Osteogenesis of Stem Cells Derived from Human Periodontal Ligaments

Objective: Our recent study showed that retinoic acid (RA) in a better inducer for osteogenic differentiation of stem cells from human exfoliated deciduous teeth than dexamethasone (DX). Multipotent stem cells have been recently isolated from human periodontal ligaments (PDL). However, the osteogenic potential of PDL stem cells has not been well investigated in response to these agents. The objective of this study was to investigate the effects of RA and DX treatments on proliferation and osteogenic differentiation of PDL stem cells. Methods: PDL stem cells were cultured in Dulbecco's modified eagle medium containing 10% fetal calf serum and antibiotics at 37^oC in a 5% CO₂ incubator until confluence. The PDL stem cells were treated with serum-free medium (control), or serum-free medium with RA (0.5, 1 or 2 µM), or DX (1, 10, and 100nM). Proliferation of PDL cells was measured at five days of treatment. The gene expressions of osteogenic markers were examined using RT-PCR at 7 days. A biochemical colorimetric assay was used to measure the alkaline phosphatase (ALP) activity of cells after 14 days of treatment. Mineralization of extracellular matrix was detected by Alizarin red staining at 21 day. Results: DX and RA inhibited the proliferation of PDL cells. The osteogenic gene expressions of PDL cells were significantly up-regulated by RA and DX. The ALP activity and gene expression of PDL cells were promoted more significantly by RA than DX. Stronger Alizarin red staining of calcium deposition was also seen on the RA treated group than the DX treated group after 21 days of cell culture. Conclusion: This study demonstrated that RA is also an effective inducer for osteogenesis of PDL stem cells. Osteogenically differentiated PDL stem cells by RA provide a potential cell source for cell-based therapy of bone defect repair.