Role of mitochondrial dysfunction and oxidative stress in sensorineural hearing loss.

Sensorineural hearing loss (SNHL) can either be genetically inherited or acquired as a result of aging, noise exposure, or ototoxic drugs. Although the precise pathophysiological mechanisms underlying SNHL remain unclear, an overwhelming body of evidence implicates mitochondrial dysfunction and oxidative stress playing a central etiological role. With its high metabolic demands, the cochlea, particularly the sensory hair cells, stria vascularis, and spiral ganglion neurons, is vulnerable to the damaging effects of mitochondrial reactive oxygen species (ROS). Mitochondrial dysfunction and consequent oxidative stress in cochlear cells can be caused by inherited mitochondrial DNA (mtDNA) mutations (hereditary hearing loss and aminoglycoside-induced ototoxicity), accumulation of acquired mtDNA mutations with age (age-related hearing loss), mitochondrial overdrive and calcium dysregulation (noise-induced hearing loss and cisplatin-induced ototoxicity), or accumulation of ototoxic drugs within hair cell mitochondria (drug-induced hearing loss). In this review, we provide an overview of our current knowledge on the role of mitochondrial dysfunction and oxidative stress in the development of SNHL caused by genetic mutations, aging, exposure to excessive noise, and ototoxic drugs. We also explore the advancements in antioxidant therapies for the different forms of acquired SNHL that are being evaluated in preclinical and clinical studies.

Apoptotic vesicles resist oxidative damage in noise-induced hearing loss through activation of FOXO3a-SOD2 pathway.

BACKGROUND: Mesenchymal stem cell (MSC) transplantation is a promising therapeutic approach for noise-induced hearing loss (NIHL). As the indispensable role of apoptosis in MSC transplantation was raised, the benefits of MSC-derived apoptotic vesicles (apoVs) in several disease models have been proved. However, whether apoVs benefit in NIHL have not been studied yet. METHODS: Female CBA/J mice and HEI-OC1 cells were used in this study. Flow cytometry, nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM) were used to characterize apoVs. Proteomic analysis was used to identify function proteins in apoVs. Immunofluorescence was used to reveal distribution pattern. Auditory brainstem response (ABR) test was used to measure the effect of apoVs treatment. DCFH-DA staining and MitoSOX staining were used to indicate oxidative damage. Western-blot and qRT-PCR were used to study the signaling pathways. RESULTS: We found that apoVs can be endocytosed by hair cells through systemic administration. Importantly, apoVs administration effectively attenuated NIHL and reduced hair cell loss by resisting oxidative damage in vivo. Further, apoVs application activated forkhead box o3 (FOXO3a)-mitochondrial superoxide dismutase 2(SOD2) pathway, which may relate to signal transduction and activators of transcription 3 (STAT3) in apoVs. CONCLUSIONS: These findings uncovered the role of apoVs in preventing NIHL and resisting oxidative damage, indicating that apoVs is a promising way for inner ear delivery and a prospective cell-free therapy for NIHL.

HMGB1 accumulation in cytoplasm mediates noise-induced cochlear damage.

Damage-associated molecular pattern molecules (DAMPs) play a critical role in mediating cochlear cell death, which leads to noise-induced hearing loss (NIHL). High-mobility group box 1 (HMGB1), a prototypical DAMP released from cells, has been extensively studied in the context of various diseases. However, whether extracellular HMGB1 contributes to cochlear pathogenesis in NIHL and the potential signals initiating HMGB1 release from cochlear cells are not well understood. Here, through the transfection of the adeno-associated virus with HMGB1-HA-tag, we first investigated early cytoplasmic accumulation of HMGB1 in cochlear hair cells after noise exposure. We found that the cochlear administration of HMGB1-neutralizing antibody immediately after noise exposure significantly alleviated hearing loss and outer hair cells (OHCs) death induced by noise exposure. In addition, activation of signal transducer and activators of transcription 1 (STAT1) and cellular hyperacetylation were verified as potential canonical initiators of HMGB1 cytoplasmic accumulation. These findings reveal the adverse effects of extracellular HMGB1 on the cochlea and the potential signaling events mediating HMGB1 release in hair cells, indicating multiple potential pharmacotherapeutic targets for NIHL.

Treatment with the Ferroptosis Inhibitor Ferrostatin-1 Attenuates Noise-Induced Hearing Loss by Suppressing Ferroptosis and Apoptosis.

Hair cell death induced by excessive reactive oxygen species (ROS) has been identified as the major pathogenesis of noise-induced hearing loss (NIHL). Recent studies have demonstrated that cisplatin- and neomycin-induced ototoxicity can be alleviated by ferroptosis inhibitors. However, whether ferroptosis inhibitors have a protective effect against NIHL remains unknown. We investigated the protective effect of the ferroptosis inhibitor ferrostatin-1 (Fer-1) on NIHL in vivo in CBA/J mice and investigated the protective effect of Fer-1 on tert-butyl hydroperoxide (TBHP)-induced hair cell damage in vitro in cochlear explants and HEI-OC1 cells. We observed ROS overload and lipid peroxidation, which led to outer hair cell (OHC) apoptosis and ferroptosis, in the mouse cochlea after noise exposure. The expression level of apoptosis-inducing factor mitochondria-associated 2 (AIFM2) was substantially increased following elevation of the expression of its upstream protein P53 after noise exposure. The ferroptosis inhibitor Fer-1was demonstrated to enter the inner ear after the systemic administration. Administration of Fer-1 significantly alleviated noise-induced auditory threshold elevation and reduced the loss of OHCs, inner hair cell (IHC) ribbon synapses, and auditory nerve fibers (ANFs) caused by noise. Mechanistically, Fer-1 significantly reduced noise- and TBHP-induced lipid peroxidation and iron accumulation in hair cells, alleviating ferroptosis in cochlear cells consequently. Furthermore, Fer-1 treatment decreased the levels of TfR1, P53, and AIFM2. These results suggest that Fer-1 exerted its protective effects by scavenging of ROS and inhibition of TfR1-mediated ferroptosis and P53-AIFM2 signaling pathway-mediated apoptosis. Our findings suggest that Fer-1 is a promising drug for treating NIHL because of its ability to inhibit noise-induced hair cell apoptosis and ferroptosis, opening new avenues for the treatment of NIHL.

A PRMT5 inhibitor protects against noise-induced hearing loss by alleviating ROS accumulation.

The aim of this study was to investigate the effect of LLY-283, a selective inhibitor of protein arginine methyltransferase 5 (PRMT5), on a noise-induced hearing loss (NIHL) mouse model and to identify a potential target for a therapeutic intervention against NIHL. Eight-week-old male C57BL/6 mice were used. The auditory brainstem response was measured 2 days after noise exposure. The apoptosis of hair cells (HCs) was detected by caspase-3/7 staining, whereas the accumulation of reactive oxygen species (ROS) was measured by 4-HNE staining. We demonstrated that the death of HCs and loss of cochlear synaptic ribbons induced by noise exposure could be significantly reduced by the presence of LLY-283. LLY-283 pretreatment before noise exposure notably decreased 4-HNE and caspase-3/7 levels in the cochlear HCs. We also noticed that the number of spiral ganglion neurons (SGNs) was notably increased after LLY-283 pretreatment. Furthermore, we showed that LLY-283 could increase the expression level of p-AKT in the SGNs. The underlying mechanism involves alleviation of ROS accumulation and activation of the PI3K/AKT pathway, indicating that LLY-283 might be a potential candidate for therapeutic intervention against NIHL.

Cochlin Deficiency Protects Aged Mice from Noise-Induced Hearing Loss.

Several studies have shown that type IV fibrocytes, located in the spiral ligament, degenerate first after noise exposure. Interestingly, this is the region where Coch expression is most abundant. As it is suggested that cochlin plays a role in our innate immune system, our goal is to investigate hearing thresholds and inner ear inflammation after noise exposure in Coch knockout (Coch-/-) mice compared to Coch wildtype (Coch+/+) mice. Animals were randomly allocated to a noise exposure group and a control group. Vestibular and auditory testing was performed at 48 h and one week after noise exposure. Whole mount staining and cryosectioning of the cochlea was performed in order to investigate hair cells, spiral ganglion neurons, inner ear inflammation, Coch expression and fibrocyte degeneration. Hearing assessment revealed that Coch+/+ mice had significantly larger threshold shifts than Coch-/- mice after noise exposure. We were unable to identify any differences in hair cells, neurons, fibrocytes and influx of macrophages in the inner ear between both groups. Interestingly, Coch expression was significantly lower in the group exposed to noise. Our results indicate that the absence of Coch has a protective influence on hearing thresholds after noise exposure, but this is not related to reduced inner ear inflammation in the knockout.

Estradiol Protects against Noise-Induced Hearing Loss and Modulates Auditory Physiology in Female Mice.

Recent studies have identified sex-differences in auditory physiology and in the susceptibility to noise-induced hearing loss (NIHL). We hypothesize that 17ß-estradiol (E2), a known modulator of auditory physiology, may underpin sex-differences in the response to noise trauma. Here, we gonadectomized B6CBAF1/J mice and used a combination of electrophysiological and histological techniques to study the effects of estrogen replacement on peripheral auditory physiology in the absence of noise exposure and on protection from NIHL. Functional analysis of auditory physiology in gonadectomized female mice revealed that E2-treatment modulated the peripheral response to sound in the absence of changes to the endocochlear potential compared to vehicle-treatment. E2-replacement in gonadectomized female mice protected against hearing loss following permanent threshold shift (PTS)- and temporary threshold shift (TTS)-inducing noise exposures. Histological analysis of the cochlear tissue revealed that E2-replacement mitigated outer hair cell loss and cochlear synaptopathy following noise exposure compared to vehicle-treatment. Lastly, using fluorescent in situ hybridization, we demonstrate co-localization of estrogen receptor-2 with type-1C, high threshold spiral ganglion neurons, suggesting that the observed protection from cochlear synaptopathy may occur through E2-mediated preservation of these neurons. Taken together, these data indicate the estrogen signaling pathways may be harnessed for the prevention and treatment of NIHL.

AZD5438-PROTAC: A selective CDK2 degrader that protects against cisplatin- and noise-induced hearing loss.

Cyclin-dependent kinase 2 (CDK2) is a potential therapeutic target for the treatment of hearing loss and cancer. Previously, we identified AZD5438 and AT7519-7 as potent inhibitors of CDK2, however, they also targeted additional kinases, leading to unwanted toxicities. Proteolysis Targeting Chimeras (PROTACs) are a new promising class of small molecules that can effectively direct specific proteins to proteasomal degradation. Herein we report the design, synthesis, and characterization of PROTACs of AT7519-7 and AZD5438 and the identification of PROTAC-8, an AZD5438-PROTAC, that exhibits selective, partial CDK2 degradation. Furthermore, PROTAC-8 protects against cisplatin ototoxicity and kainic acid excitotoxicity in zebrafish. Molecular dynamics simulations reveal the structural requirements for CDK2 degradation. Together, PROTAC-8 is among the first-in-class PROTACs with in vivo therapeutic activities and represents a new lead compound that can be further developed for better efficacy and selectivity for CDK2 degradation against hearing loss and cancer.

Primed to die: an investigation of the genetic mechanisms underlying noise-induced hearing loss and cochlear damage in homozygous Foxo3-knockout mice.

The prevalence of noise-induced hearing loss (NIHL) continues to increase, with limited therapies available for individuals with cochlear damage. We have previously established that the transcription factor FOXO3 is necessary to preserve outer hair cells (OHCs) and hearing thresholds up to two weeks following mild noise exposure in mice. The mechanisms by which FOXO3 preserves cochlear cells and function are unknown. In this study, we analyzed the immediate effects of mild noise exposure on wild-type, Foxo3 heterozygous (Foxo3+/-), and Foxo3 knock-out (Foxo3-/-) mice to better understand FOXO3's role(s) in the mammalian cochlea. We used confocal and multiphoton microscopy to examine well-characterized components of noise-induced damage including calcium regulators, oxidative stress, necrosis, and caspase-dependent and caspase-independent apoptosis. Lower immunoreactivity of the calcium buffer Oncomodulin in Foxo3-/- OHCs correlated with cell loss beginning 4 h post-noise exposure. Using immunohistochemistry, we identified parthanatos as the cell death pathway for OHCs. Oxidative stress response pathways were not significantly altered in FOXO3's absence. We used RNA sequencing to identify and RT-qPCR to confirm differentially expressed genes. We further investigated a gene downregulated in the unexposed Foxo3-/- mice that may contribute to OHC noise susceptibility. Glycerophosphodiester phosphodiesterase domain containing 3 (GDPD3), a possible endogenous source of lysophosphatidic acid (LPA), has not previously been described in the cochlea. As LPA reduces OHC loss after severe noise exposure, we treated noise-exposed Foxo3-/- mice with exogenous LPA. LPA treatment delayed immediate damage to OHCs but was insufficient to ultimately prevent their death or prevent hearing loss. These results suggest that FOXO3 acts prior to acoustic insult to maintain cochlear resilience, possibly through sustaining endogenous LPA levels.

Ultrasound Microbubbles Enhance the Efficacy of Insulin-Like Growth Factor-1 Therapy for the Treatment of Noise-Induced Hearing Loss.

The application of insulin-like growth factor 1 (IGF-1) to the round window membrane (RWM) is an emerging treatment for inner ear diseases. RWM permeability is the key factor for efficient IGF-1 delivery. Ultrasound microbubbles (USMBs) can increase drug permeation through the RWM. In the present study, the enhancing effect of USMBs on the efficacy of IGF-1 application and the treatment effect of USMB-mediated IGF-1 delivery for noise-induced hearing loss (NIHL) were investigated. Forty-seven guinea pigs were assigned to three groups: the USM group, which received local application of recombinant human IGF-1 (rhIGF-1, 10 µg/µL) following application of USMBs to the RWM; the RWS group, which received IGF-1 application alone; and the saline-treated group. The perilymphatic concentration of rhIGF-1 in the USM group was 1.95- and 1.67- fold of that in the RWS group, 2 and 24 h after treatment, respectively. After 5 h of 118 dB SPL noise exposure, the USM group had the lowest threshold shift in auditory brainstem response, least loss of cochlear outer hair cells, and least reduction in the number of synaptic ribbons on postexposure day 28 among the three groups. The combination of USMB and IGF-1 led to a better therapeutic response to NIHL. Two hours after treatment, the USM group had significantly higher levels of Akt1 and Mapk3 gene expression than the other two groups. The most intense immunostaining for phosphor-AKT and phospho-ERK1/2 was detected in the cochlea in the USM group. These results suggested that USMB can be applied to enhance the efficacy of IGF-1 therapy in the treatment of inner ear diseases.

Effect and Biocompatibility of a Cross-Linked Hyaluronic Acid and Polylactide-co-glycolide Microcapsule Vehicle in Intratympanic Drug Delivery for Treating Acute Acoustic Trauma.

The treatment of acute hearing loss is clinically challenging due to the low efficacy of drug delivery into the inner ear. Local intratympanic administration of dexamethasone (D) and insulin-like growth factor 1 (IGF1) has been proposed for treatment, but they do not persist in the middle ear because they are typically delivered in fluid form. We developed a dual-vehicle drug delivery system consisting of cross-linked hyaluronic acid and polylactide-co-glycolide microcapsules. The effect and biocompatibility of the dual vehicle in delivering D and IGF1 were evaluated using an animal model of acute acoustic trauma. The dual vehicle persisted 10.9 times longer (8.7 days) in the middle ear compared with the control (standard-of-care vehicle, 0.8 days). The dual vehicle was able to sustain drug release over up to 1 to 2 months when indocyanine green was loaded as the drug. One-third of the animals experienced an inflammatory adverse reaction. However, it was transient with no sequelae, which was validated by micro CT findings, endoscopic examination, and histological assessment. Hearing restoration after acoustic trauma was satisfactory in both groups, which was further supported by comparable numbers of viable hair cells. Overall, the use of a dual vehicle for intratympanic D and IGF1 delivery may maximize the effect of drug delivery to the target organ because the residence time of the vehicle is prolonged.

Overexpression of the receptor for advanced glycation end-products in the auditory cortex of rats with noise-induced hearing loss.

BACKGROUND: The receptor for advanced glycation end-products (RAGE) is involved in neuroinflammation. This study investigated the changes in RAGE expression following noise-induced hearing loss. METHODS: Three-week-old female Sprague-Dawley rats were exposed to 115 dB SPL white noise for 4 h daily for 3 d (noise group, n = 16). In parallel, age and sex-matched control rats were raised under standard conditions without noise exposure (control group, n = 16). After 2 h (noise immediate, n = 8) and 4 wk (noise 4-week, n = 8) of noise exposure, the auditory cortex was harvested and cytoplasmic and nuclear fractions were isolated. The gene expression levels of tumor necrosis factor alpha (TNF-α), interleukin 6 (IL6), interleukin 1 beta (IL1ß), nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), and RAGE were evaluated using real-time reverse transcription polymerase chain reaction. The protein expression levels of nuclear RAGE and cytosolic RAGE were evaluated using western blotting. Additionally, matrix metalloproteinase 9 (MMP9) was pharmacologically inhibited in the noise immediate group, and then nuclear and cytosolic RAGE expression levels were evaluated. RESULTS: The noise immediate and noise 4-week groups exhibited increased auditory thresholds at 4, 8, 16, and 32 kHz frequencies. The genes encoding the pro-inflammatory cytokines TNF-α, IL6, IL1ß, and NF- κB were increased 3.74, 1.63, 6.42, and 6.23-fold in the noise immediate group, respectively (P = 0.047, 0.043, 0.044, and 0.041). RAGE mRNA expression was elevated 1.42-fold in the noise 4-week group (P = 0.032). Cytosolic RAGE expression was increased 1.76 and 6.99-fold in the noise immediate and noise 4-week groups, respectively (P = 0.04 and 0.03). Nuclear RAGE expression was comparable between the noise and control groups. matrix metalloproteinase 9 (MMP9) inhibition reduced cytosolic RAGE expression in the noise immediate group (P = 0.004). CONCLUSIONS: Noise exposure increased the expression of cytosolic RAGE in the auditory cortex and upregulated pro-inflammatory genes, but this response could be alleviated by MMP9 inhibition.

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.

New Insights on the Effect of TNF Alpha Blockade by Gene Silencing in Noise-Induced Hearing Loss.

Noise exposure represents the second most common cause of acquired sensorineural hearing loss and we observed that tumor necrosis factor α (TNFα) was involved in this context. The effect of Tnfα gene silencing on the expression profile related to the TNFα metabolic pathway in an experimental model of noise-induced hearing loss had not previously been studied. METHODS: Single ears of Wistar rats were pretreated with Tnfα small interfering RNA (siRNA) by trans-tympanic administration 24 h before they were exposed to white noise (120 dBSPL for three hours). After 24 h of noise exposure, we analyzed the electrophysiological threshold and the amplitude of waves I, II, III, and IV in the auditory brain response click. In addition, qRT-PCR was performed to evaluate the TNFα metabolic pathway in the ears submitted or not to gene silencing. RESULTS: Preservation of the electrophysiological threshold and the amplitude of waves was observed in the ears submitted to gene silencing compared to the ears not treated. Increased anti-apoptotic gene expression and decreased pro-apoptotic gene expression were found in the treated ears. CONCLUSION: Our results allow us to suggest that the blockade of TNFα by gene silencing was useful to prevent noise-induced hearing loss.

Cis-element architecture of Nrf2-sMaf heterodimer binding sites and its relation to diseases.

Cellular detoxication is essential for health because it provides protection against various chemicals and xenobiotics. The KEAP1-NRF2 system is important for cellular defense against oxidative and electrophilic stresses as NRF2 activates the transcription of an array of cytoprotective genes, including drug-metabolizing and antioxidant enzymes, in a stress-dependent manner. The CNC family of transcription factors, including NRF2, form heterodimers with small Maf (sMaf) proteins and bind to consensus DNA sequences that have been referred to as antioxidant response element, electrophile response element, or NF-E2-binding element. These sequences are now collectively called CNC-sMaf binding element (CsMBE). In addition to forming a heterodimer with CNC proteins, sMaf proteins can form homodimers and recognize regulatory motifs called Maf recognition element (MARE). Although the CsMBE sequence substantially overlaps with that of MARE, the sequences differ. NRF2 selectively recognizes CsMBE, which is critical for cytoprotection. Recent advances in high-throughput sequencing and population-scale genome analysis provide new insights into the transcriptional regulation involved in the stress response. The integration of a genome-wide map of NRF2 occupancy with disease-susceptibility loci reveals the associations between polymorphisms in CsMBE and disease risk, information useful for the personalized medicine of the future.

The TLR-4/NF-κB signaling pathway activation in cochlear inflammation of rats with noise-induced hearing loss.

The TLR-4/NF-κB signaling pathway is involved in innate immunity and inflammation induced by trauma. The present study aimed to investigate possible TLR-4/NF-κB signaling pathway activation in the cochlea associated with acoustic trauma that might induce cochlear inflammation. A total of 72 rats were exposed to white noise at 120 dB SPL for 8 h per day repeated over 2 successive days. Auditory brainstem responses (ABR) were measured in animals before noise exposure and 0 d (PE0), 1 d (PE1), 3 d (PE3), 7 d (PE7), and 14 d (PE14) after noise exposure. At each defined time point, animals were sacrificed, and cochleae were collected to evaluate the expression levels of TLR4, MyD88, cytoplasmic NF-κB p65, IκBα, TNF-α, and IL-1ß using western blotting and NF-κB p65 transcriptional activity using an NF-κB p65 Transcription Factor Assay Kit. Cochlear localizations of TLR-4, TNF-α and IL-1ß were analyzed using immunohistochemistry in paraffin-embedded slices. The nuclear translocation of NF-κB p65 was evaluated using immunofluorescence staining in paraffin-embedded slices. DNA fragmentation was measured with a TUNEL assay in paraffin-embedded slices. We found a stable permanent threshold shift after noise exposure. After noise exposure, expression levels of TLR-4, MyD88, IκBα, TNF-α, and IL-1ß were significantly upregulated (PE3); DNA binding activity of NF-κB p65 was also significantly enhanced (PE3), while the cytoplasmic NF-κB p65 levels were unchanged. TLR-4, TNF-α, and IL-1ß immunostaining intensities were substantially enhanced in spiral ganglion cells and spiral ligament fibrocytes after noise exposure (PE3). In conclusion, the results of this study indicate that the TLR-4/NF-κB signaling pathway is activated in noise-exposed cochleae and that it participates in noise-induced cochlear inflammation.

Circadian integration of inflammation and glucocorticoid actions: Implications for the cochlea.

Auditory function has been shown to be influenced by the circadian system. Increasing evidence point towards the regulation of inflammation and glucocorticoid actions by circadian rhythms in the cochlea. Yet, how these three systems (circadian, immune and endocrine) converge to control auditory function remains to be established. Here we review the knowledge on immune and glucocorticoid actions, and how they interact with the circadian and the auditory system, with a particular emphasis on cochlear responses to noise trauma. We propose a multimodal approach to understand the mechanisms of noise-induced hearing loss by integrating the circadian, immune and endocrine systems into the bearings of the cochlea. Considering the well-established positive impact of chronotherapeutic approaches in the treatment of cardiovascular, asthma and cancer, an increased knowledge on the mechanisms where circadian, immune and glucocorticoids meet in the cochlea may improve current treatments against hearing disorders.

[The expression of P19ink4d in the pathogenesis and development of hearing loss].

Objective: This study aimed to investigate the P19ink4d expression in cochlea of mice model with noise induced hearing loss and the role of P19ink4d in the degeneration of inner ear cells. It also searched for P19ink4d gene alterations in patients with profound sensorineural deafness.Method: CBA/J mice were exposed to broad band noise at 101 dB SPL for 2 hours, auditory brainstem response (ABR) were examined to confirm noise lead to the permanent threshold shift. Immunohistochemical staining, Western blotting, and real-time polymerase chain reaction (PCR) were performed on cochlear tissues, to elucidate changes in P19ink4d expression in mice after noise exposure. For clinical evaluation, 400 children from unrelated families with severe or profound sensorineural hearing loss (SNHL) were recruited, genomic DNA was obtained from the patients and was subjected to DNA microarray to screen mutations in 4 most common genes. The sample that carried none of the common mutant alleles were subjected to PCR and sequenced to detect mutations in P19ink4d gene.Result: The ABR threshold shift of mice in the experimental group significantly increased after noise exposure and was higher than that in the null-noise group. The ABR of 1 day post noise was least among experimental groups and there is no statistical different between ABR of 7 days and 14 days post noise. The missing of outer hair cells occurred after noise exposure, while the inner hair cells hardly miss. It was found that the P19ink4d expression increased significantly in the inner ear cells 3 hours after noise exposure, then recovered in 24 hours. Western blot indicated that the amount of P19ink4d increased transitorily 3-6 h after the noise. However, no mutation existed within the coding exons of P19ink4d in the patients with profound sensorineural deafness.Conclusion: The results support the concept that P19ink4d may play an important role in the pathogenesis and development of noise induced hearing loss.

Avocado Oil Extract Modulates Auditory Hair Cell Function through the Regulation of Amino Acid Biosynthesis Genes.

Sensorineural hearing loss (SNHL) is one of the most common causes of disability, affecting over 466 million people worldwide. However, prevention or therapy of SNHL has not been widely studied. Avocado oil has shown many health benefits but it has not yet been studied in regards to SNHL. Therefore, we aimed to investigate the efficacy of avocado oil on SNHL in vitro and in vivo and elucidate its mode of action. For the present study, we used enhanced functional avocado oil extract (DKB122). DKB122 led to recovery of otic hair cells in zebrafish after neomycin-induced otic cell damage. Also, DKB122 improved auditory sensory transmission function in a mouse model of noise induced-hearing loss and protected sensory hair cells in the cochlea. In addition, RNA sequencing was performed to elucidate the mechanism involved. KEGG pathway enrichment analysis of differentially expressed genes showed that DKB122 protected House Ear Institute-Organ of Corti 1 (HEI-OC1) cells against neomycin-related alterations in gene expression due to oxidative stress, cytokine production and protein synthesis.

Use of non-invasive measures to predict cochlear synapse counts.

Cochlear synaptopathy, the loss of synaptic connections between inner hair cells (IHCs) and auditory nerve fibers, has been documented in animal models of aging, noise, and ototoxic drug exposure, three common causes of acquired sensorineural hearing loss in humans. In each of these models, synaptopathy begins prior to changes in threshold sensitivity or loss of hair cells; thus, this underlying injury can be hidden behind a normal threshold audiogram. Since cochlear synaptic loss cannot be directly confirmed in living humans, non-invasive assays will be required for diagnosis. In animals with normal auditory thresholds, the amplitude of wave 1 of the auditory brainstem response (ABR) is highly correlated with synapse counts. However, synaptopathy can also co-occur with threshold elevation, complicating the use of the ABR alone as a diagnostic measure. Using an age-graded series of mice and a partial least squares regression approach to model structure-function relationships, this study shows that the combination of a small number of ABR and distortion product otoacoustic emission (DPOAE) measurements can predict synaptic ribbon counts at various cochlear frequencies to within 1-2 synapses per IHC of their true value. In contrast, the model, trained using the age-graded series of mice, overpredicted synapse counts in a small sample of young noise-exposed mice, perhaps due to differences in the underlying pattern of damage between aging and noise-exposed mice. These results provide partial validation of a noninvasive approach to identify synaptic/neuronal loss in humans using ABRs and DPOAEs.