Enamel matrix derivative (EMDOGAIN^®) rapidly stimulates phosphorylation of the MAP kinase family and nuclear accumulation of smad2 in both oral epithelial and fibroblastic human cells

Abstract

In our previous study, we demonstrated that porcine enamel matrix derivative (EMD) induces p21WAF1/cip1 within 8 hours and subsequently arrests the cell cycle of human oral epithelial cells in G1 phase. In contrast, EMD markedly stimulates the proliferation of gingival fibroblasts without inducing p21WAF1/cip1. To investigate the mechanism of how EMD produces these differential effects, we have focused on the initial response of these two cell types to EMD. In epithelial cell cultures, EMD stimulated cytoskeletal actin polymerization within 30 min and promoted cell adhesion in our experimental system. EMD failed to stimulate either intracellular Ca2+ mobilization or cAMP production in either cell type. In both epithelial and fibroblastic cells, EMD (25-100 microgram/ml) rapidly produced dose-dependent phosphorylation of the mitogen-activated protein kinase (MAPK) family: extracellular signal response kinase (ERK), p38-MAPK (p38-K), and c-Jun-terminal kinase/stress-activated protein kinase (JNK). However, neither inhibitors of MEK (ERK kinase) nor p38-K could block EMD's anti-proliferative action on epithelial cells. On the other hand, EMD rapidly stimulated translocation of smad2 into the nucleus in both cell types. Spurred by this finding, we assayed for TGF-beta1, a ligand for one receptor associated with smad2 activation, and detected significant levels in EMD preparations. The sum of these pharmacological findings indicates that EMD contains at least one bioactive factor, which is most probably TGF-beta1 (or TGF-beta-like substances). In conjunction with the similarities in the differential growth-modulating actions between EMD and what is known for TGF-beta, we suggest that TGF-beta might act as the principal growth regulating agent of oral fibroblastic and epithelial cell types in EMD despite being present in only low levels.

Emdogain® in carcinogenesis: a systematic review of in vitro studies

“…However, one study found that EMD inhibited the adhesion of breast cancer (MCF-7) cells to bone matrix (79). The effects of EMD on intracellular cAMP levels differ in epithelial HeLa and SCC-25 cells (56,66). In HeLa cells, the production of PDGF-AB was enhanced by EMD, whereas no significant effect on TGF-β1 was observed (56).…”

“…In contrast, TGF-β1 levels in culture medium of MG-63 osteosarcoma cells were increased by EMD (39,82). In addition, EMD stimulates phosphorylation of MAP family kinases in SCC-25 cells, as does also TGF-β1 (66,83). Regarding the other cytokines, EMD enhances IL-6 production, but has no effect on the production of BMP-2 or IGF-I by MG-63 cells or HeLa cells (56,82).…”

“…EMD can also induce matrix and total protein synthesis by fibroblasts (55,62,65). The mitogenic signaling pathway by which EMD affects PDL fibroblasts has been associated with rapid phosphorylation of the MAP kinase family and nuclear accumulation of smad2, and with activation of EMD-specific receptor tyrosine kinase (RTK) and extracellular signal-regulated kinase (ERK) (63,64,(66)(67)(68). ERK activation by EMD has been shown to be related to elevated epidermal growth factor receptor expression (EGFR) by fibroblasts (68).…”

Amelogenin binds to both heparan sulfate and bone morphogenetic protein 2 and pharmacologically suppresses the effect of noggin

“…A previous report found that EMD prepared from developing porcine teeth possesses BMP-like and TGF-β-like activities [3,4], though the molecular mechanism is uncertain. To obtain detailed findings regarding the molecules contained in EMD, we performed 2-D PAGE analysis of EMD protein using a Zoom IPG Runner System (see materials and methods section).…”

“…A recent clinical review reported that EMD promotes both cementogenesis [1] and osteogenesis [2], while it has also been shown that EMD has bone morphogenetic protein (BMP) and transforming growth factor-β (TGF-β) activities [3,4], while in vitro experiments demonstrated that it stimulates osteoblast proliferation and differentiation [5,6]. EMD consists of amelogenin at nearly 90%, along with other enamel matrix proteins, such as enamelin, tuftelin, amelin, and ameloblastin [7,8].…”