Osteoblast-like cell adhesion to bone sialoprotein peptides

A number of studies have suggested that biomimetic peptides can be used in the design of a new generation of prosthetic implants to promote the successful biointegration of the implant materials. In the current study, the in vitro bioactivities of several peptides representing RGD (Arg-Gly-Asp)-containing sequences of bone sialoprotein (BSP) toward an osteoblast-like cell line (MC3T3-E1) were examined to provide insight into the molecular basis of BSP’s interaction with bone cells. BSP residues 283–288, 281–290, 278–293 and 278–302 were coated on polystyrene surfaces in 96-well non-tissue (untreated) culture plates, and their osteoblast adhesive properties compared to intact BSP and fibronectin as positive controls. BSP peptides 278–302 and 278–293 were found to be the most potent in their adhesive activity, increasing the number of adherent cells to 350% of control levels at an added concentration of 1 μM. Since these two peptides were equivalent in potency, it is suggested that the region 294–302 beyond the RGD domain is not necessary for cell binding. In comparison, peptides 283–288 and 281–290 were only active at concentrations greater than 200 μM. 50–70% of the peptide-stimulated adhesion was inhibited by the pretreatment of cell suspensions with solution phase RGD, suggesting that a portion of the peptides’ adhesive effects was specific and integrin-mediated, although other non-RGD flanking regions were probably also involved in the mechanism of adhesion. Importantly, a modified BSP peptide, in which an aspartic acid residue at position 288 of the RGD sequence was replaced by a glutamic acid residue to form RGE, was completely inactive as a cell adhesion stimulus at concentrations up to 200 μM. Thus, despite the potential role of non-RGD flanking regions, an intact RGD tripeptide was essential for all of the adhesive activity of the BSP peptides.