NCL impairment affects mobility and aggressiveness of breast cancer cells in vitro. (A and B) Immunoblot analysis using the indicated antibodies of total (A) and nuclear/cytoplasmic (B) extracts from MDA-MB-231 cells treated or transfected as indicated. Numbers under the blots indicate densitometric analysis, normalized for nuclear (DROSHA) or cytoplasmic (GAPDH) markers. (C) Vimentin, Fibronectin, and ICAM1 mRNA expression levels normalized to GADPH mRNA expression in MDA-MB-231 after 96 h of treatment and/or transfection with 10 µM of only aptamers (AS1411 or DCT), or aptamers + miRNAs, or RNA interference (si-NCL or si-Scr). Values represent the mean ± SD. *, P < 0.01. The experiments were performed in triplicate, in three independent experiments. (D) miRNA and pri-miRNA expression levels in MDA-MB-231 cells after different treatments as indicated. Controls (Ctrl) include DCT treatments and si-Scr transfection experiments. Values represent the mean ± SD. The experiments were repeated in triplicate. *, P < 0.01. All data reported are representative of at least three independent experiments. (E) Representative pictures of transwell migration assay (top) and absolute quantifications of cells migrated through the transwell measured by absorbance at 595 nm (bottom) performed on MDA-MB-231 cells transfected with si-Scr, si-Scr + miRNAs, si-NCL, and si-NCL + miRNAs or treated with 10 µM AS1411 or DCT with or without miRNAs. **, P < 0.001. (F) Matrigel invasion assay (top) and absolute quantification (bottom) performed on MDA-MB-436 cells treated as indicated. *, P < 0.01 relative to control-transfected or -treated cells. (E and F) Values represent mean ± SD from three independent sets of experiments performed in triplicate.