Fig. 6: Collagen-mediated YAP activation is predicted to relate with diversity by enhancing stemness of tumor cells.

A, B The capacity of colony formation was detected by sphere formation assays in Huh7 (liver cancer cell line). Cells with or without YAP overexpression (GFP-labeled) were treated with low (1 µL collagen with 3 µL DMEM) or high (collagen without DMEM) concentrations of collagen for 3 days. Bar, 200 µm. C, D The capacity of colony formation was detected by sphere formation assays in Huh7 cells. Cells with or without YAP knockdown were treated with low (1 µL collagen with 3 µL DMEM) or high (collagen without DMEM) concentrations of collagen for 3 days. Bar, 200 µm. E, F Staining of YAP in Huh7 cells cultivated with low or high concentrations of collagen for 3 days, 200 µm. G, H The capacity of colony formation was detected by sphere formation assays in HepG2 (liver cancer cell lines). Cells with or without ITGA2 knockdown (GFP-labeled) were treated with low (1 µL collagen with 3 µL DMEM) or high (collagen without DMEM) concentrations of collagen for 3 days. Bar, 200 µm. I Staining of YAP and ITGA2 in HepG2 cells (with or without ITGA2 knockdown) for 3 days, 200 µm. J Subcellular localization of YAP in WT and ITGA2-knockdown HepG2 cells. K ChIP experiment of CTGF and CYR61 performed with YAP antibody in HepG2 cells knockdown ITGA2. L mRNA levels of CTGF and CYR61 in WT and ITGA2-knockdown Huh7 cells with or without co-cultured with TAFs. M Staining of COL1A1, ITGA2, and YAP in tumors from liver cancer patients. Bars, 100 µm. Quantified data are presented as the means ± SD. Unpaired Student’s t tests were used for comparing two variables and one-way ANOVA was used for comparing multiple variables. At least two independent experiments were performed for all data. *P < 0.05; **P < 0.01; ***P < 0.001; n.s, no significance in comparison with the control group. See also Fig. S4.