In vitro Growth and Characterization of Stem Cells from Human Dental Pulp of Deciduous Versus Permanent Teeth

Abstract

Objective: By date investigations have indicated the presence of stem cells within the pulp tissue of both temporary and permanent human teeth. In the present study, these stem cells were compared in terms of their growth kinetics and culture requirements.

Materials and methods: Stem cells within the pulp of the human third molar (permanent tooth) and the deciduous incisor (temporary tooth) were isolated, culture-expanded and characterized. Then the proliferation potential of the cells was compared using multiple cell growth indices as PDT (Population doubling time), colonogenic activity and growth curve. Furthermore, the cultures of both cells were optimized for maximal proliferation.

Results: Stem cells of either pulp tissue appeared as fibroblastic cells capable of differentiating into osteoblastic, odontoblastic, adipocytic and chondrocytic cell lineages. In contrast to molar stem cells, those from the incisor tooth expressed neurogenic markers of ßIII Tubulin and Tau. Based on in vitro growth data, the cells from third molar tended to have a lower PDT value (20.79, SD=2.8 versus 25.55, SD=2.9 hours), higher colonogenic activity and better growth curve than those from the deciduous incisor (P<0.05). Both cells exhibited high expansion rate when being plated in a medium with 20% phosphate buffer solution at a density of 100 cells/cm(2).

Conclusion: Given the high proliferation capacity, the stem cells from the human third molar would be an appropriate candidate for use in experimental, preclinical and even clinical setups.

Keywords: Cell Differentiation; Cell Proliferation; Molar, Third; Tooth, Deciduous.

Fig 1

Pulp cell cultures from human deciduous incisors and third molar teeth. Primary cultures (A and B), The cells in confluent culture at passage 3 (C and D). Bar=100 μm

Fig 2

Flow cytometric analysis of surface antigens on stem cells derived from human deciduous incisors and third molar teeth.

Fig 3

Differentiation of human dental pulp stem cells: either cell was succeeded to differentiate along odontoblastic (A, B), chondrogenic (C, D), adipocytic (E, F) and osteoblastic (G, H) cell lineages. Bar=100 μm

Fig 4

RT-PCR analysis of gene expression in the differentiated cells isolated from human deciduous incisors and third molar teeth. Both cells were successfully expressed odontoblastic-, chondrogenic-, adipogenic- and osteogenic-related genes.

Fig 5

Colonogenic assays performed for stem cells derived from human deciduous incisors and third molar teeth. A) The cells from the third molar appeared to create significantly more colonies than their counterpart from deciduous teeth. B) Likewise, the colonies of third molar cells were larger than those of deciduous incisors. * indicates the significant difference of P<0.05.

Fig 6

Population doubling time (PDT) and growth curve determined for stem cells derived from human deciduous incisors and third molar teeth. (A) Third molar cells tended to have a shorter PDT value compared to deciduous incisors (P<0.05). * indicates the significant difference of P<0.05. (B) According to the growth curve, third molar cells reached plateau sooner than deciduous incisor cells. Both cells exhibited a lag time of a few days in culture.

Fig 7

Culture optimization for better growth of the stem cells isolated from human deciduous incisors (A) and molar teeth (B). The cells underwent considerable fold increase when are plated at low density in a medium with 20% FBS.