Figure 2

Fisetin decreases LPS-induced inflammatory mediator and cytokine levels in RAW 264.7 macrophages. RAW 264.7 macrophages (1 × 10⁵ cells/mL) were treated with fisetin (0–8 µM) 2 h before treatment with 500 ng/mL LPS. (A) Total mRNA was isolated at 6 h after 500 ng/mL LPS treatment, and RT-PCR was performed. GAPDH was used as an internal control. (B) Total proteins were isolated at 24 h and western blotting was performed. β-Actin was used as an internal control. (C) The amount of NO production in the culture medium was determined using the Griess Reagent Assay. (D) The amount of PGE₂ was determined at 24 h using an ELISA according to the manufacturer’s instructions. (E) Total mRNA was isolated at 6 h and subjected to RT-PCR for IL-6 and TNF-α. The amount of (F) IL-6 and (G) TNF-α was measured at 24 h by an ELISA. Each value indicates the mean ± SEM from three independent experiments. Significant differences among the groups were determined using the Student’s t-test (^###p < 0.001 vs. untreated cells) and one-way ANOVA with Bonferroni correction (***p < 0.001, **p < 0.05, and *p < 0.01 vs. LPS-treated cells).