Dystrophin Deficiency Creates a Pro-Ferroptotic Environment in Diaphragm of mdx Mice That Is Modified by Diet and...

Duchenne muscular dystrophy (DMD) is characterized by progressive muscle wasting with altered iron homeostasis and dysregulated lipid metabolism. Insulin resistance and obesity are common in DMD, but their impact on disease progression, particularly during glucocorticoid (GC) treatment, remains unclear. The role of ferroptosis, iron-dependent cell death, in DMD muscle pathology remains uncertain. We hypothesized that ferroptotic signaling would increase in mdx skeletal muscle, be further exacerbated by a high-fat, high-sucrose diet (HFHSD), and be attenuated by GC treatment. Male C57 and mdx mice were fed a control diet or HFHSD, with or without prednisolone, for 19 weeks before diaphragm western blot analysis. Dystrophic diaphragms had an increased abundance of transferrin (131%, p = 0.0006), transferrin receptor (72%, p = 0.0041), ferritin heavy chain (281%, p < 0.0001), acyl-CoA synthetase long-chain family member 4 (36%, p = 0.0080), and 5-lipoxygenase (101%, p = 0.0016) compared with C57. Despite these pro-ferroptotic changes, malondialdehyde and 4-hydroxynonenal were unchanged. Glutathione peroxidase 4 was robustly increased (800%, p < 0.0001), suggesting maintenance of redox balance and suppression of ferroptotic death. The HFHSD did not exacerbate ferroptotic signaling, whereas GC treatment partially offset disease-related changes in iron-handling and antioxidant proteins. These findings indicate that dystrophic diaphragms experience ferroptotic stress that is actively constrained, likely by compensatory GPX4 changes.