Yuanyuan Liu

A multi-scale controlled tissue engineering scaffold prepared by 3D printing and NFES technology

The current focus in the field of life science is the use of tissue engineering scaffolds to repair human organs, which has shown great potential in clinical applications. Extracellular matrix morphology and the performance and internal structure of natural organs are required to meet certain requirements. Therefore, integrating multiple processes can effectively overcome the limitations of the individual processes and can take into account the needs of scaffolds for the material, structure, mechanical properties and many other aspects. This study combined the biological 3D printing technology and the near-field electro-spinning (NFES) process to prepare a multi-scale controlled tissue engineering scaffold. While using 3D printing technology to directly prepare the macro-scaffold, the compositing NFES process to build tissue micro-morphology ultimately formed a tissue engineering scaffold which has the specific extracellular matrix structure. This scaffold not only takes into account the material, structu...

Composite vascular scaffold combining electrospun fibers and physically-crosslinked hydrogel with copper wire-induced grooves structure

While the field of tissue engineered vascular grafts has greatly advanced, many inadequacies still exist. Successfully developed scaffolds require mechanical and structural properties that match native vessels and optimal microenvironments that foster cell integration, adhesion and growth. We have developed a small diameter, three-layered composite vascular scaffold which consists of electrospun fibers and physically-crosslinked hydrogel with copper wire-induced grooves by combining the electrospinning and dip-coating methods. Scaffold morphology and mechanics were assessed, quantified and compared to native vessels. Scaffolds were seeded with Human Umbilical Vein Endothelial Cells (HUVECs), cultured in vitro for 3 days and were evaluated for cell viability and morphology. The results showed that composite scaffolds had adjustable mechanical strength and favorable biocompatibility, which is important in the future clinical application of Tissue-engineered vascular grafts (TEVGs).

Composite vascular repair grafts via micro-imprinting and electrospinning

Composite vascular grafts formed by micro-imprinting and electrospinning exhibited improved mechanical properties relative to those formed by electrospinning alone. The three-layered composite grafts mimic the three-layered structure of natural blood vessels. The middle layer is made by micro-imprinting poly-p-dioxanone (PPDO), while the inner and outer layers are electrospun mixtures of chitosan and polyvinyl alcohol. The graft morphology is characterized with scanning electron microscopy. For constant graft thicknesses, the PPDO increases the mechanical strength. Cells cultivated on the vascular grafts adhere and proliferate better because of the natural, biological chitosan in the inner and outer layers. Overall, the composite scaffolds could be good candidates for blood vessel repair.

Novel control of gel fraction and enhancement of bonding strength for constructing 3D architecture of tissue engineering scaffold with alginate tubular fiber

Alginate tubular fiber has been successfully prepared via coaxial fluid crosslink mode, which is potentially used for the construction of vascularized tissue engineering scaffolds (VTES). However, its elastic and smooth surface is negative for the adhesion of fibers. In this study, the gel fractions were controlled in a novel way of two-step crosslink process in order to meet the needs of each processing link. Based on such consideration, an appropriate formulation was selected to direct write single fiber, which ensured the tubular structure with enough gel portion as well as adhesion between fibers with the reserved sol. Finally, the integrity of the scaffolds had a further development within the 2nd crosslink bath process, which would help to solve the question of poor shear resistance for hydrogel scaffolds.

A novel method for fabricating engineered structures with branched micro-channel using hollow hydrogel fibers

Vascularization plays a crucial role in the regeneration of different damaged or diseased tissues and organs. Vascularized networks bring sufficient nutrients and oxygen to implants and receptors. However, the fabrication of engineered structures with branched micro-channels (ESBM) is still the main technological barrier. To address this problem, this paper introduced a novel method for fabricating ESBM; the manufacturability and feasibility of this method was investigated. A triaxial nozzle with automatic cleaning function was mounted on a homemade 3D bioprinter to coaxially extrude sodium alginate (NaAlg) and calcium chloride (CaCl2) to form the hollow hydrogel fibers. With the incompleteness of cross-linking and proper trimming, ESBM could be produced rapidly. Different concentrations of NaAlg and CaCl2 were used to produce ESBM, and mechanical property tests were conducted to confirm the optimal material concentration for making the branched structures. Cell media could be injected into the branched channel, which showed a good perfusion. Fibroblasts were able to maintain high viability after being cultured for a few days, which verified the non-cytotoxicity of the gelation and fabrication process. Thus, hollow hydrogel fibers were proved to be a potential method for fabricating micro-channels for vascularization.

Analysis of multiple types of human cells subsequent to bioprinting with electrospraying technology

The aim of the present study was to investigate bioprinting with electrospraying technology using multiple types of human cell suspensions as bio-ink, in order to lay the initial foundations for the application of the bioprinting technology in tissue engineering. In the current study, six types of human cells were selected and cultured, including human fibroblasts, human adipose-derived stem cells (hADSCs), human periodontal ligament cells (HPDLCs), adult human retinal pigment epithelial cells (ARPE-19), human umbilical vascular endothelial cells (HUVECs) and human gastric epithelial cell line (GES-1). Each cell type was divided into two groups, the experimental and control group. All the experimental group cells were electrosprayed using an electrospraying printer (voltage, 15 kV; flow rate, 150 µl/min) and collected in a petri dish placed 15 cm away from the needle (needle diameter, 0.5 mm). Subsequently, cell viability was detected by flow cytometry with a Live/Dead Viability kit. In addition, the cell morphological characteristics were observed with a phase-contrast microscope after 6 h of culturing in order to obtain adherent cells, while cell proliferation was analyzed using a Cell Counting Kit-8 assay. The control groups, without printing, were subjected to the same procedures as the experimental groups. The results of the cell viability and proliferation assays indicated a statistically significant difference after printing between the experiments and control groups only for the hADSCs (P<0.05); by contrast, no significant difference was observed in cell viability and proliferation for the other five cell types (P>0.05). In addition, there were no observable differences between all experimental and the control groups at any examined time point in the terms of cell morphological characteristics. In conclusion, bioprinting based on electrospraying technology demonstrated no distinct negative effect on cell vitality, proliferation and morphology in the present study, and thus the application of this novel technology to cell printing may provide a promising method in tissue engineering.

Novel Compound-Forming Technology Using Bioprinting and Electrospinning for Patterning a 3D Scaffold Construct with Multiscale Channels

One of the biggest challenges for tissue engineering is to efficiently provide oxygen and nutrients to cells on a three-dimensional (3D) engineered scaffold structure. Thus, achieving sufficient vascularization of the structure is a critical problem in tissue engineering. This facilitates the need to develop novel methods to enhance vascularization. Use of patterned hydrogel structures with multiscale channels can be used to achieve the required vascularization. Patterned structures need to be biocompatible and biodegradable. In this study, gelatin was used as the main part of a hydrogel to prepare a biological structure with 3D multiscale channels using bioprinting combined with selection of suitable materials and electrostatic spinning. Human umbilical vein endothelial cells (HUVECs) were then used to confirm efficacy of the structure, inferred from cell viability on different engineered construct designs. HUVECs were seeded on the surface of channels and cultured in vitro. HUVECs showed high viability and diffusion within the construct. This method can be used as a practical platform for the fabrication of engineered construct for vascularization.

Fabrication of triple-layered bifurcated vascular scaffold with a certain degree of three-dimensional structure

Constructing vascular scaffolds is important in tissue engineering. However, scaffolds with characteristics such as multiple layers and a certain degree of spatial morphology still cannot be readily constructed by current vascular scaffolds fabrication techniques. This paper presents a three-layered bifurcated vascular scaffold with a curved structure. The technique combines 3D printed molds and casting hydrogel and fugitive ink to create vessel-mimicking constructs with customizable structural parameters. Compared with other fabrication methods, the technique can create more native-like 3D geometries. The diameter and wall thickness of the fabricated constructs can be independently controlled, providing a feasible approach for vascular scaffold construction. Enzymatically-crosslinked gelatin was used as the scaffold material. The morphology and mechanical properties were evaluated. Human umbilical cord derived endothelial cells (HUVECs) were seeded on the scaffolds and cultured for 72 h. Cell viability a...

Electrohydrodynamic direct printing on hydrogel: a novel method to obtain fine fibers

In this study, we proposed a novel method to obtain fine fibers: electrohydrodynamic (EHD) direct printing PCL on hydrogel. The effects of applied voltage, flow rate, plotting speed, hydrogel viscosity on the EHD direct writing process were investigated to obtain the most appropriate set of process conditions. We also compared the fibers obtained from this method with traditional EHD which deposited on aluminum and ethanol-based collectors. We found that fibers collected on the hydrogel were thinner and exhibited pores on the surface due to the hydrogel’s facilitation of organic solvents’ volatilization in the PCL fibers, which can benefit the attachment of cells. Besides, in our previous study, we found the freeze-thaw crosslinking process could greatly increase mechanical properties of the hydrogel used in our research, so this integration will not only mimic the composition of ECM which is a composite structure with a combination of fibrous proteins within a gelatinous grounded substance but also improve the mechanical properties of the scaffold. This novel method will broaden the application of EHD technology in the field of tissue engineering and other related areas.

Bio-Electrospraying of Human Umbilical Vein Endothelial Cells with a Customized Multi-Hole Spinneret

Bio-electrospraying (BES) is becoming an attractive tool for the delivery of cells into scaffolds for tissue engineering applications. In this study, we aimed to electrospray human umbilical vein endothelial cells (HUVECs) and improve the efficiency of BES by designing a new customized multi-hole spinneret. We demonstrated that the multi-hole spinneret could produce continuous and stable jets during BES, and the efficiency was increased by 5–7 times. Morphological observations, trypan blue and sulforhodamine B assays revealed that the HUVECs electrosprayed using the multi-hole spinneret remained viable and proliferated at a rate similar to that of the controls. Thus, the new multi-hole nozzle can considerably improve output for BES without affecting cell morphology, viability, and proliferation.