Synthesis, Surface, and Cell-adhesion Properties of Polyurethanes Containing Covalently Grafted RGD-peptides

In an attempt to improve endothelial cell adhesion and growth on a polyurethane copolymer, cell adhesive RGD-containing peptides were grafted to the polymer backbone. Two peptide grafting reaction schemes, including one-step and two-step approaches, were developed. FTIR and amino acid analysis confirmed that coupling of the peptide to the polyurethane backbone was achieved by both the one-step and two-step methods. However, the two-step approach showed a higher peptide coupling efficiency and resulted in better control of the orientation of the grafted peptide. The two-step reaction scheme was used to prepare Gly-Arg-Gly-Asp-Ser-Tyr (GRGDSY), Gly-Arg-Gly-Asp-Val-Tyr (GRGDVY), and Gly-Arg-Gly-Glu-Ser-Tyr (GRGESY) peptide-grafted polyurethanes with two different peptide densities (100 and 250 mumol/g polymer). Dynamic contact angle measurements indicated that the surfaces of the peptide-grafted polyurethanes were more hydrophilic than the starting and carboxylated versions of the precursor polyurethane. In addition, the surface hydrophilicity of the peptide-grafted polymers increased with increasing bulk peptide density. Electron spectroscopy for chemical analysis suggested that the grafted peptide was present at the polymer-air interface, in vacuo, for the peptide-grafted polyurethanes. The surface peptide density appeared to correlate with the incorporated peptide density in the bulk. In vitro endothelial cell adhesion experiments showed that, without the presence of serum in culture medium, the GRGDSY- and GRGDVY-grafted polyurethanes dramatically enhanced cell attachment and spreading compared with the starting, carboxylated, and GRGESY-grafted polymers. Increasing the peptide density from 100 to 250 mumol/g polymer for the GRGDSY- and GRGDVY-grafted polyurethanes resulted in an increase in cell attachment. With approximately the same peptide density (100 or 250 mumol/g polymer), the GRGDVY-grafted polymers supported more adherent cells than did the GRGDSY-grafted polymers. Similar trends were observed in the in vitro endothelial cell growth studies using culture medium containing serum and endothelial cell growth supplement. The GRGDSY- and GRGDVY-grafted polyurethanes promoted more cell growth than did the starting polyurethane. However, the presence of adhesive serum proteins and growth factor diminished the differences between the cell-adhesive peptide grafted polymers and the GRGESY-grafted polymers.