GJB2 gene mutations are the most common causes of autosomal recessive non-syndromic hereditary deafness. For individuals suffering from severe to profound GJB2 -related deafness, cochlear implants have emerged as the sole remedy for auditory improvement. Some previous studies have highlighted the crucial role of preserving cochlear neural components in achieving favorable outcomes after cochlear implantation. Thus, we generated a conditional knockout mouse model (Cx26-CKO) in which Cx26 was completely deleted in the cochlear supporting cells driven by the Sox2 promoter. The Cx26-CKO mice showed severe hearing loss and massive loss of hair cells and Deiter’s cells, which represented the extreme form of human deafness caused by GJB2 gene mutations. In addition, multiple pathological changes in the peripheral auditory nervous system were found, including abnormal innervation, demyelination, and degeneration of spiral ganglion neurons as well as disruption of heminodes in Cx26-CKO mice. These findings provide invaluable insights into the deafness mechanism and the treatment for severe deafness in Cx26-null mice.

Noise-induced hearing loss (NIHL) is a prevalent form of adult hearing impairment, characterized by oxidative damage to auditory sensory hair cells. Although certain dihydropyridines, the L-type calcium channel blockers, exhibit protective properties against such damage, the ability of third-generation dihydropryidines like lercanidipine to mitigate NIHL remains unclear.We utilized glucose oxidase (GO)-treated OC1 cell lines and cochlear explants to evaluate the protective influence of lercanidipine on hair cells. To further investigate its effectiveness, we exposed noise-stimulated mice in vivo and analyzed their hearing thresholds. Additionally, we assessed the antioxidative capabilities of lercanidipine by examining oxidation-related enzyme expression and levels of oxidative stress markers, including 3-nitrotyrosine (3NT) and 4-hydroxynonenal (4HNE). Our findings demonstrate that lercanidipine significantly reduces the adverse impacts of GO on both OC-1 cell viability (0.3 to 2.5 µM) and outer hair cell (OHC) survival in basal turn cochlear explants (7 µM). These results are associated with increased mRNA expression of antioxidant enzyme genes (HO-1, SOD1/2, and Txnrd1), along with decreased expression of oxidase genes (COX-2, iNOS). Crucially, lercanidipine administration prior to, and following, noise exposure effectively ameliorates NIHL, as evidenced by lowered hearing thresholds and preserved OHC populations in the basal turn, 14 days post-noise stimulation at 110 dB SPL. Moreover, our observations indicate that lercanidipine’s antioxidative action persists even three days after simultaneous drug and noise treatments, based on 3-nitrotyrosine and 4-hydroxynonenal immunostaining in the basal turn. Based on these findings, we propose that lercanidipine has the capacity to alleviate NIHL and safeguard OHC survival in the basal turn, potentially via its antioxidative mechanism. These results suggest that lercanidipine holds promise as a clinically viable option for preventing NIHL in affected individuals.

Hearing loss is associated with an increased risk of Alzheimer disease (AD). However, the mechanisms of hearing loss promoting the onset of AD are poorly understood. Here we show that hearing loss aggravates cognitive impairment in both wild-type mice and mouse models of AD. Embryonic growth/differentiation factor 1 (GDF1) is downregulated in the hippocampus of deaf mice. Knockdown of GDF1 mimics the detrimental effect of hearing loss on cognition, while overexpression of GDF1 in the hippocampus attenuates the cognitive impairment induced by deafness. Strikingly, overexpression of GDF1 also attenuates cognitive impairment in APP/PS1 transgenic mice. GDF1 activates Akt, which phosphorylates asparagine endopeptidase and inhibits asparagine endopeptidase-induced synaptic degeneration and amyloid-β production. The expression of GDF1 is downregulated by the transcription factor CCAAT-enhancer binding protein-β. These findings indicate that hearing loss could promote AD pathological changes by inhibiting the GDF1 signaling pathway; thus, GDF1 may represent a therapeutic target for AD.

The GJB2 gene, encoding Connexin26 (Cx26), is one of the most common causes of inherited deafness. Clinically, mutations in GJB2 cause congenital deafness or late-onset progressive hearing loss. Recently, it has been reported that Cx26 haploid deficiency accelerates the development of age-related hearing loss (ARHL). However, the roles of cochlear Cx26 in the hearing function of aged animals remain unclear. In this study, we revealed that the Cx26 expression was significantly reduced in the cochleae of aged mice, and further explored the underlying molecular mechanism for Cx26 degradation. Immunofluorescence co-localization results showed that Cx26 was internalized and degraded by lysosomes, which might be one of the important ways for Cx26 degradation in the cochlea of aged mice. Currently, whether the degradation of Cx26 in the cochlea leads directly to ARHL, as well as the mechanism of Cx26 degradation-related hearing loss are still unclear. To address these questions, we generated mice with Cx26 knockout in the adult cochlea as a model for the natural degradation of Cx26. Auditory brainstem response (ABR) results showed that Cx26 knockout mice exhibited high-frequency hearing loss, which gradually progressed over time. Pathological examination also revealed the degeneration of hair cells and spiral ganglions, which is similar to the phenotype of ARHL. In summary, our findings suggest that degradation of Cx26 in the cochlea accelerates the occurrence of ARHL, which may be a novel mechanism of ARHL.

Clinically, thyroid-related diseases such as endemic iodine deficiency and congenital hypothyroidism are associated with hearing loss, suggesting that thyroid hormones are essential for the development of normal hearing. Triiodothyronine (T3) is the main active form of thyroid hormone and its effect on the remodeling of the organ of Corti remain unclear. This study aims to explore the effect and mechanism of T3 on the remodeling of the organ of Corti and supporting cells development during early development. In this study, mice treated with T3 at postnatal (P) day 0 or P1 showed severe hearing loss with disordered stereocilia of the outer hair cells (OHCs) and impaired function of mechanoelectrical transduction of OHCs. In addition, we found that treatment with T3 at P0 or P1 resulted in the overproduction of Deiter-like cells. Compared with the control group, the transcription levels of Sox2 and notch pathway-related genes in the cochlea of the T3 group were significantly downregulated. Furthermore, Sox2-haploinsufficient mice treated with T3 not only showed excess numbers of Deiter-like cells but also a large number of ectopic outer pillar cells (OPCs). Our study provides new evidence for the dual roles of T3 in regulating both hair cells and supporting cell development, suggesting that it is possible to increase the reserve of supporting cells.

Diabetes mellitus (DM) is a major disease threatening human health and its incidence is increasing year on year. As a chronic complication of DM, hearing loss mostly occurs undetectably. However, the mechanism of this diabetes-related hearing loss (DRHL) remains unclear and there is no effective clinical treatment. Studies of animal or human pathology show that DM causes damage to the blood vessels, spiral ganglion neurons, afferent nerve fibers, the organ of Corti, and the stria vascularis of the inner ear. In recent years, more advances in pathological research have revealed the possible mechanism of DRHL. In addition, a large number of clinical studies suggest that the duration and severity of DM are closely related to the incidence and severity of DRHL. This review focuses on the relationship between DM and hearing loss. The clinical audiological characteristics of diabetic patients, risk factors for DRHL, typical pathology, and potential interventions of DRHL are summarized. This will help reveal the pathogenesis and intervention approaches for DRHL.

Background: This study aimed to describe the distribution of the genotype and allele frequencies of GJB2 variants in the Chinese population of the Dongfeng Tongji cohort and to analyze the features of the hearing phenotype. Methods: We used data from 9910 participants in the Dongfeng Tongji cohort in 2013 and selected nine GJB2 variants. Pure tone audiometry was employed to measure hearing. Differences in genotype and allele frequencies were analyzed via chi-squared test or Fisher’s exact test. Results: Of the 9910 participants, 5742 had hearing loss. The genotype frequency of the GJB2 variant c.109G>A was statistically significantly distributed between the normal and impaired hearing groups, but not for the variant c.235delC. A higher frequency of the c.109G>A homozygous genotype was found in the hearing loss group (0.5%) than in the normal hearing group (0.1%). Patients with c.109G>A and c.235delC homozygous mutations exhibited varying degrees of hearing loss, mainly presenting sloping and flat audiogram shapes. Conclusions: A significant difference was found in the genotype frequency of the GJB2 variant c.109G>A between the case and control groups, but not in that of the variant c.235delC. Different degrees of hearing loss and various audiogram shapes were observed in patients with c.109G>A and c.235delC homozygous mutations.

Abstract GJB2 (encoding connexin26, Cx26) mutation is the most common genetic cause of hereditary deafness. Cochlear sensory hair cell (HC) death is the core pathologic phenomenon of GJB2‐related deafness. However, mechanism‐based therapy is still obscure. A targeted‐cell conditional Gjb2 knockout mouse model was established in which the Cx26 in Deiters cells and pillar cells were knocked out at birth. We explored the mechanism of HC death caused by deficiency of GJB2 in supporting cells (SCs), which exhibited moderate deafness, early HC death without SCs, and a decrease in distortion product otoacoustic emission. Here, a DNA damage response was observed in HCs during the onset of hearing loss. Apoptosis, necroptosis, and ferroptosis do not contribute to the death of HCs. We identified and demonstrated that parthanatos, a poly‐(ADP‐ribose) polymerase (PARP)‐dependent and caspase‐independent form of cell death, is a mechanism of HC degeneration. Furthermore, we observed that the use of PARP inhibitors effectively alleviated both deafness and HC loss. Our study reveals the specific mechanism of HC death caused by GJB2 deficiency. These findings demonstrate that targeting parthanatos is an HC protective strategy in the prevention of GJB2‐related hereditary deafness.

Mutations in the GJB2 gene encoding connexin26 (Cx26) protein are one of the most common causes of hereditary deafness. Previous studies have found that different Cx26-null mouse models have severe hearing loss and deformity of the organ of Corti (OC) as well as a reduction in microtubules in pillar cells (PCs). To explore the underlying mechanism of OC deformity caused by Cx26 downregulation further, we established Cx26 knockdown (KD) mouse models at postnatal days (P)0 and P8. The actin filaments contained in the pillar cells of mice in the P0 KD group were reduced by 54.85% and vinculin was increased by 22%, while the outer hair cells (OHCs) showed normal F-actin content. In the P8 KD group, PCs and OHCs of mice also showed almost normal F-actin content. The G-actin/F-actin ratio increased by 38% in the P0 KD group. No significant change was found in the mRNA or protein expression level of G-actin or the cadherin–catenin core complex in the P0 KD group at P6. Moreover, immunofluorescence showed that the intensity of LRRK2 was reduced by 97% in the P0 KD group at P6. Our results indicate that Cx26 is involved in the maturation of the cytoskeleton during the development of the OC at the early postnatal stage. The polymerization of G-actin into F-actin is prevented in Cx26 KD mice.

Thyroid hormones are essential for cochlear development and normal auditory function. In rodents, previous study reports that excessive triiodothyronine (T3) accelerates development of cochlear morphology and cause deafness. However, the mechanism of T3 induced hearing loss and its effect on the remodeling of the organ of Corti (OC) remain unclear. In this study, neonatal mice were given T3 at different time points, and then auditory brainstem responses (ABR) and morphological changes of the OC were analyzed at postnatal day (P) 18. Mice treated with T3 at P0 or P1 showed severe hearing loss with disordered stereocilia of the outer hair cells (OHCs), and impaired function of mechanoelectrical transduction of OHCs. In addition, we found that treatment with T3 at P0 or P1 resulted in overproduction of Deiter-like cells (Deiter’s cell, DC, a supporting cell in OC). These extra cells have phalangeal process structure similar to DC, and coupled with normal DC by gap junctions. Compared with the control group, the transcription levels of Sox2 and notch pathway related genes in the cochlea of T3 group were significantly down-regulated. Furthermore, T3 treatment combined with downregulation of Sox2 not only resulted in overproduction of DCs, but also led to a large number of ectopic outer Pillar cells (OPCs). This can induce a great change of morphology of OC, forming double tunnels of Corti structure. Together, our study provides new evidence for dual roles of T3 in regulating both hair cell and supporting cell development, suggesting that it is possible to increase the reserve of supporting cells.

Hearing loss is among the most common congenital sensory impairments. Genetic causes account for more than 50% of the cases of congenital hearing loss. The PTPRQ gene, encoding protein tyrosine phosphatase receptor Q, plays an important role in maintaining the stereocilia structure and function of hair cells. Mutations in the PTPRQ gene have been reported to cause hereditary sensorineural hearing loss. By using next-generation sequencing and Sanger sequencing, we identified a novel compound heterozygous mutation (c.997 G > A and c.6603-3 T > G) of the PTPRQ gene in a Chinese consanguineous family. This is the first report linking these two mutations to recessive hereditary sensorineural hearing loss. These findings contribute to the understanding of the relationship between genotype and hearing phenotype of PTPRQ-related hearing loss, which may be helpful to clinical management and genetic counseling.

To improve the etiological diagnosis of congenital hearing loss by combining whole-exome sequencing (WES) with cytomegalovirus (CMV) testing and to explore the potential benefits of adding CMV screening to newborn hearing screening, 80 children under 2 years of age with bilateral sensorineural hearing loss were recruited. Peripheral venous blood was extracted from the children for WES analysis. Saliva after mouthwash and the first urine in the morning were collected and used as samples to quantify CMV DNA copy number in urine and saliva by qPCR; among the 80 children with congenital deafness, 59 (74%) were found to have genetic variants that may cause congenital deafness, including 44 with GJB2 or SLC26A4 gene variant, 1 with STRC gene variant, and 14 with other genetic variants. A total of 12 children carried deafness gene variants associated with a syndrome; CMV test results showed that in two children, the CMV DNA copy number in saliva was >1000/mL, which indicates that they were CMV-positive, and their genetic test results were negative. A neonatal CMV test combined with genetic screening can improve the etiological diagnosis rate of congenital deafness, and the direct evidence of neonatal CMV infection deserves further verification.

Sensorineural hearing loss (SNHL) is one of the most prevalent sensory deficits in humans, and approximately 360 million people worldwide are affected. The current treatment option for severe to profound hearing loss is cochlear implantation (CI), but its treatment efficacy is related to the survival of spiral ganglion neurons (SGNs). SGNs are the primary sensory neurons, transmitting complex acoustic information from hair cells to second-order sensory neurons in the cochlear nucleus. In mammals, SGNs have very limited regeneration ability, and SGN loss causes irreversible hearing loss. In most cases of SNHL, SGN damage is the dominant pathogenesis, and it could be caused by noise exposure, ototoxic drugs, hereditary defects, presbycusis, etc. Tremendous efforts have been made to identify novel treatments to prevent or reverse the damage to SGNs, including gene therapy and stem cell therapy. This review summarizes the major causes and the corresponding mechanisms of SGN loss and the current protection strategies, especially gene therapy and stem cell therapy, to promote the development of new therapeutic methods.

Purpose Genetic testing is widely used in diagnosing genetic hearing loss in patients. Other than providing genetic etiology, the benefits of genetic testing in pediatric patients with hearing loss are less investigated. Methods From 2018-2020, pediatric patients who initially presented isolated hearing loss were enrolled. Comprehensive genetic testing, including GJB2/SLC26A4 multiplex amplicon sequencing, STRC/OTOA copy number variation analysis, and exome sequencing, were hierarchically offered. Clinical follow-up and examinations were performed. Results A total of 80 pediatric patients who initially presented isolated hearing loss were considered as nonsyndromic hearing loss and enrolled in this study. The definitive diagnosis yield was 66% (53/80) and the likely diagnosis yield was 8% (6/80) through comprehensive genetic testing. With the aid of genetic testing and further clinical follow-up and examinations, the clinical diagnoses and medical management were altered in eleven patients (19%, 11/59); five were syndromic hearing loss; six were nonsyndromic hearing loss mimics. Conclusion Syndromic hearing loss and nonsyndromic hearing loss mimics are common in pediatric patients who initially present with isolated hearing loss. The comprehensive genetic testing provides not only a high diagnostic yield but also valuable information for clinicians to uncover subclinical or pre-symptomatic phenotypes, which allows early diagnosis of SHL, and leads to precise genetic counseling and changes the medical management.

Mutations in the GJB2 gene account for approximately 20–50% of all non-syndromic hereditary deafness cases. The malformed organ of Corti (OC) was observed in different Cx26-null mouse models, which was mainly caused by the developmental arrest of pillar cells (PCs). However, the mechanism of developmental abnormalities in PCs caused by Cx26 deletion is still unclear. In this study, the ultrastructure of PCs at different postnatal days was observed in Cx26-null mice. Knockout of cochlear Cx26 led to the malformed assembly of non-centrosomal microtubule-organizing centers (MTOCs) far from the centrosome rather than near the centrosome. Additionally, the microtubule (MT) arrays emitted by abnormal non-centrosomal MTOCs were significantly reduced. In addition, we found that the protein expression of calmodulin-regulated, spectrin-associated protein2 (camsap2), a microtubule minus-end targeting protein associated with the organization of non-centrosomal MTs, was decreased in juvenile PCs in the Cx26-null group. Our results indicated that the malformation of non-centrosomal MTOCs in cochlear PCs might lead to the corresponding MTs’ failure to be captured and anchored in Cx26-null mice, which results in the deformity of OC. Additionally, this abnormal developmental process might be correlated with the reduced expression of camsap2 caused by Cx26 deletion in the early developmental stage.

Mutations in the GJB2 gene (encoding Connexin26(Cx26)) are the most common cause of hereditary deafness, accounting for about a quarter of all cases. Sensory epithelial damage is considered to be one of the main causes of deafness caused by GJB2 gene mutation. Dexamethasone (DEX) is widely used in the treatment of a variety of inner ear diseases including sudden sensorineural hearing loss (SSNHL), noise-induced hearing loss (NIHL), and deafness caused by ototoxic drugs. Whether DEX has a direct therapeutic effect on hereditary deafness, especially GJB2-related deafness, remains unclear. In this study, we revealed that DEX can effectively prevent hair cell death caused by oxidative stress in cochlear explants. Additionally, two distinct Cx26-null mouse models were established to investigate whether systemic administration of DEX alleviate the cochlear sensory epithelial injury or deafness in these models. In a specific longitudinally Cx26-null model that does not cause deafness, systemic administration of DEX prevents the degeneration of outer hair cells (OHCs) induced by Cx26 knockout. Similarly, in a targeted-Deiter's cells (DCs) Cx26-null mouse model that causes deafness, treatment with DEX can almost completely prevent OHCs loss and alleviates auditory threshold shifts at some frequencies. Additionally, we observed that DEX inhibited the recruitment of CD45-positive cells in the targeted-DCs Cx26-null mice. Taken together, our results suggest that the protective effect of dexamethasone on cochlear sensory epithelial damage and partially rescue auditory function may be related to the regulation of inner ear immune response in Cx26 deficiency mouse models.

Inherited diseases caused by connexin mutations are found in multiple organs and include hereditary deafness, congenital cataract, congenital heart diseases, hereditary skin diseases, and X-linked Charcot–Marie–Tooth disease (CMT1X). A large number of knockout and knock-in animal models have been used to study the pathology and pathogenesis of diseases of different organs. Because the structures of different connexins are highly homologous and the functions of gap junctions formed by these connexins are similar, connexin-related hereditary diseases may share the same pathogenic mechanism. Here, we analyze the similarities and differences of the pathology and pathogenesis in animal models and find that connexin mutations in gap junction genes expressed in the ear, eye, heart, skin, and peripheral nerves can affect cellular proliferation and differentiation of corresponding organs. Additionally, some dominant mutations (e.g., Cx43 p.Gly60Ser, Cx32 p.Arg75Trp, Cx32 p.Asn175Asp, and Cx32 p.Arg142Trp) are identified as gain-of-function variants in vivo, which may play a vital role in the onset of dominant inherited diseases. Specifically, patients with these dominant mutations receive no benefits from gene therapy. Finally, the complete loss of gap junctional function or altered channel function including permeability (ions, adenosine triphosphate (ATP), Inositol 1,4,5-trisphosphate (IP3), Ca2+, glucose, miRNA) and electric activity are also identified in vivo or in vitro.

Different serum thyroxine levels may influence the morphology of the inner ear during development. A well-developed organ of Corti (OC) is considered to be critical to the function of hearing. In our study, we treated mice with triiodothyronine (T3) and found that the opening of the OC occurred sooner than in control mice. We also observed an increased formation of acetylated microtubules and a decrease in the adhesion junction molecule P-cadherin the during opening of the OC. Our investigation indicates that thyroxin affects P-cadherin expression and microtubule acetylation to influence the opening of the OC.

Osteopetrosis is a rare inherited bone disease characterized by dysfunction of osteoclasts, causing impaired bone resorption and remodeling, which ultimately leads to increased bone mass and density. Hearing loss is one of the most common complications of osteopetrosis. However, the etiology and pathogenesis of auditory damage still need to be explored. In this study, we found that a spontaneous mutation of coiled-coil domain-containing 154 (CCDC154) gene, a new osteopetrosis-related gene, induced congenital deafness in mice. Homozygous mutant mice showed moderate to severe hearing loss, while heterozygous or wild-type (WT) littermates displayed normal hearing. Pathological observation showed that abnormal bony remodeling of the otic capsule, characterized by increased vascularization and multiple cavitary lesions, was found in homozygous mutant mice. Normal structure of the organ of Corti and no substantial hair cell or spiral ganglion neuron loss was observed in homozygous mutant mice. Our results indicate that mutation of the osteopetrosis-related gene CCDC154 can induce syndromic hereditary deafness in mice. Bony remodeling disorders of the auditory ossicles and otic capsule are involved in the hearing loss caused by CDCC154 mutation.

Objective:To identify the pathogenic gene mutation of two patients with non-syndromic deafness（NSHL）. Methods:Two patient with NSHL and their parents were selected in the research object. Each participant provided 3-5 mL of peripheral venous blood, which was used to establish a DNA library. Next generation sequencing was used to detect the sequence of the patient's genome, and the sequencing results were compared with the human genome sequence （GRCh）37/hg19. Sanger sequencing was used to verify the parents' genome sequence. Finally the patient's pathogenic gene mutation was confirmed.Amino acid conservatism and single nucleotide polymorphisms of the mutant sites were analyzed using a variety of databases and software. Results:The mutation was located to CDH23 gene in the chromosomal location 10q21-q22. Complex heterozygous mutations consist of c. 1343T>C and c. 7991_7993delTCA. Parents are heterozygous carriers of a single mutation. Conclusion:The next generation sequencing technology were used to screen the pathogenic gene mutation of inherited deafness. Combined with the genetic sequencing results of parents, the specific pathogenic gene mutation of deafness patients can be identified. While the pathogenicity of complex heterozygous mutation were explained by various pathogenicity analysis methods.