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 microM. 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 microM. 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 microM. 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.