Electrospinning/3D printing drug-loaded antibacterial polycaprolactone nanofiber/sodium alginate-gelatin hydrogel bilayer scaffold for skin wound repair.

Skin injuries and defects, as a common clinical issue, still cannot be perfectly repaired at present, particularly large-scale and infected skin defects. Therefore, in this work, a drug-loaded bilayer skin scaffold was developed for repairing full-thickness skin defects. Briefly, amoxicillin (AMX) was loaded on polycaprolactone (PCL) nanofiber via electrospinning to form the antibacterial nanofiber membrane (PCL-AMX) as the outer layer of scaffold to mimic epidermis. To maintain wound wettability and promote wound healing, external human epidermal growth factor (rhEGF) was loaded in sodium alginate-gelatin to form the hydrogel structure (SG-rhEGF) via 3D printing as inner layer of scaffold to mimic dermis. AMX and rhEGF were successfully loaded into the scaffold. The scaffold exhibited excellent physicochemical properties, with elongation at break and tensile modulus were 102.09 ±â€¯6.74 % and 206.83 ±â€¯32.10 kPa, respectively; the outer layer was hydrophobic (WCA was 112.09 ±â€¯4.67°), while the inner layer was hydrophilic (WCA was 48.87 ±â€¯5.52°). Meanwhile, the scaffold showed excellent drug release and antibacterial characteristics. In vitro and in vivo studies indicated that the fabricated scaffold could enhance cell adhesion and proliferation, and promote skin wound healing, with favorable biocompatibility and great potential for skin regeneration and clinical application.

A 5 + 1-Axis 3D Printing Platform for Producing Customized Intestinal Fistula Stents.

Tailored intestinal fistula stents with a hollow bent pipe structure prepared by using a three-axis bio-printing platform are often unsuitable due to low printing efficiency and quality caused by the unavoidable need for a supporting structure. Herein, a 5 + 1-axis 3D printing platform was built and developed for producing support-free intestinal fistula stents. A 3D model of the target stent shape and dimensions was treated by a dynamic slicing algorithm, which was then used to prepare a motion control code. Our printing method showed improved printing efficiency, superior stent surface properties and structure and ideal elasticity and mechanical strength to meet the mechanical requirements of the human body. Static simulations showed the importance of axial printing techniques, whereas the stent itself was shown to have excellent biocompatibility with wettability and cell proliferation tests. We present a customizable, efficient, and high-quality method with the potential for preparing bespoke stents for treating intestinal fistulas.

3D Bioprinting tissue analogs: Current development and translational implications.

Three-dimensional (3D) bioprinting is a promising and rapidly evolving technology in the field of additive manufacturing. It enables the fabrication of living cellular constructs with complex architectures that are suitable for various biomedical applications, such as tissue engineering, disease modeling, drug screening, and precision regenerative medicine. The ultimate goal of bioprinting is to produce stable, anatomically-shaped, human-scale functional organs or tissue substitutes that can be implanted. Although various bioprinting techniques have emerged to develop customized tissue-engineering substitutes over the past decade, several challenges remain in fabricating volumetric tissue constructs with complex shapes and sizes and translating the printed products into clinical practice. Thus, it is crucial to develop a successful strategy for translating research outputs into clinical practice to address the current organ and tissue crises and improve patients' quality of life. This review article discusses the challenges of the existing bioprinting processes in preparing clinically relevant tissue substitutes. It further reviews various strategies and technical feasibility to overcome the challenges that limit the fabrication of volumetric biological constructs and their translational implications. Additionally, the article highlights exciting technological advances in the 3D bioprinting of anatomically shaped tissue substitutes and suggests future research and development directions. This review aims to provide readers with insight into the state-of-the-art 3D bioprinting techniques as powerful tools in engineering functional tissues and organs.

Bioinks for Space Missions: The Influence of Long-Term Storage of Alginate-Methylcellulose-Based Bioinks on Printability as well as Cell Viability and Function.

Bioprinting is considered a key technology for future space missions and is currently being established on the International Space Station (ISS). With the aim to perform bioink production as a critical and resource-consuming preparatory step already on Earth and transport a bioink cartridge "ready to use" to the ISS, the storability of bioinks is investigated. Hydrogel blends based on alginate and methylcellulose are laden with either green microalgae of the species Chlorella vulgaris or with different human cell lines including immortilized human mesenchymal stem cells, SaOS-2 and HepG2, as well as with primary human dental pulp stem cells. The bioinks are filled into printing cartridges and stored at 4°C for up to four weeks. Printability of the bioinks is maintained after storage. Viability and function of the cells embedded in constructs bioprinted from the stored bioinks are investigated during subsequent cultivation: The microalgae survive the storage period very well and show no loss of growth and functionality, however a significant decrease is visible for human cells, varying between the different cell types. The study demonstrates that storage of bioinks is in principle possible and is a promising starting point for future research, making complex printing processes more effective and reproducible.

Cellular adhesion and chondrogenic differentiation inside an alginate-based bioink in response to tailorable artificial matrices and tannic acid treatment.

Many established bioinks fulfill important requirements regarding fabrication standards and cytocompatibility. Current research focuses on development of functionalized bioinks with an improved support of tissue-specific cell differentiation. Many approaches primarily depend on decellularized extracellular matrices or blood components. In this study, we investigated the combination of a highly viscous alginate-methylcellulose (algMC) bioink with collagen-based artificial extracellular matrix (aECM) as a finely controllable and tailorable system composed of collagen type I (col) with and without chondroitin sulfate (CS) or sulfated hyaluronan (sHA). As an additional stabilizer, the polyphenol tannic acid (TA) was integrated into the inks. The assessment of rheological properties and printability as well as hydrogel microstructure revealed no adverse effect of the integrated components on the inks. Viability, adhesion, and proliferation of bioprinted immortalized human mesenchymal stem cells (hTERT-MSC) was improved indicating enhanced interaction with the designed microenvironment. Furthermore, chondrogenic matrix production (collagen type II and sulfated glycosaminoglycans) by primary human chondrocytes (hChon) was enhanced by aECM. Supplementing the inks with TA was required for these positive effects but caused cytotoxicity as soon as TA concentrations exceeded a certain amount. Thus, combining tailorable aECM with algMC and balanced TA addition proved to be a promising approach for promoting adhesion of immortalized stem cells and differentiation of chondrocytes in bioprinted scaffolds.

Egg white improves the biological properties of an alginate-methylcellulose bioink for 3D bioprinting of volumetric bone constructs.

Three-dimensional microextrusion bioprinting has attracted great interest for fabrication of hierarchically structured, functional tissue substitutes with spatially defined cell distribution. Despite considerable progress, several significant limitations remain such as a lack of suitable bioinks which combine favorable cell response with high shape fidelity. Therefore, in this work a novel bioink of alginate-methylcellulose (AlgMC) blend functionalized with egg white (EW) was developed with the aim of solving this limitation. In this regard, a stepwise strategy was proposed to improve and examine the cell response in low-viscosity alginate inks (3%, w/v) with different EW concentrations, and in high-viscosity inks after gradual methylcellulose addition for enhancing printability. The rheological properties and printability of these cell-responsive bioinks were characterized to obtain an optimized formulation eliciting balanced physicochemical and biological properties for fabrication of volumetric scaffolds. The bioprinted AlgMC + EW constructs exhibited excellent shape fidelity while encapsulated human mesenchymal stem cells showed high post-printing viability as well as adhesion and spreading within the matrix. In a proof-of-concept experiment, the impact of these EW-mediated effects on osteogenesis of bioprinted primary human pre-osteoblasts (hOB) was evaluated. Results confirmed a high viability of hOB (93.7 ± 0.15%) post-fabrication in an EW-supported AlgMC bioink allowing cell adhesion, proliferation and migration. EW even promoted the expression of osteogenic genes, coding for bone sialoprotein (integrin binding sialoprotein/bone sialoprotein precursor (IBSP)) and osteocalcin (BGLAP) on mRNA level. To demonstrate the suitability of the novel ink for future fabrication of multi-zonal bone substitutes, AlgMC + EW was successfully co-printed together with a pasty calcium phosphate bone cement biomaterial ink to achieve a partly mineralized 3D volumetric environment with good cell viability and spreading. Along with the EW-mediated positive effects within bioprinted AlgMC-based scaffolds, this highlighted the promising potential of this novel ink for biofabrication of bone tissue substitutes in clinically relevant dimensions.

3D Printing GelMA/PVA Interpenetrating Polymer Networks Scaffolds Mediated with CuO Nanoparticles for Angiogenesis.

Biocompatible hydrogels have been considered one of the most well-known and promising in various materials used in the fabrication of tissue-engineering scaffolds. Although considerable progress has been made in recent decades, many limitations remain, such as poor mechanical and degradation properties of biomaterials. In addition, vascularization of tissue-engineering scaffold is an enduring challenge, which limited the fabrication and application of scaffold with clinically relevant dimension. To cover these challenges, in this work, a novel nanocomposite interpenetrating polymer networks (IPN) hydrogel scaffold consists of methacrylated gelatin (GelMA), poly(vinyl alcohol) (PVA), and copper oxide nanoparticles (CuONPs) is fabricated by extrusion-based 3D printing. A series of physiochemical and biological characterizations of the nanocomposite GelMA/PVA scaffolds are performed. Results showed that the mechanical and degradation properties of the nanocomposite GelMA/PVA scaffolds are obviously improved compared to GelMA scaffolds with single network. In vitro cell experiments and chick embryo angiogenesis (CEA) assay confirmed good cytocompatibility of the fabricated scaffold and its potential to promote cell migration and angiogenesis. In conclusion, altogether the results demonstrated that GelMA/PVA IPN scaffolds modified with CuONPs have great potential for fabrication of volumetric scaffolds and promote angiogenesis during tissue growth and repair.

Designing Double-Layer Multimaterial Composite Patch Scaffold with Adhesion Resistance for Hernia Repair.

Hernia repair mesh is associated with a number of complications, including adhesions and limited mobility, due to insufficient mechanical strength and nonresorbability. Among them, visceral adhesions are one of the most serious complications of patch repair. In this study, a degradable patch with an antiadhesive layer is prepared for hernia repair by 3D printing and electrospinning techniques using polycaprolactone, polyvinyl alcohol, and soybean peptide (SP). The study into the physicochemical properties of the patch is found that it has adequate mechanical strength requirements (16 N cm-1 ) and large elongation at break, which are superior than commercial polypropylene patches. In vivo and in vitro experiments show that human umbilical vein endothelial cells proliferated well on composite patches, and showed excellent biocompatibility with the host and little adhesion through a rat abdominal wall defect model. In conclusion, the results of this study show that composite patch can effectively reduce the occurrence of adhesions, while the addition of SP in the patch further enhances its biocompatibility. It is believed that a regenerative biological patch with great potential in hernia repair provides a new strategy for the development of new biomimetic biodegradable patches.

Alginate/Gelatin-Based Hydrogel with Soy Protein/Peptide Powder for 3D Printing Tissue-Engineering Scaffolds to Promote Angiogenesis.

In recent years, 3D bioprinting has attracted broad research interest in biomedical engineering and clinical applications. However, there are two issues need to be solved urgently at present, the development of ink is the first pressing thing for 3D printing tissue engineering scaffold, other thing is the promotion of angiogenesis in the scaffold. Therefore, a gelatin/sodium alginate-based hydrogel with protein-rich is developed here, which is prepared by gelatin, sodium alginate, and soy protein/soy peptide powder. The prepared inks exhibit excellent shear-thinning behavior, which contribute to extrusion-based printing; also shown good crosslinking ability by calcium chloride. The macroporous composite scaffolds are printed by 3D printing using the developed ink and the physicochemical properties of the scaffolds are evaluated. Moreover, the cytocompatibility of printed scaffold is characterized by using human umbilical vein epidermal cells, results show that the scaffolds with soy protein and soy peptide powder can promote cell attach, spread, migration, and proliferation. The further research of chicken embryo allantoic membrane assay and animal experiment are carried, and results present that the scaffold can promote the growth of neo-vessels in the scaffold, which means the developed ink with soy protein and soy peptide powder has great potential for angiogenesis.

3D printing of self-standing and vascular supportive multimaterial hydrogel structures for organ engineering.

Three dimensional printable formulation of self-standing and vascular-supportive structures using multi-materials suitable for organ engineering is of great importance and highly challengeable, but, it could advance the 3D printing scenario from printable shape to functional unit of human body. In this study, the authors report a 3D printable formulation of such self-standing and vascular-supportive structures using an in-house formulated multi-material combination of albumen/alginate/gelatin-based hydrogel. The rheological properties and relaxation behavior of hydrogels were analyzed before the printing process. The suitability of the hydrogel in 3D printing of various customizable and self-standing structures, including a human ear model, was examined by extrusion-based 3D printing. The structural, mechanical, and physicochemical properties of the printed scaffolds were studied systematically. Results supported the 3D printability of the formulated hydrogel with self-standing structures, which are customizable to a specific need. In vitro cell experiment showed that the formulated hydrogel has excellent biocompatibility and vascular supportive behavior with the extent of endothelial sprout formation when tested with human umbilical vein endothelial cells. In conclusion, the present study demonstrated the suitability of the extrusion-based 3D printing technique for manufacturing complex shapes and structures using multi-materials with high fidelity, which have great potential in organ engineering.

Adipose-Derived Stem Cells Based on Electrospun Biomimetic Scaffold Mediated Endothelial Differentiation Facilitating Regeneration and Repair of Abdominal Wall Defects via HIF-1α/VEGF Pathway.

Application of synthetic or biological meshes is the main therapy for the repair and reconstruction of abdominal wall defects, a common disease in surgery. Currently, no ideal materials are available, and there is an urgent need to find appropriate ones to satisfy clinical needs. Electrospun scaffolds have drawn attention in soft tissue reconstruction. In this study, we developed a novel method to fabricate a composite electrospun scaffold using a thermoresponsive hydrogel, poly (N-isopropylacrylamide)-block-poly (ethylene glycol), and a biodegradable polymer, polylactic acid (PLA). This scaffold provided not only a high surface area/volume ratio and a three-dimensional fibrous matrix but also high biocompatibility and sufficient mechanical strength, and could simulate the native extracellular matrix and accelerate cell adhesion and proliferation. Furthermore, rat adipose-derived stem cells (ADSCs) were seeded in the composite electrospun scaffold to enhance the defect repair and regeneration by directionally inducing ADSCs into endothelial cells. In addition, we found early vascularization in the process was regulated by the hypoxia inducible factor-1α (HIF-1α)/vascular endothelial growth factor (VEGF) pathway. In our study, overexpression of HIF-1α/VEGF in ADSCs using a lentivirus system promoted early vascularization in the electrospun scaffolds. Overall, we expect our composite biomimetic scaffold method will be applicable and useful in abdominal wall defect regeneration and repair in the future.

Topological Structure Design and Fabrication of Biocompatible PLA/TPU/ADM Mesh with Appropriate Elasticity for Hernia Repair.

The meshes for hernia repair result in many problems that are related to complications including chronic pain and limited movement due to inadequate mechanical strength, non-absorbability, or low elasticity. In this study, degradable polylactic acid (PLA), synthetic thermoplastic polyurethane (TPU), and acellular dermal matrix (ADM) powders are combined to prepare a novel PLA/TPU/ADM mesh with three different topological structures (square, circular, and diamond) by 3D printing. The physicochemical properties and structural characteristics of mesh are studied, the results show that the diamond structure mesh with the pore size of 3 mm has sufficient elasticity and tensile strength, which provides the efficient mechanical strength required for hernia repair (16 N cm-1 ) and the value more than polypropylene(PP) mesh. Besides, in vitro and in vivo experiments demonstrate human umbilical vein endothelial cells could successfully proliferate on the PLA/TPU/ADM mesh whose biocompatibility with the host is shown using a rat model of abdominal wall defect. In conclusion, the results of this study demonstrate that the PLA/TPU/ADM mesh may be considered a good choice for hernia repair as its potential to overcome the elastic and strength challenges associated with a highly flexible abdominal wall, as well as its good biocompatibility.

An integrated strategy for designing and fabricating triple-layer vascular graft with oriented microgrooves to promote endothelialization.

Compared with native blood vessels and existing vascular grafts, design and manufacture of vascular grafts with a three-dimensional topological structure is a key to induce cells and tissue growth, which remains an essential issue in both tissue engineering and regenerative medicine. This study sought to develop a novel triple-layer vascular graft (TLVG) with oriented microgrooves to investigate the mechanical property and endothelialization. The TLVGs were composed of electrospun Poly-ε-caprolactone (PCL)/thermoplastic polyurethane (TPU) as the inner layer, albumen/sodium alginate (SA) hydrogel as the middle layer, and electrospun PCL/TPU as the outer layer. In detail, a cylindrical sacrificial template was designed and printed using polyvinyl alcohol (PVA), served as the electrospinning receiving platform to form the oriented microgrooves in the inner layer of TLVGs. The highly elastic albumen/SA hydrogel and PCL/TPU nanofibers were able to simulate the elastin in blood vessels. In addition, the introduction of the albumen/SA hydrogel layer not only solves the leakage problem of a porous vascular graft but also improves the wettability of the scaffolds. The physicochemical properties and biological characteristics of TLVGs were evaluated by tensile testing, Surface wettability test, Fourier transform-infrared spectroscopy (FTIR) measurement, Live-Dead cell staining assay, and CCK-8 assay. Especially, the oriented microgrooves on the inner surface of the TLVGs can promote human umbilical vein endothelial cells (HUVECs) directed growth and migration in favor of endothelialization. All results showed that the fabricated TLVGs with excellent physicochemical properties and biocompatibility has great potential in clinic application.

Rapid fabrication of gelatin-based scaffolds with prevascularized channels for organ regeneration.

One of the biggest hindrances in tissue engineering in recent decades has been the complexity of the prevascularized channels of the engineered scaffold, which was still lower than that of human tissues. Another relative difficulty was the lack of precision molding capability, which restricted the clinical applications of the huge engineered scaffold. In this study, a promising approach was proposed to prepare hydrogel scaffold with prevascularized channels by liquid bath printing, in which chitosan/ß-sodium glycerophosphate served as the ink hydrogel, and gelation/nanoscale bacterial cellulose acted as the supporting hydrogel. Here, the ink hydrogel was printed by a versatile nozzle and embedded in the supporting hydrogel. The ink hydrogel transformed into liquid effluent at low temperature after the cross-linking of gelatin by microbial transglutaminase (mTG). No residual template was seen on the channel surface after template removal. This preparation had a high degree of freedom in the geometry of the channel, which was demonstrated by making various prevascularized channels including circular, branched, and tree-shaped networks. The molding accuracy of the channel was assessed by studying the roundness of the cross section of the molded hollow channel, and the effect of the mechanical properties by adding bacterial cellulose to the supporting hydrogel was analyzed. Human umbilical vein endothelial cells were injected into the aforementioned channels which formed a confluent and homogeneous distribution on the surface of the channels. Altogether, these results showed that this approach can construct hydrogel scaffolds with complex and accurate molding prevascularized channels, and hs great potential to resolve the urgent vascularization issue of bulk tissue-engineering scaffold.

Designed and fabrication of triple-layered vascular scaffold with microchannels.

Currently, one of the best preparation strategies for the triple-layered vascular scaffold is to imitate the three-layer structure of natural blood vessels to achieve the biofunctional characteristics of vascular transplantation. Here, we developed a combinatorial method to fabricate triple-layered vascular scaffold (TVS) by using electrospinning and coaxial 3 D printing. First, Polycaprolactone-collagen (PCL-Col) was applied to prepared the inner layer of TVS by electrospinning. Second, egg white/sodium alginate (EW/SA) blend hydrogel was extruded to form hollow filaments by coaxial 3 D printing and crosslinking mechanism, which enwound around the surface of the inner layer in a circumferential direction as the intermediate layer of TVS. Finally, electrospun PCL-Col nanofibers were wrapped on the surface of hydrogel layer as the outer layer of TVS. The morphological characterization and mechanical strength of the fabricated TVS were measured. Compared with natural blood vessels, results shown that ultimate tensile stress (UTS), strain to failure (STF), the estimated burst strength and the suture retention strength (SRS) of TVS were superior. Also, the fabricated TVS exhibits good hydrophilicity and excellent flexibility. Moreover, the biocompatibility of TVS was investigated through human umbilical vein endothelial cells (HUVECs), the results demonstrated that cells can successfully attach the surface of graft and maintain high viability. In summary, all of results demonstrated that this method could fabricate a novel triple-layered vascular scaffold, possessing appropriate mechanical properties and good biological properties, which has the potential to be used in tissue engineered vascular grafts applications.

High-resolution combinatorial 3D printing of gelatin-based biomimetic triple-layered conduits for nerve tissue engineering.

Peripheral nerve injury is a common clinical problem often requiring surgical nerve reconstruction. To this end, tissue-engineered conduit has been proved to be crucial for nerve reconstruction. Despite its progress in recent years, the design and fabrication of translational biomimetic nerve conduits is highly challenging. Therefore, this study aims to design and fabricate mechanically-tunable nerve conduits with biomimetic structural features of the human nerve suitable for nerve tissue engineering. Herein, we employed combinatorial approach comprising of electrohydrodynamic (EHD) jet printing, dip-coating, and electrospinning techniques for fabricating triple-layered conduits. The intricate structural details were achieved via high-resolution EHD jet printed PCL filaments with tunable directionality, as the innermost layer; followed by dip coating of gelatin hydrogels to form the middle layer, and lastly, wrapped with electrospun PCL nanofibers as an outer layer of the conduits. The mechanical properties, porosity, and biocompatibility of the fabricated conduits were studied and compared with control. The results of this study confirmed that the combinatorial approach has greater potential to fabricate mechanically-tunable triple-layered conduits with favorable neuronal precursor and vascular cell compatibility.

Designing vascular supportive albumen-rich composite bioink for organ 3D printing.

Bioink plays a major role in 3D printing of tissues and organs. Alginate is a widely used component for bioinks but its cellular responses are limited, which limits its clinical translation. In this study, we demonstrate the printability and cellular compatibility of composite bioink consists of sodium alginate (NaAlg) and egg white, also called albumen. The experimental conditions necessary for 3D printing composite bioink were optimized by changing different concentration ratios of Albumen/NaAlg and their various physicochemical properties were studied. The structural characteristics of the 3D printed scaffold was also investigated. In vitro experiments showed that human umbilical vein endothelial cells can successfully attach to the printed scaffold and maintain high viability during the course of study. Interestingly, vascular sprouting and neovascular network formation was observed inbetween fibers within the printed scaffold. In conclusion, the results of this study demonstrate that 3D printed Albumen/NaAlg composite bioinks with favorable biological functionality hold a great potential in tissue and organ engineering.