Modelling The Influence of Dentine Matrix on Progenitor Cell Recruitment

It is established that Dental Pulp Progenitor Cells (DPPCs) have the capacity to differentiate into odontoblasts, a process likely influenced by both the dental pulp and surrounding dentine. Attempts to isolate the individual contribution of each tissue to DPPC recruitment are difficult however, due to their complex natures and close association. Objectives: Modification of an ex vivo rat mandible model by replacing the pulp tissue with collagen gel, enabling clearer characterisation of the surrounding dentine matrix influence on DPPC recruitment. Methods: Mandibles containing incisors were extracted from 28 day old male Wistar rats, cut into 2mm sections, and dental pulps removed. Sections were treated with either 0.02M CaOH to stimulate growth factor release from the dentine or PBS as a control. The pulp cavity was subsequently re-filled with type 1 collagen gel. DPPCs, isolated from rats ubiquitously expressing green fluorescent protein (GFP), were droplet seeded onto the collagen gels using a 35 gauge micro-needle. Sections were cultured for 5 days and cell orientation observed at 1, 3 and 5 days post-seeding using confocal microscopy. Results: GFP expressing DPPCs were superficially integrated into the collagen gel present in the pulp cavity at day 1. By days 3 and 5 post seeding, DPPCs had moved deeper into the collagen gel and associated with both the cut surface of the dentine and dentine lining the pulp cavity. Both cell survival and dentine association were greater in CaOH treated mandibles. Conclusions: An existing ex vivo model has been successfully modified to permit the observation of DPPC migration in response to the release of growth factors from dentine into a simplified 3-D environment. This model may be useful in investigating the relative contributions of dental pulp and dentine matrix to the recruitment, differentiation and maintenance of DPPCs in vivo. Supported by MRC project grant No.94298.