Fig. 6

Circ_0003074 regulated KPNA4 expression through miR-516b-5p. A The schematic illustration showing the complementary sites of miR-516b-5p with KPNA4. B, C Dual-luciferase reporter assay was employed to identify the interaction between miR-516b-5p and KPNA4. D, E KPNA4 protein expression was detected by Western blotting in both MG63 cells and Saos2 cells treated with Rg3 at a dose of 0, 40, 80 or 120 µM. F–H KPNA4 expression was detected by qRT-PCR and Western blotting in hFOB1.19 cells, MG63 cells, Saos2 cells, OS tissues and adjacent normal tissues. I, J Spearman correlation analysis was used to confirm the linear correlation between KPNA4 and miR-516b-5p or circ_0003074 in expression in OS tissues. K, L The efficiency of both miR-516b-5p and KPNA4 knockdown was checked by qRT-PCR or Western blotting in MG63 cells and Saos2 cells. M, N The effects between miR-516b-5p knockdown and KPNA4 silencing on KPNA4 expression were determined by Western blotting. O, P Western blotting was carried out to investigate the effects between circ_0003074 overexpression and miR-516b-5p upregulation on KPNA4 protein production. *P < 0.05, **P < 0.01 and ***P < 0.001