Objective To study the changes in biological characteristics of a liver cancer cell line PLC / RAF /5 after repeated exposure to a chemotherapy drug,5-fluorouraci( 5-FU),and to investigate the relationship between drug-resistant liver cancer cells and liver cancer stem cells. Methods A low concentration of 5-FU( 1 μg / ml) was used to treat the human liver cancer cell line PLC / RAF /5 repeatedly to establish the PLC / RAF /5 /5-FU cell line. Morphological differences between the two types of cells were observed. The inhibitory effects of different concentrations of 5-FU( 0,0. 25,0. 5,1,1. 5,and 2 μg / ml) on the proliferation of the two types of cells were determined using the CCK-8 assay. Apoptosis of the two types of cells after exposure to different concentrations of 5-FU( 0. 5,1,and 2 μg / ml) for 48 h was analyzed using flow cytometry. The proportions of side population cells in both types of cells were measured using flow cytometry. The colony-forming ability was compared between the two types of cells by the plate colony-forming assay. The expression of Bax,Bcl-2,ABCG2,and Fox M1 proteins in both types of cells was examined by Western blot. Between-group comparison was performed by t test. Results The PLC / RAF /5 /5-FU cell line was successfully established using the chemotherapy drug 5-FU. Compared with the PLC / RAF /5cells,the PLC / RAF /5 /5-FU cells had a larger volume,fewer protrusions,a changed shape of a long shuttle,and enhanced refractivity.Moreover,compared with the parent cells,the PLC / RAF /5 /5-FU cells had a significantly lower sensitivity to the inhibitory effect of 5-FU on proliferation,a significantly lower proportion of cells at the G0 / G1 phase of the cell cycle,significantly higher proportions of cells at the S and G2 / M phases,significantly higher resistance to apoptosis,a significantly higher proportion of side population cells,and significantly enhanced proliferation( P < 0. 05). According to the results of Western blot assay,the expression of Bcl-2,Fox M1,and ABCG2 proteins in the PLC / RAF /5 /5-FU cells was significantly increased( P < 0. 01),while the expression of Bax protein in the PLC / RAF /5 cells was significantly increased( P < 0. 01). Conclusion Liver cancer stem cells are probably responsible for drug resistance of the PLC / RAF /5-FU cell line. Exposure to a low concentration of 5-FU( 1 μg / ml) can enrich liver cancer stem cell-like cells.
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