Kangxianruangan granule‑containing serum mediated inhibition of hepatic oval cell differentiation into hepatocellular carcinoma cells via the Wnt‑1/β‑catenin signaling pathway

Abstract

Hepatocellular carcinoma is a malignancy with poor clinical prognosis. Hepatic oval cells (HOCs) tend to differentiate into cancerous hepatocellular carcinoma cells (HCCs) in the tumor microenvironment. The purpose of the present study was to explore the role of kangxianruangan granule (KXRG)-containing serum in inhibiting the differentiation of HOCs into HCCs via the Wnt-1/β-catenin signaling pathway. N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) was applied to induce the transformation of the rat HOC cell line WB-F344 into HCCs. The overexpression plasmid, Wnt-1-up, was utilized to increase Wnt-1 expression. Subsequently, high, medium and low concentrations of KXRG were applied to MNNG-treated WB-F344 cells to assess the inhibitory effect of KXRG on cell differentiation. Flow cytometry was conducted to detect the cell cycle distribution, apoptotic rate and expression of cytokeratin-19 (CK-19) protein in cells. An immunofluorescence double staining protocol was used to detect the expression of Wnt-1 and β-catenin. ELISAs were performed to detect α fetoprotein in the cell supernatants.Reverse transcription-quantitative PCR and western blotting were conducted to detect the mRNA and protein expression levels of Wnt-1, β-catenin, Cyclin D1, C-myc, matrix metalloproteinase-7 (MMP-7), Axin2 and epithelial cell adhesion molecule (EpCAM) in cells. Compared with the normal group, the apoptotic rate, proportion of S phase cells, concentration of AFP in the cell supernatant, level of CK-19 protein, and mRNA and protein expression levels of Wnt-1, β-catenin, Cyclin D1, C-myc, MMP-7, Axin2 and EpCAM were all significantly increased in the model group. Addition of KXRG significantly reduced the aforementioned indicators compared with the model group. Moreover, Wnt-1 overexpression further increased the aforementioned indicators compared with the model group, whereas KXRG significantly inhibited these effects. The results indicated that KXRG inhibited the differentiation of HOCs into HCCs via the Wnt-1/β-catenin signaling pathway, which suggested the potential clinical application of KXRG for the prevention of hepatocellular carcinoma.

Shanxian granule‑containing serum mediated inhibition of hepatic oval cell EMT and malignant transformation via the Wnt/β‑catenin signaling pathway

“…By stimulating WB-F344 cells, MNNG replicated some of the conditions that occurred during the transformation of HOC to HCC cells. 21 Studies have confirmed that the abnormal activity of Wnt/β-catenin signaling pathway is one of the important causes of the occurrence and development of liver cancer, which is related to the progression and metastasis of liver cancer. 22 It has been found that Wnt/β-catenin signaling pathway is involved in the regulation of EMT and malignant transformation of WB-F344 cells.…”