Effects of Emdogain on Growth Factor Profiles of Mesenchymal Stem-Cells

OBJECTIVES: Emdogain (EMD) is widely used as a clinically effective material for inducing periodontal regeneration. Although mesenchymal stem-cells (MSC) are likely to be the source of new periodontal tissue, the mechanism of action of EMD on MSC has not yet been examined. Since growth factors (GF) have a major role in tissue regeneration, the objective of this study was to develop a flow cytometry (FCM)-based Multiplex Bead Assay (MBA) to measure the effects of EMD on GF production by MSC in vitro.

METHODS: MSC were cultured in the presence and absence of EMD for 6h, 24h and 48h. Culture supernatants obtained at these times were used for the simultaneous measurement of eight GF, including platelet-derived growth-factor-AB (PDGF-AB), keratinocyte growth-factor (KGF), osteoprotegerin (OPG), fibroblast growth-factor2 (FGF-2), bone morphogenetic protein2 (BMP-2), tissue inhibitor of metalloprotease1 (TIMP-1), angiopoietin1 (Ang-1) and vascular-endothelial growth-factor (VEGF). Tyramide Signal Amplification (TSA) was used in conjunction with the MBA to increase assay sensitivity.

RESULTS: The modified MBA was found to be able to simultaneously measure all eight mediators in the MSC culture supernatants. In the presence of EMD, MSC secreted high levels of TIMP-1 at 6h. By 48h the levels of KGF, OPG, Ang-1 and VEGF all increased significantly, with TIMP-1 being the most markedly up-regulated at this time. In contrast, BMP-2, FGF-2 and PDGF showed little response to EMD.

CONCLUSION: The use of an FCM-based MBA, combined with TSA, has enabled us to simultaneously measure multiple GF in vitro. This is the first report to demonstrate that EMD differentially affects the production of these mediators by MSC. These findings suggest that this highly sensitive procedure could also be of value in monitoring specific molecular changes in the small amounts of fluid obtained from tissues undergoing wound healing and regeneration in vivo, including gingival crevicular fluid.