Optimization of the coaxial cell printing performance of bioink and printing of tissue-engineered scaffolds with vascular-like structure / 中国组织工程研究

ABSTRACT

BACKGROUND: Cells cannot survive in the area 200 µm away from nutrients in vitro. Vascular network construction is crucial for thick tissue and organ regeneration in tissue engineering. Coaxial cell printing provides a new way to construct vascular-like channels in vitro. OBJECTIVE: To optimize the coaxial cell printing performance of bioink and to build the tissue-engineered scaffolds with vascular-like structure. METHODS: The aseptic sodium alginate solution was prepared by intermittent pasteurization and then frozen. Freeze-dried powder of aseptic silk fibroin was prepared from degummed silk and sealed. The thawed sodium alginate solution was added to the silk fibroin protein freeze-dried powder and human umbilical vein endothelial cells were added to prepare the bioink. The outer axis of the biological three-dimensional printer was connected with the bioink, and the inner axis was connected with the crosslinking agent. The scaffolds were prepared by coaxial printing, and performed by optical coherence tomography, scanning electron microscopy observation and tensile test. Coaxial scaffolds were made by freeze-preserved sodium alginate solution for 7 days with human umbilical vein endothelial cells. Coaxial scaffolds were also made by freeze-dried sodium alginate solution for 7 days with human umbilical vein endothelial cells and silk fibroin protein sealed for 6 months. The cell survival rate was detected by dead and alive staining after 24 hours of culture in vitro. Vascular-like scaffolds with series and parallel structures were designed and printed. The cell proliferation was detected after 1, 3, 7, 10, and 14 days of culture. RESULTS AND CONCLUSIONS: (1) The optical coherence tomography showed that the maximum printing height of the bioink was 9 layers and the overall thickness was about 4.4 mm. Scanning electron microscopy showed that the outer wall of hollow fiber-filament of vascular-like scaffolds presented irregular strip-shaped crimp with micron-scale internal connected pore structure, while the inner wall of hollow fiber-filament had denser pore structure. (2) The elastic modulus of silk protein freeze-dried scaffold was higher than that of sodium alginate solution (P < 0.05). (3) The cell survival rate of scaffolds treated with sodium alginate solution for 7 days was (86.7±3.4)%, and that of scaffolds treated with silk protein freeze-dried powder for 7 days was (98.1±1.2)%, indicating that the sodium alginate solution freeze- preserved for 7 days was free of bacteria and the shelf-life of silk protein could be up to 6 months. (4) The proliferation activity of cells cultured with parallel structure for 7, 10, and 14 days was higher than that with series structure (P < 0.05). (5) These results imply that the scaffolds have good biocompatibility and mechanical properties.