Clonal Characterisation of Bone Marrow and Dental Pulp Progenitors Cells

Adult stem / progenitor cells show great potential in regeneration of craniofacial structures. Much research has centred on the regenerative potential of bone marrow derived cells, but interest is growing for using dental pulp progenitor cells as an alternative source. Objectives: To compare the colony forming efficiency (CFE), population doubling (PDs) and expression of stem cell markers during cell expansion of clonal progenitor cell populations from dental pulp and bone marrow. Methods: Rat dental pulp and bone marrow were extirpated from the upper and lower incisors or the femoral bones, respectively. Cells of dental pulp were dissociated by collagenase/dispase. Mononuclear cells were separated from bone marrow using histopaque dentisty gradient centrifugation. Cell preparations were seeded at 4000 cells/cm² and progenitor cells selected by preferential adherence to fibronectin over 20 mins. Cell colonies were counted over 3-12 days, after which isolated clones were culture expanded in αMEM, 20% FCS, 100μM L-ascorbic acid 2-phosphate and PDs recorded. At early (8PDs), mid (17-18PDs) and late (30PDs) passages, cells were examined for progenitor cell markers by qPCR. Results: CFE for dental pulp progenitor cells was 0.088, whilst bone marrow was 0.007, significantly lower (n=3). However, clonal expansion of isolated colonies was 7 times more successful for bone marrow cells compared with dental pulp. For successful clonal expansions, PDs of dental pulp cells was 1.5 times lower than bone marrow. All clones expressed mesenchymal markers VCAM1, Fgfr1, Snai1, MCAM, c-kit, Nanog and Oct4. Levels differed between tissue sources and were capable of fluctuating during culture expansion. Conclusion: Progenitor cells were more readily obtainable and expanded from bone marrow. If dental pulp is to provide an alternative progenitor cell source for cell based tissue engineering, greater understanding is needed as to how the micro-environment controls survival and proliferation of these cells. Funded by MRC UK.