Spatiotemporal Developmental Upregulation of Prestin Correlates With the Severity and Location of Cyclodextrin-Induced Outer Hair Cell Loss and Hearing Loss

Abstract

2-Hyroxypropyl-beta-cyclodextrin (HPβCD) is being used to treat Niemann-Pick C1, a fatal neurodegenerative disease caused by abnormal cholesterol metabolism. HPβCD slows disease progression, but unfortunately causes severe, rapid onset hearing loss by destroying the outer hair cells (OHC). HPβCD-induced damage is believed to be related to the expression of prestin in OHCs. Because prestin is postnatally upregulated from the cochlear base toward the apex, we hypothesized that HPβCD ototoxicity would spread from the high-frequency base toward the low-frequency apex of the cochlea. Consistent with this hypothesis, cochlear hearing impairments and OHC loss rapidly spread from the high-frequency base toward the low-frequency apex of the cochlea when HPβCD administration shifted from postnatal day 3 (P3) to P28. HPβCD-induced histopathologies were initially confined to the OHCs, but between 4- and 6-weeks post-treatment, there was an unexpected, rapid and massive expansion of the lesion to include most inner hair cells (IHC), pillar cells (PC), peripheral auditory nerve fibers, and spiral ganglion neurons at location where OHCs were missing. The magnitude and spatial extent of HPβCD-induced OHC death was tightly correlated with the postnatal day when HPβCD was administered which coincided with the spatiotemporal upregulation of prestin in OHCs. A second, massive wave of degeneration involving IHCs, PC, auditory nerve fibers and spiral ganglion neurons abruptly emerged 4–6 weeks post-HPβCD treatment. This secondary wave of degeneration combined with the initial OHC loss results in a profound, irreversible hearing loss.

Multifunctional redox modulator prevents blast-induced loss of cochlear and vestibular hair cells and auditory spiral ganglion neurons

“…Under a dissection microscope, openings were made in the cochlear apex, round window, and the oval window. Samples for surface preparations of the cochlear basilar membrane and vestibular end-organs were perfused with 10% formalin in phosphate buffered saline (PBS) into the cochlear and vestibular cavities and then immersed in the fixative overnight as described previously 67 , 68 . Samples for temporal bone sections were fixed with 2.5% glutaraldehyde in 0.1 M PBS for 6 h. After rinsing with PBS, samples were immersed in 2% osmium tetroxide in 0.1 M PBS for 2 h 69 .…”

“…After fixation, samples for surface preparations of the cochlea and vestibular end-organs were decalcified (Decalcifying solution-Lite, Millipore Sigma, D0818) for 48 h. Then, the cochlear basilar membrane, macula of saccule, macula of utricle, and three crista ampullae were dissected out under a dissecting microscope, and stained with Harris hematoxylin solution, and mounted in glycerin on glass slides as described previously 68 .…”

“…The percentages of missing IHC and OHC were determined based on laboratory norms for normal adult rats 70 . The cell counts were entered into a custom computer program to generate a cochleogram showing the percentages of missing OHC and IHC as a function of percent distance from the apex of the cochlea based on laboratory norms as described previously 67 , 68 . Cochlear location was related to frequency using a rat frequency-place map 26 .…”

Hearing impairment in murine model of Down syndrome

“…DPOAEs were measured in order to assess the functional status of the outer hair cells in the cochlea; however, DPOAEs can also be negatively affected by conductive hearing loss. DPOAEs were measured in 6–8 month old mice as described previously ( Liu et al, 2020 ; Ding et al, 2021 ). Mice were anesthetized with a mixture of ketamine (50 mg/kg, i.p.)…”

“…The ABR was recorded in 3 and 9-month-old mice as described previously ( Chen et al, 2014 ; Ding et al, 2021 ). Mice were anesthetized with a mixture of ketamine (50 mg/kg, i.p.)…”