An oxygenating colloidal bioink for the engineering of biomimetic tissue constructs

Ensuring a sufficient oxygen supply is pivotal for the success of bioprinting applications since it fosters tissue integration and natural regeneration.Variation in oxygen concentration among diverse tissues necessitates the precise recreation of tissue-specific oxygen levels in imprinted constructs to support the survival of targeted cells.Although oxygen-releasing biomaterials,such as oxygen-generating microparticles(OMPs),have shown promise for enhancing the oxygen supply of microenvironments in injured tissues,whether this approach is scalable for large tissues and whether tissue-specific bioinks with varying OMP concentrations remain printable remain unknown.This study addresses this critical gap by introducing an innovative class of engineered oxygenated bioinks that combine colloidal-based microgels with OMPs.We report that incorporating nanosized calcium peroxide(nCaO_(2))and manganese oxide nanosheets(nMnO_(2))into hydrophobic polymeric microparticles enables precise modulation of oxygen release while controlling hydrogen peroxide release.Moreover,the fabrication of oxygenating and cytocompatible colloidal gels is achieved using an aqueous two-phase system.This study thoroughly evaluates the fundamental characteristics of the resulting bioink,including its rheological behaviors,printability,shape fidelity,mechanical properties,and oxygen release properties.Moreover,this study demonstrates the macroscopic scalability and cytocompatibility of printed constructs produced via cell-laden oxygenating colloidal bioinks.By showcasing the effectiveness of extrusion-based bioprinting,this study underscores how it can be used to fabricate biomimetic tissues,indicating its potential for new applications.The findings presented here advance the bioprinting field by achieving scalability with both high cell viability and the possibility of mimicking specifically oxygenated tissues.This work thereby offers a promising avenue for the development of functional tissues with enhanced physiological relevance.

Ink-structing the future of vascular tissue engineering:a review of the physiological bioink design

Three-dimensional(3D)printing and bioprinting have come into view for a plannable and standardizable generation of implantable tissue-engineered constructs that can substitute native tissues and organs.These tissue-engineered structures are intended to integrate with the patient’s body.Vascular tissue engineering(TE)is relevant in TE because it supports the sustained oxygenization and nutrition of all tissue-engineered constructs.Bioinks have a specific role,representingthenecessarymedium for printability and vascular cell growth.This review aims to understand the requirements for the design of vascular bioinks.First,an in-depth analysis of vascular cell interaction with their native environment must be gained.A physiological bioink suitable for a tissue-engineered vascular graft(TEVG)must not only ensure good printability but also induce cells to behave like in a native vascular vessel,including self-regenerative and growth functions.This review describes the general structure of vascular walls with wall-specific cell and extracellular matrix(ECM)components and biomechanical properties and functions.Furthermore,the physiological role of vascular ECM components for their interaction with vascular cells and the mode of interaction is introduced.Diverse currently available or imaginable bioinks are described from physiological matrix proteins to nonphysiologically occurring but natural chemical compounds useful for vascular bioprinting.The physiological performance of these bioinks is evaluated with regard to biomechanical properties postprinting,with a view to current animal studies of 3D printed vascular structures.Finally,the main challenges for further bioink development,suitable bioink components to create a self-assembly bioink concept,and future bioprinting strategies are outlined.These concepts are discussed in terms of their suitability to be part of a TEVG with a high potential for later clinical use.

Visible Light-Induced 3D Bioprinting Technologies and Corresponding Bioink Materials for Tissue Engineering: A Review

Three-dimensional(3D)bioprinting based on traditional 3D printing is an emerging technology that is used to precisely assemble biocompatible materials and cells or bioactive factors into advanced tissue engineering solutions.Similar technology,particularly photo-cured bioprinting strategies,plays an important role in the field of tissue engineering research.The successful implementation of 3D bioprinting is based on the properties of photopolymerized materials.Photocrosslinkable hydrogel is an attractive biomaterial that is polymerized rapidly and enables process control in space and time.Photopolymerization is frequently initiated by ultraviolet(UV)or visible light.However,UV light may cause cell damage and thereby,affect cell viability.Thus,visible light is considered to be more biocompatible than UV light for bioprinting.In this review,we provide an overview of photo curing-based bioprinting technologies,and describe a visible light crosslinkable bioink,including its crosslinking mechanisms,types of visible light initiator,and biomedical applications.We also discuss existing challenges and prospects of visible light-induced 3D bioprinting devices and hydrogels in biomedical areas.

Advanced strategies in the application of gelatin-based bioink for extrusion bioprinting

The significance of bioink suitability for the extrusion bioprinting of tissue-like constructs cannot be overemphasized.Gelatin,derived from the hydrolysis of collagen,not only can mimic the extracellular matrix to immensely support cell function,but also is suitable for extrusion under certain conditions.Thus,gelatin has been recognized as a promising bioink for extrusion bioprinting.However,the development of a gelatin-based bioink with satisfactory printability and bioactivity to fabricate complex tissue-like constructs with the desired physicochemical properties and biofunctions for a specific biomedical application is still in its infancy.Therefore,in this review,we aim to comprehensively summarize the state-of-the-art methods of gelatin-based bioink application for extrusion bioprinting.Wefirstly outline the properties and requirements of gelatin-based bioinks for extrusion bioprinting,highlighting the strategies to overcome their main limitations in terms of printability,structural stability and cell viability.Then,the challenges and prospects are further discussed regarding the development of ideal gelatin-based bioinks for extrusion bioprinting to create complex tissue-like constructs with preferable physicochemical properties and biofunctions.

Trends on physical understanding of bioink printability

Recently, 3D bioprinting is developed as an emerging approach, increasingly applied to materials for healthcare;while, the precise placement of cells and materials, and the shape fidelity of forming constructs is of great importance for successful application of 3D bioprinting. Research efforts have been made to develop new bioinks as "raw materials" with better biocompatibility and biofunctionality, but the printability of bioinks is largely ignored and still needs to be carefully examined to enable robotic bioprinting. This article aims to introduce a recent published review (Appl. Phys. Rev. 2018, 5, 041304) on the evaluation of bioink printability by Huang's research group from University of Florida. Huang et al. comprehensively reviewed the bioink printability based on the physical point of view during inkjet printing, laser printing, and microextrusion, and a series of self-consistent time scales and dimensi on less quantities were utilized to physically understand and evaluate bioink printability. This article would be helpful to know the trends on physical understanding of bioink printability.

Polyvinylpyrrolidone-based bioink:influence of bioink properties on printing performance and cell proliferation during inkjet-based bioprinting

Among the different bioprinting techniques,the drop-on-demand(DOD)jetting-based bioprinting approach facilitates contactless deposition of pico/nanoliter droplets ofmaterials and cells for optimal cell–matrix and cell–cell interactions.Although bioinks play a critical role in the bioprinting process,there is a poor understanding of the influence of bioink properties on printing performance(such as filament elongation,formation of satellite droplets,and droplet splashing)and cell health(cell viability and proliferation)during the DOD jetting-based bioprinting process.An inert polyvinylpyrrolidone(PVP360,molecular weight=360 kDa)polymerwas used in this study to manipulate the physical properties of the bioinks and investigate the influence of bioink properties on printing performance and cell health.Our experimental results showed that a higher bioink viscoelasticity helps to stabilize droplet filaments before rupturing from the nozzle orifice.The highly stretched droplet filament resulted in the formation of highly aligned“satellite droplets,”which minimized the displacement of the satellite droplets away from the predefined positions.Next,a significant increase in the bioink viscosity facilitated droplet deposition on the wetted substrate surface in the absence of splashing and significantly improved the accuracy of the deposited main droplet.Further analysis showed that cell-laden bioinks with higher viscosity exhibited higher measured average cell viability(%),as the presence of polymer within the printed droplets provides an additional cushioning effect(higher energy dissipation)for the encapsulated cells during droplet impact on the substrate surface,improves the measured average cell viability even at higher droplet impact velocity and retains the proliferation capability of the printed cells.Understanding the influence of bioink properties(e.g.,bioink viscoelasticity and viscosity)on printing performance and cell proliferation is important for the formulation of new bioinks,and we have demonstrated precise DOD deposition of living cells and fabrication of tunable cell spheroids(nL–μL range)using multiple types of cells in a facile manner.

Functionalized alginate-based bioinks for microscale electrohydrodynamic bioprinting of living tissue constructs with improved cellular spreading and alignment

Bioprinting has been widely investigated for tissue engineering and regenerative medicine applications.However,it is still difficult to reconstruct the complex native cell arrangement due to the limited printing resolution of conventional bioprinting techniques such as extrusion-and inkjet-based printing.Recently,an electrohydrodynamic(EHD)bioprinting strategy was reported for the precise deposition of well-organized cell-laden constructs with microscale filament size,whereas few studies have been devoted to developing bioinks that can be applied for EHD bioprinting and simultaneously support cell spreading.This study describes functionalized alginate-based bioinks for microscale EHD bioprinting using peptide grafting and fibrin incorporation,which leads to high cell viability(>90%)and cell spreading.The printed filaments can be further refined to as small as 30μm by incorporating polyoxyethylene and remained stable over one week when exposed to an aqueous environment.By utilizing the presented alginate-based bioinks,layer-specific cell alignment along the printing struts could be observed inside the EHD-printed microscale filaments,which allows fabricating living constructs with cell-scale filament resolution for guided cellular orientation.

The use of machine learning to predict the effects of cryoprotective agents on the GelMA-based bioinks used in extrusion cryobioprinting

Cryobioprinting has tremendous potential to solve problems to do with lack of shelf availability in traditional bioprinting by combining extrusion bioprinting and cryopreservation.In order to ensure the viability of cells in the frozen state and avoid the possible toxicity of dimethyl sulfoxide(DMSO),DMSO-free bioink design is critical for achieving successful cryobioprinting.A nontoxic gelatin methacryloyl-based bioink used in cryobioprinting is composed of cryoprotective agents(CPAs)and a buffer solution.The selection and ratio of CPAs in the bioink directly affect the survival of cells in the frozen state.However,the development of universal and efficient cryoprotective bioinks requires extensive experimentation.We first compared two commonly used CPA formulations via experiments in this study.Results show that the effect of using ethylene glycol as the permeable CPA was 6.07%better than that of glycerol.Two datasets were obtained and four machinelearning models were established to predict experimental outcomes.The predictive powers of multiple linear regression(MLR),decision tree(DT),random forest(RF),and artificial neural network(ANN)approaches were compared,suggesting an order of ANN>RF>DT>MLR.The final selected ANN model was then applied to another dataset.Results reveal that this machine-learning method can accurately predict the effects of cryoprotective bioinks composed of different CPAs.Outcomes also suggest that the formulations presented here have universality.Our findings are likely to greatly accelerate research and development on the use of bioinks for cryobioprinting.

One-pot synthesis of hydroxyapatite hybrid bioinks for digital light processing 3D printing in bone regeneration

Three-dimensional(3D)bioprinting has revolutionized tissue engineering by enabling precise fabrication with bioinks.Among these techniques,digital light processing(DLP)stands out due to its exceptional resolution,speed,and biocompatibility.However,the progress of DLP is hindered by the limited availability of suitable bioinks.Currently,some studies involve simple mixing of different materials,resulting in bioinks that lack uniformity and photopolymerization characteristics.To address this challenge,we present an innovative one-pot synthesis method for bioinks based on methacrylated gelatin/alginate with hydroxyapatite(HAP).This approach offers significant advantages in terms of efficiency and uniformity.The synthesized bioinks demonstrate excellent printability,stability,and notably enhanced mechanical properties,facilitating optimal in vitro compatibility.Additionally,the HAP-hybrid bioinks printed scaffolds demonstrated impressive bone repair capabilities in vivo compared with pure organic bioinks.In conclusion,the Gel/Alg/HAP bioinks presented herein offer an innovative solution for DLP bioprinting within the field of bone tissue engineering.Their multifaceted advantages help overcome the limitations of restricted bioink choices,pushing forward the boundaries of bioprinting technology and contributing to the progress of regenerative medicine and tissue engineering.

Bioinspired coacervate-based bioinks for construction of multiscale tissue engineering scaffolds

Engineering hydrogels that resemble biological tissues of various lengths via conventional fabrication techniques remains challenging.Three-dimensional(3D)bioprinting has emerged as an advanced approach for constructing complex biomimetic 3D architectures,which are currently restricted by the limited number of available bioinks with high printability,biomimicry,biocompatibility,and proper mechanical properties.Inspired by ubiquitous coacervation phenomena in biology,we present a unique mineral-biopolymer coacervation strategy that enables the hierarchical assembly of nanoclay and recombinant human collagen(RHC).This system was observed to undergo a coacervation transition(liquid‒liquid phase separation)spontaneously.The formed dense phase separated from its supernatant is the coacervate of clay-RHC-rich complexes,where polymer chains are sandwiched between silicate layers.Molecular dynamics simulation was first used to verify and explore the coacervation process.Then,the coacervates were demonstrated to be potential bioinks that exhibited excellent self-supporting and shear-thinning viscoelastic properties.Through extrusion-based printing,the versatility of the bioink was demonstrated by reconstructing the key features of several biological tissues,including multilayered lattice,vascular,nose,and ear-like structures,without the need for precrosslinking operations or support baths.Furthermore,the printed scaffolds were cytocompatible,elicited minimal inflammatory responses,and promoted bone regeneration in calvarial defects.

Bioprinting and regeneration of auricular cartilage using a bioactive bioink based on microporous photocrosslinkable acellular cartilage matrix

Tissue engineering provides a promising strategy for auricular reconstruction.Although the first international clinical breakthrough of tissue-engineered auricular reconstruction has been realized based on polymer scaffolds,this approach has not been recognized as a clinically available treatment because of its unsatisfactory clinical efficacy.This is mainly since reconstruction constructs easily cause inflammation and deformation.In this study,we present a novel strategy for the development of biological auricle equivalents with precise shapes,low immunogenicity,and excellent mechanics using auricular chondrocytes and a bioactive bioink based on biomimetic microporous methacrylate-modified acellular cartilage matrix(ACMMA)with the assistance of gelatin methacrylate(GelMA),poly(ethylene oxide)(PEO),and polycaprolactone(PCL)by integrating multi-nozzle bioprinting technology.Photocrosslinkable ACMMA is used to emulate the intricacy of the cartilage-specific microenvironment for active cellular behavior,while GelMA,PEO,and PCL are used to balance printability and physical properties for precise structural stability,form the microporous structure for unhindered nutrient exchange,and provide mechanical support for higher shape fidelity,respectively.Finally,mature auricular cartilage-like tissues with high morphological fidelity,excellent elasticity,abundant cartilage lacunae,and cartilage-specific ECM deposition are successfully regenerated in vivo,which provides new opportunities and novel strategies for the fabrication and regeneration of patient-specific auricular cartilage.

Biomaterials / bioinks and extrusion bioprinting

Bioinks are formulations of biomaterials and living cells, sometimes with growth factors or other biomolecules, while extrusion bioprinting is an emerging technique to apply or deposit these bioinks or biomaterial solutions to create three-dimensional (3D) constructs with architectures and mechanical/biological properties that mimic those of native human tissue or organs. Printed constructs have found wide applications in tissue engineering for repairing or treating tissue/organ injuries, as well as in vitro tissue modelling for testing or validating newly developed therapeutics and vaccines prior to their use in humans. Successful printing of constructs and their subsequent applications rely on the properties of the formulated bioinks, including the rheological, mechanical, and biological properties, as well as the printing process. This article critically reviews the latest developments in bioinks and biomaterial solutions for extrusion bioprinting, focusing on bioink synthesis and characterization, as well as the influence of bioink properties on the printing process. Key issues and challenges are also discussed along with recommendations for future research.

Tailoring bioinks of extrusion-based bioprinting for cutaneous wound healing

Extrusion-based bioprinting (EBB) holds potential for regenerative medicine. However, the widely-used bioinks of EBB exhibit some limitations for skin regeneration, such as unsatisfactory bio-physical (i.e., mechanical, structural, biodegradable) properties and compromised cellular compatibilities, and the EBB-based bioinks with therapeutic effects targeting cutaneous wounds still remain largely undiscussed. In this review, the printability considerations for skin bioprinting were discussed. Then, current strategies for improving the physical properties of bioinks and for reinforcing bioinks in EBB approaches were introduced, respectively. Notably, we highlighted the applications and effects of current EBB-based bioinks on wound healing, wound scar formation, vasculari-zation and the regeneration of skin appendages (i.e., sweat glands and hair follicles) and discussed the challenges and future perspectives. This review aims to provide an overall view of the applications, challenges and promising solutions about the EBB-based bioinks for cutaneous wound healing and skin regeneration.

3D-printable supramolecular hydrogels with shear-thinning property: fabricating strength tunable bioink via dual crosslinking

3-dimensional(3D)bioprinting technology provides promising strategy in the fabrication of artificial tissues and organs.As the fundamental element in bioprinting process,preparation of bioink with ideal mechanical properties without sacrifice of biocompatibility is a great challenge.In this study,a supramolecular hydrogel-based bioink is prepared by polyethylene glycol(PEG)grafted chitosan,α-cyclodextrin(α-CD)and gelatin.It has a primary crosslinking structure through the aggregation of the pseudo-polyrotaxane-like side chains,which are formed from the host-guest interactions betweenα-CD and PEG side chain.Apparent viscosity measurement shows the shear-shinning property of this bioink,which might be due to the reversibility of the physical crosslinking.Moreover,withβ-glycerophosphate at different concentrations as the secondary crosslinking agent,the printed constructs demonstrate different Young's modulus(p<0.001).They could also maintain the Young's modulus in cell culture condition for at least 21 days(p<0.05).By co-culturing each component with fibroblasts,CCK-8 assay demonstrate cellular viability is higher than 80%.After bioprinting and culturing,immunofluorescence staining with quantification indicate the expression of Ki-67,Paxillin,and N-cadherin is higher in day 14 than those in day 3(p<0.05).Oil red O and Nissl body specific staining reflect strength tunable bioink may have impact on the cell fate of mesenchymal stem cells(p<0.05).This work might provide new idea for advanced bioink in the application of re-establishing complicated tissues and organs.

3D Printing Bioink Preparation and Application in Cartilage Tissue Reconstruction in Vitro

Three-dimensional(3D)bioprinting technology has great potential for application in the treatment of cartilage defects.However,the preparation of biocompatible and stable bioinks is still a major challenge.In this study,decellularized extracellular matrix(dECM)of soft tissue was used as the basic material to prepare the bioink.Our results showed that this novel dECM-derived bioink had good printing performance and comprised a large number of fine nanofibers.Biological characterization revealed that the bioink was compatible with the growth of chondrocytes and that the nanofibrous structure greatly promoted cell proliferation.Histological and immunohistochemical analyses showed that the in vitro printed cartilage displayed the presence of characteristic cartilage lacunae.Thus,a new preparation method for dECM-derived bioink with potential application in generation of cartilage was developed in this study.

A structure-supporting,self-healing,and high permeating hydrogel bioink for establishment of diverse homogeneous tissue-like constructs

The ready-to-use,structure-supporting hydrogel bioink can shorten the time for ink preparation,ensure cell dispersion,and maintain the preset shape/microstructure without additional assistance during printing.Meanwhile,ink with high permeability might facilitate uniform cell growth in biological constructs,which is beneficial to homogeneous tissue repair.Unfortunately,current bioinks are hard to meet these requirements simultaneously in a simple way.Here,based on the fast dynamic crosslinking of aldehyde hyaluronic acid(AHA)/N-carboxymethyl chitosan(CMC)and the slow stable crosslinking of gelatin(GEL)/4-arm poly(ethylene glycol)succinimidyl glutarate(PEG-SG),we present a time-sharing structure-supporting(TSHSP)hydrogel bioink with high permeability,containing 1%AHA,0.75%CMC,1%GEL and 0.5%PEG-SG.The TSHSP hydrogel can facilitate printing with proper viscoelastic property and self-healing behavior.By crosslinking with 4%PEG-SG for only 3 min,the integrity of the cell-laden construct can last for 21 days due to the stable internal and external GEL/PEG-SG networks,and cells manifested long-term viability and spreading morphology.Nerve-like,muscle-like,and cartilage-like in vitro constructs exhibited homogeneous cell growth and remarkable biological specificities.This work provides not only a convenient and practical bioink for tissue engineering,targeted cell therapy,but also a new direction for hydrogel bioink development.

A facile,versatile hydrogel bioink for 3D bioprinting benefits long-term subaqueous fidelity,cell viability and proliferation

Both of the long-term fidelity and cell viability of three-dimensional(3D)-bioprinted constructs are essential to precise soft tissue repair.However,the shrinking/swelling behavior of hydrogels brings about inadequate long-term fidelity of constructs,and bioinks containing excessive polymer are detrimental to cell viability.Here,we obtained a facile hydrogel by introducing 1%aldehyde hyaluronic acid(AHA)and 0.375%N-carboxymethyl chitosan(CMC),two polysaccharides with strong water absorption and water retention capacity,into classic gelatin(GEL,5%)–alginate(ALG,1%)ink.This GEL–ALG/CMC/AHA bioink possesses weak temperature dependence due to the Schiff base linkage of CMC/AHA and electrostatic interaction of CMC/ALG.We fabricated integrated constructs through traditional printing at room temperature and in vivo simulation printing at 37C.The printed cell-laden constructs can maintain subaqueous fidelity for 30 days after being reinforced by 3%calcium chloride for only 20 s.Flow cytometry results showed that the cell viability was 91.3861.55%on day 29,and the cells in the proliferation plateau at this time still maintained their dynamic renewal with a DNA replication rate of 6.0661.24%.This work provides a convenient and practical bioink option for 3D bioprinting in precise soft tissue repair.

Collagen-based bioinks for regenerative medicine: Fabrication, application and prospective

In the field of regenerative medicine,the importance of 3D bioprinting is self-evident and nonnegligible.However,3D bioprinting technology also requires bioink with excellent performance as support material to fabricate a multi-functional bioinspired scaffold.Collagen-based bioink is regarded as an ideal 3D bioprinting ink for its excellent biocompatibility,controllable printability and cell loading property.It is an important breakthrough in regenerative medicine with the progress of collagen-based bioink,which fabricates bioinspired scaffolds with different functions and is applied in different repair scenarios.This review summarizes the different applications of collagen-based bioink and classifies them as soft tissue and hard tissue according to the target region.The applications of target region in soft tissues include skin,cartilage,heart and blood vessels,while in hard tissues include femur,skull,teeth and spine.When the collagen-based bioink is applied in repairing soft tissue,the requirements of function are higher,while the mechanical properties must be further improved in repairing hard tissue.We further summarize the characteristics of collagen-based bioink and point out the most important properties that should be considered in different repair scenarios,which can provide reference for the preparation of bioinks with different functions.Finally,we point out the main challenges faced by collagen-based bioink and prospect the future research directions.

Stem cell-laden hydrogel bioink for generation of high resolution and fidelity engineered tissues with complex geometries

Recently,3D bioprinting has been explored as a promising technology for biomedical applications with the potential to create complex structures with precise features.Cell encapsulated hydrogels composed of materials such as gelatin,collagen,hyaluronic acid,alginate and polyethylene glycol have been widely used as bioinks for 3D bioprinting.However,since most hydrogel-based bioinks may not allow rapid stabilization immediately after 3D bioprinting,achieving high resolution and fidelity to the intended architecture is a common challenge in 3D bioprinting of hydrogels.In this study,we have utilized shear-thinning and self-healing ionically crosslinked oxidized and methacrylated alginates(OMAs)as a bioink,which can be rapidly gelled by its self-healing property after bioprinting and further stabilized via secondary crosslinking.It was successfully demonstrated that stem cell-laden calcium-crosslinked OMA hydrogels can be bioprinted into complicated 3D tissue structures with both high resolution and fidelity.Additional photocrosslinking enables long-term culture of 3D bioprinted constructs for formation of functional tissue by differentiation of encapsulated human mesenchymal stem cells.

Harnessing decellularised extracellular matrix microgels into modular bioinks for extrusion-based bioprinting with good printability and high post-printing cell viability

The printability of bioink and post-printing cell viability is crucial for extrusion-based bioprinting.A proper bioink not only provides mechanical support for structural fidelity,but also serves as suitable three-dimensional(3D)microenvironment for cell encapsulation and protection.In this study,a hydrogel-based composite bioink was developed consisting of gelatin methacryloyl(GelMA)as the continuous phase and decellularised extracellular matrix microgels(DMs)as the discrete phase.A flow-focusing microfluidic system was employed for the fabrication of cell-laden DMs in a high-throughput manner.After gentle mixing of the DMs and GelMA,both rheological characterisations and 3D printing tests showed that the resulting DM-GelMA hydrogel preserved the shear-thinning nature,mechanical properties,and good printability from GelMA.The integration of DMs not only provided an extracellular matrix-like microenvironment for cell encapsulation,but also considerable shear-resistance for high post-printing cell viability.The DM sizes and inner diameters of the 3D printer needles were correlated and optimised for nozzle-based extrusion.Furthermore,a proof-of-concept bioink composedg of RSC96 Schwann cells encapsulated DMs and human umbilical vein endothelial cell-laden GelMA was successfully bioprinted into 3D constructs,resulting in a modular co-culture system with distinct cells/materials distribution.Overall,the modular DM-GelMA bioink provides a springboard for future precision biofabrication and will serve in numerous biomedical applications such as tissue engineering and drug screening.