Fig. 4

miR-21 overexpression restores stem cell-like characteristics of gastric cancer cells which were inhibited by Bmi-1 knockdown. a miR-21 overexpression restores microsphere formation of Bmi-1 knockdown MKN45 cells. Microsphere formation in control cells (Ctrli), Bmi-1 gene interference cells (Bmi-1i), and cells with Bmi-1 interference and simultaneous overexpression of miR-21 (Bmi-1i+miR-21) was detected by serum-free suspension culture (upper panel) and quantified (lower panel). b miR-21 overexpression increases drug resistance in MKN45 cells which was inhibited by Bmi-1 knockdown. Cell viability in Ctrli, Bmi-1i, and Bmi-1i+miR-21 MKN45 cells treated with different doses of EPI for 48 h was determinated with CCK-8 method. c miR-21 overexpression enhances migration ability in MKN45 cells which was inhibited by Bmi-1 knockdown. Migration ability in Ctrli, Bmi-1i, and Bmi-1i+miR-21 MKN45 cells was tested by Transwell migration assay. d miR-21 overexpression in Bmi-1 knockdown MKN45 cells was confirmed by QRT-PCR. Fold change of miR-21 in Ctrli, Bmi-1i, and Bmi-1i+miR-21 MKN45 cells was determinated by QRT-PCR. e miR-21 overexpression enhances tumorigenecity in MKN45 cells which was inhibited by Bmi-1 knockdown. In vivo tumorigenecity in Ctrli, Bmi-1i, and Bmi-1i+miR-21 MKN45 cells was detected by subcutaneously cancer cells injected SCID mice model. f miR-21 overexpression enhances implantation metastasis of MKN45 cells which was inhibited by Bmi-1 knockdown. In vivo implantation metastasis potential in Ctrli, Bmi-1i, and Bmi-1i+miR-21 MKN45 cells was detected by intraperitoneal cancer cells injected SCID mice model. g miR-21 overexpression restores the expression of stem cell markers and miR-21 target genes (PTEN, P53) in Bmi-1 knockdown MKN45 cells. The expression of Bmi-1, stem cell markers and miR-21 target genes (PTEN-AKT, P53) in Ctrli, Bmi-1i and Bmi-1i+miR-21 MKN45 cells was analyzed by Western blot. Error bars in all panels represent the mean ± SD (*P < 0.05, **P < 0.01)