Micro/nano topological modification of TiO2 nanotubes activates Thy-1 signaling to control osteogenic differentiation of stem cells

Micro/nano topological modification is critical for improving the in vivo behaviors of bone implants, regulating multiple cellular functions. Titania (TiO₂) nanotubes show the capacity of promoting osteoblast-related cell differentiation and induce effective osseointegration, serving as a model material for studying the effects of micro/nano-topological modifications on cells. However, the intracellular signaling pathways by which TiO₂ nanotubes regulate the osteogenic differentiation of stem cells are not fully defined. Thy-1 (CD90), a cell surface glycoprotein anchored by glycosylphosphatidylinositol, has been considered a key molecule in osteoblast differentiation in recent years. Nevertheless, whether the micro/nano topology of the implant surface leads to changes in Thy-1 is unknown, as well as whether these changes promote osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). Here, TiO₂ nanotubes of various diameters were prepared by adjusting the anodizing voltage. qPCR and immunoblot were carried out to assess the mechanism by which TiO₂ nanotubes regulate Thy-1. The results revealed Ti plates harboring TiO₂ nanotubes ∼100-nm diameter (TNT-100) markedly upregulated Thy-1. Subsequently, upregulated Thy-1 promoted the activation of Fyn/RhoA/MLC Ⅱ/F-actin axis, which enhanced the nuclear translocation of YAP. After Thy-1 knockdown by siRNA, the Fyn/RhoA/MLC Ⅱ/F-actin axis was significantly inhibited and TiO₂ nanotubes showed decreased effects on osteogenic differentiation. Therefore, Thy-1 upregulation might be a major mechanism by which micro/nano-topological modification of TiO₂ nanotubes promotes osteogenic differentiation in BMSCs. This study provides novel insights into the molecular mechanism of TiO₂ nanotubes, which may help design improved bone implants for clinical application.