Figure 5
Effects of E 2 in RL95-2 cells are mediated by ER. RL95-2 cells were plated at 0.2 × 106 cells/well/ml and maintained in CD-F12 for 48 hours prior to experiments. Media was replaced with media containing vehicle, 10-6 M ICI 182, 780 (A), or 10-6 M RU486 (B). After 2 hours, media was replaced with the indicated combinations of ethanol vehicle, 10-8 M E2, 10-7 M P, 10-6 M ICI 182, 780, and 10-6 M RU486. Cells remained in the indicated media for a total of 66 hours. Cells were stimulated with ethanol vehicle, Poly I:C (10 μg/ml), or PMA/I (10 ng/ml and 500 ng/ml) 18 hours prior to harvest. A total of 100 μl of cell-free supernatant was used to detect IL-6, IL-8, and IP-10 by ELISA (A, B, and data not shown). Experiments were performed using triplicate wells for each treatment condition and repeated three times. Data representative of three experiments are shown. Error bars indicate standard deviation of three samples. Statistical significance (*) was determined by one-way ANOVA and the Tukey post-hoc test (P < 0.05).