CA1 induced dental follicle stem cells co-culture with dental pulp stem cells and loaded three-dimensional printed PCL/β-TCP scaffold: a novel strategy for alveolar cleft bone regeneration.

BACKGROUND

Bone tissue engineering for alveolar clefts is in the early stages of development, and more research is needed to determine the optimal cell types, growth factors and delivery methods for the therapy.

METHODS

We co-cultured Carbonic anhydrase 1 (CA1) induced dental follicle stem cells (DFSCs) with dental pulp stem cells (DPSCs). In vitro, the Lentivirus vector overexpressing CA1 (LV-CA1) gene was constructed, transfected into DFSCs, and co-cultured with DPSCs indirectly. Osteoblast biomarkers in differentiated DFSCs were detected using quantitative real-time polymerase chain reaction and Western blotting. In vivo, establish a rat alveolar cleft model, transplanted stem cell Polycaprolactone/β-tricalcium phosphate (PCL/β-TCP) three-dimensional printed composite scaffold and samples were collected at 4 and 8 weeks postoperatively. The osteogenic effect was evaluated through micro computed tomography and histomorphometric analysis.

RESULTS

In vitro, the activity of DFSCs in the LV-CA1+Co-culture group was increased, and the mRNA and protein expressions of CA1, Alkaline phosphatase (ALP), Bone morphogenetic proteins 2 (BMP2), and Runt-related transcription factor 2 (RUNX2) were amplified to varying degrees (P < 0.05). In vivo, micro-CT displayed at 4 and 8 weeks postoperatively, the LV-CA1+Co-culture group had a considerably higher percentage of new bone development (39.1% and 56.9%) (P < 0.05) than the other two groups. Histomorphometric analysis displayed the LV-CA1+Co-culture group had more newly formed bone trabeculae and immature collagen.

CONCLUSION

A strategy based on a novel osteogenic gene CA1 and dental-derived mesenchymal stem cells co-culture is applied to the alveolar cleft, providing a novel idea for the application of bone tissue engineering in alveolar cleft bone grafting.