Effect of hydroxyapatite and pulsed electromagnetic field on osteogenic differentiation of human stem cells seeded onto 3D-printed polycaprolactone scaffolds for future bone graft transplantation

Bone graft transplantation is a risky procedure mainly due to postoperative complications. Tissue engineering is an alternative to regenerating bone tissue by seeding mesenchymal stem cells into 3D scaffolds by osteoinductive signaling. Poly(ε-caprolactone) (PCL) is one of the most used biopolymers for fabricating scaffolds for bone tissue engineering. PCL is enriched with hydroxyapatite (HA) to promote the production of the bone-mineralized matrix. Moreover, the application of pulsed electromagnetic fields (PEMFs) on scaffolds loaded with HA can significantly benefit collagen and HA networks favoring the interaction between organic and mineral phases of bones. In this context, we investigated, for the first time, the effect exerted by a PEMF exposure on the osteogenic differentiation of human adipose stem cells (hASCs) seeded onto 3D printed PCL-HA scaffolds and cultured in growth and osteogenic medium (OM). After 21 days of culture, the results showed excellent colonization of scaffolds and the deposition of extracellular matrix (ECM) secreted by hASCs. Immunofluorescence staining confirmed the deposition of collagen type I and osteopontin both on scaffolds. Gene expression analysis indicated that both HA crystal and PEMF treatment were able to trigger osteogenic differentiation, as documented by the increase of osteogenic markers. These results suggest that HA, osteogenic factors, and PEMFs converge on the same signaling pathways as there are no additive effects and no change in protein synthesis between scaffolds with HA or treated with PEMFs for cells cultured in OM. The proposed study lays the foundations for exploring new approaches in bone graft transplantation.