Mechanism of ferroptosis mediated by the bromodomain protein subfamily 4/nuclear factor-kappa B signaling pathway in chemotherapy resistance of triple-negative breast cancer
Objective:To investigate the mechanism of ferroptosis mediated by the bromodomain protein subfamily 4(BRD4)/nuclear factor-kappa B(NF-κB)signaling pathway in the malignant biological behavior of breast cancer.Methods:Doxorubicin-resistant MDA-MB-231 cells(MDA-MB-231/DOX)were divided into control group(Con group),DOX group,and si-BRD4+DOX group.In the si-BRD4+DOX group,MDA-MB-231/DOX cells were transfected with si-BRD4 for 48 hours and were then treated with 1 μmol/L DOX for 24 hours.Colony formation assay and 5-ethynyl-2'-deoxy-uridine(EdU)analysis were used to evaluate the proliferation ability of cells,and FerroOrange and liperfloo were used to measure the concentration of ferrous ion and the level of lipid peroxidation.A total of 15 female BALB/c-nu mice were randomly divided into control group,DOX group,and DOX+Si-BRD group,with 5 mice in each group,and a subcutaneous inoculation model of MDA-MB-231/DOX cells was established.The methods of qRT-PCR,Western blotting,and immunohistochemistry were used to measure the expression of the BRD4/NF-κB signaling pathway.Results:Compared with the si-NC group,the si-BRD4#1 group and the si-BRD4#2 group had significant reductions in the number of cell clones,positive EDU staining,and glutathione level in MDA-MB-231 and BT549 cells(P＜0.001),as well as significant increases in the level of ferrous ion,the level of reactive oxygen species(ROS),and oxidized glutathione/glutathione ratio(P＜0.01).Compared with the paren-tal cells(MDA-MB-231),there were increases in the mRNA and protein expression levels of BRD4 in MDA-MB-231/DOX cells.Compared with the Con group,the DOX group had significant increases in the expression levels of IKβ-α,NF-κB,and BRD4 and the level of ROS(P＜0.05)and significant reductions in the number of cell clones and positive EDU staining(P＜0.05);compared with the DOX group,the si-BRD4+DOX group had significant reductions in the expression levels of IKβ-α and NF-κB,the number of cell clones,and positive EDU staining(P＜0.05)and a significant increase in the level of ROS(P＜0.05).Compared with the control group,the DOX group had significant reductions in tumor weight and volume(P＜0.05)and a significant increase in the number of TUNEL-stained cells in tumor tissue(P＜0.05).In addition,compared with the DOX group,the BRD4+DOX group had further reductions in tumor weight and volume(P＜0.001)and a significant increase in the number of TUNEL-stained cells(P＜0.001).Compared with the DOX group,the BRD4+DOX group had significant reductions in Ki-67,BRD-4,and NF-κB(P＜0.01)and a significant increase in 4-hydroxynonenal(P＜0.01)in tumor tissue.Conclusion:BRD4 is an important drug resistance factor,which can inhibit ferroptosis induced by chemotherapy in breast cancer by promoting the activation of the NF-κB signaling pathway.
bromodomain protein subfamily 4nuclear factor-kappa Bferroptosisbreast cancer
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