Rapid mineralization of hydrothermally induced TiO2 coating on titanium and early osteogenic responses in vitro

This study evaluated hydrothermally induced nanostructured TiO 2 coatings (HT-TiO 2 ) on Grade 1 titanium (Ti) with a focus on surface mineralization behavior and early cellular responses. Rapid mineralization (RM) was achieved by immersion in modified simulated body fluid (m-SBF). Ti discs were subjected to hydrothermal treatment (HT) to produce a TiO 2 coating, then mineralized in m-SBF for 2, 4, or 6 h. Surface characterization was conducted using SEM, EDX, XRD, and FTIR. Cellular responses were assessed using pre-osteoblastic cells and mesenchymal stem cell (MSC)‑like cells in vitro. Cell proliferation on Ti substrates was evaluated over 7 days, and alkaline phosphatase (ALP) activity in MSC-like cells was measured at days 7 and 14 across four groups: NC (non-coated), HT-TiO 2 , RM4 (HT-TiO 2 with 4-h RM), and RM6 (HT-TiO 2 with 6-h RM) discs. Cell attachment and spreading were evaluated by fluorescence microscopy at days 3 and 7. The intensity of the Ca–P layer and the molar composition increased with mineralization time (2–6 h). All surfaces supported pre-osteoblast proliferation without significant differences between groups. ALP activity was significantly elevated in the HT-TiO 2 and RM6 groups compared to controls, indicating early osteogenic activity in a subset of cells. Fluorescence imaging showed flattened cell morphology on HT-TiO 2 -discs and more elongated morphology on RM discs at day 3, while comparable cell confluency was observed on all surfaces by day 7. Overall, hydrothermally-induced nanostructured TiO 2 coatings on Ti surfaces can be rapidly mineralized, and the resulting Ca–P layer supports cell attachment, proliferation, and early osteogenic activity in vitro. These findings highlight the potential of surface‑driven mineralization strategies for modulating early cellular responses to titanium implants.