OBJECTIVES:To explore the correlations between genotype and phenotype, disease progression, and outcomes of interventions in patients with auditory neuropathy (AN) caused by OTOF gene variants. DESIGN:Patients with AN associated with OTOF variants were identified using whole-exome or panel sequencing. Variants were interpreted according to the American College of Medical Genetics and Genomics guidelines. Audiometric tests were conducted, including auditory brainstem response, distortion product otoacoustic emission (DPOAE), behavioral audiometry/pure-tone audiometry, etc. The genetic and audiological characteristics of the patients were analyzed. The disease progression and intervention of the patients were followed up, and phenotype differences were analyzed in combination with genotype. RESULTS:A total of 43 AN patients were identified with OTOF gene variants, including seven novel variants. The pure-tone average was 89.20 ± 17.81 dB HL, with 91.11% having severe hearing loss or greater. The auditory steady-state response and DPOAE results deteriorated with increasing disease duration. Twenty-five patients underwent cochlear implantation, with the Category of Auditory Performance score of 7.00 (5.00, 7.50). Patients with biallelic loss-of-function variants showed a trend toward worse hearing but a higher DPOAE response rate. Similarly, patients with a single allele variant causing protein truncation also had a higher DPOAE extraction rate. CONCLUSIONS:Seven novel mutations of the OTOF gene were identified, which enriched the spectrum of OTOF gene variants. Most of the AN patients with OTOF variants showed severe to profound hearing loss, with the progression of the disease course. The cochlear implantation effects were good, which were related to the intervention age and duration. There is a correlation trend between OTOF variants' genotype and phenotype.

Background Inhibition of cardiac hypertrophy and improving cardiac function are the main aims for treating heart failure. Sanweidoukou decoction (SD-3) is a traditional Chinese remedy known for its heart-protective properties and effectiveness in treating cardiovascular and cerebrovascular conditions. However, the mechanisms of action of SD-3 have not been clearly established. We investigated the protective effects of water extraction of SD-3 against heart failure and elucidated on its potential molecular mechanisms. Methods Cardiac failure models were established in rats and H9c2 cells using isoproterenol (ISO). Cardiac function and fibrosis were evaluated via echocardiography and histological staining. The underlying mechanisms were analyzed through RNA-Seq, while ROS levels, mitochondrial function, and AMPK/PPARα pathway-related indicators were measured. Molecular docking was employed to validate the binding between active components and target proteins. Results Findings indicated that SD-3 reduced cardiac hypertrophy, improved cardiac function, and inhibited the expression of genes associated with hypertrophy in both laboratory settings and living organisms. Bioinformatic analyses identified a number of genes enriched in metabolic pathway, especially for AMPK and PPARα pathway. Mechanistically, it could entail the suppression of the AMPK/PPARα pathway, reduction in ROS levels, prevention of mitochondrial membrane potential collapse, and the opening of mitochondrial permeability transition pores, all of which contribute to a decreased apoptotic response in heart failure. AMPK-Pisatin and PPARα-sesamin had the lowest free binding energy values, showing good binding capacity and therapeutic potential. Conclusions SD-3 exerts protective effects against heart failure by inhibiting mitochondrial dysfunction and the AMPK /PPARα pathway.

De novo mutations (DNMs) are significant genetic factors contributing to sporadic hearing loss (HL) and complex HL syndromes. To analyze the genetic counseling characteristics and interpretation of pathogenic DNMs for sporadic HL, we retrospectively analyze the clinical information of probands and their parents from 410 sporadic HL core pedigrees enrolled in the "Chinese Deafness Genome Project (CDGP)" between October 2015 and October 2023. We apply family trio-based genome sequencing (targeted gene capture and high throughput sequencing, mitochondrial genome sequencing, and copy number variants analysis) and validate the samples of their unaffected-parents. Homologous allele sequencing is used to identity by descent (IBD) in the DNM family trios. The results reveal that 7.3% (30 cases) of the probands in these sporadic hearing loss core pedigrees carry 17 types of autosomal dominant gene de novo single nucleotide variants (SNVs), insertions/deletions (Indels), and one type of de novo copy number variation, encompassing all types of DNM. Among them, WFS1 c.2051C>T, ATP1A3 c.2452G>A, and ACTG1 c.94C>T are common DNM in sporadic HL. The genotype C>T transversion exhibit a high number (34.6%). Clinical feature analyses also show that 56.7% (17/30) of the probands have non-syndromic HL, but more than half of them (52.9%, 9/17) carry pathogenic genotypes clearly associated with "syndromic HL", possibly exhibiting temporary "mimic" non-syndromic HL phenotypic characteristics. The average parental ages at childbirth for the 30 probands are 29.4 years for fathers and 28.3 years for mothers, with 13.3% of fathers or mothers aged ≥35 years. Additionally, among the family structure of the proband of genetic counseling, 63.3% are single-child families with a clear desire for another child, and 16.7% of the probands' parents seek prenatal genetic counseling for conceiving a "second child". During genetic counseling, it is essential to test the "family proband-parents' trios" core pedigree as a unit to analyze the genetic contribution of DNMs to HL. Furthermore, there is a certain correlation between the occurrence of DNMs and increasing parental age at childbirth. Therefore, for families with a history of DNM-associated sporadic HL, it is necessary to collect clinical information such as the parental age at childbirth and obstetric history of hearing-healthy parents. For these families planning another child, it is recommended to undergo prenatal diagnosis for the identified DNM pathogenic variations after conception and pay attention to the pregnancy outcome.

Auditory neuropathy (AN) represents a clinical manifestation of OPA1-related diseases. The diagnostic process of these diseases is challenging owing to the broad spectrum of intermediate phenotypes and diverse inherited patterns. The aim of this study was to comprehensively delineate the feature of OPA1-related patients in a Chinese AN cohort, encompassing the incident rate, inherited pattern, detailed audiological characteristics, and genotype-phenotype correlation. Between 2003 and 2020, 452 unrelated probands with a diagnosis of AN were referred to our laboratory for molecular genetic investigation with high-throughput sequencing. Sanger sequencing was performed on the probands and their parents to verify the genetic results. Patients diagnosed as AN by clinical evaluation, auditory brainstem responses, otoacoustic emission and/or cochlear microphonic. Comprehensive auditory evaluations were conducted on OPA1-related patients, and some of them were performed a follow-up study. We identified seven probands (1.55

OBJECTIVES:Baicalein, a flavonoid derived from the roots of Scutellaria baicalensis Georgi, demonstrates multifarious pharmacological effects due to its high antioxidant activity. However, the latent mechanisms remain insufficiently resolved. In the present research, we evaluated the therapeutic effects of baicalein on isoprenaline (ISO)-induced heart failure and investigated the possible underlying mechanisms. METHODS:Toxicity was analyzed in zebrafish embryos and mouse atrial myocytes HL-1. The MTT assay was used to evaluate the effectiveness of baicalein. DCFH-DA was used as a fluorescence probe to detect intracellular reactive oxygen species (ROS). Superoxide dismutase (SOD), malondialdehyde (MDA), and glutathione peroxidase (GSH-Px) levels were measured using SOD, MDA and GSH-Px commercial kits. Adult BALB/c mice were randomized into six groups of ten animals each. Cardiac function was analyzed by echocardiographic images. Structural changes were analyzed by hematoxylin & eosin (HE) staining, Masson staining and TUNEL staining. The mechanism of baicalein was investigated by analyzing relative signaling pathways through western blotting. RESULTS:Our studies show that baicalein both significantly reduces ISO-induced oxidative stress, apoptosis and cardiac fibrosis in vitro and vivo, this phenomenon was related to mitochondrial fusion/fission balance and inhibiting GRP78/CHOP pathway. CONCLUSIONS:Our results suggested that baicalein controls mitochondrial fusion/fission balance and inhibits GRP78/CHOP pathway, thus exerting therapeutic effects in ISO-induced heart failure in HL-1 cells and BALB/c mice. These results suggested that baicalein may be a potential therapeutic agent for heart failure.

Objective:To investigate the clinical characteristics and treatment outcomes of patients with blast-induced hearing loss（BIHL）. Methods:The clinical features, laboratory parameters, audiometric profiles, and treatment efficacy of patients with blast induced hearing loss and those with idiopathic sudden hearing loss（ISHL） were analyzed using t-tests, Wilcoxon rank-sum tests, and chi-square tests, with a significance level set at P<0.05. Results:A total of 59 patients in the BIHL group and 117 patients in the ISHL group were included in this study. The mean age of the BIHL group was（39.07±14.49） years, comprising 45 males and 14 females. After the blast, 21 patients went to the hospital within the initial 14-day period, and an additional 38 patients seeking admission thereafter. In the BIHL group, 33 patients had unilateral hearing loss with PTA of （50.30±28.85） dB HL, while 26 had bilateral hearing loss with a PTA of（44.54±26.22） dB HL. In comparison, among the ISHL group, 112 patients had unilateral hearing loss with a PTA of（56.28±14.19） dB HL, and 5 had bilateral involvement with a PTA of（56.25±35.14） dB HL. The effective treatment rate within 14 days for the BIHL group was 31.8%, while for the ISHL group, the effective rate within 14 days was 77.0%. Conclusion:Blast-induced hearing loss is caused by exposure to high-intensity noise. The overall treatment effectiveness during hospitalization is lower compared to idiopathic sudden hearing loss, and the treatment window is shorter. Therefore, greater emphasis should be placed on prevention.

Objectives:To investigate the clinical and audiological features in auditory neuropathy (AN) patients with different image performance of cochlear nerve (CN) on MRI. Methods:AN patients were selected from the Multicenter Study on Clinical Diagnosis and Intervention of AN (2003-2023). Subjects were divided into four groups by CN size on MRI: bilateral AN with CN normalization (BANcnn), bilateral CN deficiency (BANcnd), unilateral AN with CND (UANcnd), and CN normalization (UANcnn) groups. Clinical and audiological features were analyzed, alongside differential analysis using the Internal auditory meatus (IAM) grading system. Results:Totally 132 cases were included. The rate of CND was 35.6% (47/132), with 32.7% (36/110) and 50.0% (11/22) in bilateral and unilateral AN patients. The proportion of IAM I to IV grades increased progressively. Clinical and audiological characteristics differed between bilateral and unilateral AN patients. For bilateral AN patients, the age of onset and diagnosis of BANcnd were older, which were 11.8 ± 7.2 years and 19.5 ± 11.1 years, respectively, compared to patients in BANcnn group. UANcnd patients exhibited more severe hearing loss at all frequencies compared to UANcnn. Prognostic analysis indicated that AN patients with CND had no lower intervention rates. Conclusion:The detection rate of CND was 35.6%, predominantly in unilateral AN patients and IAM IV was the most common type. Some clinical and audiological indicators were associated with imaging features of CN but varied between bilateral and unilateral AN. Variation in image performance may not be related to prognosis in AN patients.

OBJECTIVE:Benign paroxysmal positional vertigo is a common cause of vertigo in adults but has been studied less extensively in children. Interestingly, the reported diagnostic frequency of benign paroxysmal positional vertigo in paediatric vertigo showed high variability. This systematic review and meta-analysis aimed to evaluate the proportion of benign paroxysmal positional vertigo in childhood vertigo and explore relevant factors to accurately diagnose BPPV in children. DESIGN:Systematic review and meta-analysis. SETTING AND PARTICIPANTS:PubMed, EMBASE database and Cochrane library (from 1 January 1962, to 12 April 2022) were searched for eligible studies analysing the proportion of benign paroxysmal positional vertigo in children with vertigo. Only papers published in English were included. MAIN OUTCOME MEASURES:The pooled proportion of paediatric BPPV in childhood vertigo, and its corresponding 95% confidence interval (CI), were calculated. Additional subgroup analysis was performed by year of publication, testing techniques, department, regions, age and registering comorbidities of relevance for balance. RESULTS:Ultimately, 26 studies were eligible for our meta-analysis. These studies involved 4043 childhood vertigo cases, of which 460 were BPPV. Based on the results of a random-effects model-based meta-analysis, the pooled proportion of benign paroxysmal positional vertigo in childhood vertigo was 5.5%. However, a subgroup analysis revealed a significantly higher pooled proportion of benign paroxysmal positional vertigo, with 9.9% in the videonystagmography subgroup and 17.1% in the comorbidity subgroup. Moreover, the proportion of BPPV varies among children with vertigo in different age groups. CONCLUSION:In conclusion, the pooled proportion of BPPV in vertigo children was 5.5% across all samples. The present study revealed that age, applying VNG in positional testing, and comorbidity diagnosis may be closely related to the diagnosis of BPPV in children. Additionally, to prevent missed diagnosis of benign paroxysmal positional vertigo, diagnostic BPPV tests should not be neglected in children suffering from other vertigo-associated diseases.

Hearing is one of the most vital sensory functions in human beings and a crucial means of perceiving and acquiring information from the natural environment. The advancement of human society is closely linked to the development of language, with hearing serving as the foundation for verbal communication. As individuals age, the deterioration of the auditory system becomes a significant factor contributing to sensory impairments in the elderly. In addition to hearing loss, the aging of the auditory system is also associated with cognitive decline and psychosocial disorders, which severely impact the quality of life for older adults. Currently, there are no effective treatments or interventions available for addressing the aging of the auditory system. Therefore, identifying biomarkers of the auditory system aging is of great significance. The Aging Biomarker Consortium of China has conducted a comprehensive evaluation of aging biomarkers in the auditory system, focusing on three dimensions: morphological, functional, and humoral biomarkers. This initiative aims to establish a foundation for assessing the degree of aging in the auditory system and to promote the management of auditory health in an aging society, ultimately enhancing the auditory health of the elderly population both in China and globally.

Exposure to different types of noise for varying durations can lead to damage in different parts of the auditory pathway, such as synapse and hair cell loss, resulting in temporary or permanent threshold shifts. Noise-induced hearing loss (NIHL) involves intricate mechanisms in its pathogenesis, with oxidative stress and glutamate excitotoxicity considered important factors. Currently, there is a lack of effective drugs for NIHL. This study exhibited noise-induced hidden hearing loss (NIHHL) and noise-induced permanent hearing loss in CBA/CaJ mouse models and found that pre-administration of NADH via intraperitoneal injection effectively mitigated noise-induced damage in both models. NIHHL mice exhibited threshold recovery at 14 days post-exposure but mild sustained synapse loss. NIHL mice developed persistent threshold elevation with irreversible ribbon synapse and outer hair cells (OHC) damage. NADH pretreatment mitigated temporary threshold shifts in the NIHHL model, while in the NIHL model, it reduced threshold elevation, attenuated synapse and OHC loss, and preserved stereocilia bundle integrity. Furthermore, pretreatment with NADH significantly reduced cochlear Ca²⁺ levels and delayed the time to peak Ca²⁺ concentration in NIHL mice. Concurrently, upregulated mGluR expression and dynamic changes in P-PKC levels were observed, with P-PKC exhibiting a trend consistent with Ca²⁺ fluctuations. Additionally, cochlear ROS levels decreased, while NRF2 was rapidly upregulated and underwent nuclear translocation in OHCs, accompanied by enhanced mRNA expression of its downstream antioxidant factors. These results indicated that the mitigation of hearing loss by NADH pretreatment in NIHL mice may involve both the mGluR-PKCCa²⁺ and NRF2 pathways.

OBJECTIVES:Over the past 30 years, significant advancements have been made in auditory neuropathy (AN) research. The study aimed to explore auditory function and the natural history of AN from neonates to adulthood based on a large Chinese AN patient population. DESIGN:This retrospective case series included 645 patients diagnosed with AN at our institution, the Chinese PLA General Hospital, over a 27-year period from January 1997 to December 2023. All patients had an absent or abnormal auditory brainstem response (ABR), normal or slightly altered otoacoustic emissions and/or cochlear microphonics (CM), and binaural onset. RESULTS:The study enrolled patients who were from 29 provinces and 11 different ethnic groups in China. The diagnostic rate of AN has ranged from 1 case per year to 44 diagnoses per year. The study assessed the typical auditory characteristics of individuals at various ages at diagnosis. Statistically significant differences were found in the elicitation rates of ABR, distortion product otoacoustic emissions, acoustic stapedius reflex threshold, pure-tone average, 40 Hz auditory event-related potential, and progress of speech discrimination scores in AN patients from neonates to adulthood; however, the elicitation of CM ranged from 96.36 to 100% from neonates to adulthood. A relatively large proportion of AN patients had hearing loss predominantly affecting the lower frequency range. Follow-up results showed that patients with AN had stable abnormal or absent ABR, distortion product otoacoustic emissions gradually disappeared, but CM remained stable, and poor speech discrimination scores persisted. Distinct clinical profiles were observed in patients with AN carrying pathogenic or likely pathogenic mutations, including OTOF, AIFM1, ATP1A3, WFS1 , and OPA1 . OTOF and ATP1A3 mutation carriers had early onset and poor hearing thresholds, while AIFM1 had later onset and better hearing thresholds by multiple comparisons. Interventions included hearing aids with mainly bilateral interventions, and cochlear implants with predominantly unilateral intervention. Cochlear implant recipients showed lower diagnostic and onset ages and superior aided hearing thresholds compared with hearing aid users. CONCLUSIONS:Advances in disease awareness, hearing screening, and diagnostic methods have enabled early detection and diagnosis of AN. Follow-up results highlighted the importance of CM in evaluating the function of outer hair cells. AN patients experienced a higher frequency of hearing loss progression while maintaining poor, unchangeable speech recognition ability. This study elucidates the multifaceted nature of AN, thereby establishing a more comprehensive understanding of the diagnosis and management of AN. Moreover, it serves as a pivotal reference for guiding counseling and evaluating the prognoses of clinical AN patients.

The prevalence of RNA binding proteins (RBPs) in regulating alternative splicing (AS) in age-related sensorineural hearing loss (SNHL) is unclear. To address this question, we comprehensively analyzing RNA-seq data from a mouse model to identify deregulated AS events (RASEs) and differentially expressed RNA binding proteins (DERBPs) associated with age-related SNHL, and constructed the networks between these two levels of changes. We have identified 198 and 187 RASEs related to severe and mild symptoms, respectively. Genes harboring these RASEs are significantly enriched in positive regulation of GTPase activity and cytoskeleton organization. We further identified 25 and 14 DERBPs related to severe and mild symptoms, respectively, and constructed the regulatory network between DERBPs and RASEs. Among these DERBPs, the significantly increased expression of Isg15 and Myom1, the decreased expression of Acan, as well as some of their regulated RASEs including two in Uap1 also demonstrated in cochlear transcriptomes obtained from the age-related SNHL mouse model constructed in this study. Analysis of human pluripotent stem cell-derived macrophages containing ISG15 knockout samples revealed that ISG15 is a key splicing regulator, and can directly regulate the alternative splicing of UAP1 in human cells. These findings prove insights into the involvement of a large number of alternative splicing events driven by RNA-binding proteins (RBPs) in the pathogenesis of age-related SNHL, and suggest their potentials as therapeutic targets and disease markers.

Noise-induced hearing loss(NIHL)is an acquired sensorineural hearing loss induced by long-term noise exposure.The susceptibility of exposed people may vary even in the same noise environment.With the de-velopment of sequencing techniques,genes related to oxidative stress,immunoinflammatory,ion homeostasis,energy metabolism,DNA damage repair and other mechanisms in NIHL have been reported continuously.And some genes may interact with noise exposure indexes.In this article,population studies on NIHL-related gene polymorphisms and gene-environment interactions in the past 20 years are reviewed,aimed to providing evidence for the construction of NIHL-related risk prediction models and the formulation of individualized interventions.

Auditory neuropathy spectrum disorder (ANSD) is a hearing impairment involving disruptions to inner hair cells (IHCs), ribbon synapses, spiral ganglion neurons (SGNs), and/or the auditory nerve itself. The outcomes of cochlear implants (CI) for ANSD are variable and dependent on the location of lesion sites. Discovering a potential therapeutic agent for ANSD remains an urgent requirement. Here, 293T stable transfection cell lines and patient induced pluripotent stem cells (iPSCs)-derived auditory neurons carrying the apoptosis inducing factor (AIF) p.R422Q variant were used to pursue a therapeutic regent for ANSD. Nicotinamide adenine dinucleotide (NADH) is a main electron donor in the electron transport chain (ETC). In 293T stable transfection cells with the p.R422Q variant, NADH treatment improved AIF dimerization, rescued mitochondrial dysfunctions, and decreased cell apoptosis. The effects of NADH were further confirmed in patient iPSCs-derived neurons. The relative level of AIF dimers was increased to 150.7 % (P = 0.026) from 59.2 % in patient-neurons upon NADH treatment. Such increased AIF dimerization promoted the mitochondrial import of coiled-coil-helix-coiled-coilhelix domain-containing protein 4 (CHCHD4), which further restored mitochondrial functions. Similarly, the content of mitochondrial calcium (mCa2+) was downregulated from 136.7 % to 102.3 % (P = 0.0024) in patientneurons upon NADH treatment. Such decreased mCa2+ levels inhibited calpain activity, ultimately reducing the percentage of apoptotic cells from 30.5 % to 21.1 % (P = 0.021). We also compared the therapeutic effects of gene correction and NADH treatment on hereditary ANSD. NADH treatment had comparable restorative effects on functions of ANSD patient-specific cells to that of gene correction. Our findings offer evidence of the molecular mechanisms of ANSD and introduce NADH as a potential therapeutic agent for ANSD therapy.

Non-syndromic sensorineural hearing loss is characterized by genetic heterogeneity, leading to potential clinical misdiagnosis. TMC1 , a unique causative gene associated with deafness, exhibits variants with autosomal dominant and recessive inheritance patterns. TMC1 codes for the transmembrane channel-like 1 (TMC1), a key component of the mechano-electrical transduction (MET) machinery for hearing. However, the molecular mechanism of Ca2+ regulation in MET, which is essential for sound perception, remains unclear. CIB2, another MET component associated with deafness, can bind with calcium and may play a role in MET gating. Our study reveals that the TMC1-CIB2 complex undergoes a Ca2+-induced conformational change, highlighting the crucial role of Ca2+ in MET. We identified a vertebrate-specific binding site on TMC1, named CR3, that interacts with apo CIB2, with the binding interface linked to hearing loss. Using an in-vivo animal model, we demonstrated that disruption of the calcium-sensing site of CIB2 perturbs the MET channel conductivity, confirming its role as a Ca2+ sensor for MET. Additionally, after systematically analyzing the hearing-loss variants, we observed that dominant mutations of TMC1 are often located near the ion pore or at the binding interfaces with the Ca2+ sensor CIB2. This provides a detailed mechanism of the genetic heterogeneity of TMC1 in hearing loss at an atomic level. ### Competing Interest Statement The authors have declared no competing interest.

Auditory neuropathy (AN) is a unique type of language developmental disorder, with no precise rate of genetic contribution that has been deciphered in a large cohort. In a retrospective cohort of 311 patients with AN, pathogenic and likely pathogenic variants of 23 genes were identified in 98 patients (31.5% in 311 patients), and 14 genes were mutated in two or more patients. Among subgroups of patients with AN, the prevalence of pathogenic and likely pathogenic variants was 54.4% and 56.2% in trios and families, while 22.9% in the cases with proband-only; 45.7% and 25.6% in the infant and non-infant group; and 33.7% and 0% in the bilateral and unilateral AN cases. Most of the OTOF gene (96.6%, 28/29) could only be identified in the infant group, while the AIFM1 gene could only be identified in the non-infant group; other genes such as ATP1A3 and OPA1 were identified in both infant and non-infant groups. In conclusion, genes distribution of AN, with the most common genes being OTOF and AIFM1 , is totally different from other sensorineural hearing loss. The subgroups with different onset ages showed different genetic spectrums, so did bilateral and unilateral groups and sporadic and familial or trio groups.

Mutations in AIFM1, encoding for apoptosis-inducing factor (AIF), cause AUNX1, an X-linked neurologic disorder with late-onset auditory neuropathy (AN) and peripheral neuropathy. Despite significant research on AIF, there are limited animal models with the disrupted AIFM1 representing the corresponding phenotype of human AUNX1, characterized by late-onset hearing loss and impaired auditory pathways. Here, we generated an Aifm1 p.R450Q knock-in mouse model (KI) based on the human AIFM1 p.R451Q mutation. Hemizygote KI male mice exhibited progressive hearing loss from P30 onward, with greater severity at P60 and stabilization until P210. Additionally, muscle atrophy was observed at P210. These phenotypic changes were accompanied by a gradual reduction in the number of spiral ganglion neuron cells (SGNs) at P30 and ribbons at P60, which coincided with the translocation of AIF into the nucleus starting from P21 and P30, respectively. The SGNs of KI mice at P210 displayed loss of cytomembrane integrity, abnormal nuclear morphology, and dendritic and axonal demyelination. Furthermore, the inner hair cells and myelin sheath displayed abnormal mitochondrial morphology, while fibroblasts from KI mice showed impaired mitochondrial function. In conclusion, we successfully generated a mouse model recapitulating AUNX1. Our findings indicate that disruption of Aifm1 induced the nuclear translocation of AIF, resulting in the impairment in the auditory pathway.

Hereditary deafness is a common neurosensory disorder, and 148 non-syndromic deafness genes have been identified to date. Gene therapy has been used to treat a variety of genetic diseases, but no gene therapy drug for hereditary deafness has been approved for clinical use. At present, several clinical trials of gene therapy for hereditary deafness are underway. However, few normative documents have been issued to guide the standardization of gene therapy for hearing loss, and this document is the first global gene therapy guideline for hereditary hearing loss. The guidelines were jointly developed and drafted by experienced audiologists, virologists and biologists who are vigorously involved in inner ear gene therapy research in the Hearing, Speech and Communication Subsociety of Biophysical Society of China, Audiology Development Foundation Of China and Audiology Subsociety of Jiangsu Medical Association. These guidelines cover preclinical research and clinical practice of gene therapy for hereditary deafness, including indications, key points of pre-clinical research, patient selection criteria, pre-clinical preparation, drug efficacy, drug safety evaluation criteria, ethical review, etc. We hope that the guidelines will promote the standardization of clinical practice related to gene therapy for hereditary deafness in China and around the world.

Objective:To dentify the genetic and audiological characteristics of families affected by late-onset hearing loss due to GSDMEgene mutations, aiming to explore clinical characteristics and pathogenic mechanisms for providing genetic counseling and intervention guidance. Methods:Six families with late-onset hearing loss from the Chinese Deafness Genome Project were included. Audiological tests, including pure-tone audiometry, acoustic immittance, speech recognition scores, auditory brainstem response, and distortion product otoacoustic emission, were applied to evaluate the hearing levels of patients. Combining with medical history and physical examination to analyze the phenotypic differences between the probands and their family members. Next-generation sequencing was used to identify pathogenic genes in probands, and validations were performed on their relatives by Sanger sequencing. Pathogenicity analysis was performed according to the American College of Medical Genetics and Genomics Guidelines. Meanwhile, the pathogenic mechanisms of GSDME-related hearing loss were explored combining with domestic and international research progress. Results:Among the six families with late-onset hearing loss, a total of 30 individuals performed hearing loss. The onset of hearing loss in these families ranged from 10 to 50 years（mean age: 27.88±9.74 years）. In the study, four splicing mutations of the GSDME were identified, including two novel variants: c. 991－7C>G and c. 1183+1G>T. Significantly, the c. 991－7C>G was a de novo variant. The others were previously reported variants: c. 991-1G>C and c. 991-15_991-13del, the latter was identified in three families. Genotype-phenotype correlation analysis revealed that probands with the c. 991－7C>G and c. 1183+1G>T performed a predominantly high-frequency hearing loss. The three families carrying the same mutation exhibited varying degrees of hearing loss, with an annual rate of hearing deterioration exceeding 0.94 dB HL/year. Furthermore, follow-up of interventions showed that four of six probands received intervention（66.67%）, but the results of intervention varied. Conclusion:The study analyzed six families with late-onset non-syndromic hearing loss linked to GSDME mutations, identifying four splicing variants. Notably, c. 991－7C>G is the first reported de novo variant of GSDME globally. Audiological analysis revealed that the age of onset generally exceeded 10 years,with variable effectiveness of interventions.