Age-DependentIn VivoConversion of Mouse Cochlear Pillar and Deiters' Cells to Immature Hair Cells by Atoh1 Ectopic Expression

Abstract

Unlike non-mammalian vertebrates, mammals cannot convert inner ear cochlear supporting cells (SCs) into sensory hair cells (HCs) after damage, thus causing permanent deafness. Here, we achieved in vivo conversion of two SC subtypes, pillar cells (PCs) and Deiters’ cells (DCs), into HCs by inducing targeted expression of Atoh1 at neonatal and juvenile ages using novel mouse models. The conversion only occurred in ~10% of PCs and DCs with ectopic Atoh1 expression and started with reactivation of endogenous Atoh1, followed by expression of 11 HC and synaptic markers; a process that took at least 3 weeks in vivo. These new HCs resided in the outer HC region, formed stereocilia, contained mechanoelectrical transduction channels, and survived for more than 2 months in vivo; however, they surprisingly lacked prestin and oncomodulin expression and mature HC morphology. In contrast, adult PCs and DCs no longer responded to ectopic Atoh1 expression, even after outer HC damage. Finally, permanent Atoh1 expression in endogenous HCs did not affect prestin expression, but caused cell loss of mature HCs. Together our results demonstrate that in vivo conversion of PCs and DCs into immature HCs by Atoh1 is age-dependent, and resembles normal HC development. Therefore combined expression of Atoh1 with additional factors holds therapeutic promise to convert PCs and DCs into functional HCs in vivo for regenerative purposes.

Spontaneous regeneration of cochlear supporting cells after neonatal ablation ensures hearing in the adult mouse

“…The present findings also provide a framework for interpreting our previous observation that ectopic Atoh1 overexpression in IBCs and IPhCs in the neonatal cochlea converts them into HCs without altering organ of Corti morphology (54,55). Based on our current findings, we propose that, after the original IBCs/IPhCs convert to HCs due to Atoh1 expression, they are replenished rapidly by transdifferentiation of GER cells.…”

“…GER cells are immediate neighbors of IBCs, IPhCs, and IHCs and may be less differentiated than SCs in the organ of Corti. GER cells exhibit higher levels of Wnt signaling (e.g., β-catenin and Lgr5) (38)(39)(40)52) and lower levels of Notch signaling (53) and respond readily to Atoh1 (atonal homolog 1) ectopic expression (54)(55)(56)(57). GER cells also express Nestin, a stem/progenitor cell marker in the central nervous system (58).…”

Genetic rescue of Muenke syndrome model hearing loss reveals prolonged FGF-dependent plasticity in cochlear supporting cell fates

“…Indeed, immature mouse pillar cells can be induced in cultured cochleae to convert to hair cells when both Notch and FGF signaling are blocked (Doetzlhofer et al 2009), and following noise damage, Notch inhibition in vivo has a limited ability to convert residual cochlear support cells into functional hair cells (Mizutari et al 2013). Furthermore, induced expression of Atoh1 in immature but not mature Deiters' cells promotes hair cell differentiation (Liu et al 2012). Interestingly, mature Deiters' cells continue to express Fgfr3 well past the stage when cochlear FGF expression is extinguished (Hayashi et al 2010).…”

Atoh1 in sensory hair cell development: constraints and cofactors

“…However, this ability is confined to early developmental stages, with the competence of the inner ear epithelium to respond to Atoh1 strongly tapering off shortly after birth (19,20). A second temporal aspect is that although Atoh1 is best described for its role in early hair cell specification, temporally controlled knock-out studies have suggested that it is also involved in later aspects of hair cell differentiation and survival (10,21).…”