Objectives: It is suggested that TGF-beta play an important role in periodontal development and tissue regeneration. However, the molecular mechanism of TGF-beta signaling as well as its regulation of gene expression in osteogenic differentiation of PDLSCs is still poorly known. The aim of the present study was to verify the roles of Smad-3 signaling induced by TGF-beta 1 on human periodontal stem cells. Material and methods: Human periodontal ligament stem cells were obtained from adult patients with a clinically healthy periodontium, using the explant culture and collection of multiple colony technique. The effect of TGF-beta 1 on PDLSCs was evaluated with mRNA and protein expressions of collagen 1 (COL1), alkaline phosphatase (ALP), osteocalcin (OCN). Furthermore, Smad-3 expression induced by TGF-beta 1 was detected by quantitative RT-PCR. Levels of phosphorylated Smad-3 (p-Smad3) were detected by flow cytometry analysis and immunofluorescence observation. On the other hand, the PDLSCs were also submitted to the combination of TGF-beta 1 with Smad3 inhibitor (SIS3), and gene expressions of p-Smad3 and COL1, ALP, OCN were detected. Results: Certain dose of TGF-beta 1 can significantly enhance COL1, ALP, OCN gene expression of PDLSCs and showed concentration-dependent characteristics. TGF-beta 1 can induce increased expression of Smad3 and p-Smad3, and p-Smad3 translocated into nucleus as indicated by immunofluorescence observation. Smad3 inhibitor (SIS3) completely inhibited Smad-3 activity. Furthermore, SIS3 significantly inhibited the osteogenic activity of PDLSCs enhanced by TGF-beta 1. Conclusion: These findings suggest that TGF-â1 may function as an osteo-promotive cytokine, stimulating the osteoblastic differentiation of periodontal ligament stem cells mainly through Smad-3 signal transducers and activator of transcription pathways were involved in this process.
Key words: human periodontal ligament stem cells, osteogenic differentiation, TGF-beta signaling, Smad-3