diameter and Cyclic Mechanical Stimulation on the Beating Frequency of Myocardial Cell-Laden Fibers.

Atrioventricular block (AVB) is a severe disease for pediatric patients. The repetitive operations needed in the case of the pacemaker implantation to maintain the electrical signal at the atrioventricular node (AVN) affect the patient's life quality. In this study, we present a method of biofabrication of multi-cell-laden cylindrical fibrin-based fibers that can restore the electrical signal at the AVN. We used human umbilical vein smooth muscle cells (HUVSMCs), human umbilical vein endothelial cells (HUVECs) and induced pluripotent stem cell cardiomyocytes (iPSC-CMs) cultivated either statically or dynamically to mimic the native AVN. We investigated the influence of cell composition, construct diameter and cyclic stretch on the function of the fibrin hydrogels in vitro. Immunohistochemistry analyses showed the maturity of the iPSC-CMs in the constructs through the expression of sarcomeric alpha actinin (SAA) and electrical coupling through Connexin 43 (Cx43) signal. Simultaneously, the beating frequency of the fibrin hydrogels was higher and easy to maintain whereas the concentration of iPSC-CMs was higher compared with the other types of cylindrical constructs. In total, our study highlights that the combination of fibrin with the cell mixture and geometry is offering a feasible biofabrication method for tissue engineering approaches for the treatment of AVB.

A novel 3D biofabrication strategy to improve cell proliferation and differentiation of human Wharton's jelly mesenchymal stromal cells for cell therapy and tissue engineering.

Purpose: Obtaining sufficient numbers of cells in a short time is a major goal of cell culturing in cell therapy and tissue engineering. However, current bidimensional (2D) culture methods are associated to several limitations, including low efficiency and the loss of key cell differentiation markers on cultured cells. Methods: In the present work, we have designed a novel biofabrication method based on a three-dimensional (3D) culture system (FIBRIAGAR-3D). Human Wharton's jelly mesenchymal stromal cells (HWJSC) were cultured in 3D using 100%, 75%, 50%, and 25% concentrations of fibrin-agarose biomaterials (FA100, FA75, FA50 and FA25 group) and compared with control cells cultured using classical 2D systems (CTR-2D). Results: Our results showed a significant increase in the number of cells generated after 7days of culture, with cells displaying numerous expansions towards the biomaterial, and a significant overexpression of the cell proliferation marker KI67 was found for the FA75 and FA100 groups. TUNEL and qRT-PCR analyses demonstrated that the use of FIBRIAGAR-3D was not associated with an induction of apoptosis by cultured cells. Instead, the 3D system retained the expression of typical phenotypic markers of HWJSC, including CD73, CD90, CD105, NANOG and OCT4, and biosynthesis markers such as types-I and IV collagens, with significant increase of some of these markers, especially in the FA100 group. Finally, our analysis of 8 cell signaling molecules revealed a significant decrease of GM-CSF, IFN-g, IL2, IL4, IL6, IL8, and TNFα, suggesting that the 3D culture system did not induce the expression of pro-inflammatory molecules. Conclusion: These results confirm the usefulness of FIBRIAGAR-3D culture systems to increase cell proliferation without altering cell phenotype of immunogenicity and opens the door to the possibility of using this novel biofabrication method in cell therapy and tissue engineering of the human cornea, oral mucosa, skin, urethra, among other structures.

Combining rotary wet-spinning biofabrication and electro-mechanical stimulation for thein vitroproduction of functional myo-substitutes.

In this work, we present an innovative, high-throughput rotary wet-spinning biofabrication method for manufacturing cellularized constructs composed of highly-aligned hydrogel fibers. The platform is supported by an innovative microfluidic printing head (MPH) bearing a crosslinking bath microtank with a co-axial nozzle placed at the bottom of it for the immediate gelation of extruded core/shell fibers. After a thorough characterization and optimization of the new MPH and the fiber deposition parameters, we demonstrate the suitability of the proposed system for thein vitroengineering of functional myo-substitutes. The samples produced through the described approach were first characterizedin vitroand then used as a substrate to ascertain the effects of electro-mechanical stimulation on myogenic maturation. Of note, we found a characteristic gene expression modulation of fast (MyH1), intermediate (MyH2), and slow (MyH7) twitching myosin heavy chain isoforms, depending on the applied stimulation protocol. This feature should be further investigated in the future to biofabricate engineered myo-substitutes with specific functionalities.

Injectable rBMSCs-laden hydrogel microspheres loaded with naringin for osteomyelitis treatment

Osteomyelitis, caused by purulent bacteria invading bone tissue, often occurs in long bones and seriously affects the physical and mental health and working ability of patients; it can even endanger life. However, due to bone cavity structure, osteomyelitis tends to occur inside the bone and thus lacks an effective treatment; anti-inflammatory treatment and repair of bone defects are necessary. Here, we developed injectable hydrogel microspheres loaded with naringin and bone marrow mesenchymal stem cells, which have anti-inflammatory and osteogenic properties. These homogeneous microspheres, ranging from 200 to 1000 μm, can be rapidly fabricated using an electro-assisted bio-fabrication method. Interestingly, it was found that microspheres with relatively small diameters (200 μm) were more conducive to the initial cell attachment, growth, spread, and later osteogenic differentiation. The developed microspheres can effectively treat tibial osteomyelitis in rats within six weeks, proving their prospects for clinical application.

Biomimetic hydrogel scaffolds embedded with porous microtubes as perfusion channels

Despite the remarkable progress in biofabrication, the vascularization of large-sized tissue scaffolds remains a great challenge for tissue engineering. The lack of effective capillary vessels in the scaffold can result in cellular necrosis due to the lack of oxygen and nutrients. Integrating capillary vessels in scaffolds is critical to maintaining cellular metabolic functions and the ultimate success of engineering large-scale tissues and organs. This paper demonstrates a novel hybrid biofabrication method that combines coaxial electrospinning and extrusion-based bioprinting to fabricate microtube-embedded hydrogel scaffolds. The porous microtubes mimicked the capillary morphology and were embedded between layers of 3D printed sodium alginate scaffold to function as a microchannel diffusion system. The dye diffusion test showed that the microtubes enhanced the mass transport within the scaffold.

Multiscale Anisotropic Scaffold Integrating 3D Printing and Electrospinning Techniques as a Heart-on-a-Chip Platform for Evaluating Drug-Induced Cardiotoxicity.

Cardiac safety assessments are significant in drug discovery, as drug-induced cardiotoxicity (DIC) is the primary cause of drug attrition. Despite heart-on-a-chip (HoC) technology becoming an increasingly popular tool for evaluating DIC, its development remains a challenge owing to the anisotropic cardiac structure of the native myocardium. Herein, an anisotropic multiscale cardiac scaffold is presented via a hybrid biofabrication method by combining 3D printing with electrospinning technology, where the 3D-printed micrometer-scale scaffold frames enable mimicking the interwoven myocardium anatomical structure and the branched-aligned electrospun nanofibers network is able to directionally guide cellular arrangements. The in vitro 3D bioengineered cardiac tissues are then fabricated by encapsulating three-layer multiscale scaffolds within a photocurable methacrylated gelatin hydrogel shell. It is demonstrated that such an anisotropic multiscale structure could contribute to enhancing cardiomyocyte maturation and synchronous beating behavior. More attractively, with the integration of 3D bioengineered cardiac tissues and a self-designed microfluidic perfusion system, a 3D anisotropic HoC platform is established for evaluating DIC and cardioprotective efficacy. Collectively, these results indicate that the HoC model developed by integrating the 3D bioengineered cardiac tissues could effectively recapitulate the clinical manifestations, thereby highlighting their efficacy as a valuable preclinical platform for testing drug efficacy and cardiotoxicity.

Engineering synthetic poly(ethylene) glycol-based hydrogels compatible with injection molding biofabrication.

Hydrogel injection molding is a biofabrication method that is useful for the rapid generation of complex cell-laden hydrogel geometries, with potential utility in biomanufacturing products for tissue engineering applications. Hydrogel injection molding requires that hydrogel polymers have sufficiently delayed crosslinking times to enable injection and molding prior to gelation. In this work, we explore the feasibility of injection molding synthetic poly(ethylene) glycol (PEG)-based hydrogels functionalized with strain promoted azide-alkyne cycloaddition click chemistry functional groups. We evaluate the mechanical properties of a PEG-based hydrogel library, including time to gelation and successful generation of complex geometries via injection molding. We evaluate the binding and retention of adhesive ligand RGD within the library matrices and characterize the viability and function of encapsulated cells. This work demonstrates the feasibility of injection molding synthetic PEG-based hydrogels for tissue engineering applications, with potential utility in the clinic and biomanufacturing.

Three-dimensional biofabrication of nanosecond laser micromachined nanofibre meshes for tissue engineered scaffolds.

There is a high demand for bespoke grafts to replace damaged or malformed bone and cartilage tissue. Three-dimensional (3D) printing offers a method of fabricating complex anatomical features of clinically relevant sizes. However, the construction of a scaffold to replicate the complex hierarchical structure of natural tissues remains challenging. This paper reports a novel biofabrication method that is capable of creating intricately designed structures of anatomically relevant dimensions. The beneficial properties of the electrospun fibre meshes can finally be realised in 3D rather than the current promising breakthroughs in two-dimensional (2D). The 3D model was created from commercially available computer-aided design software packages in order to slice the model down into many layers of slices, which were arrayed. These 2D slices with each layer of a defined pattern were laser cut, and then successfully assembled with varying thicknesses of 100 μm or 200 μm. It is demonstrated in this study that this new biofabrication technique can be used to reproduce very complex computer-aided design models into hierarchical constructs with micro and nano resolutions, where the clinically relevant sizes ranging from a simple cube of 20 mm dimension, to a more complex, 50 mm-tall human ears were created. In-vitro cell-contact studies were also carried out to investigate the biocompatibility of this hierarchal structure. The cell viability on a micromachined electrospun polylactic-co-glycolic acid fibre mesh slice, where a range of hole diameters from 200 μm to 500 μm were laser cut in an array where cell confluence values of at least 85% were found at three weeks. Cells were also seeded onto a simpler stacked construct, albeit made with micromachined poly fibre mesh, where cells can be found to migrate through the stack better with collagen as bioadhesives. This new method for biofabricating hierarchical constructs can be further developed for tissue repair applications such as maxillofacial bone injury or nose/ear cartilage replacement in the future.

Swelling-Dependent Shape-Based Transformation of a Human Mesenchymal Stromal Cells-Laden 4D Bioprinted Construct for Cartilage Tissue Engineering.

3D bioprinting is usually implemented on flat surfaces, posing serious limitations in the fabrication of multilayered curved constructs. 4D bioprinting, combining 3D bioprinting with time-dependent stimuli-induced transformation, enables the fabrication of shape-changing constructs. Here, a 4D biofabrication method is reported for cartilage engineering based on the differential swelling of a smart multi-material system made from two hydrogel-based materials: hyaluronan and alginate. Two ink formulations are used: tyramine-functionalized hyaluronan (HAT, high-swelling) and alginate with HAT (AHAT, low-swelling). Both inks have similar elastic, shear-thinning, and printability behavior. The inks are 3D printed into a bilayered scaffold before triggering the shape-change by using liquid immersion as stimulus. In time (4D), the differential swelling between the two zones leads to the scaffold's self-bending. Different designs are made to tune the radius of curvature and shape. A bioprinted formulation of AHAT and human bone marrow cells demonstrates high cell viability. After 28 days in chondrogenic medium, the curvature is clearly present while cartilage-like matrix production is visible on histology. A proof-of-concept of the recently emerged technology of 4D bioprinting with a specific application for the design of curved structures potentially mimicking the curvature and multilayer cellular nature of native cartilage is demonstrated.

Biofabrication of Sodium Alginate Hydrogel Scaffolds for Heart Valve Tissue Engineering.

Every year, thousands of aortic valve replacements must take place due to valve diseases. Tissue-engineered heart valves represent promising valve substitutes with remodeling, regeneration, and growth capabilities. However, the accurate reproduction of the complex three-dimensional (3D) anatomy of the aortic valve remains a challenge for current biofabrication methods. We present a novel technique for rapid fabrication of native-like tricuspid aortic valve scaffolds made of an alginate-based hydrogel. Using this technique, a sodium alginate hydrogel formulation is injected into a mold produced using a custom-made sugar glass 3D printer. The mold is then dissolved using a custom-made dissolving module, revealing the aortic valve scaffold. To assess the reproducibility of the technique, three scaffolds were thoroughly compared. CT (computed tomography) scans showed that the scaffolds respect the complex native geometry with minimal variations. The scaffolds were then tested in a cardiac bioreactor specially designed to reproduce physiological flow and pressure (aortic and ventricular) conditions. The flow and pressure profiles were similar to the physiological ones for the three valve scaffolds, with small variabilities. These early results establish the functional repeatability of this new biofabrication method and suggest its application for rapid fabrication of ready-to-use cell-seeded sodium alginate scaffolds for heart valve tissue engineering.

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.

Digital Light Processing Bioprinting Advances for Microtissue Models.

Digital light processing (DLP) bioprinting has been widely introduced as a fast and robust biofabrication method in tissue engineering. The technique holds a great promise for creating tissue models because it can replicate the resolution and complexity of natural tissues and constructs. A DLP system projects 2D images onto layers of bioink using a digital photomask. The resolution of DLP bioprinting strongly depends on the characteristics of the projected light and the photo-cross-linking response of the bioink microenvironment. In this review, we present a summary of DLP fundamentals with a focus on bioink properties, photoinitiator selection, and light characteristics in resolution of bioprinted constructs. A simple guideline is provided for bioengineers interested in using DLP platforms and customizing technical specifications for its design. The literature review reveals the promising future of DLP bioprinting for disease modeling and biofabrication.

Applications of Tissue Decellularization Techniques in Ventricular Myocardial Biofabrication.

Ischemic heart disease is the leading cause of death around the world, and though the advent of coronary revascularization has revolutionized its treatment, many patients who sustain ischemic injury to the heart will go on to develop heart failure. Biofabrication of ventricular myocardium for replacement of irreversibly damaged ischemic myocardium is sought after as a potential therapy for ischemic heart failure, though challenges in reliably producing this biomaterial have limited its clinical application. One method that shows promise for generation of functional myocardium is the use of tissue decellularization to serve as a scaffold for biofabrication. This review outlines the methods, materials, challenges, and prospects of tissue decellularization techniques for ventricular myocardium biofabrication. Decellularization aims to preserve the architecture and composition of the extracellular matrix of the tissue it is applied to, allowing for the subsequent implantation of stem cells of the desired cell type. Decellularization can be achieved with multiple reagents, most of which have detergent properties. A variety of cell types can be implanted in the resulting scaffold, including cardiac progenitor cells, and embryonic or induced pluripotent stem cells to generate a range of tissue, from patches to beating myocardium. The future of this biofabrication method will likely emphasize patient specific tissue engineering to generate complex 3-dimensional constructs that can replace dysfunctional cardiac structures.

A Bioprinting Process Supplemented with In Situ Electrical Stimulation Directly Induces Significant Myotube Formation and Myogenesis (Adv. Funct. Mater. 51/2021)

Bioprinting In article number 2105170, In-Bo Han, Geun Hyung Kim, and co-workers propose a bioprinting process, in situ electrical stimulation-enforced printing, and the approach is used to successfully align muscle tissue structures. The in situ stimulated adipose stem cells in the constructs fully remodel, restoring myofibers and muscle function with accelerated innervation/vascularization in the VML model. The biofabrication method could provide valuable insights into the current directions of 3D bioprinting.

Lithiophilic Carbon Nanofiber/Graphene Nanosheet Composite Scaffold Prepared by a Scalable and Controllable Biofabrication Method for Ultrastable Dendrite-Free Lithium-Metal Anodes.

Li metal is regarded as a promising anode for high-energy-density Li batteries, while the limited cycle life and fast capacity decay caused by notorious Li dendrite growth seriously impedes its application. Herein, a robust and highly lithiophilic bacterial cellulose-derived carbon nanofiber@reduced graphene oxide nanosheet (BC-CNF@rGO) composite scaffold is fabricated as a host for dendrite-free Li metal anode through an in situ biofabrication method. The abundant lithiophilic functional groups, conductive 3D network, and excellent mechanical property can effectively regulate uniform Li nucleation and deposition, enable fast reaction kinetics, and alleviate volume change. As a result, the BC-CNF@rGO skeleton achieves exceptional Li plating/stripping performance with a high average Coulombic efficiency of 98.3% over 800 cycles, and a long cycle life span of 5000h at 2mAcm-2 @1mAhcm-2 with a low overpotential of ≈15mV for lithium plating. Furthermore, full cells coupling BC-CNF@rGO-Li anode with LiFePO4 cathode achieves an unprecedented cycling stability with a long cycle life of 3000 cycles at 1C. This work sheds light on a promising material design and fabrication strategy for realizing high performance Li metal batteries.

A biofabrication method to align cells within bioprinted photocrosslinkable and cell-degradable hydrogel constructs via embedded fibers

The extracellular matrix (ECM) is composed of biochemical and biophysical cues that control cell behaviors and bulk mechanical properties. For example, anisotropy of the ECM and cell alignment are essential in the directional properties of tissues such as myocardium, tendon, and the knee meniscus. Technologies are needed to introduce anisotropic behavior into biomaterial constructs that can be used for the engineering of tissues as models and towards translational therapies. To address this, we developed an approach to align hydrogel fibers within cell-degradable bioink filaments with extrusion printing, where shear stresses during printing align fibers and photocrosslinking stabilizes the fiber orientation. Suspensions of hydrogel fibers were produced through the mechanical fragmentation of electrospun scaffolds of norbornene-modified hyaluronic acid, which were then encapsulated with meniscal fibrochondrocytes, mesenchymal stromal cells, or cardiac fibroblasts within gelatin-methacrylamide bioinks during extrusion printing into agarose suspension baths. Bioprinting parameters such as the needle diameter and the bioink flow rate influenced shear profiles, whereas the suspension bath properties and needle translation speed influenced filament diameters and uniformity. When optimized, filaments were formed with high levels of fiber alignment, which resulted in directional cell spreading during culture over one week. Controls that included bioprinted filaments without fibers or non-printed hydrogels of the same compositions either with or without fibers resulted in random cell spreading during culture. Further, constructs were printed with variable fiber and resulting cell alignment by varying print direction or using multi-material printing with and without fibers. This biofabrication technology advances our ability to fabricate constructs containing aligned cells towards tissue repair and the development of physiological tissue models.