关键词:

肝癌, 顺铂耐药, RNF31, 上皮间质转化, AMPK/mTOR

Abstract:

Objective　To explore the role and potential mechanism of ringfinger protein 31 (RNF31) in the regulation of cisplatin resistance in hepatoma cells. Methods　The samples of cancer lesion tissues and adjacent normal tissues from 40 patients with liver cancer who underwent surgical treatment in Shaanxi Provincial Cancer Hospital from July 2020 to December 2023, as well as human normal liver cell line IHHA-1, human liver cancer cell line HepG2, and human liver cancer cisplatin-resistant cell line HepG2/DDP, were collected for the study. The expressions of RNF31 in liver cancer tissues, liver cancer cells, and cisplatin-resistant cells were detected by the quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. The RNF31 over-expressed cell line was constructed. The effects on the proliferation, half maximal inhibitory concentration (IC₅₀), and apoptosis of the HepG2/DDP cells were detected by the cell counting kit-8 (CCK-8) and flow cytometry. The expression levels of E-cadherin, N-cadherin, vimentin, and proteins related to the adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) / mammalian target of rapamycin (mTOR) signaling pathway were detected by the Western blot. t test was used for the statistical analysis. Results　The expression levels of RNF31 mRNA and protein in the liver cancer tissues were lower than those in the adjacent tissues (0.45±0.03 vs. 1.01±0.02 and 0.36±0.03 vs. 1.00±0.02; both P<0.05). The expression levels of RNF31 mRNA and protein in the HepG2 cells (0.49±0.06 and 0.46±0.05) and HepG2/DDP cells (0.30±0.05 and 0.28±0.05) were lower than those in the IHA-1 cells (1.00±0.02 and 1.01±0.02) (all P<0.05). Compared with the control group, the over-expression of RNF31 significantly inhibited the proliferation of the HepG2/DDP cells (1.74±0.23 vs. 1.21±0.15) and the IC₅₀ value (8.01±0.52 vs. 3.53±0.25), and increased the cells' apoptosis rate (5.11±1.34 vs. 23.99±2.06) (all P<0.05). Compared with the control group, the over-expression of RNF31 increased the expression level of E-cadherin protein (1.02±0.03 vs. 2.36±0.18; P<0.05) in the HepG2/DDP cells, and decreased the expression levels of N-cadherin (1.01±0.02 vs. 0.43±0.05; P<0.05) and vimentin (1.01±0.03 vs. 0.32±0.05; P<0.05) proteins. The over-expression of RNF31 significantly reduced the protein expression levels of AMPK (1.02±0.02 vs. 0.42±0.05; P<0.05) and mTOR (1.01±0.02 vs. 0.50±0.06; P<0.05). The over-expression of AMPK significantly reduced the apoptosis rate of the HepG2/DDP cells, and increased the IC₅₀ value, and its regulatory effect could be inhibited by the over-expression of RNF31. Conclusion　RNF31 may reduce cisplatin resistance in hepatoma cells regulating the epithelial-mesenchymal transition mediated by the AMPK-mTOR  singnaling pathway.

Key words:

Hepatocellular carcinoma, Cisplatin resistance, RNF31, Epithelial mesenchymal transition, AMPK/mTOR