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

Age-related hearing loss (ARHL), also known as presbycusis, is a prevalent condition among older adults and affects a substantial proportion of the global aging population. The underlying mechanisms of ARHL remain unclear, and this study aimed to explore the role of superenhancers (SEs) and the transcription factor Sp1 in regulating hair cell (HC) aging and ferroptosis, a form of regulated cell death associated with iron metabolism.

We utilized a combination of bioinformatics analysis, including transcriptional regulatory element enrichment analysis (TREA) and SE prediction, with SEdb 2.0 to identify key transcriptional regulators and their target genes. Experimental validation was performed using auditory brainstem response (ABR) measurements, immunofluorescence staining, Western blot analysis and quantitative real-time PCR (RT‒qPCR) in mouse and cell models. Additionally, we employed CUT&Tag assays to map Sp1 binding sites and performed statistical analyses using SPSS Statistics 25 and GraphPad Prism.

Our study revealed that reduced binding of Sp1 to the Fth1 superenhancer triggered HC ferroptosis and the progression of ARHL. We identified Sp1 as a key upstream transcriptional regulator whose binding to the Fth1 SE decreased with aging, leading to reduced Fth1 gene transcription and increased intracellular iron levels. This phenomenon resulted in cellular iron overload, subsequent ferroptosis, and increased reactive oxygen species (ROS) levels, ultimately promoting HC and cochlear aging. In vivo studies with the SE inhibitor JQ-1 confirmed the importance of SE activity in maintaining auditory function.

This study provides evidence for the role of Sp1 and Fth1 in the regulation of HC aging and ARHL. These findings suggest that manipulating SE sites and inhibiting ferroptosis may offer novel therapeutic strategies for treating ARHL. Understanding the interplay between SEs, Sp1, Fth1 and ferroptosis reveals novel targets for AAV gene therapy to preserve hearing in aging populations by modulating iron homeostasis during sensory cell senescence.