Fig. 5.

Enhanced osteogenesis and osteoimmunomodulation of the ABL@ZnTA nanointerface in vitro. (A) Schematic diagram of osteogenic differentiation evaluation under different conditions. Condition i): MC3T3 cells are directly seeded on the different coating samples stimulated with LPS. Condition ii): MC3T3 cells are seeded on the culture plate and incubated in a macrophage-conditioned medium stimulated with H₂O₂. (B) Relative mRNA expression of osteogenesis-related genes encoding RUNX 2, ALP, and COL 1 on day 3 with LPS stimulus by qRT-PCR analysis, n = 3. (C) Relative mRNA expression of osteogenesis-related genes encoding OCN, ALP, and SOD 1, detected on day 3 in macrophage-conditioned medium stimulated with H₂O₂ by qRT-PCR analysis, n = 3. The alkaline phosphatase and Alizarin Red S staining images (D) of MC3T3 on various functionalized coating samples at each checkpoint and (E) quantitative analysis of calcium nodules, n = 3. (F) Schematic diagram and SEM of in vitro mineralization of coated Ti soaked in simulated body fluid for 40 days. (G) Schematic diagram of in vitro antibacterial evaluation of coated Ti implants. (H) Top: Representative agar plate images of S. aureus after incubation with different samples; Bottom: SEM morphologies of S. aureus incubated with different samples (arrowheads indicate the cracked and shrinking membranes). (I) Statistical analysis of colony counts after 24 h of incubation on coated Ti implants, n = 3. Data are presented as mean ± SD. Statistical significance was calculated via Student's t-test or one-way ANOVA with Tukey posthoc test (*, P < 0.05; **, P < 0.01; and ***, P < 0.001).