Emdogain Stimulates the Proliferation of Human Gingival Fibroblasts via ERK

Introduction: Emdogain ® (EMD), a formulation of Enamel Matrix Proteins (EMPs), promotes periodontal (bone, PDL, cementum) regeneration and was suggested to promote gingival healing as well. Objective: To examine the effect of EMD on the proliferation of primary human gingival fibroblasts (hGFs) and to characterize the signaling pathway(s) underlying this effect. Methods: Primary cultures of hGFs, derived from healthy donors during periodontal procedures, were allowed to attach for 24 hours, serum starved for additional 24 h and were stimulated with EMD in the absence or presence of fetal calf serum (FCS, (0.2-10)%). Cell number was quantified by crystal violet staining and proliferation by thymidine incorporation and FACS analysis. ERK (extracellular regulated kinase) activation (phosphorylation) was demonstrated by Western blotting. Results: In the absence of FCS, EMD (50microg/ml) induced hGF entry into S phase and DNA synthesis but not completion of the cell cycle (i.e. no increase in cell number). In the presence of low FCS concentrations (0.2-0.5%), EMD synergistically induced completion of the cell cycle, manifested as increased DNA synthesis and cell number. The mitogenic response to EMD was associated with activation of ERK1/2, which occurred in two waves, peaking after 15 minutes and 4 to 6 hours following stimulation. Both the EMD-induced proliferation and ERK activation were abolished by pretreatment with U0126, a specific MEK1/2 inhibitor, and the inhibition of the second wave was sufficient to abrogate mitogenesis. Conclusions: EMD stimulates GF proliferation by synergizing with one or more growth factors present in FCS via activation of the ERK cascade. Thus, in addition to its documented actions on PDL and bone cells, EMD may promote gingival healing by stimulating GF proliferation.