Targeted Inhibition Of YAP1-induced Apoptosis Of Hepatocelular Carcinoma HepG2 Cells By CA3 And Its Mechanism

ObjectiveTo investigate the effect of CA3（CIL56）on the proliferation and apoptosis of Hep G2 cells of liver cancer by targeting the inhibition of YAP1,and to further investigate its anti-tumor mechanism.MethodsHepatocellular carcinoma Hep G2 cells were placed in 1640 medium containing 10%fetal bovine serum（FBS）and incubated at 37℃in a incubator containing 5%CO₂.Hep G2 cells were cultured in vitro,and CA3 concentrations（0.25,0.5,1.0,2.0,4.0mmol/L）were added to the experimental group for 24and 48 hours.The enhanced CCK8 reagent was used to detect the effect of CA3 at different concentrations on Hep G2 cell proliferation.The apoptosis rate of Hep G2 cells was detected by flow cytometry after CA3 treatment at different concentrations for 48h.The changes of reactive oxygen species（ROS）in Hep G2 cells were detected after CA3 treatment at different concentrations for48h.Western Blot analysis of the expression of YAP1,Bcl-2,Bax and Caspase-3 in Hep G2 cells after CA3 treatment at different concentrations for48h.Results1.The enhanced CCK8 results showed that the proliferation rate of cells decreased with the increase of drug concentration after the same time of action of CA3 on Hep G2 cells.After 24 and 48 h of treatment,there was a statistically significant difference in cell proliferation rate between the drug treatment group and the control group（P<0.05）.After CA3 treatment for 48h,the difference within the group was significant.To further explore its mechanism,0.5mmol/L,1.0mmol/L and 2.0mmol/L concentrations were selected for follow-up study.2.Flow cytometry results showed that the apoptosis rate of Hep G2 cells at different concentrations of CA3 increased with the increase of drug concentration for 48 h,and the apoptosis rate increased from 15.56%in the control group to 58.48%in 2.0mmol/L,with statistically significant difference（P<0.05）.3.Reactive oxygen species（ROS）assay results showed that after 48h of CA3 treatment on Hep G2 cells at different concentrations,intracellular ROS content increased significantly with the increase of CA3 drug concentration.4.Western blot results showed that after 48h of treatment with CA3 at different concentrations on Hep G2 cells,compared with the control group,the expression levels of YAP1 and Bcl-2 decreased and the expression levels of Bax and Caspase-3 increased with the increase of drug concentration,with statistically significant differences（P<0.05）.ConclusionCA3 can inhibit the proliferation and induce apoptosis of Hep G2 cells,and the mechanism may be related to the increase of intracellular reactive oxygen production and the regulation of the expression of YAP1,Bcl-2,Bax and Caspase-3 proteins.