Immortalization of Dental Papilla Cells Differentiating into Odontoblast in vitro

Abstract

Dental papilla cells are composed by cranial neural crest-derived mesenchymal progenitors that can differentiate into odontoblasts via epithelial-mesenchymal interaction. However the differentiation potential of these cells remains largely unknown. We therefore attempted to establish immortalized mouse dental papilla cells, and investigated whether they possess odontoblast progenitors. Cells growing out from the dental papilla tissue of mouse incisors at postnatal 1 day were immortalized by human papillomaviruses type 16 E6 gene deleted with PDZ domain binding motif (MDP^E6). MDP^E6 was successfully immortalized and continuously grown beyond population doubling 80 without changing cell morphology or proliferation rate. To investigate the differentiation potential of MDP^E6, it was treated with osteogenic differentiation medium for 21 days. MDP^E6 showed high alkaline phosphatase activity and mineralized nodule formation. RT-PCR analysis showed that MDP^E6 expressed osteoblast-phenotype related genes, such as bone sialoprotein, osteocalcin or osterix. However, expression of odontoblast marker genes such as dentin sialoprotein(DSP)and dentin matrix protein 1 was not observed in this condition. These data suggest that MDP^E6 did not induce the expression of DSP by osteogenic differentiation medium. The odontoblast differentiation potential of MDP^E6 was assessed by treatment with bFGF. As a result, MDP^E6 showed an odontoblast-like phenotype that showed a prolonged columnar structure and induced the expression of DSP. DSP expression levels were found to increase by 3-fold over the levels in plate cultures without bFGF. Our results suggest that MDP^E6 possesses odontoblast progenitors that can differentiate into odontoblasts by bFGF signaling. Thus, MDP^E6 provides a useful cell culture system for investigating the molecular mechanisms of odontogenesis.