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Fig. 2 | BMC Genomics

Fig. 2

From: The highly expressed 5’isomiR of hsa-miR-140-3p contributes to the tumor-suppressive effects of miR-140 by reducing breast cancer proliferation and migration

Fig. 2

Effect of 5’ isomiR-140-3p overexpression on cell viability, cell cycle and cell migration. a and b MCF10A, MDA-MB-231 and MDA-MB-468 cells were transfected with miRNA mimics (hsa-miR-140-3p and 5’isomiR-140-3p), or mimic-ctrl2 (as negative control), then were incubated for 72 h. a WST-1 reagent was added to the cells and absorbance at 450 nm was measured. Results were normalized to the negative control. Data are presented as average of 6 biological replicates ± standard deviation indicated as error bars. b MCF10A, MDA-MB-231 and MDA-MB-468 cells were transfected with miRNA mimics, 24 h after transfection starved in serum-free media for 24 h and allowed to re-enter the cell cycle for 24 h in full growth medium. Cells were incubated with BrdU for 30 min before fixation and lysis and stained with anti-BrdU-FITC and 7-AAD for subsequent FACS analysis. Data are presented as average and standard deviation of three biological replicates. c Cells were transfected in 6-well plates with miRNA mimics (hsa-miR-140-p3 and 5’ isomiR-140-3p) or mimic miRNA negative control. 48 h later, cells were reseeded in transwell inserts (with 8.0 μm polycarbonate membrane) in starvation medium. The lower well compartment had full growth medium to stimulate cell migration across the insert membrane. After 20 h of migration, transwells were removed and cells were trypsinized from the lower surface of the membrane and counted using flow cytometry (FACSCalibur, BD Biosciences). The absolute number of cells migrating was normalized to the total cell number and values are presented as percentage of cells migrating. Values represent the average of 3 biological replicates ± standard deviation indicated as error bars (*** P ≤ 0.001, ** P ≤ 0.01, * P ≤ 0.05 compared to control, unpaired t test)

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