Figure 3.

Effects of the exposure to P25/lipopolysaccharide (LPS) on nitric oxide production and Nos2 expression in macrophages. Raw264.7 cells were incubated for 24 h in Dulbecco’s modified Eagle’s medium (DMEM) + 10% fetal bovine serum (FBS) in the presence of P25 (128 µg/ml), LPS (10 ng/ml), the mixture of P25 and LPS (128 µg/ml + 10 ng/ml, respectively), P25/LPS (the pellet of the spun mixture, re-suspended at the original volume, with nominal doses of 128 µg/ml, for TiO₂ NP, and 40 pg/ml for LPS), the undiluted (SUP, nominal LPS dose 10 ng/ml), or diluted (D.SUP, nominal LPS dose of 40 pg/ml) supernatant of the spun mixture. (A) At the end of the incubation, the culture medium was harvested to determine nitrite concentration. (B) The same cell monolayers were lysed to evaluate Nos2 expression through WB analysis. The experiment has been performed twice with comparable results. (C) Raw264.7 cells were incubated for 24 h in the presence of the indicated doses of LPS in DMEM + 10% FBS. At the end of the incubation, the culture medium was harvested to determine nitrite concentration. For (A,C), data are means of four independent determinations ± SD. For (A), ***, p < 0.001 vs. cells treated with P25; ns, not significant vs. LPS alone, as evaluated by one-way ANOVA for multiple comparisons with Bonferroni correction. For (C), *, ***, p < 0.05, p < 0.001 vs. LPS-untreated cells; ns, not significant vs. LPS-untreated cells, as evaluated by one-way ANOVA for multiple comparisons with Bonferroni correction.