Fig 4. Effect of metformin on breast cancer cell migration and EMT.

(A) Scratch assay was performed. MDA-MB-231 cell monolayers were scratched and incubated with 2mM metformin for 24 hrs. After the incubation, the cell monolayers were photographed, and represented photos at 0 hr and 24 hr for both control and metformin-treated cells were shown here. (B) The unfilled gap areas between two ends of the scratch were measured and subsequently plotted. Significant inhibition of cell migration was observed in case of metformin treated well as compared to control. Values represent mean ± SEM of triplicate measurements, ***p < .001 vs. control at 0 hr and ***p < .001 vs. control at 24 hr. (C) RT-PCR analysis was performed using total RNA and gene specific primers. Increased level of epithelial markers E-cadherin and keratin 19 were seen in metformin (2 mM) treated cells. On the other hand, decreased expression of mesenchymal markers vimentin and N-cadherin, and decreased expression of transcription factors Zeb1 and Zeb2 were found in metformin (2mM) treated cells. (D) Bars showed densitometry analysis (ratio of concerned gene/ GAPDH), * p < .05 and **p < .01 vs. control.