Genes that are primarily expressed in cochlear glia-like supporting cells (GLSs) have not been clearly associated with progressive deafness. Herein, we present a deafness locus mapped to chromosome 3p25.1 and an auditory neuropathy spectrum disorder (ANSD) gene, TMEM43, mainly expressed in GLSs. We identify p.(Arg372Ter) of TMEM43 by linkage analysis and exome sequencing in two large Asian families segregating ANSD, which is characterized by inability to discriminate speech despite preserved sensitivity to sound. The knock-in mouse with the p.(Arg372Ter) variant recapitulates a progressive hearing loss with histological abnormalities in GLSs. Mechanistically, TMEM43 interacts with the Connexin26 and Connexin30 gap junction channels, disrupting the passive conductance current in GLSs in a dominant-negative fashion when the p.(Arg372Ter) variant is introduced. Based on these mechanistic insights, cochlear implant was performed on three subjects, and speech discrimination was successfully restored. Our study highlights a pathological role of cochlear GLSs by identifying a deafness gene and its causal relationship with ANSD. Significance Auditory neuropathy spectrum disorder (ANSD) is a confounding auditory disease in which the subjects respond to sound but have difficulties in speech discrimination. Herein, we examined two Asian families with hereditary late-age–onset ANSD. By linkage analysis and exome sequencing, we identified the TMEM43-p.(Arg372Ter) variant as the etiology of the disease. To examine the mechanism of TMEM43 on ANSD, we generated a knock-in mouse with the p.(Arg372Ter) variant that recapitulated the progressive hearing loss phenotype of the two families. We discovered that variation in TMEM43 impairs the connexin-linked function of mediating passive conductance current in cochlear glial cells. Based on the pathology involving cochlear glial cells, we performed cochlear implant on the human patients, and their speech discrimination ability was restored.

Enlarged vestibular aqueduct (EVA) is one of the most common congenital inner ear malformations and accounts for 1–12% of sensorineural deafness in children and adolescents. Multiple genetic defects contribute to EVA; therefore, early molecular diagnosis is critical for EVA patients to ensure that the most effective treatment strategies are employed. This study explored a new genetic diagnosis method for EVA and applied it to clinic diagnoses of EVA patients. Using next-generation sequencing technology, we set up a multiple polymerase chain reaction enrichment system for target regions of EVA pathogenic genes (SLC26A4, FOXI1, and KCNJ10). Forty-six EVA samples were sequenced by this system. Variants were detected in 87.0% (40/46) of cases, including three novel variants (SLC26A4 c.923_929del, c.1002-8C>G, and FOXI1 c.519C>A). Biallelic potential pathogenic variants were detected in 27/46 patient samples, leading to a purported diagnostic rate of 59%. All results were verified by Sanger sequencing. Our target region capture system was validated to amplify and measure SLC26A4, FOXI1, and KCNJ10 in one reaction system. The result supplemented the mutation spectrum of EVA. Thus, this strategy is an economic, rapid, accurate, and reliable method with many useful applications in the clinical diagnosis of EVA patients.

The ABCC1 gene belongs to the ATP-binding cassette membrane transporter superfamily, which plays a crucial role in the efflux of various endogenous and exogenous substances. Mutations in ABCC1 can result in autosomal dominant hearing loss. However, the specific roles of ABCC1 in auditory function are not fully understood. Through immunofluorescence, we found that ABCC1 was expressed in microvascular endothelial cells (ECs) of the stria vascularis (StV) in the murine cochlea. Then, an Abcc1 knockout mouse model was established by using CRISPR/Cas9 technology to elucidate the role of ABCC1 in the inner ear. The ABR threshold did not significantly differ between WT and Abcc1^-/- mice at any age studied. After noise exposure, the ABR thresholds of the WT and Abcc1^-/- mice were significantly elevated. Interestingly, after 14 days of noise exposure, ABR thresholds largely returned to pre-exposure levels in WT mice but not in Abcc1^-/- mice. Our subsequent experiments showed that microvascular integrity in the StV was compromised and that the number of outer hair cells and the number of ribbons were significantly decreased in the cochleae of Abcc1^-/- mice post-exposure. Besides, the production of ROS and the accumulation of 4-HNE significantly increased. Furthermore, StV microvascular ECs were cultured to elucidate the role of ABCC1 in these cells under glucose oxidase challenge. Notably, 30 U/L glucose oxidase (GO) induced severe oxidative stress damage in Abcc1^-/- cells. Compared with WT cells, the ROS and 4-HNE levels and the apoptotic rate were significantly elevated in Abcc1^-/- cells. In addition, the reduced GSH/GSSG ratio was significantly decreased in Abcc1^-/- cells after GO treatment. Taken together, Abcc1^-/- mice are more susceptible to noise-induced hearing loss, possibly because ABCC1 knockdown compromises the GSH antioxidant system of StV ECs. The exogenous antioxidant N-acetylcysteine (NAC) may protect against oxidative damage in Abcc1^-/- murine cochleae and ECs.