Fig 1: Effect of miR-335 transfection on U266B1 cells. SOX4 and SOX4 downstream genes and gene products were investigated at 72 h. (A) RT-qPCR assay. The mRNA modulation (green indicates the up-modulation and orange the down) was generated by normalizing miR-335 gene expression with sham-transfected cells. Statistical analysis was performed between three different control and treated cell values (p < 0.0005 = *** evaluated by t-test). (B) Western blot. Empty plasmid transfected cells, sham-, and miR-335-transfected cells were tested for SOX4, AKT/pAKT (Ser473), PI3K, PTEN, TGF-ß, and HIF1-a. (C) Example of flow cytometry 7AAD analysis of U266B1 cells treated as indicated in panel B. (D) Cell mortality (7AAD positivity) served in miR-335- vs. empty plasmid-transfected cells. Values were normalized with sham-transfected cells. Data are representative of three independent experiments. (p < 0.005 = ** evaluated by t-test).