Role Of Telomere In Apoptosis And Proliferation Of L02 And HepG2 Cells Induced By As₂O₃

Objective: To investigate the role of telomere, telomerase reverse transcriptase (hTERT)mRNA, telomerase activity and telomere-binding proteins expression in the process of apoptosis and proliferation of L02 and HepG2 cells induced by arsenic.Methods: After cell lines L02 and HepG2 were treated with 12μmol/L As2O3, the apoptosis was detected with flow cytometry. Expressions of hTRET mRNA, telomerase activity, telomere-binding proteins TRF1,TRF2 and PCNA were analyzed by RT-PCR, PCR based silver staining telomeric repeat amplification protocol (TRAP-PCR) and western-blot. The cell chromosome analysis was used to observe the chromosome aberration rates after cell lines L02 and HepG2 were treated with 0.625μmol/LAs2O3 for 2w, 4w, 6w. The expressions of telomerase reverse transcriptase (hTERT) mRNA and telomerase activity were measured by RT-PCR and TRAP-PCR. The expressions of telomere-binding proteins TRF1, TRF2 and PCNA were measured by western blot.Result: Flow cytometry analysis revealed that L02 cells with 12μmol/L As2O3 for 24, 48, 72h, the apoptosis rates were 5.81%±1.00, 8.65%±2.05, 13.15%±1.06 respectively. Treated HepG2 cells with 12μmol/L As2O3 for 24, 48, 72h, the percentage of apoptotic cells were 9.74%±1.22, 21.89%±2.14, 42.72%±0.83 respectively, the apoptosis rates of treated groups were significantly higher than the control group(P<0.05). RT-PCR analysis revealed that treated L02 and HepG2 cells with 12μmol/L As2O3 for72h, the expression of hTRETmRNA was decreased than the control group(P<0.05). TRAP-PCR analysis showed that treated L02 and HepG2 cells with 12μmol/L As2O3 for72h, the expression of telomerase activity was decreased than the control group(P<0.05). Western-blot analysis showed treated L02 and HepG2 cells with 12μmol/L As2O3 for72h, the expressions of telomere-binding proteins were up-regulated than control group(P<0.05). The chromosome aberration rates of double minute chromosomes and chromosome fusion of L02 and HepG2 cells were increased than the control group with 0.625μmol/ LAs2O3 for 6w(P<0.05). RT-PCR and TRAP-PCR analysis revealed that treated L02 and HepG2 cells with 0.625μmol/LAs2O3 for 6w, the expression of hTRETmRNA was increased than the control group(P<0.05), the expression of telomerase activity was increased than the control group(P<0.05). Western-blot analysis showed treated L02 and HepG2 cells with 0.625μmol/L As2O3 for 6w, the expressions of telomere-binding proteins were down-regulated and PCNA was up-regulated than the control group(P<0.05)Conclusion: High concentration arsenic could induce L02 and HepG2 cells apoptosis. Low concentration arsenic could induce L02 and HepG2 cell chromosome aberration and proliferation. Chromosome aberration could make genome instability and was the base of cell carcinogenesis. Alterations of telomerase reverse transcriptase mRNA, telomerase activity and telomere-binding proteins expressions in this course may, in part, explain the paradoxical phenomena of arsenic actions in carcinogenic and anticancer effects.