Fig. 3

circFIRRE affects OS proliferation, migration and invasion in vitro. A Relative mRNA expression of circFIRRE and linear FIRRE in both MG63 and U2OS cells was examined by RT-qPCR after stable infection of either shRNAs lentivirus (sh-circFIRRE-1 and -2) or scramble shRNA lentivirus (sh-N.C.) (n=3 in each group). B-C CCK8 assay was applied to measure the cell viability influenced by circFIRRE knockdown at different time points in both MG63 and U2OS (n=6 at each time point). D-F EdU assay was performed to detect cell proliferation after circFIRRE silencing in MG63 and U2OS. S-phase entry was visualized by EdU incorporation (red); DAPI-stained nuclei (blue). Scar bar=200 μm. Image quantification conducted as described in methods (n=5 in each group). G-I Cell migration and invasion were detected by transwell assay after circFIRRE silencing in MG63 and U2OS cells. Scar bar=400 μm. Image quantification conducted as described in methods (n=5 in each group). J Relative expression of circFIRRE and linear FIRRE in both MG63 and U2OS cells was detected by RT-qPCR after transfection of circFIRRE overexpression vectors or scramble vectors (n=3 in each group). K-L CCK8 assay was applied to measure the cell viability after circFIRRE overexpression at different time points in MG63 and U2OS (n=6 at each time point). M-R EdU and Transwell assays were employed to detect cell proliferation, migration and invasion after circFIRRE overexpression in MG63 and U2OS. Scar bars=200 μl and 400 μm. Values are presented as mean ± SD; the bar charts, line charts, error bars and dots represent the quantitative analysis of 3 independent experiments; two-way ANOVA was used; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; ns = not significant