Ammonium tetrathiomolybdate improves auditory and vestibular function after gentamicin exposure via the NRF2-GPX4 axis.

Gentamicin-induced inner-ear injury can cause disabling vestibular dysfunction and hearing loss; however, the regulated death programs driving vestibular and cochlear hair cell loss remain incompletely defined. In this study, we identify ferroptosis as a shared and central mechanism of gentamicin-induced vestibular and cochlear toxicity in mice. Gentamicin exposure induced robust lipid peroxidation in both cochlear and vestibular tissues and altered the expression of ferroptosis-associated proteins, most notably glutathione peroxidase 4 (GPX4), a key antioxidant enzyme that was markedly downregulated and closely associated with hair cell degeneration. We further identified ammonium tetrathiomolybdate (TM) as a novel agonist of nuclear factor erythroid 2-related factor 2 (NRF2) and evaluated its therapeutic potential via semicircular canal injection. Mechanistically, TM promoted NRF2 nuclear accumulation and upregulated downstream antioxidant programs, restoring GPX4 expression and suppressing lipid peroxidation and ferroptosis-linked signaling in inner-ear hair cells. Structurally and functionally, TM preserved cochlear and vestibular hair cell integrity after gentamicin exposure and significantly improved auditory and vestibular performance. Together, these findings establish ferroptosis as a convergent mechanism underlying gentamicin-induced cochleovestibular injury and identify TM as a pathway-directed candidate for mitigating gentamicin-induced cochleovestibular ototoxicity.