Evaluation of the effect of 3D-bioprinted gingival fibroblast-encapsulated ADM scaffolds on keratinized gingival augmentation

Background and Objectives: The keratinized gingiva plays an important role in maintaining healthy periodontal and peri--implant tissue. Acellular dermal matrix (ADM), as a substitute biomaterial, has a porous structure and good biocompatibility. 3D-bioprinting has the potential for tissue engineering because it enables precise loading of cells layer--by--layer. Herein, we bioprinted ADM scaffold encapsulating gingival fibroblasts (GFs) and evaluated its efficacy in keratinized gingiva augmentation in vivo to assess its potential for clinical periodontal tissue regeneration. Methods: GFs were extracted from the gingiva of beagles and transfected with a green fluorescent protein (GFP). The ADM scaffold (ADM cell-free group) was constructed using ADM, gelatin, and sodium alginate mixed at an appropriate ratio via 3D-bioprinting. The ADM cell scaffold (ADM cell group) was established by adding extra GFs in the same manner. Six beagles were divided into blank control, ADM cell-free, and ADM cell groups; and implant surgery was performed. The keratinized gingiva was clinically and histologically evaluated at baseline and after 2 months. Results: GFs transfected with GFPs expressed green fluorescence and were present in new tissue in the ADM cell group and not observed in the ADM cell-free group. At 2 months after surgery, the keratinized gingival augmentation in the ADM cell group was significantly more than that in the ADM cell--free group. Attached gingival augmentation was also observed more in the ADM cell group than that in the ADM cell--free group. Histological staining showed that the tissue in the ADM cell group displayed a more integrated structure and higher expression of COL I, COL III, and VEGF--A than those in the ADM cell--free group. Conclusion: 3D-bioprinted GF- -encapsulated ADM scaffolds increased the amount of keratinized gingiva in vivo, suggesting that 3D-bioprinting has great potential for oral soft tissue regeneration.