The Wnt/β-catenin signaling pathway is regulated by titanium with nanotopography to induce osteoblast differentiation

Wnt/beta-catenin signal transduction is involved in the homeostatic control of bone mass. It is well established that a titanium surface with nanotopography (Ti-Nano) favors osteoblast differentiation by modulating different signaling pathways. However, few studies have investigated the participation of the Wnt/beta-catenin pathway in the osteogenic effect of nanoscale topographies. In this study, we aimed to determine whether the Wnt/beta-catenin signaling pathway is involved in the elevated osteogenic potential of Ti-Nano. MC3T3-E1 cells were cultured on Ti-Nano and machined Ti (Ti-Control) for evaluation of the expression of Wnt/beta-catenin signaling pathway-related genes. Based on the results to real-time PCR, the Wnt receptor Fzd4 was selected and silenced by CRISPRi. The resulting cells were cultured on both Ti surfaces, and several events involved in osteoblast differentiation were evaluated. The results revealed that Fzd4 gene silencing, corresponding to negative modulation of Wnt/beta-catenin, inhibits expression of the osteoblast phenotype. It is worthy of note that this inhibitory effect on osteoblast differentiation was more pronounced in cells grown on Ti-Nano compared with those grown on Ti-Control. By disrupting Fzd4 gene expression, we have shown that the elevated osteogenic potential of Ti-Nano is due to activation of the Wnt/beta-catenin signaling pathway, which reveals a new mechanism to explain osteoblast differentiation induced by nanotopography. Such an understanding of the intracellular machinery involved in surface guiding of osteoblast fate may contribute to the development of smart biomaterials to modulate the process of implant osseointegration.