Regeneration of Cochlear Hair Cells and Hearing Recovery through Hes1 Modulation with siRNA Nanoparticles in Adult Guinea Pigs

Abstract

Deafness is commonly caused by the irreversible loss of mammalian cochlear hair cells (HCs) due to noise trauma, toxins, or infections. We previously demonstrated that small interfering RNAs (siRNAs) directed against the Notch pathway gene, hairy and enhancer of split 1 (Hes1), encapsulated within biocompatible poly(lactic-co-glycolic acid) nanoparticles (PLGA NPs) could regenerate HCs within ototoxin-ablated murine organotypic cultures. In the present study, we delivered this sustained-release formulation of Hes1 siRNA (siHes1) into the cochleae of noise-injured adult guinea pigs. Auditory functional recovery was measured by serial auditory brainstem responses over a nine-week follow-up period, and HC regeneration was evaluated by immunohistological evaluations and scanning electron microscopy. Significant HC restoration and hearing recovery were observed across a broad tonotopic range in ears treated with siHes1 NPs, beginning at three weeks and extending out to nine weeks post-treatment. Moreover, both ectopic and immature HCs were uniquely observed in noise-injured cochleae treated with siHes1 NPs, consistent with de novo HC production. Our results indicate that durable cochlear HCs were regenerated and promoted significant hearing recovery in adult guinea pigs through reversible modulation of Hes1 expression. Therefore, PLGA-NP-mediated delivery of siHes1 to the cochlea represents a promising pharmacologic approach to regenerate functional and sustainable mammalian HCs in vivo.

Inner ear organoids: new tools to understand neurosensory cell development, degeneration and regeneration

“…In the organ of Corti of adults, however, the activity of the Atoh1, Wnt and the Notch pathway is reduced, and the effects of Notch inhibition by GSI are consequently minimal compared with the neonatal situation (Basch et al, 2016b;Hartman et al, 2009;Maass et al, 2015). Moreover, responsiveness to GSI-treatment in the adult inner ear may depend on damage; expression of Notch downstream effectors increases after trauma, which causes overt hair cell loss (Batts et al, 2009;Du et al, 2018;Mizutari et al, 2013), but not after exposure to noise that induces less extensive hair cell death (Maass et al, 2015). Activation of canonical Wnt signaling though β-catenin stabilization also results in cell cycle re-entry of supporting cells (Li et al, 2015;Roccio et al, 2015;Samarajeewa et al, 2018) and prolongs the time window for GSI-induced hair cell differentiation.…”

“…Although Wnt and Notch pathway components are expressed throughout early (P0) and late (P8) neonatal stages, targets related to cell proliferation and cell cycle progression are downregulated (Samarajeewa et al, 2018). Unlike the results in the newborn cochlea and vestibular organs (Bramhall et al, 2014;Jeon et al, 2011;Lin et al, 2011;McLean et al, 2017;Yamamoto et al, 2006), drugs and siRNAs targeting Wnt and Notch signaling show limited effects in adult models of hearing loss, such as from noise damage (Du et al, 2018;Mizutari et al, 2013;Tona et al, 2014). The generation of new hair cells through Atoh1 overexpression in some animal studies have demonstrated promise for the recovery of hearing (Izumikawa et al, 2005), although others have shown only marginal functional improvements (Atkinson et al, 2014).…”

Perspectives on Human Hearing Loss, Cochlear Regeneration, and the Potential for Hearing Restoration Therapies

“…The other used guinea pigs as an experimental model, but only tested three animals [ 124 ]. Lastly, siRNA against the NOTCH effector Hes1 have been introduced into the cochleae of guinea pigs after noise exposure [ 125 ]. This intervention drove improvements of 8–12 dB from an initial threshold shift of 60–80 dB, when compared to scrambled RNA controls.…”

Therapeutic Potential of Wnt and Notch Signaling and Epigenetic Regulation in Mammalian Sensory Hair Cell Regeneration

“…Although, the newly generated hair cells exhibit an immature phenotype and lineage-tracing studies are still needed to confirm the origins of these new hair cells. 38 , 39 …”

“… 19 , 22 , 23 , 25 , 27 , 28 , 29 , 30 , 31 Given their roles in development, activation of canonical Wnt signaling following birth can promote supporting cell proliferation and limited hair cell induction, and inhibition of the Notch pathway can promote the transdifferentiation of supporting cells to hair cells. 34 , 35 , 36 , 37 , 38 , 39 , 46 , 47 , 49 , 50 , 52 , 53 , 54 , 55 There has been some success in hair cell regeneration following Wnt and Notch modulation in the mature mammalian cochlea. 34 , 52 However, it is important to note that many of these newly generated hair cells show limited survival and do not appear to fully differentiate, lacking mature stereocilia, and they may fail to form proper synaptic connections with auditory neurons.…”