Osteogenic Differentiation of Stem Cells from Human Exfoliated Deciduous Teeth

Objective: The osteogenesis of mesenchymal cells derived from bone marrow and adipose tissues can be promoted by retinoic acid (RA) and dexamethasone (DX), however, the osteogenic potential of dental stem cells has not been well investigated in response to these agents. The objective of this study was to investigate if RA and DX could promote the proliferation and osteogenic differentiation of stem cells from human exfoliated deciduous teeth (SHED). Methods: SHED were cultured in Dulbecco's modified eagle medium containing 10% fetal calf serum and antibiotics at 37^oC in a 5% CO₂ atmosphere until confluence. The SHED were treated with serum-free medium (control), or serum-free medium with 0.5, 1 or 2 µl of RA, or 1, 10, and 100nM of DX. SHED proliferation was measured after five days of treatment. The gene expression of osteogenic markers was examined using RT-PCR. Alkaline phosphatase (ALP) activity was determined using a biochemical colorimetric assay from cell lysates obtained after two weeks of treatment. The ALP quantity was normalized to total protein content measured using the Bradford protein assay. After twenty one days of treatment, calcium deposition was detected by Alizarin red staining. Results: The proliferation of SHED was inhibited by DX and RA dose-dependently. The ALP gene expression of SHEDs was significantly up-regulated by RA in a dose-dependent manner, but not by DX. The ALP activity was consistent with the SHED mineralizing gene expression promoted by RA. The Alizarin red staining of calcium deposition was seen on the RA treated SHED after twenty one days of cell culture. Significantly less mineralizing gene expression and Alizarin red staining were observed on the DX treated SHED. Conclusion: RA can be an effective inducer of SHED osteogenesis, suggesting these cells in combination with RA have the therapeutic potential to help repair congenital cleft palate bone defects.