ABSTRACT
Objective To investigate the effects of sodium arsenite (NaAsO2) on the expressions of Cyclin D1 and Cyclin dependent kinase 4 (CDK4) in human normal liver cells (L-02).Methods L-02 cells were exposed to different doses of NaAsO2 (0,50,100,150 μmol/L) for 24 h,flow cytometry was used to detect the cell cycle,real time quantitative PCR and Western blotting were used to detect the Cyclin D1,CDK4 mRNA and protein expression.Results Cell cycle detection:compared with the control group G0-G1 phase [(60.33 ± 0.40)％],except 50 μmol/L NaAsO2 group [(54.58 ± 0.40)％],the numbers of cells in 100 and 150 μmol/L NaAsO2 groups [(64.60 ± 0.62)％,(83.13 ± 0.25)％] were increased,the differences were statistically significant (all P ＜ 0.05);cell proportion of S phase in the control group,50,100 and 150 μmol/L NaAsO2 groups [(34.35 ± 0.30)％,(29.89 ± 0.32)％,(29.50 ± 0.44)％,(11.93 ± 0.12)％] were decreased,the differences were statistically significant (all P ＜ 0.05);cell proportion of G2-M phase was compared between the control group,50,100 and 150 μmol/L NaAsO2 groups [(5.32 ± 0.11)％,(15.54 ± 0.14)％,(5.90 ± 0.20)％,(4.93 ± 0.15)％],the difference was statistically significant (F =908.359,P ＜ 0.05).Cyclin D1 and CDK4 detection:the expressions of Cyclin D1 mRNA (0.48 ± 0.17,1.00 ± 0.31,1.00 ± 0.21,2.06 ± 0.53) and protein (0.65 ± 0.02,0.64 ± 0.05,0.71 ± 0.10,0.84 ± 0.05) were compared betwee the control group,50,100 and 150 μmol/L NaAsO2 groups,the differences were statistically significant (F =167.886,46.575,all P ＜ 0.05);the expressions of CDK4 mRNA (1.04 ± 0.19,1.00 ± 0.21,1.29 ± 0.22,2.31 ± 0.31) and protein (0.67 ± 0.04,0.74 ± 0.03,0.83 ± 0.07,0.94 ± 0.04) were compared betwee the control group,50,100 and 150 μ mol/L NaAsO2 groups,the differences were statistically significant (F =95.171,145.848,all P ＜ 0.05).Conclusions Treatment of L-02 cells with NaAsO2 has changed the expressions of Cyclin D1,CDK4 mRNA and protein,which leads to L-02 cell cycle arrested at G0-G1 phase,ultimately leads to cell damage.
ABSTRACT
Objective To investigate the effects of sodium arsenite (NaAsO2) on cell survival circumstance,reactive oxygen species (ROS) and cell apoptosis in human normal hepatic cells (L-02).Methods L-02 cells were exposed to different doses of NaAsO2 (0,50,100,150 μmol/L) for 24 h.MTT assay was used to detect the survival of L-02 cells,and flow cytometry (FCM) was used to detect the ROS levels and the early (Q4),late (Q2) apoptosis of L-02 cells.Results Cell survival rate:cell survival rate was compared between groups,the difference was statistically significant (F =350.51,P ＜ 0.05),the cell survival rates of 50,100 and 150 μmol/L NaAsO2 groups [(87.30 ± 3.74)％,(49.03 ± 4.72)％,(13.44 ± 4.01)％] were significantly lower than that of the control group [(100.00 ± 0.00)％,all P ＜ 0.05];compared with 50 μmol/L NaAsO2 group,the cell survival rates of 100 and 150 μmol/L NaAsO2 groups were significantly decreased (all P ＜ 0.05);compared with 100 μmol/L NaAsO2 group,the cell survival rate of 150 μmol/L NaAsO2 group was significantly decreased (P ＜ 0.05).The ROS levels:ROS levels were compared between groups,the difference was statistically significant (F =407.78,P ＜ 0.05),the ROS levels of 100 and 150 μ mol/L NaAsO2 groups (3 212.00 ± 221.93,5 521.33 ± 179.63) were significantly higher than that of the control group (1 691.67 ± 73.98,all P＜ 0.05);compared with 50 μmol/L NaAsO2 group (1 927.67 ± 62.45),the ROS levels of 100 and 150 μmol/L NaAsO2 groups were significantly increased (all P ＜ 0.05);compared with 100 μmol/L NaAsO2 group,the ROS level of 150 μ mol/L NaAsO2 group was significantly increased (P ＜ 0.05).Cell apoptosis:cell apoptosis rates of Q2,Q4 and Q2 + Q4 were compared between groups,the differences were statistically significant (F =256.84,26.53,63.89,all P ＜ 0.05);excecpt the cell apoptosis rate of Q4 in 50 μ mol/L NaAsO2 group [(5.43 ± 0.57) ％],the cell apoptosis rates of Q2 [(5.67 ± 0.21)％] and Q2 + Q4 [(11.10 ± 0.40) ％] in 50 μ mol/L NaAsO2 group,the cell apoptosis rates of Q2 [(13.60 ± 0.79) ％],Q4 [(7.37 ± 2.01) ％] and Q2 + Q4 [(20.97 ± 2.38) ％] in 100 μmol/L NaAsO2 group,the cell apoptosis rate of Q2 [(13.47 ± 0.78) ％],Q4 [(16.97 ± 3.45) ％] and Q2 + Q4 [(30.43 ± 3.84) ％] in 150 μmol/L NaAsO2 group were significantly higher than those of the control group [Q2:(3.47 ± 0.12) ％,Q4:(2.90 ± 0.90) ％,Q2 + Q4:(6.37 ± 1.00) ％,all P ＜ 0.05];compared with 50 μmol/L NaAsO2 group,the cell apoptosis rates of Q2,Q4 and Q2 + Q4 in 100 and 150 μmol/L NaAsO2 groups were increased,except the cell apoptosis rate of Q4 in 100 μ mol/L NaAsO2 group,the differences were statistically significant (all P＜0.05);the cell apoptosis rates of Q4 and Q2 + Q4 in 150 μmol/L NaAsO2 group compared with 100 μmol/L NaAsO2 group were significantly increased (all P ＜ 0.05).Conclusions NaAsO2 can induce L-02 cells to increase ROS levels,and inhibit L-02 cell proliferation.In addition,NaAsO2 can induce early apoptosis and late apoptosis in L-02 cells.