Sustained release of melatonin from TiO₂ nanotubes for modulating osteogenic differentiation of mesenchymal stem cells <i>in vitro</i>

Abstract

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.