Objective To investigate the expression of protein phosphatase 2 A catalytic subunit in hepatoma cells after acute and chronic injury induced by hydrogen peroxide and aflatoxin B1. Methods Hepatoma cells( HepG2/Huh-7 cells) were treated with different concentrations of hydrogen peroxide( 0,50,100,200,400,and 800 μmol/L) for 2,4,6,and 8 hours or aflatoxin B1( 0,1. 25,2. 5,5,10,20,and 40 μmol/L) for 24 and 48 hours. An inverted microscope was used to observe cell morphology,resazurin assay was used to measure the activity of hepatoma cells,and Western blot was used to measure the expression of demethylated PP2 Ac( dem-PP2 Ac) and total PP2 Ac protein. The independent samples t-test was used for comparison between two groups,a one-way analysis of variance was used for comparison between multiple groups,and the two-way analysis of variance was used for factorial design data. Results HepG2/Huh-7 cells were sensitive to hydrogen peroxide and aflatoxin B1,and with the increases in the concentration of hydrogen peroxide and AFB1 concentration or the prolonged incubation time,there were changes in cell morphology and a reduction in the number of cells. At 2,4,6,and 8 hours of the treatment with hydrogen peroxide at a concentration of 100 μmol/L or above,the activity of HepG2/Huh-7 cells gradually decreased with the increase in the concentration of hydrogen peroxide,and the one-way analysis of variance showed a significant difference between suchcells and the control group( all P < 0. 05). The two-way analysis of variance showed no significant difference in IC50 between the two cell lines during these four periods of time( P > 0. 05). At 24 and 48 hours of the treatment with aflatoxin B1 at a concentration of 2. 5 μmol/L or above,the activity of HepG2/Huh-7 cells gradually decreased with the increase in the concentration of aflatoxin B1,and the one-way analysis of variance showed a significant difference between such cells and the control group( all P < 0. 05). The two-way analysis of variance showed no significant difference in IC50 between the two cell lines during these two periods of time( P > 0. 05). After HepG 2 cells were induced by 200 μmol/L hydrogen peroxide for 2 hours,this group of cells had significantly higher expression levels of total PP2 Ac protein and dem-PP2 Ac than the control group( t =-5. 553 and-2. 990,P = 0. 005 and 0. 040); after Huh-7 cells were induced by 400 μmol/L hydrogen peroxide for 2 hours,this group of cells had significantly higher expression levels of total PP2 Ac protein and dem-PP2 Ac than the control group( t = 0. 073 and-5. 149,P = 0. 002 and 0. 007). After HepG 2 cells were induced by 10 μmol/L aflatoxin B1 for 2 hours,this group of cells had significantly higher expression levels of total PP2 Ac protein and dem-PP2 Ac than the control group( t =-4. 561 and-5. 788,P = 0. 010 and 0. 004); after Huh-7 cells were induced by 20 μmol/L aflatoxin B1 for 2 hours,this group of cells had significantly higher expression levels of total PP2 Ac protein and dem-PP2 Ac than the control group( t =-5. 120 and-6. 144,P = 0. 007 and 0. 004).Conclusion Morphological changes,a reduction in cell activity,and increases in the expression of total PP2 Ac and dem-PP2 Ac are observed after acute and chronic injury of hepatoma cells induced by hydrogen peroxide and aflatoxin B1.
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