葫芦素B通过核因子E2相关因子2/溶质载体家族7成员11/谷胱甘肽过氧化物酶4信号通路诱导舌鳞状细胞癌CAL-27细胞铁死亡OA

Objective This study aims to observe whether Cucurbitacin B(CuB)induces ferroptosis in CAL-27 cells derived from tongue squamous cell carcinoma and explore its possible mechanism.Methods CAL-27 cells were treated with different concentrations of CuB(0,5,10,15,20,and 30 μmol/L).Cell proliferation activity was detected using the cell counting kit-8(CCK-8)method,and the median in-hibition concentration(IC50)was calculated.Cell clo-ning and wound healing assays were used to detect the effects of different concentrations of CuB on the prolif-eration and migration of CAL-27 cells.CAL-27 cells were treated with different concentrations of CuB,and the changes in intracellular reactive oxygen species(ROS),Fe2+,GSH,and malondialdehyde(MDA)levels were detected.Western blot(WB)was used to detect the expression levels of nuclear factor erythroid 2-related factor 2(Nrf2),glutathione peroxidase 4(GPX4),and solute carrier family 7 member 11(SLC7A11)proteins.Results CCK-8 results showed that CuB at concentrations of 5,10,15,20,and 30 μmol/L signifi-cantly inhibited the proliferation activity of CAL-27 cells,with an IC50 of 13.93 μmol/L(P<0.001).The plate cloning as-say demonstrated that CuB inhibited the cloning ability of CAL-27 cells(P<0.01).The wound healing assay revealed that CuB significantly shortened the migration distance of CAL-27 cells and inhibited their migration ability(P<0.01).The re-sults of ROS,GSH,MDA,and Fe2+measurements indicated that,compared with the control group,the fluorescence inten-sity of ROS and Fe2+in CAL-27 cells significantly increased after CuB intervention at different concentrations.Mean-while,the intracellular GSH content significantly decreased,and the MDA content significantly increased,which induced cellular oxidative damage.These changes could be reversed by the ferroptosis inhibitor Fer-1(P<0.01).WB results showed that CuB intervention at different concentrations significantly downregulated the expression levels of Nrf2,SLC7A11,and GPX4 proteins(P<0.05).Furthermore,the ferroptosis inhibitor Fer-1 reversed the protein expression of Nrf2,SLC7A11 and GPX4(P<0.05).Conclusion CuB can induce ferroptosis in CAL-27 cells derived from tongue squamous cell carcinoma,and its mechanism may be related to the ability of CuB to regulate the signaling pathway in-volving Nrf2,SLC7A11,and GPX4.