Biocompatibility evaluation of a 3D-bioprinted alginate-GelMA-bacteria nanocellulose (BNC) scaffold laden with oriented-growth RSC96 cells

Peripheral nerve injury can cause various degrees of damage to the morphological structure and physiological function of the peripheral nerve. At present, compared with “gold standard” autologous nerve transplantation, tissue engineering has certain potential for regeneration and growth; however, achieving oriented guidance is still a challenge. In this study, we used 3D bioprinting to construct a nerve scaffold of RSC96 cells wrapped in sodium alginate/gelatin methacrylate (GelMA)/bacterial nanocellulose (BNC) hydrogel. The 5% sodium alginate+5% GelMA+0.3% BNC group had the thinnest lines among all groups after printing, indicating that the inherent shape of the scaffold could be maintained after adding BNC. Physical and chemical property testing (Fourier transform infrared, rheometer, conductivity, and compression modulus) showed that the 5% alginate+5% GelMA+0.3% BNC group had better mechanical and rheological properties. Live/dead cell staining showed that no mass cell death was observed on days 1, 3, 5, and 7 after printing. In the 5% alginate+5% GelMA group, the cells grew and formed linear connections in the scaffold. This phenomenon was more obvious in the 5% alginate+5% GelMA+0.3% BNC group. In the 5% alginate+5% GelMA+0.3% BNC group, S-100β immunofluorescence staining and cytoskeleton staining showed oriented growth. Polymerase chain reaction (PCR) array results showed that mRNA levels of related neurofactors ASCL1, POU3F3, NEUROG1, DLL1, NOTCH1 and ERBB2 in the 5%GelMA+0.3%BNC group were higher than those of other groups. Four weeks after implantation in nude mice, RSC96 cells grew and proliferated well, blood vessels grew, and S-100β immunofluorescence was positive. These results indicate that a 3D-bioprinted sodium alginate/GelMA/BNC composite scaffold can improve cell-oriented growth, adhesion and the expression of related factors. This 3D-bioprinted composite scaffold has good biocompatibility and is expected to become a new type of scaffold material in the field of neural tissue engineering. • 3D-bioprinted alginate-GelMA-bacteria nanocellulose scaffold containing RSC96 cells. • 5% alginate+5% GelMA+0.3% bacteria nanocellulose hydrogel has better mechanical and rheological properties. • The scaffold can improve cell-oriented growth, adhesion and the expression of related mRNA levels.