Figure 2
Effects of E 2 and P on cytokine and chemokine production and proliferation in RL95-2 cells. RL95-2 cells were plated at 0.2 × 106 cells/well/ml and maintained in charcoal/dextran-treated DMEM/F12 (CD-F12) for 48 hours prior to experiments. Media was replaced with media containing either 10-8 M E2 or 10-7 M P as determined by dose response. Cells remained in hormone-containing media for 66 hours total and were stimulated with 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 C). RL95-2 cells were plated at 0.2 × 106 cells/well/ml, harvested, and counted after 18 hours of stimulation with vehicle (black bars) or Poly I:C (10 μg/ml, gray bars). The mean of three cell counts of triplicate wells counted for each time point from one experiment is shown (D). Experiments were performed using triplicate wells for each treatment condition and repeated three times. Representative data is 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).