Fabrication of Bone-derived Decellularized Extracellular Matrix/ceramic-based Biocomposites and Their Osteo/odontogenic Differentiation Ability for Dentin Regeneration

The goal of this study was to fabricate bioactive cell-laden biocomposites supplemented with bone-derived decellularized extracellular matrix (dECM) with calcium phosphate ceramic, and to assess the effect of the biocomponents on the osteogenic and odontogenic differentiation of human dental pulp stem cells (hDPSCs). By evaluating the rheological properties and selecting printing parameters, mechanically stable cell-laden 3D biocomposites with high initial cell-viability (>90%) and reasonable printability (≈0.9) were manufactured. The cytotoxicity of the biocomposites was evaluated via MTT assay and nuclei/F-actin fluorescent analyses, while the osteo/odontogenic differentiation of the hDPSCs was assessed using histological and immunofluorescent analyses and various gene expressions. Alkaline phosphate activity and alizarin red staining results indicate that the dECM-based biocomposites exhibit significantly higher osteogenic activities, including calcification, compared to the collagen-based biocomposites. Furthermore, immunofluorescence images and gene expressions demonstrated upregulation of dentin matrix acidic phosphoprotein-1 and dentin sialophosphoprotein in the dECM-based biocomposites, indicating acceleration of the odontogenic differentiation of hDPSCs in the printed biocomposites. The hDPSC-laden biocomposite was implanted into the subcutaneous region of mice, and the biocomposite afforded clear induction of osteo/odontogenic ectopic hard tissue formation 8 weeks post-transplantation. From these results, we suggest that the hDPSC-laden biocomposite is a promising biomaterial for dental tissue engineering.