Fig. 3
MenSCs reverse sorafenib resistance in HCC-SR cells. A, B The regulation of BNIP3 and BNIP3L expression by MenSCs in HCC-SR cells was, respectively, determined using immunoblotting (protein) and qRT-PCR (mRNA) analyses. MenSCs significantly upregulated BNIP3 and BNIP3L expression in HepG2-SR and Huh7-SR cells. Cells were cultured under normoxia or hypoxia (1%O2) for 48 h. C, D Cell viability and apoptosis of HCC-SR cells were determined using CCK-8 and Annexin V/PI analyses, respectively. Combination therapy resulted in a further reduction in cell viability (C) and an increase in apoptosis (D) in HepG2-SR and Huh7-SR cells compared to sorafenib alone. Cells were cultured under hypoxia for 48 h. Sorafenib: 10 µM. E Cell proliferation of HCC-SR cells was determined using clone formation assay. Combination therapy resulted in a further reduction in cell proliferation compared to sorafenib alone. F Cell proliferation ability was determined using clone formation assay after the levels of BNIP3 or BNIP3L in HCC-SR cells were modulated as indicated. Knocking down BNIP3 or BNIP3L rescued the decreased proliferation observed after combination therapy in HepG2-SR and Huh7-SR cells. Cells in E, F were cultured in complete DMEM medium containing 6 µM sorafenib for one week; and cells receiving combination therapy were cocultured with MenSCs during the test. G Cell viability was determined using CCK-8 analysis after the levels of BNIP3 and BNIP3L in HCC-SR cells were modulated as indicated. Knocking down BNIP3 and BNIP3L rescued the decreased cell viability observed after combination therapy in HepG2-SR and Huh7-SR cells. Cells were cultured under hypoxia for 48 h. Sorafenib: 10 µM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. ns represents not statistically significant. Full-length blots are presented in Additional file 6