Fig 2: Hsa_circ_104348 regulated miR-187-3p to affect proliferation, migration, invasion, and apoptosis of HCC cells, and may have effect on Wnt/β-catenin signaling pathway.Huh7 cells were co-transfected with sh-circRNA or sh-NC and miR-187-3p inhibitor or inhibitor NC: A and B Cell proliferation abilities were determined by CCK-8 assay (A) and colony formation assay (B). C and D Cell apoptosis was determined by EdU staining (C) and flow cytometry assay (D). The scale bars were 100 μm. E and F Cell migration and invasion abilities were checked by wound healing assay (E) and transwell assay (F). The scale bars were 50 μm. G The expression of E-cadherin, N-cadherin was determined by IF. The scale bars were 20 μm. H The expression of RTKN2, β-catenin, PCNA, CyclinD1, c-Myc, Bcl-2, Bax, MMP-7, E-cadherin, N-cadherin was determined by western blot. Data were expressed as mean ± SD.

Fig 3: Overexpression of lncRNA GHET1 activated the expression of cyclin D1, c-Myc and MMP-7. A The expression of lncRNA GHET1 in SW480 cells was detected with the RT-PCR. B, C The expression of cyclin D1, c-myc and MMP-7 was determined with the western blotting. ***p < 0.001 vs. control; #p < 0.05, ##p < 0.01, ###p < 0.001 vs. siPITX2-2

Fig 4: Down-regulation of HMGB3 by siRNA enhanced the expression of Wnt/β-catenin pathway proteins. (A) Down-regulation of HMGB3 shows no influences on the methylation degree and expression of miR-216a, but causes a significant decrease in HMGB3 protein. miR-216a levels were measured by qRT-PCR. HMGB3 proteins were detected by western blot, and the representative images were shown. The unprocessed raw data were presented in Supplementary Figure 2 . (N = 3, *P < 0.05). (B) Down-regulation of HMGB3 causes increase of Caspase 3, Caspase 9, E-cadherin, Bax and decrease of Bcl2, N-cadherin, compared with NC siRNA group (N = 3, *P < 0.05). See Supplementary Figure 3 for the raw data of western blot. (C) Down-regulation of HMGB3 inhibits cell migration and invasion (N = 3, *P < 0.05). (D) Down-regulation of HMGB3 suppresses Wnt/β-catenin pathway proteins β-catenin, c-myc, and MMP7, compared with NC siRNA group (N = 3, *P < 0.05). See Supplementary Figure 4 for the raw data of western blot. (E) Down-regulation of HMGB3 weakens cell viability, compared with NC siRNA group (N = 3, *P < 0.05).

Fig 5: PLCD1 suppresses ERK1/2/β-catenin/MMP7 signalling(A) PLCD1-expressing cells and vector-only control cells (MDA-MB-231 and MCF-7) and BT-549 cells transfected with siN.C. and siPLCD1 for 72 h. PLCD1, pERK1/2, ERK1/2, pGSK-3β, GSK-3β, active-β-catenin, β-catenin, and MMP7 protein levels were measured by immunoblotting with β-actin as an internal control, and supernatant MMP7 levels were also measured by ELISA. Data were based on at least three independent assays, and representative images were shown on the left. Means ±SD, **p<0.01. (B) Expression of related proteins was detected by immunoblotting at the indicated time points (0 h, 1 h, 12 h, 24 h) after treatment with U0126 in MDA-MB-231 and MCF-7 cells. Supernatant MMP7 levels were also measured by ELISA. Data were based on at least three independent assays, and representative images were shown. Means ±SD, **p<0.01. (C) Cells transfected with siN.C. or siPLCD1 were treated with or without U0126 for 1 h, and protein levels were measured by immunoblotting in BT-549 cells. Supernatant MMP7 levels were also measured by ELISA. Data were based on at least three independent assays, and representative images were shown. Means ±SD, **p<0.01.