Superenhancer-mediated ferroptosis in age-related hearing loss: cochlear epigenomics.

Sp1 was identified as a key upstream transcriptional regulator whose binding to the Fth1 superenhancer decreases with aging.

• Bioinformatics analysis including transcriptional regulatory element enrichment analysis (TREA) and SE prediction with SEdb 2.0 was used to identify Sp1 as a key transcriptional regulator.
• CUT&Tag assays were employed to map Sp1 binding sites and confirm reduced binding to the Fth1 superenhancer in aging conditions.
• Reduced Sp1 binding led to decreased Fth1 gene transcription in hair cells during aging.

Decreased Fth1 transcription resulting from reduced Sp1-superenhancer binding led to increased intracellular iron levels and cellular iron overload.

• Fth1 (ferritin heavy chain 1) is a key gene involved in iron metabolism and storage.
• Reduced Fth1 expression was associated with elevated intracellular iron concentrations in hair cells.
• Cellular iron overload was followed by subsequent ferroptosis and increased reactive oxygen species (ROS) levels.
• Experimental validation was performed using Western blot analysis and quantitative real-time PCR (RT-qPCR) in mouse and cell models.

Iron overload-induced ferroptosis and increased ROS levels promoted hair cell and cochlear aging.

• Ferroptosis is described as a form of regulated cell death associated with iron metabolism.
• Increased ROS levels were observed downstream of cellular iron overload in aging hair cells.
• Immunofluorescence staining was used to validate ferroptosis and ROS accumulation in cochlear hair cells.
• The findings were validated in both mouse models and cell models.

In vivo treatment with the superenhancer inhibitor JQ-1 confirmed the importance of superenhancer activity in maintaining auditory function.

• JQ-1 is described as a SE inhibitor used in in vivo studies.
• Auditory brainstem response (ABR) measurements were used to assess auditory function following JQ-1 treatment.
• JQ-1 treatment confirmed that superenhancer activity is necessary for maintaining normal hearing.
• Statistical analyses were performed using SPSS Statistics 25 and GraphPad Prism.

The study identifies the Sp1–Fth1 superenhancer axis and ferroptosis pathway as potential targets for AAV gene therapy to preserve hearing in aging populations.

• The authors propose that manipulating superenhancer sites and inhibiting ferroptosis may offer novel therapeutic strategies for treating ARHL.
• AAV gene therapy targeting iron homeostasis modulation during sensory cell senescence is suggested as a therapeutic approach.
• The interplay between superenhancers, Sp1, Fth1, and ferroptosis is described as revealing novel targets for intervention.
• The study highlights modulation of iron homeostasis as a mechanism to preserve hearing during aging.