DDX58 variant triggers IFN-ß-induced autophagy in trabecular meshwork and influences intraocular pressure.

Singleton-Merten syndrome (SMS) is a rare immunogenetic disorder affecting multiple systems, characterized by dental dysplasia, aortic calcification, glaucoma, skeletal abnormalities, and psoriasis. Glaucoma, a key feature of both classical and atypical SMS, remains poorly understood in terms of its molecular mechanism caused by DDX58 mutation. This study presented a novel DDX58 variant (c.1649A>C [p.Asp550Ala]) in a family with childhood glaucoma. Functional analysis showed that DDX58 variant caused an increase in IFN-stimulated gene expression and high IFN-ß-based type-I IFN. As the trabecular meshwork (TM) is responsible for controlling intraocular pressure (IOP), we examine the effect of IFN-ß on TM cells. Our study is the first to demonstrate that IFN-ß significantly reduced TM cell viability and function by activating autophagy. In addition, anterior chamber injection of IFN-ß remarkably increased IOP level in mice, which can be attenuated by treatments with autophagy inhibitor chloroquine. To uncover the specific mechanism underlying IFN-ß-induced autophagy in TM cells, we performed microarray analysis in IFN-ß-treated and DDX58 p.Asp550Ala TM cells. It showed that RSAD2 is necessary for IFN-ß-induced autophagy. Knockdown of RSAD2 by siRNA significantly decreased autophagy flux induced by IFN-ß. Our findings suggest that DDX58 mutation leads to the overproduction of IFN-ß, which elevates IOP by modulating autophagy through RSAD2 in TM cells.

The NLRP3 inflammasome as a target for sensorineural hearing loss.

Sensorineural hearing loss is the most common type of hearing loss in adults and occurs due to damage of the inner ear caused by a range of factors including ageing, excessive noise, toxins, and cancer. Auto-inflammatory disease is also a cause of hearing loss and there is evidence that inflammation could contribute to hearing loss in other conditions. Within the inner ear there are resident macrophage cells that respond to insults and whose activation correlates with damage. The NLRP3 inflammasome is a multi-molecular pro-inflammatory protein complex that forms in activated macrophages and may contribute to hearing loss. The aim of this article is to discuss the evidence for the NLRP3 inflammasome and associated cytokines as potential therapeutic targets for sensorineural hearing loss in conditions ranging from auto-inflammatory disease to tumour-induced hearing loss in vestibular schwannoma.

Regenerative Therapy Approaches and Encountered Problems in Sensorineural Hearing Loss.

Hearing loss is one of the most important public health matters worldwide, severely affecting people's social, psychological, and cognitive development. The perception of sound, movement, and balance in vertebrates depends on a special sensory organ called the cochlea, which contains hair cells and supporting cells in the inner ear. Genetic factors, epigenetics, the use of ototoxic drugs (some antibiotics and chemotherapeutics), noise, infections, or even aging can cause loss of hair cells and their related primary neurons, leading to sensorineural hearing loss. Although a sensorineural hearing loss, also known as permanent hearing loss, is treated with hearing aids and cochlear implants, treatment methods are limited. Since even the best implant cannot exhibit the characteristics of the original ear, the permanent sensory deficit will be permanent. For this reason, it has become important to develop regenerative treatment methods to regenerate and replace lost or damaged hair cells and neurons. Developments in stem cell technology have led to promising studies in regenerating damaged/lost hair cells or neurons with endogenous or exogenous cell-based therapies. Epigenetic mechanisms can turn hearing-related genes on and off and determine which proteins to copy. In addition, due to gene silencing, gene replacement, and CRISPR/CAS9 technology, gene therapy methods have accelerated, and studies have been carried out to treat dominant and recessive mutations that cause genetic-induced hearing loss or increase hair cell regeneration. In this paper, potential gene therapy and stem cell applications in the acquisition of cochlear function, which causes sensorineural hearing loss, and the difficulties encountered in these applications are compiled from a bioengineering perspective.

Aucubin protects mouse cochlear hair cells from cisplatin-induced ototoxicity via activation of the PI3K/AKT/STAT3 pathway.

Cisplatin is commonly used to treat cancers and is associated with a significant risk of irreversible sensorineural hearing loss. However, no effective preventive strategies are available for cisplatin-induced HL. Therefore, significant efforts have been made to discover new drugs protecting cochlear hair cells from cisplatin-induced damage. We found that a new phytochemical, aucubin, attenuated cisplatin-induced apoptosis, the production of reactive oxygen species, and mitochondrial dysfunction in House Ear Institute Organ of Corti 1 cells and cochlear hair cells. Moreover, aucubin attenuated cisplatin-induced sensorineural hearing loss and hair cells loss in vivo. Furthermore, RNA sequencing analysis revealed that the otoprotective effects of aucubin were mainly mediated by increased STAT3 phosphorylation via the PI3K/AKT pathway. Inhibition of the STAT3 signaling pathway with the inhibitor S3I-201 or siRNA disrupted the protective effects of aucubin on cisplatin-induced apoptosis. In conclusion, we identified an otoprotective effect of aucubin. Therefore, aucubin could be used to prevent cisplatin-induced ototoxicity.

Murine cytomegalovirus employs the mixed lineage kinases family to regulate the spiral ganglion neuron cell death and hearing loss.

Cytomegalovirus (CMV)-induced sensorineural hearing loss (SNHL) is a worldwide epidemic. Recent studies have shown that the degree of spiral ganglion neuron (SGN) loss is correlated with hearing loss after CMV infection. We aimed to better understand the pathological mechanisms of CMV-related SGN death and to search for intervention measures. We found that both apoptosis and pyroptosis are involved in CMV-induced SGN death, which may be caused by the simultaneous activation of the p53/JNK and NLRP3/caspase-1 signaling pathways, respectively. Moreover, considering that mixed lineage kinase family (MLK1/2/3) are host restriction factors against viral infection and upstream regulators of the p53/JNK and inflammatory (including NLRP3-caspase1) signaling pathways, we further demonstrated that the MLKs inhibitor URMC-099 exhibited a protective effect against CMV-induced SGN death and hearing loss. These results indicate that MLKs signaling may be a key regulator and promising novel target for preventing apoptosis and even pyroptosis during the CMV infection of SGN cells and for treating hearing loss.

Gene therapy with a synthetic adeno-associated viral vector improves audiovestibular phenotypes in Pjvk-mutant mice.

Recessive PJVK mutations that cause a deficiency of pejvakin, a protein expressed in both sensory hair cells and first-order neurons of the inner ear, are an important cause of hereditary hearing impairment. Patients with PJVK mutations garner limited benefits from cochlear implantation; thus, alternative biological therapies may be required to address this clinical difficulty. The synthetic adeno-associated viral vector Anc80L65, with its wide tropism and high transduction efficiency in various inner ear cells, may provide a solution. We delivered the PJVK transgene to the inner ear of Pjvk mutant mice using the synthetic Anc80L65 vector. We observed robust exogenous pejvakin expression in the hair cells and neurons of the cochlea and vestibular organs. Subsequent morphologic and audiologic studies demonstrated significant restoration of spiral ganglion neuron density and hair cells in the cochlea, along with partial recovery of sensorineural hearing impairment. In addition, we observed a recovery of vestibular ganglion neurons and balance function to WT levels. Our study demonstrates the utility of Anc80L65-mediated gene delivery in Pjvk mutant mice and provides insights into the potential of gene therapy for PJVK-related inner ear deficits.

Head and neck radiotherapy causes significant disruptions of cochlear ribbon synapses and consequent sensorineural hearing loss.

PURPOSE: To investigate the possible effects of head and neck radiotherapy on hearing function in mice. METHOD: Adult C57BL/6J mice were irradiated to the head and neck once with cobalt-60 rays at doses of 10 Gy or 20 Gy. Hearing function was estimated by the detection of auditory brainstem response (ABR) thresholds and the suprathreshold function of cochlear was indicated by the peak amplitudes and latencies of wave I. The mice were tested on days 1, 7, 14, and 21 after radiation treatment, and untreated mice in littermates served as controls. The cochlear pre-synaptic ribbons were labeled using an anti-RIBEYE/CtBP2 antibody, and the synaptic vesicle membrane was traced using anti-vesicular glutamate transporter 3 (VGLUT-3) antibody. The number and size of the pre-synaptic ribbons were counted along the cochlear axis from the apex to the base. The expression of VGLUT-3 was measured by the intensity of immunofluorescence. Hematoxylin and eosin (H&E) staining was also performed to evaluate the structural changes in the cochlea. RESULTS: Compared with the controls, mice treated with 10 Gy and 20 Gy doses on days 1, 7, 14, and 21 were found to have significant disruptions in ABR thresholds and amplitudes (p < 0.05). Moreover, mice in the 20 Gy group, compared with the 10 Gy group, showed greater hearing loss and suprathreshold deficits (p < 0.05). Quantitative analysis revealed a decrease in the number and size of CtBP2-positive puncta in both the 10 Gy and 20 Gy groups compared with the controls (p < 0.05); in the 20 Gy group, the number and size of CtBP2-positive puncta were less than those in the 10 Gy group (p < 0.05). We observed a significant disruption in the expression of VGLUT-3 in the group treated with 20 Gy. However, compared with the control group, both immunofluorescence and H&E staining revealed no significant changes in the number of hair cells or the array for the 10 or 20 Gy treatments (p > 0.05). CONCLUSION: Radiation therapy targeting the head and neck can cause sensorineural hearing loss via disruption specific to the cochlear ribbon synapses. To our knowledge, this is the first study to demonstrate that cochlear ribbon synapses may be a subcellular target of radiation-induced hearing loss.

Activation of Rictor/mTORC2 signaling acts as a pivotal strategy to protect against sensorineural hearing loss.

The Food and Drug Administration­approved drug sirolimus, which inhibits mechanistic target of rapamycin (mTOR), is the leading candidate for targeting aging in rodents and humans. We previously demonstrated that sirolimus could treat ARHL in mice. In this study, we further demonstrate that sirolimus protects mice against cocaine-induced hearing loss. However, using efficacy and safety tests, we discovered that mice developed substantial hearing loss when administered high doses of sirolimus. Using pharmacological and genetic interventions in murine models, we demonstrate that the inactivation of mTORC2 is the major driver underlying hearing loss. Mechanistically, mTORC2 exerts its effects primarily through phosphorylating in the AKT/PKB signaling pathway, and ablation of P53 activity greatly attenuated the severity of the hearing phenotype in mTORC2-deficient mice. We also found that the selective activation of mTORC2 could protect mice from acoustic trauma and cisplatin-induced ototoxicity. Thus, in this study, we discover a function of mTORC2 and suggest that its therapeutic activation could represent a potentially effective and promising strategy to prevent sensorineural hearing loss. More importantly, we elucidate the side effects of sirolimus and provide an evaluation criterion for the rational use of this drug in a clinical setting.

Perilymph metabolomic and proteomic MALDI-ToF profiling with porous silicon chips: A proof-of-concept study.

INTRODUCTION: Sensorineural hearing losses (SNHLs) are a significant public health issue, and the hearing loss field is desperately in need of effective therapy. Pathophysiological mechanisms are not yet clearly understood in the absence of validated methods to assess the inner ear content. Proteomic and metabolomic analysis of perilymph is opening new research perspectives for SNHLs. We aimed to demonstrate the feasibility of an innovative mass spectrometry (MS) strategy using porous silicon chips (PSCs) to investigate the low molecular weight (LMW) protein and metabolite content of human perilymph. Our second objective was to stratify perilymph samples according to their MS profiles and compare these results with clinical data. MATERIAL AND METHODS: Perilymph samples obtained during cochlear implant surgery from patients with SNHLs were retrieved from a validated biobank. To focus on LMW entities, we used a PSC enrichment protocol before MALDI-ToF MS analysis. PSCs were used as a LMW molecular preanalytical stabilizer and amplifier. Patients' clinical data and SNHL characteristics were retrieved retrospectively from medical charts. RESULTS: We successfully acquired and compared 59 exploitable MS profiles out of 71 perilymph samples. There was a good correlation between duplicates. Comparing both ears from the same patient, we found good reproducibility even when there was a one-year interval between samplings. We identified three distinct groups when comparing the samples' metabolomic profiles and four homogeneous groups comparing their LMW proteome profiles. Clinical data analysis suggested that some groups shared clinical or preanalytical characteristics. CONCLUSION: This proof-of-concept study confirms that LMW proteome and metabolome content of perilymph can be analyzed with PSCs. Based on protein profiles, we managed to stratify perilymp samples according to their molecular composition. These results must be confirmed with a larger population, and sampling methods require improvement, but this approach seems promising. In the future, this approach may pave the way for companion test strategies to precisely diagnose and define potential molecular targets for audioprotective therapies.

Mechanism of NLRX1 Mediating JNK Apoptosis Pathway to Participate in Functional Impairment of Cochlear Hair Cells in Mice with Senile Deafness.

To explore the mechanism of cochlear hair cell damage and study the prevention and treatment of sensorineural hearing loss, the effect of NLRX1 gene expression on the functional damage of cochlear hair cells in presbycusis was comprehensively analyzed. In the in vivo detection, C57BL/6 mice of different ages were used as experimental subjects. Cochlear tissues were taken after the hearing test of mice, and the number of cells and protein changes in NLRX1 immunofluorescence staining were detected. In the in vitro detection, the cochlear hair cell HEI-OE1 was used as the experimental object, and the cell proliferation activity was detected after overexpression or silencing of NLRX1.In the in vivo and in vitro experiments, the expression of JNK pathway-related proteins was simultaneously detected. The results of in vivo experiments showed that the hearing threshold of 270d -old mice was substantially greater than that of 15d-, 30d-, and 90d-old mice (P <0.05). I addition, with increasing age, the expression of p-JNK, Bcl-2, Bax, and Caspase-3 in the mouse cochlea gradually increased (P<0.05).In vitro experimental results showed that cell proliferation activity decreased after overexpression of NLRX1, and the expression of p-JNK, Bcl-2, Bax, and Caspase-3 was substantially decreased (P<0.05). Silencing NLRX1 can inhibit the above phenomenon, indicating that NLRX1 can inhibit the proliferation of hair cells in old mice through the activation of the JNK apoptosis pathway, thereby promoting the occurrence of sensorineural hearing loss.

Mutational signatures in GATA3 transcription factor and its DNA binding domain that stimulate breast cancer and HDR syndrome.

Transcription factors (TFs) play important roles in many biochemical processes. Many human genetic disorders have been associated with mutations in the genes encoding these transcription factors, and so those mutations became targets for medications and drug design. In parallel, since many transcription factors act either as tumor suppressors or oncogenes, their mutations are mostly associated with cancer. In this perspective, we studied the GATA3 transcription factor when bound to DNA in a crystal structure and assessed the effect of different mutations encountered in patients with different diseases and phenotypes. We generated all missense mutants of GATA3 protein and DNA within the adjacent and the opposite GATA3:DNA complex models. We mutated every amino acid and studied the new binding of the complex after each mutation. Similarly, we did for every DNA base. We applied Poisson-Boltzmann electrostatic calculations feeding into free energy calculations. After analyzing our data, we identified amino acids and DNA bases keys for binding. Furthermore, we validated those findings against experimental genetic data. Our results are the first to propose in silico modeling for GATA:DNA bound complexes that could be used to score effects of missense mutations in other classes of transcription factors involved in common and genetic diseases.

Sex Differences in the Triad of Acquired Sensorineural Hearing Loss.

The triad of noise-generated, drug-induced, and age-related hearing loss is the major cause of acquired sensorineural hearing loss (ASNHL) in modern society. Although these three forms of hearing loss display similar underlying mechanisms, detailed studies have revealed the presence of sex differences in the auditory system both in human and animal models of ASNHL. However, the sexual dimorphism of hearing varies among noise-induced hearing loss (NIHL), ototoxicity, and age-related hearing loss (ARHL). Importantly, estrogen may play an essential role in modulating the pathophysiological mechanisms in the cochlea and several reports have shown that the effects of hormone replacement therapy on hearing loss are complex. This review will summarize the clinical features of sex differences in ASNHL, compare the animal investigations of cochlear sexual dimorphism in response to the three insults, and address how estrogen affects the auditory organ at molecular levels.

Sensorineural hearing loss may lead to dementia-related pathological changes in hippocampal neurons.

Presbycusis contributes to cognitive decline and Alzheimer's disease. However, most research in this area involves clinical observations and statistical modeling, and few studies have examined the relationship between hearing loss and the molecular changes that lead to cognitive dysfunction. The present study investigated whether hearing loss contributes to dementia in the absence of aging and noise using a mouse model of severe bilateral hearing loss induced by kanamycin (1000 mg/kg) and furosemide (400 mg/kg). Immunohistochemistry, silver staining, immunofluorescence analysis, and Western blotting were used to observe pathological changes in different regions of the hippocampus in animals with hearing loss. Changes in the cognitive function of animals with hearing loss were assessed using the Morris water maze test. The results showed that neurons began to degenerate 60 days after hearing loss, and this degeneration was accompanied by structural disorganization and decreased neurogenesis. The level of phosphorylated tau increased over time. Increases in escape latency and distance traveled during the training phase of the Morris water maze test were observed 90 days after hearing loss. Activated microglia and astrocytes with increased levels of inflammatory cytokines, such as tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) were detected in the hippocampus. These results suggest that hearing loss alone causes neuronal degeneration, inhibition of neurogenesis, increased tau protein phosphorylation, and increased neuroinflammation in the hippocampus. Early intervention in individuals with hearing loss may reduce the risk of cognitive decline.

Deficiency of Klc2 Induces Low-Frequency Sensorineural Hearing Loss in C57BL/6 J Mice and Human.

The transport system in cochlear hair cells (HCs) is important for their function, and the kinesin family of proteins transports numerous cellular cargos via the microtubule network in the cytoplasm. Here, we found that Klc2 (kinesin light chain 2), the light chain of kinesin-1 that mediates cargo binding and regulates kinesin-1 motility, is essential for cochlear function. We generated mice lacking Klc2, and they suffered from low-frequency hearing loss as early as 1 month of age. We demonstrated that deficiency of Klc2 resulted in abnormal transport of mitochondria and the down-regulation of the GABAA receptor family. In addition, whole-genome sequencing (WGS) of patient showed that KLC2 was related to low-frequency hearing in human. Hence, to explore therapeutic approaches, we developed adeno-associated virus containing the Klc2 wide-type cDNA sequence, and Klc2-null mice delivered virus showed apparent recovery, including decreased ABR threshold and reduced out hair cell (OHC) loss. In summary, we show that the kinesin transport system plays an indispensable and special role in cochlear HC function in mice and human and that mitochondrial localization is essential for HC survival.

Evaluation of DNA damages in congenital hearing loss patients.

In the current study, we aimed to compare the level of genetic damages measured as micronucleus (MN), nucleoplasmic bridge (NPB), and nuclear bud formation (NBUD) in congenital hearing loss patients (n = 17) and control group (n = 24). The cytokinesis-blocked micronucleus assay (CBMN) was applied to the blood samples to measure the frequency of the markers in both groups. The frequencies of MN of hearing loss patients were found to be consistently significantly higher than those obtained for the control group (p < 0.0001). Similarly, we found significantly higher frequency of NPB in patients was obtained for the patient group (p < 0.0001). Finally, the frequencies of NBUD in patients is significantly higher than the level measured in the control group (p < 0.0001). Furthermore, the age-adjusted MNL, BNMN, NPB, and NBUD frequencies in the patients were significantly higher than those obtained in the control group. We observed that the frequency of MN in patients was positively correlated with NBUD frequency which may indicate a common mechanism for these biomarkers in the patient group. We found, for the first time, that there were statistically significant higher levels of MN, NPB, and NBUD in sensorineural hearing loss patients. Since the markers we evaluated were linked with crucial diseases, our findings might suggest that sensorineural hearing loss patients are susceptible to several crucial diseases, especially cancer. Furthermore, the results demonstrated the significance of the MN, NPB, and NBUD level and thus provides a potential marker for the diagnosis of congenital hearing loss patients.

Expression of α2-Na/K-ATPase in C57BL/6J Mice Inner Ear and Its Relationship with Age-related Hearing Loss.

K+ cycling in the cochlea is critical to maintain hearing. Many sodium-potassium pumps are proved to participate in K+ cycling, such as Na/K-ATPase. The α2-Na/K-ATPase is an important isoform of Na/K-ATPase. The expression of α2-Na/K-ATPase in the cochlea is not clear. In this study, we used C57BL/6 mice as a model of presbycusis and implemented immunohistochemistry staining and quantitative real time-PCR, and the α2-Na/K-ATPase expression pattern was confirmed in the inner ear. It was found α2-Na/K-ATPase was expressed widely in cochlea and its mRNA and protein expression was gradually reduced with aging (4-, 14-, 26- and 48-weeks old mice). We suspected that, the down-regulation of α2-Na/K-ATPase expression might be associated with the remodeling of K+ cycling, degeneration of morphological structure and decrease of hearing function in aging C57 mice. In conclusion, we speculated that the reduction of α2-Na/K-ATPase might play an important role in the pathogenesis of age-related hearing loss.

GRß Regulates Glucocorticoid Resistance in Sudden Sensorineural Hearing Loss.

BACKGROUND: In recent years, the incidence of sudden deafness has gradually increased, with a very limited understanding of its etiology and pathogenesis. Glucocorticoids are the first choice of the treatment, but some hormone-resistant patients are not sensitive to glucocorticoid therapy. The pathogenesis is not yet known. In this study, we aim to construct the HEI-OC1 cell line stably overexpressing Glucocorticoid Receptor Beta (GRß), and identify its exact role in the cases of glucocorticoidresistant sudden deafness. METHODS: We used the endotoxin lipopolysaccharide-stimulated cochlear hair cells (HEI-OC1) to investigate the relationship of inflammation factor IL-2, TNF alpha, and SRp30c with the high expression GRß. We built a stable GRß high expression HEI-OC1 cell line and clarified its effects on the therapeutic effect of dexamethasone. MTT assay, colony formation assay, CCK-8 assay, Western blot, and RT-qPCR were utilized for characterizations. RESULTS: Dexamethasone reduced the LPS-induced inflammatory response from HEI-OC1 cells (p<0.05), detected by MTT assay. Dexamethasone could protect HEI-OC1 cells, but its protective effect was weakened due to the transfection of SRp30c over-expression plasmid (p<0.05). The transfection of SRp30c over-expression plasmid in HEI-OC1 cells could elevate the expressions of GRß (p<0.05). CONCLUSION: We clarified the mechanisms of high expression of GRß in glucocorticoid-resistant sudden sensorineural hearing loss, and proved that the inhibition of SRp30c may act as a new treatment way of glucocorticoid-resistant sudden sensorineural hearing loss.

Regulator of G Protein Signalling 4 (RGS4) as a Novel Target for the Treatment of Sensorineural Hearing Loss.

We and others have previously identified signalling pathways associated with the adenosine A1 receptor (A1R) as important regulators of cellular responses to injury in the cochlea. We have shown that the "post-exposure" treatment with adenosine A1R agonists confers partial protection against acoustic trauma and other forms of sensorineural hearing loss (SNHL). The aim of this study was to determine if increasing A1R responsiveness to endogenous adenosine would have the same otoprotective effect. This was achieved by pharmacological targeting of the Regulator of G protein Signalling 4 (RGS4). RGS proteins inhibit signal transduction pathways initiated by G protein-coupled receptors (GPCR) by enhancing GPCR deactivation and receptor desensitisation. A molecular complex between RGS4 and neurabin, an intracellular scaffolding protein expressed in neural and cochlear tissues, is the key negative regulator of A1R activity in the brain. In this study, Wistar rats (6-8 weeks) were exposed to traumatic noise (110 dBSPL, 8-16 kHz) for 2 h and a small molecule RGS4 inhibitor CCG-4986 was delivered intratympanically in a Poloxamer-407 gel formulation for sustained drug release 24 or 48 h after noise exposure. Intratympanic administration of CCG-4986 48 h after noise exposure attenuated noise-induced permanent auditory threshold shifts by up to 19 dB, whilst the earlier drug administration (24 h) led to even better preservation of auditory thresholds (up to 32 dB). Significant improvement of auditory thresholds and suprathreshold responses was linked to improved survival of sensorineural tissues and afferent synapses in the cochlea. Our studies thus demonstrate that intratympanic administration of CCG-4986 can rescue cochlear injury and hearing loss induced by acoustic overexposure. This research represents a novel paradigm for the treatment of various forms of SNHL based on regulation of GPCR.

Adenosine A2B receptor: A pathogenic factor and a therapeutic target for sensorineural hearing loss.

Over 466 million people worldwide are diagnosed with hearing loss (HL). About 90% of HL cases are sensorineural HL (SNHL) with treatments limited to hearing aids and cochlear implants with no FDA-approved drugs. Intriguingly, ADA-deficient patients have been reported to have bilateral SNHL, however, its underlying cellular and molecular basis remain unknown. We report that Ada-/- mice, phenocopying ADA-deficient humans, displayed SNHL. Ada-/- mice cochlea with elevated adenosine caused substantial nerve fiber demyelination and mild hair cell loss. ADA enzyme therapy in these mice normalized cochlear adenosine levels, attenuated SNHL, and prevented demyelination. Additionally, ADA enzyme therapy rescued SNHL by restoring nerve fiber structure in Ada-/- mice post two-week drug withdrawal. Moreover, elevated cochlear adenosine in untreated mice was associated with enhanced Adora2b gene expression. Preclinically, ADORA2B-specific antagonist treatment in Ada-/- mice significantly improved HL, nerve fiber density, and myelin compaction. We also provided genetic evidence that ADORA2B is detrimental for age-related SNHL by impairing cochlear myelination in WT aged mice. Overall, understanding purinergic molecular signaling in SNHL in Ada-/- mice allows us to further discover that ADORA2B is also a pathogenic factor underlying aged-related SNHL by impairing cochlear myelination and lowering cochlear adenosine levels or blocking ADORA2B signaling are effective therapies for SNHL.