Bone regeneration using aged donor cells and visible light-curable hydrogel: An in vitro and in vivo evaluation.

BACKGROUND

Transplantation therapy combining scaffolds and cells can be used for extensive bone regeneration (e.g., to manage severe alveolar bone defects). Gelatin methacryloyl (GelMA) may be a suitable scaffold for cell transplantation. We developed a novel GelMA that polymerizes under visible light, using riboflavin as a photoinitiator (GelMA-RF). Here we investigated the efficacy of bone-regenerative therapy combining GelMA-RF and immature osteoblasts in rats.

METHODS

Rat alveolar bone immature osteoblasts (RAOBs) from 70-week-old rats were encapsulated in GelMA-RF (aged RAOBs + GelMA-RF) and transplanted into palatal (control, RAOBs alone, and RAOBs + GelMA-RF groups; n = 8, respectively) or femoral (control, GelMA-RF alone, RAOBs alone, and RAOBs + GelMA-RF groups; n = 8, respectively) bone defects. Macroscopic assessment, micro-computed tomography, and histological analyses were performed, and mineralization (aged and young RAOBs with or without osteogenic differentiation (OD) groups; n = 8, respectively) and the calcium/phosphorus ratio (aged and young RAOBs in OD+ or OD- groups; n = 8, respectively) for RAOBs under 3D-culture conditions were evaluated. Bone differentiation-related gene expression (aged and young RAOBs in OD+ or OD- groups; n = 8, respectively) was analyzed by quantitative polymerase chain reaction.

RESULTS

Transplantation of RAOBs encapsulated in GelMA-RF into palatal or femoral defects resulted in significantly earlier bone-like tissue formation compared with controls. RAOBs showed sufficient mineralization in a 3D-culture environment. Bone differentiation-related gene expression was significantly increased in 3D cultures.

CONCLUSIONS

Taken together, GelMA-RF, particularly when combined with RAOBs, provides a supportive scaffold for bone differentiation and may represent a novel bone-regeneration therapy for complex/extensive bone defects, even when combined with immature alveolar-bone osteoblasts from aged rats.

PLAIN LANGUAGE SUMMARY: Regenerative medicine combining scaffold materials and cell transplantation shows promise for widespread bone regeneration. This study focused on gelatin methacryloyl (GelMA) as a scaffold material. We developed GelMA (GelMA-RF) that gelated under visible light, avoiding the disadvantage of UV irradiation in conventional methods, and used it as a scaffold for cell transplantation. Furthermore, considering the increasing need for regenerative medicine in elderly patients, this study focused on using immature-osteoblasts derived from aged individuals as transplant cells to verify the bone regenerative capacity. In this study, immature-osteoblasts isolated from the alveolar bone of aged rats were encapsulated in GelMA-RF and transplanted into bone defects, confirming early bone regeneration sufficiently. Furthermore, in vitro analysis confirmed bone formation-related gene expression and calcification capacity, demonstrating that GelMA-RF is a suitable material for bone regeneration. These findings suggest that even alveolar bone immature-osteoblasts derived from aged individuals, when combined with GelMA-RF, hold promise as a novel therapeutic approach for regenerating complex and extensive bone defects.