Fig. 2.
Effect of dexamethasone (DEX) and/or mifepristone (RU486) on P3-mediated reporter gene expression. HepG2 cells (8 × 105 cells/well) were seeded in a 48-well plate 24 h prior to transfection. For each well, cells were incubated for 12 h with 2 μg of plasmid DNA mixed with 4 μg of linear polyethelynimine as transfection reagent and then maintained in serum-containing medium for 24 h. Cells were then treated for additional 24 h with DMSO (carrier solution) with or without DEX or DEX/RU486 in combination. Luciferase activity in each well was determined using luciferase assay. a Effect of co-transfection of HepG2 cells with increasing amount of pCMV-hGRα plasmid DNA on luciferase gene expression from either pGL3-3kP3-Luc or pGL3-3kP1-Luc plasmids. Of the 2 μg of plasmid DNA used for transfection, 1 μg accounts for reporter plasmid while the other accounts for increasing amount of pCMV-hGR (0, 0.2, 0.4, 0.8, or 1 μg) and decreasing amount of empty vector (pGL3-Luc without promoter; 1, 0.8, 0.6, 0.2, or 0 μg). b Blockade of DEX-enhanced luciferase gene expression by RU486. HepG2 cells were transfected with equal amount (1 μg) of pGL3-3kP3-Luc and pCMV-hGRα for 12 h and maintained in serum-containing medium for 24 h. Cells were then treated for another 24 h with DMSO (carrier solution) with or without DEX (1 μM) or increasing amount of RU486 (0.1, 1, or 10 μM) before luciferase assay. Values represent the mean ± SD of three independent transfections. *p ≤ 0.05; **p ≤ 0.01, significantly different from cells transfected without pCMV-hGRα; #(p ≤ 0.01), significantly different from DEX treatment without RU486. Filled diamonds, pGL3-3kP3-Luc/DEX; empty squares, pGL3-3kP3-Luc/DMSO; filled triangles, pGL3-3kP1-Luc/DEX