Mechanism study of Chaetocin as a TrxR1 inhibitor inducing apoptosis in ovarian cancer cells
Objective Investigate the role of Thioredoxin Reductase 1(TrxR1)and the associated molecular pathways in Chaetocin-induced apoptosis in ovarian cancer cells in vitro.Methods Ovarian cancer cell line OVCAR-3 was cultured in vitro.The CCK-8 assay was used to evaluate the effects of Chaetocin on cell proliferation and TrxR1 activity in OVCAR-3 cells.Molecular docking and molecular dynamics analysis were performed to investigate the interaction between Chaetocin and TrxR1.Transfected the lentiviral vector(TrxR1-OE)overexpressing TrxR1 and the empty vector(Vec)into OVCAR-3 cells,and divided them into:control group transfected with Vec(Vec group),TrxR1-OE group overexpressing TrxR1,and OVCAR-3 cell group transfected with TrxR1 or Vec treated with Chaetocin(Chaetocin + TrxR1-OE group and Chaetocin + Vec group).DCFH-DA assay was used to measure the expression of reactive oxygen species(ROS)in OVCAR-3 cells of each group.Annexin V-FITC/PI assay was performed to determine the apoptosis rate of cells in each group.Western blot analysis was conducted to detect the expression of apoptosis-related proteins Cleaved-PARP,Bax,Cleaved-caspase-3,as well as the ratio of p-JNK/JNK and p-c-JUN/c-JUN in the JNK/c-Jun signaling pathway.Results Chaetocin significantly inhibited the proliferation of OVCAR-3 cells and directly interacted with TrxR1 to inhibit its activity.Compared to the Vec group,there were no significant changes in ROS levels,apoptosis rate,expression of Cleaved-PARP,Bax,Cleaved-caspase-3,and the ratio of p-JNK/JNK and p-c-JUN/c-JUN in the TrxR1-OE group(P>0.05).Compared with the Vec group,the Vec +Chaetocin group and TrxR1-OE +Chaetocin group showed significant increases(P<0.05).Compared with the Vec +Chaetocin group,the TrxR1-OE +Chaetocin group showed a significant decrease in ROS content,cell apoptosis rate,expression of Cle-PARP,Bax,Cle-caspase-3 in cells,and p-JNK/JNK,p-c-JUN/c-JUN ratios,with statistical significance(P<0.05).Conclusion Chaetocin can induce the accumulation of reactive oxygen species(ROS)and promote caspase-dependent apoptosis in ovarian cancer cells by activating the JNK/c-JUN pathway and inhibiting TrxR1 activity in vitro.
Chaetocinthioredoxin reductase 1(TrxR1)ovarian cancerJNK/c-JUN signal
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