Figure 3.

COUP-TFII is down-regulated by miR-302a. A, Representative Western blot images show levels of COUP-TFII in normal endometrial stromal cells treated with IL-1β (upper panel), TNF-α (middle panel), and TGF-β1 (lower panel) in the presence or absence of actinomycin D (Act D; 1 μM) or cycloheximide (CHX; 1 μM) for 24 hours. These experiments were repeated three times using different batches of cells. B, Representative Western blot (upper panel) and quantitative result (lower panel) show levels of COUP-TFII in normal endometrial stromal cells treated with miR-302a mimics (Mimic). *, Significant difference from control. C, Schematic drawing (upper panel) shows construct of luciferase reporter system containing COUP-TFII 3′UTR. The two seed regions of miR-302a (WT) and mutated sequences (Mut1, Mut2, and Mut1+2) are shown. Lower panel shows relative luciferase activity in endometrial stromal cells transfected with wild-type or miR-302a seed sequence-mutated reporter constructs cotransfected with control miRNA (Con) or miR-302a. *, Significant difference from control miRNAs; #, significant difference compared with wild-type constructs treated with miR-302a. D and E, Quantitative results show miR-302a levels in endometrial stromal cells treated with different doses of IL-1β (D) or 1 ng/ml IL-1β for different lengths of time (E). *, Significant difference from control. F, Quantitative results show levels of miR-302a in normal endometrial stromal cells (Nor; n = 21) and endometriotic stromal cells (Endo; n = 36). *, Significant difference from normal cells by a Student's t test.