Temozolomide Drives Ferroptosis via a DMT1-Dependent Pathway in Glioblastoma Cells
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Purpose: Temozolomide is used in first-line treatment for glioblastoma. However, chemoresistance to temozolomide is commonin glioma patients. In addition, mechanisms for the anti-tumor effects of temozolomide are largely unknown. Ferroptosis is a formof programmed cell death triggered by disturbed redox homeostasis, overloaded iron, and increased lipid peroxidation. The presentstudy was performed to elucidate the involvement of ferroptosis in the anti-tumor mechanisms of temozolomide. Materials and Methods: We utilized the CCK8 assay to evaluate cytotoxicity. Levels of lactate dehydrogenase (LDH), malondialdehyde(MDA), iron, and glutathione (GSH) were measured. Flow cytometry and fluorescence microscope were used to detectthe production of reactive oxygen species (ROS). Western blotting, RT-PCR and siRNA transfection were used to investigate molecularmechanisms. Results: Temozolomide increased the levels of LDH, MDA, and iron and reduced GSH levels in TG905 cells. Furthermore, wefound that ROS levels and DMT1 expression were elevated in TG905 cells treated with temozolomide and were accompanied bya decrease in the expression of glutathione peroxidase 4, indicating an iron-dependent cell death, ferroptosis. Our results alsoshowed that temozolomide-induced ferroptosis is associated with regulation of the Nrf2/HO-1 pathway. Conversely, DMT1 knockdownby siRNA evidently blocked temozolomide-induced ferroptosis in TG905 cells. Conclusion: Taken together, our findings indicate that temozolomide may suppress cell growth partly by inducing ferroptosis bytargeting DMT1 expression in glioblastoma cells.
Abstract
Purpose: Temozolomide is used in first-line treatment for glioblastoma. However, chemoresistance to temozolomide is commonin glioma patients. In addition, mechanisms for the anti-tumor effects of temozolomide are largely unknown. Ferroptosis is a formof programmed cell death triggered by disturbed redox homeostasis, overloaded iron, and increased lipid peroxidation. The presentstudy was performed to elucidate the involvement of ferroptosis in the anti-tumor mechanisms of temozolomide. Materials and Methods: We utilized the CCK8 assay to evaluate cytotoxicity. Levels of lactate dehydrogenase (LDH), malondialdehyde(MDA), iron, and glutathione (GSH) were measured. Flow cytometry and fluorescence microscope were used to detectthe production of reactive oxygen species (ROS). Western blotting, RT-PCR and siRNA transfection were used to investigate molecularmechanisms. Results: Temozolomide increased the levels of LDH, MDA, and iron and reduced GSH levels in TG905 cells. Furthermore, wefound that ROS levels and DMT1 expression were elevated in TG905 cells treated with temozolomide and were accompanied bya decrease in the expression of glutathione peroxidase 4, indicating an iron-dependent cell death, ferroptosis. Our results alsoshowed that temozolomide-induced ferroptosis is associated with regulation of the Nrf2/HO-1 pathway. Conversely, DMT1 knockdownby siRNA evidently blocked temozolomide-induced ferroptosis in TG905 cells. Conclusion: Taken together, our findings indicate that temozolomide may suppress cell growth partly by inducing ferroptosis bytargeting DMT1 expression in glioblastoma cells.
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