The Effects Of Low-dose As₂O₃ Combined With AZT On The Autophagy Of Human Hepatocellular Carcinoma HepG2 Cells And Its Possible Mechanism

Objective:To investigate the effects of low-dose arsenic trioxide?As₂O₃?combined with3?-azido-3?-deoxythymidine?AZT?on the autophagy of human liver cancer Hep G2 cells,and explore its possible mechanism.Methods:The Hep G2 cells was divided into blank control group,As₂O₃?2?mol/L?alone group,AZT?20?mol/L?alone group and As₂O₃?2?mol/L?combined with AZT?20?mol/L?group respectively.The formation of autophagosomes in control and combination group was observed by projection electron microscopy.The accumulation of acidic vesicular and autolysosomes in different groups was observed by acridine orange staining.The expression level of Beclin1,Lc3 and P62 mRNAs in different group was detected by Real-time fluorescent quantitative-PCR.The expression level of BECLIN1,LC3?,LC3?/?and P62proteins in different group was detected by western blotting.Results:Projection electron microscopy showed that a large number of autophagosomes were observed in the As₂O₃ combined with AZT grou.Acridine orange staining showed As₂O₃combined with AZT could significantly increase the accumulatio n of acidic vesicular and autolysosomes.Real-time fluorescent quantitative-PCR showed that As₂O₃ combined with AZT could significantly up-regulate the expression level of Beclin1,Lc3 mRN As and down-regulate the expression level of P62 mRNAs.Western blotting showed that As₂O₃combined with AZT could significantly up-regulate the expression level of BECLIN1,LC3?,LC3?/?protein and down-regulate the expression of P62 protein compared with the blank control,As₂O₃ and AZT alone group in human liver cancer HepG2 cells.Conclusions:The level of autophagy in human liver cancer HepG2 cells treated with As₂O₃combined with AZT was significantly higher than that in blank control,As₂O₃ and AZT alone group,this effect may be related to the up-regulation of Beclin1.