Dynasore Blocks Ferroptosis through Combined Modulation of Iron Uptake and Inhibition of Mitochondrial Respiration
Abstract
:1. Introduction
2. Materials and Methods
2.1. Reagents
2.2. Cell Lines and Culture Conditions
2.3. Transfection with Small Interfering RNA (siRNA)
2.4. Cell Viability Assays
2.5. Fluorescence-Activated Cell Sorting (FACS) Assays
2.6. Western Blotting
2.7. Relative Intracelluar Iron Quantification
2.8. GSH Measurement
2.9. Lipid ROS Quantification
2.10. Mitochondrial ROS Quantification
2.11. General Cellular ROS Quantification
2.12. Seahorse Assay in Intact Cells
2.13. Seahorse Assay on Isolated Mitochondria
2.14. DPPH Assay
2.15. Time-Lapse Cell Death Assays
2.16. Quantification and Statistical Analysis
3. Results
3.1. Dynasore Blocks Transferrin Receptor Uptake and Ferroptosis
3.2. Inhibition of Dynamin 1- and 2-Regulated Iron Uptake is Insufficient to Block Ferroptosis
3.3. Dynasore Protects Neuronal Cells from Ferroptosis and Blocks Mitochondrial ROS Formation and Respiration
3.4. Dynasore Functions as a Broadly Effective Radical-Trapping Agent
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
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Clemente, L.P.; Rabenau, M.; Tang, S.; Stanka, J.; Cors, E.; Stroh, J.; Culmsee, C.; von Karstedt, S. Dynasore Blocks Ferroptosis through Combined Modulation of Iron Uptake and Inhibition of Mitochondrial Respiration. Cells 2020, 9, 2259. https://doi.org/10.3390/cells9102259
Clemente LP, Rabenau M, Tang S, Stanka J, Cors E, Stroh J, Culmsee C, von Karstedt S. Dynasore Blocks Ferroptosis through Combined Modulation of Iron Uptake and Inhibition of Mitochondrial Respiration. Cells. 2020; 9(10):2259. https://doi.org/10.3390/cells9102259
Chicago/Turabian StyleClemente, Laura Prieto, Malena Rabenau, Stephan Tang, Josefina Stanka, Eileen Cors, Jenny Stroh, Carsten Culmsee, and Silvia von Karstedt. 2020. "Dynasore Blocks Ferroptosis through Combined Modulation of Iron Uptake and Inhibition of Mitochondrial Respiration" Cells 9, no. 10: 2259. https://doi.org/10.3390/cells9102259