Fig 2: SFN promotes HCC cell proliferation, migration, and invasion in vitro and tumor growth in vivo. The efficiency of overexpression or knockdown of SFN was determined by RT–qPCR and WB in SMMC-7721 (a and b) and MHCC-97H (c and d) cell lines. Wound healing assays (e and f) and transwell migration assays (g and h) were used to explore the migratory ability of HCC cells with upregulated or downregulated SFN expression. (g and h) The invasion capacity of HCC cells with upregulated or downregulated SFN expression was examined by transwell invasion assays. (i and j) Colony formation assays were utilized to investigate the proliferation ability of HCC cells with altered SFN expression. (k and l) CCK-8 assays were used to evaluate the proliferation capacity of MHCC-97H cells with SFN knockdown and SMMC-7721 cells with SFN overexpression. (m) Xenograft mouse models were used to analyze the role of SFN in tumor growth. Images of subcutaneous tumors derived from MHCC-97H cells with SFN knockdown or SMMC-7721 cells with SFN overexpression in nude mice. (n and o) The weight and volume of the tumors in each xenografted mouse in the model. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001. All data are from three independent experiments.

Fig 3: SFN induces EMT and activates Wnt/ß-catenin signaling in HCC cells. (a) The protein levels of E-cadherin, N-cadherin, vimentin, Snail, MMP2, and MMP9 in MHCC-97H cells infected with lentiviruses expressing shSFN#1, shSFN#2, and shNC, as determined by WB. (b) The protein levels of E-cadherin, N-cadherin, vimentin, Snail, MMP2, and MMP9 in SMMC-7721 cells infected with lentiviruses carrying the SFN overexpression plasmid or Vector, as determined by WB. (c) The protein levels of ß-catenin, non-phospho-ß-catenin (active), GSK-3ß, phospho-GSK-3ß (Ser9), c-Myc, and Axin2 in MHCC-97H cells infected with lentiviruses expressing shSFN#1, shSFN#2, or shNC, as determined by WB. (d) The protein levels of ß-catenin, non-phospho ß-catenin (active), GSK-3ß, phospho-GSK-3ß (Ser9), c-Myc, and Axin2 in SMMC-7721 cells infected with lentiviruses carrying the SFN overexpression plasmid or Vector, as determined by WB. (e) The protein level of ß-catenin in the nucleus of MHCC-97H cells infected with lentiviruses expressing shSFN#1, shSFN#2, or shNC, as determined by WB. (f) The protein levels of ß-catenin in the nucleus of SMMC-7721 cells infected with lentiviruses carrying the SFN overexpression plasmid or Vector, as determined by WB. (g) TOP/FOP luciferase reporter activity in MHCC-97H cells with SFN knockdown. (h) TOP/FOP luciferase reporter activity in SMMC-7721 cells overexpressing SFN. All data are from three independent experiments.

Fig 4: SFN gene expression is closely associated with poor prognosis. (a and b) The OS and DFS of HCC patients in the low (n = 17) and high (n = 17) SFN expression cohorts. (c–f) The OS, PFS, RFS, and DSS of patients with HCC in the Kaplan–Meier plotter online database.

Fig 5: SFN gene expression is upregulated in HCC cells and tissues. (a) SFN mRNA expression in 377 HCC samples compared with 50 nontumor samples was analyzed in the TCGA database. (b, c, d, and e) SFN mRNA expression in HCC tissues in four Oncomine datasets (Roessler liver, Mas liver, Roessler liver 2, Wurmbach liver). (f) SFN mRNA expression was measured in five kinds of HCC cell lines and normal liver cells (L02) by RT?qPCR in three independent experiments. (g) SFN mRNA expression was measured in 34 paired fresh HCC tissues and adjacent nontumor tissues by RT?qPCR. (h) SFN protein expression was measured in five HCC cell lines and in normal liver cells (L02) by WB in three independent experiments. (i) Representative WB images from 34 paired fresh HCC tissues and adjacent nontumor tissues. (j) SFN protein expression in HCC tissues and nontumor tissues in the Human Protein Atlas. **P < 0.01, ***P < 0.001, ****P < 0.0001.