Titanium dioxide nanotubes increase purinergic receptor P2Y6 expression and activate its downstream PKCα–ERK1/2 pathway in bone marrow mesenchymal stem cells under osteogenic induction

Abstract: Background: Titanium dioxide (TiO₂) nanotubes can improve the osseointegration of pure titanium implants, but this exact mechanism has not been fully elucidated. The purinergic receptor P2Y6 is expressed in bone marrow mesenchymal stem cells (BMSCs) and participates in the regulation of bone metabolism. However, it is unclear as to whether P2Y6 is involved in the osteogenic differentiation of BMSCs induced by TiO₂ nanotubes. Methods TiO₂ nanotubes were prepared on the surface of titanium specimens using the anodizing method. The surface morphology of the nanotubes was observed by a scanning electron microscope, and the elemental analysis was carried out by X-ray energy dispersive spectroscopy. Quantitative reverse transcriptase polymerase chain reaction and western blotting were used to detect the expression of P2Y6, markers of osteogenic differentiation, and PKCα–ERK1/2. Results: The average inner diameter of the TiO₂ nanotubes increases with an increase in voltage (voltage range of 30–90V), and the expression of P2Y6 in BMSCs could be upregulated by TiO₂ nanotubes in osteogenic culture. Inhibition of P2Y6 expression partially inhibited the osteogenic effect of TiO₂ nanotubes and downregulated the activity of the PKCα–ERK1/2 pathway. The osteogenic effect of TiO₂ nanotubes when combined with P2Y6 agonists was more pronounced. Conclusions: TiO₂ nanotubes can promote the P2Y6 expression of BMSCs during osteogenic differentiation and promote osteogenesis by activating the PKCα–ERK1/2 pathway.