Repairing Craniofacial Tissues using Dental Pulp or Bone Marrow Progenitors

Bone marrow progenitors are a major clinical tissue source for bone repair, but may not be the most practical, with only a small percentage of progenitor clones truly multipotent. Dental pulp progenitors may represent an alternative source. Objectives: Compare osteogenic differentiation capacity of clonal populations from bone marrow stem cells (BMSCs) and dental pulp stem cells (DPSCs), and understand which progenitor source could provide rapid and effective bone formation for repair. Methods: DPSCs and BMSCs were extirpated from 21-28 day old rat incisors or femurs respectively. Pulp cells were dissociated by collagenase/dispase. Mononuclear marrow cells were isolated using histopaque density-gradient centrifugation. Cell preparations were seeded at 4000cells/cm² and progenitors selected by preferential adherence to fibronectin over 20 minutes. Isolated clones were culture expanded in αMEM, 20% FCS, and 100μM L-ascorbate. Osteogenic capacity was investigated with 10nM Dexamethasone and 10mM β-Glycerophosphate media supplements. Mineralising nodules were examined using Alizarin Red staining. Osteoblast formation was analysed by RUNX2 and osteocalcin expression before and after treatment using PCR, and osteocalcin and bone sialoprotein synthesis by immunocytochemistry. Results: Clonal expansion of BMSCs was four times more successful than DPSCs, and had a 1.5 times higher proliferative capability. Clonogenic frequency of DPSCs however was 12 times greater. Both BMSCs and DPSCs were capable of osteoblast differentiation and showed Alizarin Red staining, osteocalcin, and bone sialoprotein within the extracellular matrix, and upregulated RUNX2 and osteocalcin at gene levels. However, DPSCs displayed a lower capacity for differentiation, and failed to form mineralised nodules characteristic of osteogenic differentiation. Conclusions: Progenitor populations appear to mimic their native microenvironments. The greater proliferative capacity and osteogenic differentiation of BMSCs is likely related to continual bone remodelling experienced throughout adulthood, unlike the tooth where DPSCs responds solely to traumatic damage to produce a reparative mineralised matrix. JH funded by MRCUK Studentship.