Fig. 4

Sponging miR-149-5p is essential for the protumor effects of circUCK2(2,3) both in vitro and in vivo. A and B Colony formation assays (A) and EdU flow cytometry assays (B) in SNU398 cells with WT circUCK2(2,3) or 149-mut circUCK2(2,3) overexpression. C Transwell migration and invasion assays in SNU398 cells with WT circUCK2(2,3) or 149-mut circUCK2(2,3) overexpression. D–F SNU398 cells overexpressing WT circUCK2(2,3) or 149-mut were injected into nude mice subcutaneously. Seven days after injection, tumor volumes were measured three times weekly by using a caliper to draw tumor growth curve (D) (means ± SD, two-way ANOVA). After 23 days of injection, subcutaneous tumors were dissected and imaged (E). Tumor weight comparison among VC, 149-mut, and WT groups (F). G SNU398 cells with WT circUCK2(2,3) or 149-mut overexpression were introduced into nude mice intravenously. Eight weeks after the initial injection, lungs were dissected and lung micrometastases were detected by H&E staining between each group (scale bar, 100 μm and 25 μm). Statistical analyses were performed using paired Student’s t-tests (ns, no significance; **p < 0.01; ***p < 0.001). VC vector control, 149-mut miR-149-5p binding site mutant, WT wild type, DAPI 4′,6-diamidino-2-phenylindole, EdU 5-ethynyl-2′-deoxyuridine