Figure 1.
Time Course Analysis of LPS-Stimulated Macrophages
(A) Diagram to depict the basic experimental design. Murine RAW 264.7 macrophage cells were seeded at a density of 1 × 105 cells per square centimeter and cultured for 20 h before stimulation with a dose of 10 ng/mL LPS with or without the addition of the anti-inflammatory corticosteroid dexamethasone (1 μg/mL). Cells and conditioned media were assayed at 2, 4, and 6 h after stimulation. (B) qRT-PCR assay for the expression of TNF-α, IL-1β, IL-6, and iNOS. Individual PCR reactions were normalized against the internal control GAPDH, and transcript levels in untreated macrophages were expressed as 1 U. Data represent mean ± SD of three independent experiments. *p < 0.0001 between the LPS and LPS+Dexa groups derived from one-way ANOVA after multiple comparisons, Tukey post hoc test. (C) TNF-α and IL-6 secretion in conditioned media assessed by ELISA. Data represent mean ± SD of an experiment run in quadruplicate. *p < 0.0001 between the LPS and LPS+Dexa groups derived from one-way ANOVA after multiple comparisons, Tukey post hoc test. (D) Reproducibility of the IL-6 ELISA in conditioned media at 4 h post-LPS stimulation. Data represent mean ± SD of eight independent experiments. *p < 0.0001 between the LPS and LPS+Dexa groups derived from one-way ANOVA after multiple comparisons, Tukey post hoc test.