Fig. 2

Lenti-viral transfection of 520d-5p to HMV-I cells in vitro. a A representative phenotype of 520d/HMV-I is presented (x100 magnification). Spheroid-like cells comprising a small fraction of the cell population are presented (top). Analysis of GFP expression confirmed the effective induction (more than 99 %) of 520d/HMV-I cells (x100 magnification) (middle). miR-520d-5p induced the expression of Nanog in HMV-I cells (x100 magnification) (bottom). The phenotypic changes were not sufficient to assess whether they were malignant or benign. b Significant 520d-5p expression was confirmed in 520d/HMV-I cells (*, P < 0.01 by the Mann-Whitney U test). The relative expression of 520d-5p in 520d/HMV-I cells compared with mock/HMV-I cells is presented. All expression data are standardized to the β-actin expression level (n = 4). c Fluorescence activated cell sorting (FACS) analysis revealed that GFP-positive 520d/HMV-I cells (right) exhibited increased DNA content in the S phase compared with GFP-positive mock/HMV-I cells (left). d The invasive abilities of HMV-I, mock-transfected HMV-I (mock/HMV-I) and HMV-I cells treated with miR-520d-5p (520d/HMV-I) were estimated with a migration assay using a fibronectin membrane (10 μg/ml). Most of the 520d/HMV-I cells did not pass through the membrane