Fig. 8

Protein oxidation and aggregation are induced by AgNPs and Ag⁺ at different levels in SUM159 and iMEC cells in proportion to the relative sensitivity of the cells to each toxicant. To assess protein oxidation, SUM159 and iMEC cells were treated with A AgNPs or B Ag⁺ for 24 h, stained with DCP-NEt2C, and fluorescence was measured using flow cytometry. Data are representative of three independent experiments. Statistical analysis was performed by two-way ANOVA followed by post hoc tukey test. Statistical differences are indicated (*p < 0.05; **p < 0.01). SUM159 and iMEC cells were treated with C AgNPs or D Ag⁺ for 24 h and protein aggregation was measured by confocal microscopy. E Fluorescence images were quantified using ImageJ. Data is plotted as total cell fluorescence ± SEM relative to untreated controls of at least 10 cells per image for 3 images. Statistical analysis was performed by one-way ANOVA followed by post-hoc Sidaks test. Significant differences are indicated (**p < 0.01; *p < 0.05; ****p < 0.0001). Western blotting was used to detect proteotoxic stress responses in F SUM159 cells or G iMEC cells exposed to AgNPs (untreated, 18.25, 37.5 and 75 μg/mL, left to right) or Ag⁺ (untreated, 0.5, 1, 2 μg/mL, left to right) for 24 h. GAPDH was used as a loading control. Data are representative of a minimum of two independent experiments