MEPE's diverse effects on mineralization

Abstract

Matrix extracellular phosphoglycoprotein (MEPE) is an inhibitor of mineralization in situ and in cell cultures where altered expression is associated with oncogenic osteomalacia and hypophosphatemic rickets. The purpose of this study was to determine whether the intact protein or the peptide(s) originating from this protein was responsible for the inhibition. The ability of the intact protein and the acidic, serine- and aspartate-rich MEPE-associated motif (ASARM) peptide to promote or inhibit de novo hydroxyapatite formation and growth of hydroxyapatite seed crystals, in both phosphorylated and dephosphorylated forms, was assessed at room temperature in a dynamic gel diffusion system at 3.5 and 5 days. The most effective nucleator concentration was also examined when associated with fibrillar type I collagen. The phosphorylated intact protein was an effective promoter of mineralization in the gelatin gel diffusion system, while the ASARM peptide was an effective inhibitor. When dephosphorylated both the intact protein and the ASARM peptide had no effect on mineralization. Associated with collagen fibrils, some of the effect of the intact protein was lost. This study demonstrates the importance of posttranslational modification for the site-specific activity of MEPE and its ASARM peptide.

Fig. 1

a Effect of the ASARM peptide on de novo formation (left) and seeded growth (right) of hydroxyapatite in the gelatin gel system at 5.0 days. b In the gelatin gel system, when dephosphorylated, the ASARM peptide has no detectable effect on seeded growth. Values (experimental/control) are mean (SD) for three to five determinations at each concentration and normalized to controls run at the same time. * P < 0.05 relative to control (dotted line = 1.00)

Fig. 2

a There is a dose-dependent increase in calcium and phosphate accumulation in the gelatin gel system with increasing intact MEPE concentrations, with the observed maximum occurring at 25 μg/mL. With seeded growth, the 25 μg/mL concentration also stimulated mineral formation, but there was a statistically significant decrease at 50 μg/mL. b When dephosphorylated, intact MEPE has no effect on mineral accumulation. Values (experimental/control) are mean (SD) at 5 days for three to five determinations at each concentration and normalized to controls run at the same time. * P < 0.05 relative to control (dotted line = 1.00)

Fig. 3

When collagen fibrils were coated with intact MEPE, there was a slight increase in the mineral yield relative to control (* P < 0.05), but the yield in terms of calcium uptake was significantly less (** P < 0.05) than that due to 25 μg/mL MEPE in the absence of fibrillar collagen. Values (experimental/control) are mean (SD) for three to five determinations at each concentration at day 5 and normalized to controls run at the same time. * P < 0.05 relative to control, ** P < 0.05 vs. group indicated by arrow