Fig. 4

miR-103a-2-5p and miR-660-3p regulate Wnt2b/β-catenin signaling. a Western blotting for β-catenin and β-tubulin in three sorafenib-resistant cell lines compared with their parental control cell lines. b Box-plot (left) and gene expression level (right) of CTNNB1 expression in TCGA HCC tumor and matched TCGA normal liver tissues along with GTEx data. c Overall survival analysis of HCC patients (N = 182) from TCGA project with high or low CTNNB1 expression levels (defined by RNA sequencing with group cutoff in 75%/25% quartile). d Western blot for Wnt2b, β-catenin and β-tubulin in three sorafenib-resistant cell lines with or without si-circRNA-SORE. e Correlation of relative RNA expression of circRNA-SORE and protein expression of CTNNB1 by immunohistochemistry. The slides were reviewed and scored by an experienced pathologist without the knowledge of patient outcome. f Western blot for Wnt2b, β-catenin, β-tubulin with or without miR-103a-2-5p/miR-660-3p mimics in HepG2-SR. g qPCR analysis for downstream genes of β-catenin. h Western blot for β-catenin and β-tubulin (upper) and CCK8 assays (lower) for HepG2-SR cells transfected with si-circRNA-SORE and over-expressing β-catenin. i Western blot for β-catenin and β-tubulin (upper) and CCK8 assays (lower) for HepG2 cells transfected with si-β-catenin and over-expressing circRNA-SORE. j-k Renilla and firefly dual luciferase reporter assay using Wnt2b 3′-UTR constructs containing wild-type or mutant seed sequences in HepG2-SR cells transfected with negative control (NC) or miR-103a-2-5p/miR-660-3p miRNA mimics. Three different independent experiments with three technical repetitions were performed. Data are expressed as the mean ± SEM. Statistical analyses used Student’s t-test, and p < 0.05 was considered statistically significant. * p < 0.05 and ** p < 0.01. NS; not statistically significant