RGD peptide immobilized on TiO2 nanotubes for increased bone marrow stromal cells adhesion and osteogenic gene expression

Recently, TiO₂ nanotube layers are widely used in orthopedics and dental applications because of their good promotion effect on bone cells. Furthermore, peptide sequences such as arginine–glycine–aspartic acid are used to modify Ti implant for binding to cell surface integrins through motif. In this study, a cellular adhesive peptide of arginine–glycine–aspartic acid–cysteine (RGDC) was immobilized onto anodized TiO₂ nanotubes on Ti to examine its in vitro responses on rat bone marrow stromal cells (BMSCs). Materials were characterized by scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy techniques. High-resolution C1s scans suggested the presence of RGDC on the surface and SEM images confirmed the nanotubes were not destroyed after modification. BMSCs adhesion and osteogenic gene expression were detected in TiO₂ nanotube layers with and without RGDC modification by fluorescence microscopy, confocal laser scanning microscopy, SEM, and realtime polymerase chain reaction (Real-time PCR). Results showed that the TiO₂ nanotube layers immobilized with RGDC increased BMSCs adhesion compared to nonfunctionalized nanotubes after 4 h of cultivation. Furthermore, the osteogenic gene expression of BMSCs was dramatically enhanced on the TiO₂ nanotube layers immobilized with RGDC (10 mM) compared to the TiO₂ nanotube layers immobilized with RGDC (1 mM) and non-functionalized anodized Ti. Our results from in vitro study provided evidence that Ti anodized to possess nanotubes and then further functionalized with RGDC should be further studied for the design of better biomedical implant surfaces.