Combination of miR-520d-3p and EphA2 siRNA treatment shows enhanced EphA2 inhibition and antitumor efficiency in vitro. Combination of miR-520d-3p and.

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Combination of miR-520d-3p and EphA2 siRNA treatment shows enhanced EphA2 inhibition and antitumor efficiency in vitro. Combination of miR-520d-3p and EphA2 siRNA treatment shows enhanced EphA2 inhibition and antitumor efficiency in vitro. A, immunoblotting of EphA2 and GAPDH in HeyA8 and SKOV3ip1 cells after treatment with miR-520d-3p, different EphA2-targeting siRNAs, or a combination of both (1–6). B and C, representative images showing the effect of different combination treatments (1–4) on SKOV3ip1 and HeyA8 migration (B) and invasion (C) using Transwell migration assay (left). Cells were counted in 10 random fields per well at ×40 after 6 hours for migration and 24 hours for invasion, and the percentage migratory or percentage invasive cells were calculated compared with control treatment. A representative experiment is shown at right. The experiment was carried out in duplicate at three independent times. D, representative images showing the effect of rescue treatment with anti-miR-520d-3p in different combinations (1–6) on SKOV3ip1 migration using Transwell migration assay (left). Absorbance was measured at 590 nm after 24 hours and the percentage migratory cells was calculated compared with control treatment. The data from one representative experiment are shown at right. The experiment was carried out in triplicate at three independent times. Statistical significance was determined by unpaired, two-tailed Student t test when compared with empty clones for all analyses. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001. Data are mean ± SD. NS, not significant. Masato Nishimura et al. Cancer Discovery 2013;3:1302-1315 ©2013 by American Association for Cancer Research