Available online 26 November 2022

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. TiO2 nanotubes were prepared on the surface of titanium specimens using the anodizing method and characterized their features. 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. A rat femoral defect model was established to evaluate the osseointegration effect of TiO2 nanotubes combined with P2Y6 agonists. The results showed that the average inner diameter of the TiO2 nanotubes increased 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. When using in vitro and in vivo experiments, the osteogenic effect of TiO2 nanotubes when combined with P2Y6 agonists was more pronounced. TiO2 nanotubes promoted the P2Y6 expression of BMSCs during osteogenic differentiation and promoted osteogenesis by activating the PKCα-ERK1/2 pathway. The combined application of TiO2 nanotubes and P2Y6 agonists may be an effective new strategy to improve the osseointegration of titanium implants.

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. For the first time, this study revealed the relationship between TiO2 nanotubes and purine receptor P2Y6, and further explored its mode of action, which may provide clues as to the regulatory role of TiO2 nanotubes on osteogenic differentiation of BMSCs. These findings will help to develop novel methods for guiding material design and biosafety evaluation of nano implants.

TiO2 nanotubes

Bone marrow mesenchymal stem cells

Purinergic receptor

PKCα-ERK1/2 signal pathway

Osteogenic differentiation

bone morphogenetic protein

bone marrow mesenchymal stem cells

energy dispersive spectroscopy

extracellular signal regulated kinase

fibroblast growth factor 2

mitogen-activated protein kinase

phosphate buffered saline

quantitative reverse transcriptase polymerase chain reaction

runt-related transcription factor 2

scanning electron microscope

transforming growth factor-beta

wingless-type MMTV integration site

© 2022 The Author(s). Published by Elsevier Ltd on behalf of Acta Materialia Inc.