Insights Into Ferroptosis: Targeting Glycolysis to Treat Graves' Orbitopathy

Context: Oxidative stress plays an indispensable role in pathogenesis of Graves' orbitopathy (GO). Ferroptosis is a newly discovered form of cell death resulting from lipid peroxidation. Little is known about the role of ferroptosis in GO. Objective: We aimed to identify the divergent role of ferroptosis in the GO and control orbital fibroblasts (OFs). Methods: Orbital fat/connective tissues and serum immunoglobulins (Igs) were collected from GO and control subjects. Cell viability and lipid peroxidation were measured to evaluate ferroptosis sensitivity. Pyruvate dehydrogenase kinase 2 (PDK2) level and oxygen consumption rate were quantified to assess glycolysis status. Results: Primary OFs were cultured from orbital tissues. Ferroptosis was induced by cystine deprivation and/or erastin treatment. The GO OFs possessed stronger resistance to ferroptosis than the control OFs. Selenium, a potential ferroptosis inhibitor, protected the control OFs from ferroptosis. Both transcriptomic and proteomic analyses indicated glycolytic shift in the GO OFs. Metabolic profiling, PDK2 quantification, and oxygen consumption assay confirmed enhanced glycolysis in the GO OFs. Inhibition of glycolysis by PDK2 knockdown and dichloroacetic acid (DCA) promoted ferroptosis sensitivity in the GO OFs. The ferroptosis-sensitizing effects of DCA were also observed when the GO OFs were treated with GO-Igs. IGF1R overexpression in the GO OFs contributed to glycolysis shift. IGF1R inhibitory antibodies facilitated ferroptosis induction in the GO OFs, but the effects were less remarkable under GO-Igs treatment. Conclusion: These study findings establish that glycolysis facilitates ferroptosis resistance in the GO OFs, providing insights into the therapeutic role of glycolysis for GO treatment.