Novel Ex Vivo Modelling of Dental Pulp Progenitor Cell Behaviour

Objectives: Much consideration has been given to therapeutically targeting dental pulp progenitor cells (DPPCs) present in the dental pulp to stimulate reparative dentinogenesis. A novel ex vivo culture model of DPPC migration and behaviour within the pulp of a rat incisor is described. Methods: Mandibles, including incisors, were extracted from 28 day old male Wistar rats, cut into 2mm sections and cultured in α-MEM supplemented with 20% FBS. After 24 hours 1,000 DPPCs clonally selected from human dental pulps and labelled with PKH-26 fluorescent cell tracker dye were injected in 1μl volume directly into the incisor pulps within each rat mandible section. Injection was performed using a 35 gauge microsyringe. Confocal images showing injected DPPCs in each mandible section were taken at 0, 3 and 5 days after injection and their distribution over time observed. Results: Labelled DPPCs were observed within the incisor pulps at all time points. At day 0, DPPCs were present in the incisor pulp as a densely packed cluster near the site of injection. At both days 3 and 5, DPPCs were seen to diffuse from the injection site into the incisor pulp, spreading out considerably over time, reaching the widest distribution at day 5. Labelled DPPCs were not observed in any area of the mandible section outside of the incisor pulp. No fluorescent cells were observed in sham controls injected with ml of culture media. Conclusions: This novel ex vivo model demonstrates that it is possible to observe, over time, the movement of injected DPPCs through cultured dental pulp. This opens up numerous possibilities to study DPPC recruitment and differentiation in an environment which closely parallels an in vivo situation. Such a model would be highly useful in developing therapies targeting DPPC mediated reparative dentinogenesis. Research supported by MRC, grant reference: 94298