Cell-Free bone regeneration using dental MSCs secretomes from pulp and gingiva in rabbit tibial defects

Abstract

Mesenchymal stem cells (MSCs)-secretome represent a promising cell-free strategy for bone regeneration, overcoming cell therapies' drawbacks. This study compares the bone repair capabilities of secretomes derived from human dental pulp stem cells (hDPSCs) and human gingival stem cells (hGMSCs) in a rabbit tibial defect model. Secretomes were prepared, and levels of RUNX, osterix, and alkaline phosphatase (ALP) levels were quantified using ELISA. Six-mm defects were created in rabbit tibiae treated with either hDPSCs or hGMSCs secretomes, collagen scaffolds, or left untreated. The mineral apposition rate (MAR) was assessed using fluorescent labeling. Histomorphometric analysis (including bone area percentage, mature/immature bone and bone marrow quantification) and qRT-PCR for osteocalcin were conducted at 3 and 6 weeks. Results showed significantly higher concentrations of RUNX, osterix, and ALP in hDPSCs secretome compared to hGMSCs. Additionally, defects treated with hDPSCs secretome exhibited a higher MAR and greater new bone formation, accelerated maturation (higher mature bone area; p < 0.05), and reduced bone marrow spaces at 3 weeks than those treated with hGMSCs, collagen, or control groups. By 6 weeks, both secretomes achieved comparable bone maturation (83-85% bone area), significantly surpassing controls. Both secretomes upregulated osteocalcin gene expression. These findings show the therapeutic potential of dental MSCs secretomes, and particularly hDPSCs secretomes, as a cell-free, clinically relevant method for improving bone regeneration. This approach addresses limitations associated with traditional bone grafting and possibly presents a new pathway for regenerative therapies in dentistry and orthopedics.