Fiber-Based Mini Tissue with Morphology-Controllable GelMA Microfibers.

The use of microscale fibers could facilitate nutrient diffusion in fiber-based tissue engineering and improve cell survival. However, in order to build a functional mini tissue such as muscle fibers, nerve conduits, and blood vessels, hydrogel microfibers should not only mimic the structural features of native tissues but also offer a cell-favorable environment and sufficient strength for tissue functionalization. Therefore, an important goal is to fabricate morphology-controllable microfibers with appropriate hydrogel materials to mimic the structural and functional complexity of native tissues. Here, gelatin methacrylate (GelMA) is used as the fiber material due to its excellent biological performance, and a novel coaxial bioprinting method is developed to fabricate morphology-controllable GelMA microfibers encapsulated in calcium alginate. By adjusting the flow rates, GelMA microfibers with straight, wavy, and helical morphologies could be obtained. By varying the coaxial nozzle design, more complex GelMA microfibers such as Janus, multilayered, and double helix structures could be fabricated. Using these microfibers, mini tissues containing human umbilical cord vein endothelial cells are built, in which cells gradually migrate and connect to form lumen resembling blood vessels. The merits of cytocompatibility, structural diversity, and mechanical tunability of the versatile microfibers may open more avenues for further biomedical research.

A bioengineered trachea-like structure improves survival in a rabbit tracheal defect model.

A practical strategy for engineering a trachea-like structure that could be used to repair or replace a damaged or injured trachea is an unmet need. Here, we fabricated bioengineered cartilage (BC) rings from three-dimensionally printed fibers of poly(ɛ-caprolactone) (PCL) and rabbit chondrocytes. The extracellular matrix (ECM) secreted by the chondrocytes combined with the PCL fibers formed a "concrete-rebar structure," with ECM deposited along the PCL fibers, forming a grid similar to that of native cartilage. PCL fiber-hydrogel rings were then fabricated and alternately stacked with BC rings on silicone tubes. This trachea-like structure underwent vascularization after heterotopic transplantation into rabbits for 4 weeks. The vascularized bioengineered trachea-like structure was then orthotopically transplanted by end-to-end anastomosis to native rabbit trachea after a segment of trachea had been resected. The bioengineered trachea-like structure displayed mechanical properties similar to native rabbit trachea and transmural angiogenesis between the rings. The 8-week survival rate in transplanted rabbits was 83.3%, and the respiratory rate of these animals was similar to preoperative levels. This bioengineered trachea-like structure may have potential for treating tracheal stenosis and other tracheal injuries.

Perfusable Vessel-on-a-Chip for Antiangiogenic Drug Screening with Coaxial Bioprinting.

Vessel-on-a-chips, which can be used to study microscale fluid dynamics, tissue-level biological molecules delivery and intercellular communication under favorable three-dimensional (3D) extracellular matrix microenvironment, are increasingly gaining traction. However, not many of them can allow for long-term perfusion and easy observation of angiogenesis process. Since angiogenesis is necessary for the expansion of tumor, antiangiogenic drugs play a significant role in cancer treatment. In this study, we established an innovative and reliable antiangiogenic drug screening chip that was highly modularly integrated for long-term perfusion (up to 10 days depending on the hydrogel formula) and real-time monitoring. To maintain an unobstructed flow of cell-laden tubes for subsequent perfusion culture on the premise of excellent bioactivities, a polycaprolactone stent inspired by coronary artery stents was introduced to hold up the tubular lumen from the inside, while the perfusion chip was also elaborately designed to allow for convenient observation. After 3 days of perfusion screening, distinct differences in human umbilical vein endothelial cell sprouting were observed for a gradient of concentrations of bevacizumab, which pointed to the effectiveness and reliability of the drug screening perfusion system. Overall, a perfusion system for antiangiogenic drug screening was developed, which can not only conduct drug evaluation, but also be potentially useful in other vessel-mimicking scenarios in the area of tissue engineering, drug screening, pharmacokinetics, and regenerative medicine.

3D printed high-resolution scaffold with hydrogel microfibers for providing excellent biocompatibility.

Melt electrowriting (MEW) can print high-resolution scaffolds with the ultrafine fibers from 800 nm to 20 µm. However, the cell seeding efficiency relatively low due to the large pore size of the MEW scaffold. Here, we reported a method to solve this dilemma by electrospinning a gelatin methacrylate (GelMA) hydrogel fibers membrane (HFM) on the MEW scaffold. This composite scaffold can own the controlled structures and porosity and excellent cell seeding performance. We systematically investigate the fabrication, morphology, and biocompatibility of composite scaffolds. The implanting of human umbilical vein endothelial cells(HUVES) showed excellent adhesion and biocompatibility on the composite scaffold. Moreover, the cells migrated gradually into the MEW scaffold along the GelMA HFM to form the cell sheet. We hold the opinion that the composite scaffolds have potential applications in the field of tissue engineering repair.

Directly coaxial 3D bioprinting of large-scale vascularized tissue constructs.

Three-dimensional (3D) bioprinting of soft large-scale tissues in vitro is still a big challenge due to two limitations, (i) the lack of an effective way to print fine nutrient delivery channels (NDCs) inside the cell-laden structures above the millimetre level; (ii) the need for a feasible strategy to vascularize NDCs. Here, a novel 3D bioprinting method is reported to directly print cell-laden structures with effectively vascularized NDCs. Bioinks with desired tissue cells and endothelial cells (ECs) are separately and simultaneously printed from the outside (mixed with GelMA) and inside (mixed with gelatin) of a coaxial nozzle. As a result, the printed large-scale tissue consists of sheath-core fibers. At the same time, when the core fibers are dissolved to generate channels, the ECs deposit and adhere to the channels automatically. With this method, 3D cell-laden, vascularized tissue constructs (⩾1 cm) with a long-term culture (⩾20 d) are firstly reported. Specifically, vascularized cancer tissue constructs and osteogenic tissue constructs were generated. Considering the above advantages, this advanced bioprinting strategy has significant potential for building large-scale vascularized tissue constructs for applications in tissue engineering, and possibly even in regenerative medicine and organ repair.

Synchronous 3D Bioprinting of Large-Scale Cell-Laden Constructs with Nutrient Networks.

Maintaining an adequate supply of nutrients/oxygen is a major challenge in the biofabrication of large tissue constructs. However, building preformed nutrient networks may be an effective strategy for engineering thick tissues. Here, a novel way for bioprinting large-scale tissue constructs with intentional nutrient networks is presented. A special nozzle is developed which can print bioink and sacrificial ink half and half synchronously in a single filament. Nutrient networks of these bioprinted constructs are formed by subsequently dissolving away gelatin, which allows for effective oxygen, nutrient, and waste diffusion, facilitating the cell activity and the generation of functional tissues. Due to the cell-laden bioink and sacrificial ink working together and promoting each other's printability to support themselves, complex soft cell-laden constructs with nutrient networks can easily be printed. Furthermore, two different cell types (osteoblast, human umbilical vein endothelial cells) encapsulated in the bioprinted large-scale constructs (≥1 cm) with nutrient networks show enhanced cell viability and spreading within a period of culture. It is envisioned that the advanced bioprinting technology may have significant potentials in facilitating the engineering of complex structures for tissue-specific needs, and bioprinting large-scale tissue constructs with nutrient networks toward applications in organ transplantation and repair.

Micro/nanofabrication of brittle hydrogels using 3D printed soft ultrafine fiber molds for damage-free demolding.

Hydrogels are very popular in biomedical areas for their extraordinary biocompatibility. However, most bio-hydrogels are too brittle to perform micro/nanofabrication. An effective method is cast molding; yet during this process, many defects occur as the excessive demolding stress damages the brittle hydrogels. Here, we propose a brand-new damage-free demolding method and a soft ultrafine fiber mold (SUFM) to replace the traditional mold. Both mechanical and finite element analysis (FEA) reveal that SUFMs have obvious advantages especially when the contact area between hydrogel and mold gets larger. By means of a high-resolution 3D printing called electrohydrodynamic (EHD) printing, SUFMs with various topological structures can be achieved with the fiber diameter ranging from 500 nm to 100 µm, at a low cost. Microfluidics and cell patterns are implemented as the demonstration for potential applications. Owing to the tiny scale of microstructures and the hydrophilicity of hydrogels, significant capillary effect occurs which can be utilized to deliver liquid and cells autonomously and to seed cells into those ultrafine channels evenly. The results open up a new avenue for a wider use of hydrogels in biomedical devices, tissue engineering, hydrogel-based microfluidics and wearable electronics; the proposed fabrication method also has the potential to become a universal technique for micro/nanofabrication of brittle materials.

3D printed multi-scale scaffolds with ultrafine fibers for providing excellent biocompatibility.

It is a dilemma that both strength and biocompatibility are requirements for an ideal scaffold in tissue engineering. The normal strategy is mixing or coating another material to improve the biocompatibility. Could we solve this dilemma by simply adjusting the scaffold structure? Here, a novel multi-scale scaffold was designed, in which thick fibers provide sufficient strength for mechanical support while the thin fibers provide a cell-favorable microenvironment to facilitate cell adhesion. Moreover, we developed a promising multi-scale direct writing system (MSDWS) for printing the multi-scale scaffolds. By switching the electrostatic field, scaffolds with fiber diameters from 3 µm to 600 µm were fabricated using one nozzle. Using this method, we proved that PCL scaffolds could also have excellent biocompatibility. BMSCs seeded on the scaffolds readily adhered to the thin fibers and maintained a high proliferation rate. Moreover, the cells bridged across the pores to form a cell sheet and gradually migrated to the thick fibers to cover the entire scaffold. We further combined the scaffolds with hydrogel for 3D cell culture and found that the fibers enhanced the strength and induced cell migration. We believe that the multi-scale scaffolds fabricated by an innovative 3D printing system have great potential for tissue engineering.

Bioprinting of Cell-Laden Microfiber: Can It Become a Standard Product?

Hydrogel microfibers have many fascinating applications as microcarriers for drugs, factors, and cells, such as 3D cell culture, building micro-organoids, and transplantation therapy due to their simple structures. It is unknown whether cell-laden fiber can become a standard-use product like woundplast. Here, from the technical and practical view, the elements required for user-oriented microfibers are first discussed: i) the materials used should promote cell functionalization and be easily processed; ii) follow a manufacturing method for mass fabrication; iii) have the ability to be stored long-term and be available for immediate use. Here, it is demonstrated that bioactive microfibers can be simply fabricated with coaxial bioprinting using gelatin methacrylate due to its tunable biological and mechanical properties. Additionally, programmed microfibers and 3D constructs with controllable composition can also be fabricated. These microfibers can be used to directly build organoids and complex co-culture tissue models. In the present study, vascular organoid, angiogenic sprouts, and tumor angiogenesis are demonstrated. It is also demonstrated, for the first time, that the cell-laden microfibers can be stored long-term via cryopreservation. These results show that cell-laden structures can be developed as a novel type of organoid product, which will open more avenues for tissue engineering and clinical organ repair.

Three-Dimensional Printed Wearable Sensors with Liquid Metals for Detecting the Pose of Snakelike Soft Robots.

Liquid metal (LM)-based flexible sensors, which utilize advanced liquid conductive materials to serve as sensitive elements, are emerging as a promising solution to measure large deformations. Nowadays, one of the biggest challenges for precise control of soft robots is the detection of their real-time positions. Existing fabrication methods are unable to fabricate flexible sensors that match the shape of soft robots. In this report, we first described a novel 3D printed multifunction inductance flexible and stretchable sensor with LMs, which is capable of measuring both axial tension and curvature. This sensor is fabricated with a developed coaxial LM 3D printer by coprinting of silicone rubber and LMs. Because of the solenoid shape, this sensor can be easily installed on snakelike soft robots and can accurately distinguish different degrees of tensile and bending deformations. We determined the structural parameters of the sensor and proved its excellent stability and reliability. As a demonstration, we used this sensor to measure the curvature of a finger and feedback the position of an endoscope, a typical snakelike structure. Because of its bending deformation form consistent with the actual working status of the soft robot and unique shape, this sensor has better practical application prospects in the pose detection.