Fig 1: IL27RA promotes HCC cell proliferation in vitro and in vivo. (A and B) The effects of silencing or overexpressing IL27RA on the proliferation of HCC cells were assessed via the CCK-8 assay. (C–F) A colony formation assay was performed. Left panel: representative images; right panel: quantification analysis. (G, H) Hepa 1–6 cells with stable overexpression or knockdown of IL27RA were implanted subcutaneously into the axillae of C57BL/6 mice. After 27 days, the tumors were excised and photographed (n = 5). (I, J and K–N) H&E staining and IHC staining of Ki67 are shown (n = 5 for each group). All cells were treated with either recombinant murine IL-27 (100 ng ml⁻1) or PBS for 24 h. The data are presented as the mean ± SDs. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001; ns, not significant.
Fig 2: IL27RA promoted liver cancer progression in a TGFβR1 signalling-dependent manner. Huh-7 and SNU398 cells stably transfected with IL27RA overexpression and knockdown of TGFβR1 were used for immunoblotting analysis. (B) The effect of silencing TGFβR1 on the proliferation of HCC cells overexpressing IL27RA was measured via the CCK8 assay. (C, D) Colony formation assay: The left panel shows representative images, and the right panel presents the quantification analysis. (E–G) Hepa 1‒6 cells with stable overexpression of IL27RA and knockdown of TGFβR1 were injected through the hepatic portal vein into C57BL/6 mice. Representative liver images are shown, and the liver/body weight ratio was determined (right panel). H&E staining revealed liver cancer nodules (n = 10). (F) The ratio of liver weight to total body weight was determined (n = 4). (G) Kaplan‒Meier survival analysis was performed to evaluate the overall survival of the mice (n = 6). (H–I) Transwell migration assays were performed to assess the migration ability of HCC cells overexpressing IL27RA following the silencing of TGFβR1. (J–M) Wound healing analysis was performed to assess the migration of HCC cells at 0 and 48 h. Representative images (left panel) and quantification (right panel) are shown as indicated. (N) Hepa 1‒6 cells with stable overexpression of IL27RA and knockdown of TGFβR1 were injected into the spleens of C57BL/6 mice. After 30 days, the livers of the mice were excised and photographed, and a representative liver is shown. H&E staining revealed liver metastatic nodules (n = 10). (O) The number of liver metastatic nodules was determined and analysed via Student’s t- test (n = 4). (P) Kaplan‒Meier survival analysis was performed to evaluate the OS of the mice (n = 6). All human HCC cells were treated with rmIL-27 (100 ng/ml) for 24 h. The data are presented as the mean ± SDs. **p < 0.01; ***p < 0.001; ****p < 0.0001.
Fig 3: Expression of IL27RA in hepatocellular carcinoma and its prognostic significance. (A–C) TCGA database analysis revealed that the transcription level of IL27RA was greater in liver cancer tissues than in paracarcinoma tissues and that high expression of IL27RA was associated with poorer overall survival. (D) The expression levels of IL27RA in HCC tissues (T) and paired noncancerous tissues (N) were detected by Western blotting. (E) The expression levels of IL27RA were analysed via Western blotting in HepaRG cells and other liver cancer cell lines. (F) Immunohistochemical staining of IL27RA in human HCC tissues was performed; representative images are presented. (G) The expression levels of IL27RA in HCC and nontumor tissue were statistically analysed, the results were statistically significant (p < 0.0001). (H) Kaplan–Meier analysis demonstrated that patients with high expression of IL27RA had shorter overall survival. The data are presented as the mean ± SDs.
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