Fig. 2. Midazolam inhibits lipopolysaccharide (LPS)–induced nuclear factor-κB (NF-κB) activation. (  A ) Effect of midazolam on the IκB-α degradation in the RAW264.7 cells. IκB-α levels were determined by Western blot. β-Actin is shown as a loading control. (  B ) Effect of midazolam on the NF-κB luciferase activity in human embryonic kidney 293 cells. Data are presented as percent fold change of basal luciferase activity. *  P < 0.05  versus basal, #  P < 0.05  versus basal, **  P < 0.05  versus tumor necrosis factor α (TNF-α) alone. Data represent the mean ± SEM for four separate experiments. (  C ) Effect of midazolam on the NF-κB p65 translocation induced by lipopolysaccharide in RAW264.7 cells. RAW264.7 cells were treated with lipopolysaccharide (300 ng/ml) for 30 min in the presence of absence of midazolam (30 μm). Cells then were fixed and subjected to immunofluorescent microscopy after staining with NF-κB p65 antibody (  green ) and propidium iodide (PI) to visualize nuclei (  red ) as described in the Materials and Methods.