Figure 6

COX-2 and EP4 mediated regulation of miR655. (A) COX-2 protein expression in MCF7, MCF7-Mock, MCF7-COX-2 and MCF7-miR655 cells. MiR655 expression in (B) MCF7-COX-2 and (C) SKBR3-COX-2 cells treated with COX-2 inhibitor (NS-398, 10 µM) and EP4 antagonist (ONO-AE3-208, 10 µM). (D) Comparison of fold changes in miR655 expression relative to controls (vehicle treatment) in a panel of breast cancer cell lines treated with an EP ligand (PGE2; 10 µM), EP4 receptor agonist (PGE1OH; 10 µM), or vehicle (DMSO). Comparison of miR655 expression after stimulation with PGE2 and PGE1OH followed by treatment with two PI3K inhibitors, Wortmannin (WT), LY-240-002 (LY) both at (10 µM) and ERK inhibitor U0126 (10 µM) in (E) MCF7 and in (F) T47D cells. To test intermediary role of NF-κB in EP4 mediated regulation of miR655 in (G) MCF7 and (H) T47D cells, the cells were treated with NF-κB inhibitor BAY-11-7082 (10 µM) or vehicle (DMSO) along with PGE2 and PGE1OH. Quantitative data are presented as means of triplicate experiments ± SEM. *Indicates p < 0.05, **p < 0.01, ***p < 0.005.