Fig 1: ß-Catenin deletion represses E2-induced Snail expression and EMT in vitro. A, Representative Western blot analysis showing the levels of ß-catenin, Snail, E-cadherin and vimentin in ß-catenin siRNA- and/or Snail siRNA-transfected Ishikawa cells cultured with E2 for 48 h. B, Representative photomicrographs of the migration and invasion of ß-catenin siRNA- and/or Snail siRNA-transfected Ishikawa cultured with E2 for 48 h. (*P < .05 compared to siCON group, n = 3). Data are shown as the mean ± SD (Nod-ph-ß-cat = dephosphorylated ß-catenin; siCON = scrambled siRNA; si-SN = Snail siRNA)
Fig 2: E2 stimulates ß-catenin and Snail colocalization in the nucleus and induced Snail transcription through ß-catenin. A, Representative confocal microscopy images of EECs immunostained for ß-catenin(green) and Snail (red). B, The binding of ß-catenin to the Snail proximal promoter in response to E2 treatment was determined by ChIP assay and quantified by PCR.C-F, Snail promoter activity was measured under the following conditions. C, Ishikawa cells were transfected with different concentrations of ß-catenin overexpression plasmid, ranging from 0 to 200 ng, and then incubated with DMSO or E2 (*P < .05 compared to E2(-)+ß-catenin(-)group, n = 3). D, Ishikawa cells were transfected with different concentrations of ß-catenin siRNA, ranging from 0 to 100 pmol, and then incubated with DMSO or E2 (*P < .05 compared to E2(-)+siß-cat(-) group, #P < .05 compared to E2(+)+siß-cat(-) group, n = 3). E, ß-catenin/TCF-3 overexpression plasmid-transfected Ishikawa cells were cotransfected with different length wild-type Snail promoters (*P < .05 compared to Luc(-2084~+50)+pcDNA3.0 group, #P < .05 compared to Luc(-2084~+50)+ß-catenin + TCF3 group, n = 3). F, ß-catenin overexpression plasmid- or ß-catenin siRNA-transfected Ishikawa cells were cotransfected with wild-type or mutant Snail promoters and then incubated with DMSO or E2 (*P < .05 compared to WT group, #P < .05 compared to E2 + MUT group, n = 3). The results are expressed as the mean ± SD. (WT = wild type; MUT = mutant; CON = control; siß-cat = ß-catenin siRNA)
Fig 3: SNAI1 reversed the partial impact of miR-133a. (A) RT-qPCR and western blot analysis of SNAI1 expression level in U-87MG ATCC and U251 cells after transfected with miR-153 mimic and SNAI1. (B) The invasive ability of U-87MG ATCC and U251 cells following transfection with miR-153 mimic and SNAI1 compared with only transfection with miR-153. (C) Kaplan-Meier curves between miR-153 expression and glioma patients. *P<0.05; **P<0.01; ***P<0.001. OS, overall survival; DFS, disease-free survival.
Fig 4: Effect of circ_0004712/miR-148a-3p on cell migration. A, Cell migration of E2-treated orE2-untreated ishikawa and End1/E6E7 cells after NC, si-circ and/or miR mimics transfection was analysed by transwell assay, respectively. B, Protein expression of E-cadherin and N-cadherin in E2-treated orE2-untreated ishikawa and End1/E6E7 cells after NC, si-circ and/or miR mimics transfection was analysed by Western blot assay. C, Protein expression of ß-catenin, p-ß-catenin and Snail in E2-treated or E2-untreated ishikawa and End1/E6E7 cells after NC, si-circ and/or miR mimics transfection was analysed by Western blot assay. Results show as mean ± SD. **Means compared with control group, P < 0.05. ##Means compared with E2-treated group, P < 0.05
Fig 5: miR-153 expression was negatively associated with SNAI1. (A) miR-153 expression level in glioma and paracancerous tissues (normal). (B) miR-153 expression level in glioma cells U-87MG ATCC and U251 and normal immortalized gliocyte HEB cells. (C) SNAI1 mRNA level in glioma cells U-87MG ATCC and U251 and normal immortalized gliocyte HEB cells. (D) Correlation between miR-153 and SNAI1. **P<0.01; ***P<0.001.
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