Fig. 4.

Effect of CuET on the ERK pathway, and the proliferation, migration and invasion properties of different HCC cells.

A. Qualitative and quantitative analyses of p-ERK levels in MHCC-97H/REGO, MHCC-97H, SMCC-7721, and MHCC-LM3 cells by western blot. Total protein was extracted after the cells were treated with or without CuET (0.1 μM) for 24 h. B. The viability of MHCC-97H/REGO, MHCC-97H, SMCC-7721 and MHCC-LM3 cells exposed to the indicated concentrations of CuET for 24 h was determined by the CCK-8 assay. Cells were plated in 96-well plates at a density of 10³ cells per well. C. Colony formation assay of MHCC-97H/REGO, MHCC-97H, SMCC-7721 and MHCC-LM3 cells. Cells were plated at a density of 10³ cells per well and treated with vehicle or CuET at the indicated concentration for 24 h. The treatment-containing medium was then replaced with medium alone for 6 days. D. Cell cycle distributions of different CuET-treated HCC cell lines. Qualitative and quantitative analyses of the expression of cyclin B1 in different HCC cell lines treated with or without CuET (0.1 μM) for 24 h by western blotting. E. Wound healing experiments were performed on different HCC cell lines for 0 h, 24 h and 48 h. Cells were treated with or without CuET (0.1 μM). F. Transwell assays of different HCC cell lines for 24 h. Cells were treated with or without CuET (0.1 μM). G. Qualitative and quantitative analyses of different HCC cell lines treated with or without CuET (0.1 μM) were performed via western blotting to detect changes in the protein levels of EMT-related molecules. n.s: no significant difference, *p < 0.05, **p < 0.01, ***p < 0.001 (one-way ANOVA).