Figure 3

miR-378a-3p involvement in apoptosis, cell proliferation and migration of RMS cells. (A) FACS analysis. Increase of apoptotic cells (Annexin V-positive) in miR-378a-3p compared to miR-Control positive RH30, 72 h after transfection. Results in the bar chart are the means ± standard deviation (SD) of three separate experiments. (B) Immunoblot analysis, 72 h after transfection. Reduction of phosho-AKT (pAKT) levels and up-regulation of cleaved caspase-3 in RH30 positive cells in comparison to miR-Ctr transfected cells. Total AKT andcaspase-3 levels were similar in both samples. Tubulin was used as loading control. (C) MTT assay. Alteration of the cell viability/proliferation rate in miR-378a-3p transfected RH30 compared with mock control cells. Results are means ± SD of sextuplicate points of two separate experiments. (D) Migration assay. miR-378a-3p transfected RH30 cells compared to negative control cells showed decreased ability to migrate through Boyd chamber membranes. Representative fields of migrated cells through the membrane, stained with crystal violet dye (20× magnification). Data was obtained from the average migration cell number per field (n =8) ± SD of triplicate experiments performed twice. Asterisks denote data in which miR-378a-3p was significantly different from control (*, p < 0.05; **, p < 0.01; ***, p < 0.001).