Fig. 3

HERC2 promoted the stemness of HCC cells. (A) HERC2 knockout Huh7 cells were treated with 50 ng/ml IL-6 for 24 h and then subjected to RNA-seq analysis. GSEA revealed enrichment of the somatic stem cell division-related gene signature in HERC2 knockout cells. B-D HERC2 knockout and overexpression cells were treated with 50 ng/ml IL-6 for 24 h. B RT-qPCR analysis was used to evaluate the mRNA levels of cancer stem cell-related genes. C Flow cytometry analysis was used to determine CD133 expression. D An immunofluorescence assay was conducted for CD133 detection. E CD133-positive and -negative cells were isolated from the Huh7 and Hep3B cell lines, respectively, through magnetic cell sorting. Western blot analysis was performed to detect HERC2 levels. (F) Cells were cultured under 100 × N2, 50 × B27, 20 ng/ml EGF, 10 nmol FGF, 5 μg/ml insulin, and 0.4% BSA conditions for 7 days, scale bars = 100 μm. G Cells were treated with 20 μM sorafenib for 24 h. A flow cytometry assay was used to determine the percentage of apoptotic cells. H The correlation between HERC2 expression and levels of cancer stem cell-related genes (e.g., CD133, CD44, CD90, Epcam, OCT3/4, Bmi1, Nanog, and Zeb1) based on TCGA datasets (n = 369). I The levels of HERC2 in liver tissues from sorafenib-sensitive and -resistant patients were analyzed according to GSE109211 datasets. *p < 0.05, **p < 0.01, ***p < 0.001. Data from one representative experiment of three independent experiments are presented