The Effect of Bone and Dentin Matrix Derivatives on the Differentiation of Human Dental Pulp Stem Cells for Osteogenesis and Dentinogenesis in a Scaffold-Free Culture

Purpose The most important challenges in tissue engineering are to find an effective growth/differentiating factor for stem cell induction while retaining high cell vitality and convert to correct end cell linage. In this study, for the first time, self-administered dentin matrix derivatives were used as a growth/differentiation factor for dentin regeneration. Methods In the fourth passage, colonies of human dental pulp stem cells (hDPSCs) were extracted; then, the purified cells were evaluated based on the percentage of vital cells by the MTT method. Afterward, hDPSCs in the first group were cultured with self-administered dentin derivative signal (DDS), while those in the second group with demineralized freeze-dried bone allograft (DFDBA) as a signal for 3, 7, and 10 days, and then compared with group 3 (negative control) that contained only hDPSC colonies. Three groups were evaluated based on cell differentiation and expression of osteoblast and odontoblast cell markers by immunocytochemistry (ICC) staining. Results In the first group, hDPSCs were differentiated into odontoblast and in the second group into osteoblast. The mean expression percentage of the dentin sialophosphoprotein (DSPP) marker in differentiated cells in the first group was significant at days 3, 7, and 10 ( p ≤ 0.0001). Also, the mean expression of the bone sialoprotein (BSP) marker was significant in the second group on days 3, 7, and 10 ( p ≤ 0.0001). Conclusion hDPSCs in scaffold-free culture enable to differentiate to odontoblasts and osteoblasts through DDS and DFDBA. Both the DSPP and BSP markers were increasingly expressed with time and effective in cell induction. Future in vivo studies can recommend to investigate the role of such natural growth factors in regenerative endodontic and dentistry in clinics. Lay Summary This study evaluated the differentiation capability of dental pulp stem cells by two bioactive materials, self-administered dentin derivative signal (DDS) and demineralized freeze-dried bone allograft (DFDBA), as induction signals. The challenge in this regard is to find a simple, suitable, and efficient method of DDS preparation, retaining proteins undamaged and ensuring enough disinfected particles for cell exposure. For this purpose, we investigated freeze-dried methods that are commonly applied in material preparation, and tested DDS with the scaffold-free stem cell’s culture comparing with DFDBA that exhibits great induction potential to be used for tissu