Sustained release of melatonin from TiO 2 nanotubes for modulating osteogenic differentiation of mesenchymal stem cells in vitro.

To control the sustained release of melatonin and modulate the osteogenic differentiation of mesenchymal stem cells (MSCs), melatonin was firstly loaded onto TiO2 nanotubes by direct dropping method, and then a multilayered film was coated by a spin-assisted layer-by-layer technique, which was composed of chitosan (Chi) and gelatin (Gel). Successful fabrication was characterized by field emission scanning electron microscopy, atomic force microscope, X-ray photoelectron spectroscopy and contact angle measurement, respectively. The efficient sustained release of melatonin was measured by UV-visible-spectrophotometer. After 2 days of culture, well-spread morphology was observed in MSCs grown on the Chi/Gel multilayer-coated melatonin-loaded TiO2 nanotube substrates as compared to different groups. After 4, 7, 14 and 21 days of culture, the multilayered-coated melatonin-loaded TiO2 nanotube substrates increased cell proliferation, increased alkaline phosphatase (ALP) and mineralization, increased expression of mRNA levels for runt-related transcription factor 2 (Runx2), ALP, osteopontin (OPN) and osteocalcin (OC), indicative of osteoblastic differentiation. These results demonstrated that Chi/Gel multilayer-coated melatonin-loaded TiO2 nanotube substrates promoted cell adhesion, spreading, proliferation and differentiation and could provide an alternative fabrication method for titanium-based implants to enhance the osteointegration between bone tissues and implant surfaces.