Co-overexpression of Atoh1, Pou4f3, and Gfi1 enhances the transdifferentiation of supporting cells into hair cells in the neonatal mouse utricle.

Hair cells (HCs) are essential for vestibular function, and irreversible damage to vestibular HCs in mammals is closely associated with vertigo. The stimulation of HC regeneration through exogenous gene delivery represents an ideal therapeutic approach for restoring vestibular function. Overexpression of Atoh1, Pou4f3, and Gfi1 (collectively referred to as APG) has demonstrated efficacy in promoting HC regeneration in the cochlea. However, the effects of APG on vestibular HC regeneration remain unclear. Here, we used adeno-associated virus-inner ear (AAVie) as a carrier to deliver APG to the utricles of neonatal mice and assessed the morphology and number of HCs and supporting cells (SCs) by immunofluorescence staining. GLAST;Rosa26 mouse line was used to trace SCs. The results showed that APG overexpression resulted in substantial SC transdifferentiation into HCs in the neonatal mouse utricle. Furthermore, APG overexpression maintained SC number by facilitating SC proliferation. Continuous Atoh1 overexpression caused stereocilia damage, which was alleviated by APG overexpression. This study highlights the potential of regulating multiple transcription factors to promote vestibular HC regeneration.