Isolation and characterisation of stem cells from human dental pulp

Objectives: Therapies based upon cell replacement and tissue engineering, underpinned by stem cell biology, are emerging as potentially powerful treatment strategies. The aim of this study was to investigate the use human dental pulp tissue as a source of pluripotent stem cells. Methods: Fifty dental pulps were extirpated from healthy permanent teeth extracted at Leeds Dental Institute and were divided into six transverse segments in an apical-coronal direction to investigate any site-specific differences in stem cell potential. Primary cells were isolated using standard organ culture methods. Cells were cultured in basal media or under osteogenic, chondrogenic and adipogenic conditions and assessed by enumeration of colony forming units fibroblastic (CFU-F) formation, histological staining (alkaline phosphatase - ALP, von Kossa, Oil Red), biochemical assays (ALP, DNA content, sGAG) and flow cytometry analysis. Results: The majority of human dental pulp derived stem cells (hDPDSCs) demonstrated classic spindle-shaped fibroblast-like morphology. The proliferative potential of both the dental pulp derived CFU-F and primary cell cultures varied from tooth-to-tooth and from site-to-site within individual pulps. Cells derived from the middle segments of the pulp demonstrated stronger ALP positive activity compared with those derived from the more apical and coronal regions. Microscopical analyses of hDPSDCs following culture in specialised media indicated osteogenic, chondrogenic, neural and epithelial morphologies whereas adipogenesis was low. Mineralized deposits were observed when the cells were cultured under osteogenic conditions. hDPSDCs had surface marker phenotypes similar to human mesenchymal stem cells (CD45-CD34-CD133-CD105+CD73+CD166+). Conclusion: This study, together with the work of others, indicates the potential for using human dental pulp as a source of pluripotent stem cells for future tissue replacement therapies and tissue engineering. The reasons for the observed site-specific differences in proliferative and osteogenic potential are unknown but they may reflect differences in the relative contributions of the different tissue compartments at these locations.