In vitro cementoblast-like differentiation of postmigratory neural crest-derived p75+ stem cells with dental follicle cell conditioned medium

• Cranial neural crest-derived cells (CNCCs) take part in tooth morphogenesis.
• positive (p75+) CNCCs are fibroblast-like and resemble mesenchymal stem cells.
• p75+ CNCCs in dental follicle cell medium (DFCCM/dNCP) appear like cementoblasts.
• DFCCM/dNCP-treated p75+ cells express cementoblast specific mineralization markers.
• p75+ cells are pure stem cells and able to differentiate to neuronal cells.

Cranial neural crest-derived cells (CNCCs) play important role in epithelial–mesenchymal interactions during tooth morphogenesis. However, the heterogeneity of CNCCs and their tendency to spontaneously differentiate along smooth muscle or osteoblast lineages in vitro limit further understanding of their biological properties. We studied the differentiation properties of isolated rat embryonic postmigratory CNCCs, expressing p75 neurotrophin receptor (p75NTR). These p75NTR positive (p75+) CNCCs, isolated using fluorescence activated cell sorter, exhibited fibroblast-like morphology and characteristics of mesenchymal stem cells. Incubation of p75+ CNCCs in dental follicle cell conditioned medium (DFCCM) combined with dentin non-collagenous proteins (dNCPs), altered their morphological features to cementoblast-like appearance. These cells also showed low proliferative activity, high ALP activity and significantly increased calcified nodule formation. Markers related to mineralization or specific to cementoblast lineage were highly expressed in dNCPs/DFCCM-treated p75+ cells, suggesting their differentiation along cementoblast-like lineage. p75+ stem cells selected from postmigratory CNCCs represent a pure stem cell population and could be used as a stem cell model for in vitro studies due to their intrinsic ability to differentiate to neuronal cells and transform from neuroectoderm to ectomesenchyme. They can provide a potential stem cell resource for tooth engineering studies and help to further investigate mechanisms of epithelial–mesenchymal interactions in tooth morphogenesis.