Elevated Labile Iron Contributes to Membrane Repair Deficits in FSHD

Facioscapulohumeral muscular dystrophy (FSHD) is an autosomal dominant muscle disease caused by expression of the transcription factor DUX4, which we previously found to be associated with plasma membrane (PM) repair deficits. However, the molecular mechanisms associated with poor membrane repair are not clear. Here, we show PM injury itself results in mild but significant increase in DUX4 mRNA expression, as well dysregulation of genes enriched in pathways critical for membrane repair. We find these dysregulated genes to be sensitive classifiers of FSHD status in human muscle biopsies, indicating their potential utility as novel biomarkers. In addition, FSHD myoblasts show dysregulation of genes in the ferroptosis cell death signaling pathway in response to membrane damage. Subsequent experiments indicated FSHD myoblasts present with hallmark features of ferroptotic stress, including elevated labile ferrous (Fe2+) iron, and lipid peroxidation at baseline and post-injury. Furthermore, we demonstrate that the expression of ferroptosis biomarkers is elevated in FSHD muscle biopsies, which is predictive of the presence of inflammation and positively correlates with the degree of fatty infiltration in FSHD skeletal muscle. Increasing labile iron and lipid peroxidation worsens membrane repair in FSHD myoblasts, while treatment with the iron chelator 2,2′-Bipyridyl and ferroptosis inhibitor ferrostatin-1 improve repair. These data are the first to identify signs of ferroptotic stress in FSHD myoblasts, and demonstrate the potential therapeutic benefit of iron chelation and ferroptosis inhibition to improve membrane repair capacity in FSHD.