Figure 4.

Restoration of CCND2 and E2F3 expression reverses the effects of miR-145. (A) SKOV3 and ES-2 cells were transfected with CCND2 and E2F3 overexpression plasmids, and the expression was then examined by western blot analysis. The 2 cells lines were also cotransfected with miR-145 mimic + CCND2 and E2F3 overexpression plasmids or the control vector. (B) Cell viability was measured by Cell Counting Kit-8 assay. (C) SKOV3 and ES-2 cells were stained with propidium iodide and subjected to flow cytometry to determine the distribution of cells at each phase of the cell cycle, and were compared with the NC group. The proportion of G1 phase decreased in the cells transfected with CCND2 and E2F3 overexpression plasmids. (D) Quantification of cell cycle distribution results presented as the mean ± standard error of the mean of 3 independent experiments. *P<0.05 and **P<0.01. (E) Wound healing assay was performed at 24 h post-transfection (magnification, 200×). (E-a) SKOV3 cells at 0 h. Following 24 h, the migration of SKOV3 cells transfected with (E-b) NC was not significantly altered compared with cells transfected with (E-c) miR-145, E2F3 and CCND2. The results indicated that the effects of miR-145 were reversed by CCND2 and E2F3. (F) Invasion assays were performed. Experiments were performed in triplicate. *P<0.05 and **P<0.01 vs. the control. CCND2, cyclin D2; E2F3, E2F transcription factor 3; miR, microRNA; NC, negative control.