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Differentiation of stem cells in the dental follicle - PubMed

Differentiation of stem cells in the dental follicle

S Yao et al. J Dent Res. 2008 Aug.

Abstract

The dental follicle (DF) differentiates into the periodontal ligament. In addition, it may be the precursor of other cells of the periodontium, including osteoblasts and cementoblasts. We hypothesized that stem cells may be present in the DF and be capable of differentiating into cells of the periodontium. Stem cells were identified in the DF of the rat first mandibular molar by Hoechst staining, alkaline phosphatase staining, and expression of side-population stem cell markers. These cells were shown to be able to differentiate into osteoblasts/cementoblasts, adipocytes, and neurons. Treating the DF cell population with doxorubicin, followed by incubation in an adipogenesis medium, suggested that the adipocytes originated from stem cells. Thus, a possibly puripotent stem cell population is present in the rat DF.

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Figures

Figure 1
Figure 1

DF cells grown in a stem cell medium form cell clusters after 2 wks of culture (A), and the clusters stained positive for alkaline phosphatase (B). In contrast, DF cells grown in the normal MEM medium did not form cell clusters, and no alkaline phosphatase staining was seen (C). (This Figure appears in color in the online version.)

Figure 2
Figure 2

Fluorescent staining of DF cells with Hoechst 33342. Note that the fluorescence staining of nuclei was weaker in some cells (arrows), putative stem cells, as compared with the majority with bright fluorescence (A). RT-PCR to detect gene expression of BCRP. Note that BCRP expression was detected in the DF of the rat first mandibular molar at post-natal days 5, 7, 9, and 11 in vivo, as well as in the cultured DF cells (B).

Figure 3
Figure 3

Differentiation of DF stem cells into various cell types. Alizarin red staining (A) and von Kossa staining (B) to determine osteogenic differentiation: The red staining in Alizarin red and black staining in von Kossa indicate the deposition of mineralization (A,B). No such staining was seen in the controls not subjected to osteogenic induction (C-a, control for Alizarin red staining; C-b, control for von Kossa staining). Oil Red O stain to detect adipogenesis (D). Note the stained adipocytes (arrow) in the induction treatment (D) vs. no adipocytes in the control without adipogenic induction (E). Multipolar neurons (arrow) were seen in the neuron induction treatment (F), while no neurons were seen in the control maintained in the stem cell medium (G). Immunostaining for neurofilament 200, a late neuron differentiation marker (H). Note that the heavy staining was seen in the induced neurons (H), while no staining appeared in the control cells maintained in the growth medium only (I). No immunostaining was seen when primary antibody was substituted with IgG in the immunostaining controls (J).

Figure 4
Figure 4

Treatment of the DF cells with doxorubicin (DOX). The number of surviving cells was greatly reduced as the duration of DOX (1 µM) incubation was increased from 2 hrs (B), to 4 hrs (C), and to 6 hrs (D), as compared with the control without DOX treatment (A). The placement of cells treated with DOX for 4 hrs in an adipogenesis induction medium resulted in the majority of the cells forming Oil Red O-positive adipocytes (arrow) (E), whereas in the controls not treated with DOX, but placed in adipogenesis induction medium, the majority of the cells remained undifferentiated, with only a few forming adipocytes (arrow) (F). The results suggest that DOX kills the non-stem cells, and that adipocytes are derived from the stem cells. (This Figure appears in color in the online version.)

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