Effect of modified three-dimensional-printed titanium scaffold on osteogenic differentiation of adipose-derived mesenchymal stem cells
Wang Jing1, Yang Jiuju1, Wang Ningning1, Liu Chao2
1Department of Stomatology, Cangzhou Medical College, Cangzhou 061001, Hebei Province, China; 2Department of Oral and Maxillofacial Surgery, Qilu Hospital of Shandong University, Jinan 250012, Shandong Province, China
Abstract: BACKGROUND: The three-dimensional (3D)-printed titanium scaffold overcomes the disadvantages of traditional titanium scaffolds, such as poor controllability of the internal structure and mismatch in shape. However, due to the relatively large biological inertia of titanium, it is difficult to combine with surrounding tissues quickly and stably after implantation in the body.
OBJECTIVE: To modify the surface of 3D-printed titanium by sandblasting acid etching and anodic oxidation and observe its effect on the adhesion, proliferation, and osteogenic differentiation of adipose-derived mesenchymal stem cells.
METHODS: Micro-nano porous structure was constructed on the surface of 3D-printed titanium by sandblasting acid etching and anodic oxidation. The surface was characterized by scanning electron microscopy. Adipose-derived mesenchymal stem cells at passage 3 were inoculated in the Ti group (group A), Ti+osteogenic induction group (group B), and modified Ti group (group C). Cytoskeleton and CCK-8 assay were used to detect cell adhision and proliferation. Alkaline phosphatase staining and Alizarin red staining were used to observe osteogenesis. Real-time PCR was applied to analyze the expression of osteogenic genes. Immunofluorescence staining was employed to observe the expression of osteocalcin and osteopontin in cells.
RESULTS AND CONCLUSION: (1) Scanning electron microscopy showed that micro and submicron scale pits and grooves could be seen on the surface of the modified 3D-printed titanium scaffold. There were nanoscale pores with a diameter of 70-100 nm inside the scaffold and the pores were connected with each other. (2) Under a confocal microscope, there were few pseudopods and antennae in groups A and B, and they had not been spread; the cells in group C were completely spread on the surface of the material, and a large number of obvious pseudopods and antennae were visible, which were closely attached to the surface of the material. CCK-8 assay showed that modified 3D-printed titanium scaffolds could promote the proliferation of adipose-derived mesenchymal stem cells. (3) Alkaline phosphatase staining and Alizarin red staining exhibited that alkaline phosphatase activity and the mineralization ability were higher in the groups B and C than those in the group A (P < 0.05). (4) Osteocalcin, RUNX2, alkaline phosphatase, and type I collagen mRNA expression levels were higher in the groups B and C than those in the group A (P < 0.05). Osteopontin and osteocalcin protein expression levels were higher in the groups B and C than those in the group A (P < 0.05). (5) These results suggest that 3D-printed titanium scaffold modified by sandblasting acid etching and anodic oxidation has good biocompatibility and can promote the adhesion, proliferation and osteogenic differentiation of adipose-derived mesenchymal stem cells.
Key words: 3D-printed titanium, adipose-derived mesenchymal stem ells, adhesion, proliferation, biocompatibility, osteogenic protein, osteogenic gene, osteogenic differentiation