Fig. 1.

Sorafenib induces ABCC5 expression in human HCC cells. (A) Data from GSE96793 showed that the expression of ABCC members were markedly increased in sorafenib-resistant cells compared with untreated cells. (B) HuH7 cells were treated with sorafenib (3 µM) for 24 h, and the mRNA expression levels of ABCC members were examined by qRT-PCR (n = 3, *P < 0.05 versus the untreated group). (C) HuH7 cells were treated with sorafenib (1.5 μM and 3 µM) for 24 h, and Western blot analysis evaluated the protein expression of ABCC1, ABCC3 and ABCC5. (D) Human HCC cell lines (HuH7, HepG2 and Sk-Hep-1 cells) were treated with sorafenib (1.5 μM and 3 µM) for 24 h, and the mRNA expression of ABCC5 was examined by qRT-PCR (n = 3, *P < 0.05 versus the untreated group). (E) HepG2 and Sk-Hep-1 cells were treated with sorafenib (1.5 μM and 3 µM) for 24 h, and Western blot analysis detected the expression of ABCC5. (F) The generation of acquired resistance to sorafenib in HuH7 and Sk-Hep-1 HCC cell lines. (G) CCK-8 assays examined the effect of sorafenib on HCC cell activity, and SPSS was used to calculate the drug concentration required to cause death of one-half of HCC cells. (H) Flow cytometry detected the effects of sorafenib (5 μM and 10 μM) on the apoptosis of HCC cells. (I, J) The expression of ABCC5 in HuH7-SR and Sk-Hep-1-SR cell lines was determined by qRT-PCR and Western blot assays. (K) HuH7-SR cells enhanced the anti-cancer activity of sorafenib in vivo. Nude mice were injected with HuH7-SR and HuH7 cells (2 × 10⁶ cells/mouse) into the subcutaneous space on their left and right flanks and were treated with sorafenib (10 mg/kg/i.p., once every other day) at day seven for two weeks (n = 6 mice/group). (L) The tumor weight of each mice was calculated. (M) H&E staining and immunohistochemical staining (expression of ABCC5) were used to visualize the tumor tissue of mice. Representative figures were shown. Scale bars, 50 μm. *P < 0.05, ** P < 0.01, # P > 0.05.