RESUMEN
Gasdermin E (GSDME), a member of the gasdermin protein family, is associated with post-lingual hearing loss. All GSDME pathogenic mutations lead to skipping exon 8; however, the molecular mechanisms underlying hearing loss caused by GSDME mutants remain unclear. GSDME was recently identified as one of the mediators of programmed cell death, including apoptosis and pyroptosis. Therefore, in this study, we injected mice with GSDME mutant (MT) and examined the expression levels to assess its effect on hearing impairment. We observed loss of hair cells in the organ of Corti and spiral ganglion neurons. Further, the N-terminal release from the GSDME mutant in HEI-OC1 cells caused pyroptosis, characterized by cell swelling and rupture of the plasma membrane, releasing lactate dehydrogenase and cytokines such as interleukin-1ß. We also observed that the N-terminal release from GSDME mutants could permeabilize the mitochondrial membrane, releasing cytochromes and activating the mitochondrial apoptotic pathway, thereby generating possible positive feedback on the cleavage of GSDME. Furthermore, we found that treatment with disulfiram or dimethyl fumarate might inhibit pyroptosis and apoptosis by inhibiting the release of GSDME-N from GSDME mutants. In conclusion, this study elucidated the molecular mechanism associated with hearing loss caused by GSDME gene mutations, offering novel insights for potential treatment strategies.
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Apoptosis , Piroptosis , Piroptosis/genética , Animales , Ratones , Mutación con Ganancia de Función , Pérdida Auditiva/genética , Pérdida Auditiva/patología , Humanos , Ganglio Espiral de la Cóclea/metabolismo , Ganglio Espiral de la Cóclea/patología , Órgano Espiral/metabolismo , Órgano Espiral/patología , Células Ciliadas Auditivas/metabolismo , Células Ciliadas Auditivas/patología , GasderminasRESUMEN
The apex or apical region of the cochlear spiral within the inner ear encodes for low-frequency sounds. The disposition of sensory hair cells on the organ of Corti is largely variable in the apical region of mammals, and it does not necessarily follow the typical three-row pattern of outer hair cells (OHCs). As most underwater noise sources contain low-frequency components, we expect to find most lesions in the apical region of the cochlea of toothed whales, in cases of permanent noise-induced hearing loss. To further understand how man-made noise might affect cetacean hearing, there is a need to describe normal morphological features of the apex and document interspecific anatomic variations in cetaceans. However, distinguishing between apical normal variability and hair cell death is challenging. We describe anatomical features of the organ of Corti of the apex in 23 ears from five species of toothed whales (harbor porpoise Phocoena phocoena, spinner dolphin Stenella longirostris, pantropical spotted dolphin Stenella attenuata, pygmy sperm whale Kogia breviceps, and beluga whale Delphinapterus leucas) by scanning electron microscopy and immunofluorescence. Our results showed an initial region where the lowest frequencies are encoded with two or three rows of OHCs, followed by the typical configuration of three OHC rows and three rows of supporting Deiters' cells. Whenever two rows of OHCs were detected, there were usually only two corresponding rows of supporting Deiters' cells, suggesting that the number of rows of Deiters' cells is a good indicator to distinguish between normal and pathological features.
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Cóclea , Pérdida Auditiva Provocada por Ruido , Animales , Biomarcadores/metabolismo , Cóclea/patología , Células Ciliadas Auditivas Externas/metabolismo , Células Ciliadas Auditivas Externas/patología , Pérdida Auditiva Provocada por Ruido/metabolismo , Humanos , Órgano Espiral/patología , BallenasRESUMEN
OBJECTIVE: Age-related hearing loss (AHL), characterized by degeneration of cochlea structures, is the most common sensory disorder among the elderly worldwide. The calcium channel is considered to contribute to normal hearing. However, the role of the T-type voltage-activated calcium channel, Cav3.1, remains unclear in AHL. Here, we investigate the age-related change of Cav3.1 expression in the cochlea and D-gal-induced senescent HEI-OC1 cells. METHODS: Cochleae from C57BL/6 mice at 2 months and 12 months of age were assessed. Senescence in House Ear Institute-Organ of Corti 1 (HEI-OC1) cells was induced by D-gal treatment. The immunofluorescence technique was employed to investigate the distribution of Cav3.1 in vivo and in vitro. Quantitative assessment was achieved by Western blotting and real-time PCR. RESULTS: In comparison with 2-month-old animals, 12-month old C57BL/6 mice exhibited great loss of hair cells and elevated auditory brainstem threshold. The Cav3.1 was located in hair cells, spiral ganglion cells, lateral walls, and the expression of Cav3.1 protein and mRNA decreased in the aged cochleae. D-gal-induced senescence assay confirmed the down-regulation of Cav3.1 expression in senescent HEI-OC1 cells. CONCLUSION: Our results show that age-related down-regulated expression of Cav3.1 in the cochleae is associated with AHL and may contribute to the pathogenesis of AHL.
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Canales de Calcio Tipo T/genética , Cóclea/metabolismo , Presbiacusia/genética , Animales , Cóclea/diagnóstico por imagen , Cóclea/patología , Modelos Animales de Enfermedad , Regulación de la Expresión Génica/genética , Células Ciliadas Auditivas/metabolismo , Células Ciliadas Auditivas/patología , Humanos , Ratones , Órgano Espiral/diagnóstico por imagen , Órgano Espiral/metabolismo , Órgano Espiral/patología , Presbiacusia/patología , Ganglio Espiral de la Cóclea/diagnóstico por imagen , Ganglio Espiral de la Cóclea/metabolismo , Ganglio Espiral de la Cóclea/patologíaRESUMEN
Hearing loss is a debilitating disease that affects 10% of adults worldwide. Most sensorineural hearing loss is caused by the loss of mechanosensitive hair cells in the cochlea, often due to aging, noise, and ototoxic drugs. The identification of genes that can be targeted to slow aging and reduce the vulnerability of hair cells to insults is critical for the prevention of sensorineural hearing loss. Our previous cell-specific transcriptome analysis of adult cochlear hair cells and supporting cells showed that Clu, encoding a secreted chaperone that is involved in several basic biological events, such as cell death, tumor progression, and neurodegenerative disorders, is expressed in hair cells and supporting cells. We generated Clu-null mice (C57BL/6) to investigate its role in the organ of Corti, the sensory epithelium responsible for hearing in the mammalian cochlea. We showed that the deletion of Clu did not affect the development of hair cells and supporting cells; hair cells and supporting cells appeared normal at 1 month of age. Auditory function tests showed that Clu-null mice had hearing thresholds comparable to those of wild-type littermates before 3 months of age. Interestingly, Clu-null mice displayed less hair cell and hearing loss compared to their wildtype littermates after 3 months. Furthermore, the deletion of Clu is protected against aminoglycoside-induced hair cell loss in both in vivo and in vitro models. Our findings suggested that the inhibition of Clu expression could represent a potential therapeutic strategy for the alleviation of age-related and ototoxic drug-induced hearing loss.
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Clusterina/deficiencia , Células Ciliadas Auditivas/fisiología , Pérdida Auditiva Sensorineural/prevención & control , Presbiacusia/prevención & control , Animales , Umbral Auditivo , Secuencia de Bases , Sistemas CRISPR-Cas , Senescencia Celular , Clusterina/biosíntesis , Clusterina/genética , Clusterina/fisiología , Sinergismo Farmacológico , Potenciales Evocados Auditivos del Tronco Encefálico , Furosemida/administración & dosificación , Furosemida/toxicidad , Células Ciliadas Auditivas/efectos de los fármacos , Pérdida Auditiva Sensorineural/inducido químicamente , Kanamicina/administración & dosificación , Kanamicina/toxicidad , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Técnicas de Cultivo de Órganos , Órgano Espiral/patología , Emisiones Otoacústicas Espontáneas , ARN Mensajero/biosíntesis , ARN Mensajero/genéticaRESUMEN
OBJECTIVES/HYPOTHESIS: Intracellular entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) depends on the interaction between its spike protein with the cellular receptor angiotensin-converting enzyme 2 (ACE2) and depends on Furin-mediated spike protein cleavage and spike protein priming by host cell proteases, including transmembrane protease serine 2 (TMPRSS2). As the expression of ACE2, TMPRSS2, and Furin in the middle and inner ear remain unclear, we analyzed the expression of these proteins in mouse ear tissues. STUDY DESIGN: Animal Research. METHODS: We performed immunohistochemical analysis to examine the distribution of ACE2, TMPRSS2, and Furin in the Eustachian tube, middle ear spaces, and cochlea of mice. RESULTS: ACE2 was present in the nucleus of the epithelium of the middle ear and Eustachian tube, as well as in some nuclei of the hair cells in the organ of Corti, in the stria vascularis, and the spiral ganglion cells. ACE2 was also expressed in the cytoplasm of the stria vascularis. TMPRSS2 was expressed in both the nucleus and cytoplasm in the middle spaces, with the expression being stronger in the nucleus in the mucosal epithelium of the middle ear spaces and Eustachian tube. TMPRSS2 was present in the cytoplasm in the organ of Corti and stria vascularis and in the nucleus and cytoplasm in the spiral ganglion. Furin was expressed in the cytoplasm in the middle ear spaces, Eustachian tube, and cochlea. CONCLUSIONS: ACE2, TMPRSS2, and Furin are diffusely present in the Eustachian tube, middle ear spaces, and cochlea, suggesting that these tissues are susceptible to SARS-CoV-2 infection. LEVEL OF EVIDENCE: NA Laryngoscope, 131:E2013-E2017, 2021.
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Enzima Convertidora de Angiotensina 2/genética , COVID-19/patología , Oído Interno/patología , Oído Medio/patología , Trompa Auditiva/patología , Furina/genética , Expresión Génica/genética , Serina Endopeptidasas/genética , Animales , Cóclea/patología , Epitelio/patología , Inmunohistoquímica , Ratones , Membrana Mucosa/patología , Órgano Espiral/patología , Ganglio Espiral de la Cóclea/patología , Estría Vascular/patología , Hueso Temporal/patologíaRESUMEN
Noise-induced hearing loss (NIHL) can lead to secondary changes that induce neural plasticity in the central auditory pathway. These changes include decreases in the number of synapses, the degeneration of auditory nerve fibers, and reorganization of the cochlear nucleus (CN) and inferior colliculus (IC) in the brain. This study investigated the role of microRNAs (miRNAs) in the neural plasticity of the central auditory pathway after acute NIHL. Male Sprague-Dawley rats were exposed to white band noise at 115 dB for 2 h, and the auditory brainstem response (ABR) and morphology of the organ of Corti were evaluated on days 1 and 3. Following noise exposure, the ABR threshold shift was significantly smaller in the day 3 group, while wave II amplitudes were significantly larger in the day 3 group compared to the day 1 group. The organ of Corti on the basal turn showed evidence of damage and the number of surviving outer hair cells was significantly lower in the basal and middle turn areas of the hearing loss groups relative to controls. Five and three candidate miRNAs for each CN and IC were selected based on microarray analysis and quantitative reverse transcription PCR (RT-qPCR). The data confirmed that even short-term acoustic stimulation can lead to changes in neuroplasticity. Further studies are needed to validate the role of these candidate miRNAs. Such miRNAs may be used in the early diagnosis and treatment of neural plasticity of the central auditory pathway after acute NIHL.
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Núcleo Coclear , Pérdida Auditiva Provocada por Ruido/metabolismo , Colículos Inferiores , MicroARNs/metabolismo , Plasticidad Neuronal , Animales , Núcleo Coclear/metabolismo , Núcleo Coclear/patología , Potenciales Evocados Auditivos del Tronco Encefálico , Colículos Inferiores/metabolismo , Colículos Inferiores/patología , Masculino , Órgano Espiral/patología , Ratas , Ratas Sprague-DawleyRESUMEN
Cells of the immune system are present in the adult cochlea and respond to damage caused by noise exposure. However, the types of immune cells involved and their locations within the cochlea are unclear. We used flow cytometry and immunostaining to reveal the heterogeneity of the immune cells in the cochlea and validated the presence of immune cell gene expression by analyzing existing single-cell RNA-sequencing (scRNAseq) data. We demonstrate that cell types of both the innate and adaptive immune system are present in the cochlea. In response to noise damage, immune cells increase in number. B, T, NK, and myeloid cells (macrophages and neutrophils) are the predominant immune cells present. Interestingly, immune cells appear to respond to noise damage by infiltrating the organ of Corti. Our studies highlight the need to further understand the role of these immune cells within the cochlea after noise exposure.
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Cóclea/inmunología , Cóclea/lesiones , Pérdida Auditiva Provocada por Ruido/inmunología , Inmunidad Adaptativa , Animales , Linfocitos B/inmunología , Linfocitos B/patología , Cóclea/patología , Modelos Animales de Enfermedad , Potenciales Evocados Auditivos del Tronco Encefálico/inmunología , Femenino , Pérdida Auditiva Provocada por Ruido/patología , Pérdida Auditiva Provocada por Ruido/fisiopatología , Inmunidad Innata , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/patología , Antígenos Comunes de Leucocito/metabolismo , Macrófagos/inmunología , Macrófagos/patología , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Transgénicos , Neutrófilos/inmunología , Neutrófilos/patología , Órgano Espiral/inmunología , Órgano Espiral/lesiones , Órgano Espiral/patología , RNA-Seq , Linfocitos T/inmunología , Linfocitos T/patologíaRESUMEN
Ear development requires the transcription factors ATOH1 for hair cell differentiation and NEUROD1 for sensory neuron development. In addition, NEUROD1 negatively regulates Atoh1 gene expression. As we previously showed that deletion of the Neurod1 gene in the cochlea results in axon guidance defects and excessive peripheral innervation of the sensory epithelium, we hypothesized that some of the innervation defects may be a result of abnormalities in NEUROD1 and ATOH1 interactions. To characterize the interdependency of ATOH1 and NEUROD1 in inner ear development, we generated a new Atoh1/Neurod1 double null conditional deletion mutant. Through careful comparison of the effects of single Atoh1 or Neurod1 gene deletion with combined double Atoh1 and Neurod1 deletion, we demonstrate that NEUROD1-ATOH1 interactions are not important for the Neurod1 null innervation phenotype. We report that neurons lacking Neurod1 can innervate the flat epithelium without any sensory hair cells or supporting cells left after Atoh1 deletion, indicating that neurons with Neurod1 deletion do not require the presence of hair cells for axon growth. Moreover, transcriptome analysis identified genes encoding axon guidance and neurite growth molecules that are dysregulated in the Neurod1 deletion mutant. Taken together, we demonstrate that much of the projections of NEUROD1-deprived inner ear sensory neurons are regulated cell-autonomously.
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Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Células Ciliadas Auditivas/metabolismo , Fibras Nerviosas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Animales , Apoptosis/genética , Axones/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Diferenciación Celular/genética , Epitelio/metabolismo , Eliminación de Gen , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Células Ciliadas Auditivas/patología , Células Ciliadas Auditivas/ultraestructura , Ratones Noqueados , Modelos Biológicos , Mutación/genética , Proteínas del Tejido Nervioso/genética , Órgano Espiral/patología , Factores de Transcripción SOXB1/metabolismo , Ganglio Espiral de la Cóclea/metabolismoRESUMEN
Acid-base homeostasis is critical for normal growth, development, and hearing function. The sodium-hydrogen exchanger 6 (NHE6), a protein mainly expressed in early and recycling endosomes, plays an important role in regulating organellar pH. Mutations in NHE6 cause complex, slowly progressive neurodegeneration. Little is known about NHE6 function in the mouse cochlea. Here, we found that all NHE isoforms were expressed in wild-type (WT) mouse cochlea. Nhe6 knockout (KO) mice showed significant hearing loss compared to WT littermates. Immunohistochemistry in WT mouse cochlea showed that Nhe6 was localized in the organ of Corti (OC), spiral ganglion (SG), stria vascularis (SV), and afferent nerve fibres. The middle and the inner ears of WT and Nhe6 KO mice were not different morphologically. Given the putative role of NHE6 in early endosomal function, we examined Rab GTPase expression in early and late endosomes. We found no change in Rab5, significantly lower Rab7, and higher Rab11 levels in the Nhe6 KO OC, compared to WT littermates. Because Rabs mediate TrkB endosomal signalling, we evaluated TrkB phosphorylation in the OCs of both strains. Nhe6 KO mice showed significant reductions in TrkB and Akt phosphorylation in the OC. In addition, we examined genes used as markers of SG type I (Slc17a7, Calb1, Pou4f1, Cal2) and type II neurons (Prph, Plk5, Cacna1g). We found that all marker gene expression levels were significantly elevated in the SG of Nhe6 KO mice, compared to WT littermates. Anti-neurofilament factor staining showed axon loss in the cochlear nerves of Nhe6 KO mice compared to WT mice. These findings indicated that BDNF/TrkB signalling was disrupted in the OC of Nhe6 KO mice, probably due to TrkB reduction, caused by over acidification in the absence of NHE6. Thus, our findings demonstrated that NHEs play important roles in normal hearing in the mammalian cochlea.
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Endosomas/metabolismo , Pérdida Auditiva/metabolismo , Neuronas/fisiología , Órgano Espiral/metabolismo , Intercambiadores de Sodio-Hidrógeno/metabolismo , Ganglio Espiral de la Cóclea/fisiología , Animales , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Pérdida Auditiva/genética , Concentración de Iones de Hidrógeno , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Órgano Espiral/patología , Fosforilación , Proteínas Tirosina Quinasas/metabolismo , Transducción de Señal , Intercambiadores de Sodio-Hidrógeno/genéticaRESUMEN
Sensorineural hearing loss due to aging, noise exposure, trauma or drug ototoxicity is irreversible because cochlear hair cells and neurons cannot regenerate. Recently, therapeutic strategies involving nanoparticles have been developed as innovative drug delivery systems. Thermodynamically stable liquid crystalline nanoparticles based on the polar lipid glycerol monooleate (GMO NP, cubosomes), nontoxic and able to encapsulate both hydrophilic and hydrophobic compounds, were produced and tested for biocompatibility in an immortalized Organ of Corti derived cell line (OC-k3), through cell viability and cytomorphological assays, and Western blot expression profiles of apoptotic markers. Overall, the GMO NP were biocompatible in OC-k3 at the doses and time tested, supporting previous data obtained in a neuronal cell line (PC12). The results encourage further tests on GMO NP-mediated drug release with improved target specificity and could be useful to develop innovative therapies against sensorineural hearing loss.
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Materiales Biocompatibles/toxicidad , Portadores de Fármacos , Glicéridos/toxicidad , Nanopartículas , Órgano Espiral/efectos de los fármacos , Animales , Materiales Biocompatibles/química , Línea Celular , Supervivencia Celular/efectos de los fármacos , Composición de Medicamentos , Glicéridos/química , Cristales Líquidos , Ratones , Órgano Espiral/metabolismo , Órgano Espiral/patología , Medición de RiesgoRESUMEN
Although human cytomegalovirus (HCMV) is a known cause of sensorineural hearing loss in infants with congenital HCMV (cCMV) infections, mechanisms that contribute to sensorineural hearing loss (SNHL) in infants with cCMV infection are not well defined. Using a murine model of CMV infection during auditory development, we have shown that peripheral infection of newborn mice with murine CMV (MCMV) results in focal infection of the cochlea and virus-induced cochlear inflammation. Approximately 50%-60% of infected mice exhibited increased auditory brainstem response (ABR) thresholds across a range of sound frequencies. Histological analyses of the cochlea in MCMV-infected mice with elevated ABR thresholds revealed preservation of hair cell (HC) number and morphology in the organ of Corti. In contrast, the number of spiral ganglion neurons (SGN), synapses, and neurites connecting the cochlear HC and SGN nerve terminals were decreased. Decreasing cochlear inflammation by corticosteroid treatment of MCMV-infected mice resulted in preservation of SGN and improved auditory function. These findings show that virus-induced cochlear inflammation during early auditory development, rather than direct virus-mediated damage, could contribute to histopathology in the cochlea and altered auditory function without significant loss of HCs in the sensory epithelium.
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Cóclea/inmunología , Pérdida Auditiva Sensorineural/inmunología , Audición/fisiología , Inflamación , Activación Viral , Animales , Animales Recién Nacidos , Cóclea/patología , Cóclea/virología , Citomegalovirus/genética , Infecciones por Citomegalovirus/patología , Infecciones por Citomegalovirus/virología , Modelos Animales de Enfermedad , Potenciales Evocados Auditivos del Tronco Encefálico , Femenino , Células Ciliadas Auditivas/patología , Pérdida Auditiva Sensorineural/patología , Pérdida Auditiva Sensorineural/virología , Inflamación/inmunología , Inflamación/patología , Masculino , Ratones , Ratones Endogámicos BALB C , Neuronas/patología , Órgano Espiral/patología , Ganglio Espiral de la Cóclea/patología , Sinapsis , Activación Viral/genéticaRESUMEN
OBJECTIVES: The aim of our study was to investigate the effects of folic acid on cisplatin-induced ototoxicity. MATERIALS AND METHODS: Thirty Wistar albino rats were divided into five groups. Group I received intraperitoneal cisplatin (IP) 10 mg/kg/day and IP folic acid 10 mg/kg/day; Group II received IP cisplatin 10 mg/kg/day and IP physiological saline; Group III received IP cisplatin 10 mg/kg/day and intratympanic (IT) folic acid 0.15 mL/day; Group IV received IP cisplatin 10 mg/kg/day and IT physiological saline; and Group V received IT folic acid 0.15 mL/day. Before and after drug administration, plasma homocysteine, folic acid levels, and auditory brainstem evoked responses (ABR) were measured. The rats were then sacrificed, and the inner ears were processed for electron microscopy. RESULTS: The differences of ABR thresholds in Group I compared to Group II were significantly smaller at 4 kHz, 8 kHz, and 16 kHz, whereas they were smaller but not statistically significant at 12 kHz in ABR. The differences of ABR thresholds in Group III compared to Group IV were significantly smaller at 12 kHz, and smaller but not statistically significant at 4 kHz, 8 kHz, and 16 kHz. Cisplatin treatment resulted in the degeneration of the cells of the organ of Corti, stria vascularis, and spiral ganglion. The cells of the organ of Corti, stria vascularis, and spiral ganglion showed a partially preserved morphology in both Group I and Group III. CONCLUSION: Our study results suggests that folic acid is a potential agent in preventing cisplatin-induced ototoxicity.
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Antineoplásicos/toxicidad , Cisplatino/toxicidad , Ácido Fólico/farmacología , Ototoxicidad/prevención & control , Complejo Vitamínico B/farmacología , Animales , Antineoplásicos/administración & dosificación , Cisplatino/administración & dosificación , Cóclea/patología , Esquema de Medicación , Potenciales Evocados Auditivos/fisiología , Potenciales Evocados Auditivos del Tronco Encefálico/fisiología , Ácido Fólico/metabolismo , Células Ciliadas Auditivas/ultraestructura , Pérdida Auditiva/prevención & control , Homocisteína/metabolismo , Masculino , Microscopía Electrónica , Órgano Espiral/patología , Ototoxicidad/patología , Ototoxicidad/fisiopatología , Ratas Wistar , Umbral Sensorial/fisiologíaRESUMEN
OBJECTIVES: This study aimed to evaluate the effect of quercetin on cochlear function and morphology, and its possible protective effect against acute cisplatin-induced ototoxicity in rats. MATERIALS AND METHODS: This prospective and controlled animal study was conducted on Wistar albino rats divided into four groups. Otoacoustic emission measures were performed three days after the first infiltration in Group 1 (saline), 2 (cisplatin), and 3 (quercetin). This interval was five days for Group 4 (cisplatin+quercetin). At the end of the study, the rats were decapitated with deep anesthesia, and histological changes in the cochleas were observed by light microscopy. RESULTS: Group 2 (cisplatin) revealed significant differences between the first and second measures in all frequencies. When compared to other group, the difference of the changes in Group 2 statistically significantly decreased, especially in higher frequencies. Morphologically, there were no acute changes in Group 1 and Group 3. Outer hair cell loss and the degeneration of stria vascularis and spiral ganglion were observed in both Groups 2 and 4; the damages in the latter were lesser. CONCLUSION: Quercetin does not have negative effect on cochlea, and it has protective effect on cisplatin-induced ototoxicity.
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Antineoplásicos/toxicidad , Antioxidantes/farmacología , Cisplatino/toxicidad , Ototoxicidad/prevención & control , Quercetina/farmacología , Análisis de Varianza , Animales , Femenino , Órgano Espiral/efectos de los fármacos , Órgano Espiral/patología , Ototoxicidad/patología , Ratas Wistar , Estría Vascular/efectos de los fármacos , Estría Vascular/patologíaRESUMEN
OBJECTIVES: Auditory neuropathy due to toxicity mechanism of pyridoxine has not yet been fully documented. Therefore, the present study explored a direct mechanism underlying the effects of pyridoxine on auditory neuropathy in organ of Corti (OC) explants ex vivo and cochlear neuroblast cell line, VOT-33 in vitro. METHODS: Primary OC explants containing spiral ganglion neurons and cultured VOT-33 cells were treated with pyridoxine. RESULTS: In nerve fiber of primary OC explants, pyridoxine decreased staining for NF200, a neuro-cytoskeletal protein. We also found that pyridoxine-induced VOT-33 apoptosis, as indicated by accumulation of the sub-G0/G1 fraction, caspase-3 activation, and PARP cleavage. In addition, pyridoxine induced reactive oxygen species (ROS) generation and alteration of mitochondrial membrane potential transition (MPT), including Bcl-2 family protein expression and consequently Ca2+ accumulation and changes of endoplasmic reticulum (ER) stress-related protein expression such as phospho-PERK, caspase-12, Grp78, and CHOP. CONCLUSION: Pyridoxine preferentially induced severe cell death on nerve fiber in primary OC explants and markedly increased apoptotic cell death via mitochondria-mediated ER stress in VOT-33 cells.
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Estrés del Retículo Endoplásmico/efectos de los fármacos , Pérdida Auditiva Central/etiología , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Órgano Espiral/efectos de los fármacos , Piridoxina/farmacología , Complejo Vitamínico B/farmacología , Animales , Apoptosis/efectos de los fármacos , Técnicas de Cultivo de Célula , Línea Celular , Chaperón BiP del Retículo Endoplásmico , Ratones , Órgano Espiral/patología , Ratas , Ratas Sprague-DawleyRESUMEN
Cisplatin-induced early-onset ototoxicity is linked to hearing loss. The mechanism by which cisplatin causes ototoxicity remains unclear. The purpose of this study was to identify the involvement of receptor-interacting protein kinase (RIP)3-dependent necroptosis in cisplatin-induced ototoxicity in vitro and in vivo. Sprague-Dawley rats (SD, 8 week) were treated via intraperitoneal (i.p.) injection with cisplatin (16 mg/kg for 1 day), and their hearing thresholds were measured by the auditory brainstem response (ABR) method. Hematoxylin and eosin (H & E) staining, immunohistochemistry, and western blots were performed to determine the effect of cisplatin-induced ototoxicity on cochlear morphology. Inhibitor experiments with necrostatin 1 (Nec-1) and Z-VAD were also performed in HEI-OC1 cell line. H&E stains revealed that the necroptotic changes were increased in the organ of Corti (OC) and spiral ganglion neurons (SGNs). Moreover, immunohistochemistry and western blot analysis showed that cisplatin treatment increased the protein levels of RIP3 in both OCs and SGNs. The treatment of Nec-1, a selective RIP1 inhibitor, resulted in markedly suppression of cisplatin-induced cell death in HEI-OC1 cells, whereas Z-VAD treatment did not change the cisplatin-induced cell death. Our results suggest that RIP3-dependent necroptosis was substantial in cisplatin-induced ototoxicity; inner cochlear regions, the OCs, and SGNs were especially sensitive to necroptosis.
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Cisplatino/efectos adversos , Ototoxicidad/metabolismo , Ototoxicidad/patología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Animales , Pérdida Auditiva/inducido químicamente , Pérdida Auditiva/patología , Masculino , Necroptosis , Órgano Espiral/efectos de los fármacos , Órgano Espiral/patología , Ratas Sprague-Dawley , Ganglio Espiral de la Cóclea/efectos de los fármacos , Ganglio Espiral de la Cóclea/patologíaRESUMEN
Hair cells do not undergo spontaneous regeneration when they are damaged in the mammalian organ of Corti, leading to irreversible hearing loss. Previous studies have shown that 24-diamino-5-phenylthiazole (DAPT), an inhibitor of Notch signaling, plays a major role in inner ear development. However, whether DAPT influences antibiotic-induced hair cell damage remains uncertain. The present study aimed to investigate whether DAPT exerts protective or regenerative effects on neomycin-damaged hair cells. A histological analysis was carried out to assess the number and morphological changes of hair cells in cultured organ of Corti explants. Our results showed that in-vitro treatment with DAPT induced extra hair cells, whereas no newly generated supporting cells were found. We also found that DAPT was effective for preventing hair cell loss when cotreatment with neomycin was performed, suggesting that DAPT exerted protective effects on neomycin ototoxicity. In addition, DAPT treatment for 2-4 days following neomycin damage induced supernumerary hair cells. These findings indicate that inhibition of Notch signaling is a possible strategy for the treatment of hair cell loss caused by aminoglycoside antibiotics.
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Antibacterianos/toxicidad , Diaminas/farmacología , Células Ciliadas Auditivas/efectos de los fármacos , Neomicina/toxicidad , Órgano Espiral/efectos de los fármacos , Tiazoles/farmacología , Animales , Animales Recién Nacidos , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Células Ciliadas Auditivas/patología , Células Ciliadas Auditivas/fisiología , Ratones , Ratones Endogámicos ICR , Técnicas de Cultivo de Órganos , Órgano Espiral/patología , Órgano Espiral/fisiología , Receptores Notch/antagonistas & inhibidores , Receptores Notch/fisiologíaRESUMEN
Transmembrane proteins that mediate the cellular uptake or efflux of thyroid hormone potentially provide a key level of control over neurodevelopment. In humans, defects in one such protein, solute carrier SLC16A2 (MCT8) are associated with psychomotor retardation. Other proteins that transport the active form of thyroid hormone triiodothyronine (T3) or its precursor thyroxine (T4) have been identified in vitro but the wider significance of such transporters in vivo is unclear. The development of the auditory system requires thyroid hormone and the cochlea is a primary target tissue. We have proposed that the compartmental anatomy of the cochlea would necessitate transport mechanisms to convey blood-borne hormone to target tissues. We report hearing loss in mice with mutations in Slc16a2 and a related gene Slc16a10 (Mct10, Tat1). Deficiency of both transporters results in retarded development of the sensory epithelium similar to impairment caused by hypothyroidism, compounded with a progressive degeneration of cochlear hair cells and loss of endocochlear potential. Administration of T3 largely restores the development of the sensory epithelium and limited auditory function, indicating the T3-sensitivity of defects in the sensory epithelium. The results indicate a necessity for thyroid hormone transporters in cochlear development and function.
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Sistemas de Transporte de Aminoácidos Neutros/genética , Sordera/genética , Eliminación de Gen , Células Ciliadas Auditivas/metabolismo , Proteínas de Transporte de Membrana/genética , Animales , Sordera/diagnóstico , Sordera/terapia , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Potenciales Evocados Auditivos , Femenino , Técnica del Anticuerpo Fluorescente , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Células Ciliadas Auditivas/patología , Masculino , Ratones , Ratones Noqueados , Transportadores de Ácidos Monocarboxílicos , Órgano Espiral/metabolismo , Órgano Espiral/patología , Simportadores , Hormonas Tiroideas/sangre , Hormonas Tiroideas/metabolismo , Hormonas Tiroideas/farmacologíaRESUMEN
HYPOTHESIS: This study evaluates the degree of preservation of hair cells, supporting cells, and innervating dendritic processes after cochlear implantation in the human using immunohistochemical methods. BACKGROUND: Surgical insertion of a cochlear implant electrode induces various pathologic changes within the cochlea including insertional trauma, foreign body response, inflammation, fibrosis, and neo-osteogenesis. These changes may result in loss of residual acoustic hearing, adversely affecting the use of hybrid implants, and may result in loss of putative precursor cells, limiting the success of future regenerative protocols. METHODS: Twenty-eight celloidin-embedded temporal bones from 14 patients with bilateral severe to profound sensorineural hearing loss and unilateral cochlear implants were studied. Two sections including the modiolus or basal turn from each temporal bone were stained using antineurofilament, antimyosin-VIIa, and antitubulin antibodies in both the implanted and unimplanted ears. RESULTS: Inner and outer hair cells: Immunoreactivity was reduced throughout the implanted cochlea and in the unimplanted cochlea with the exception of the apical turn.Dendritic processes in the osseous spiral lamina: Immunoreactivity was significantly less along the electrode of the implanted cochlea than in the other segments.Inner and outer pillars, inner and outer spiral bundles, and Deiters' cells: Immunoreactivity was similar in the implanted and unimplanted cochleae. CONCLUSION: Insertion of a cochlear implant electrode may significantly affect the inner and outer hair cells both along and apical to the electrode, and dendritic processes in the osseous spiral lamina along the electrode. There was less effect on pillar cells, Deiters' cells, and spiral bundles.
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Implantación Coclear , Implantes Cocleares/efectos adversos , Pérdida Auditiva Sensorineural/cirugía , Órgano Espiral/patología , Pérdida Auditiva Sensorineural/patología , Humanos , Inmunohistoquímica , MasculinoRESUMEN
Mutations in the GJB2 gene [which encodes connexin 26 (Cx26)] are the most common causes of hereditary hearing loss in humans, and previous studies showed postnatal development arrest of the organ of Corti in different Cx26-null mouse models. To explore the pathological changes and the mechanism behind the cochlear abnormalities in these mice further, we established transgenic mouse models by conditional knockdown of cochlear Cx26 at postnatal day (P) 0 and P8. Auditory brainstem responses were recorded and the morphological features in the organ of Corti were analyzed 18â days after Cx26 knockdown. Mice in the P0 knockdown group displayed severe hearing loss at all frequencies, whereas mice in the P8 knockdown group showed nearly normal hearing. In the P8 knockdown group, the organ of Corti displayed normal architecture, and no ultrastructural changes were observed. In the P0 knockdown group, the phalangeal processes of Deiter's cells did not develop into finger-like structures, and the formation of microtubules in the pillar cells was significantly reduced; moreover, the amount of acetylated α-tubulin was reduced in pillar cells. Our results indicate that Gjb2 participates in postnatal development of the cytoskeleton in pillar cells during structural maturation of the organ of Corti. In P0 knockdown mice, the reduction in microtubules in pillar cells might be responsible for the failure of the tunnel of Corti to open, and the malformed phalangeal processes might negatively affect the supporting framework of the organ of Corti, which would be a new mechanism of Gjb2-related hearing loss.
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Cóclea/anomalías , Cóclea/crecimiento & desarrollo , Conexinas/deficiencia , Citoesqueleto/metabolismo , Técnicas de Silenciamiento del Gen , Animales , Animales Recién Nacidos , Recuento de Células , Cóclea/patología , Conexina 26 , Conexinas/metabolismo , Citoesqueleto/ultraestructura , Modelos Animales de Enfermedad , Pérdida Auditiva/patología , Integrasas/metabolismo , Ratones Noqueados , Órgano Espiral/patología , Órgano Espiral/ultraestructuraRESUMEN
Identification of the causal effects of specific proteins on recurrent and partially reversible hearing loss has been difficult because of the lack of an animal model that provides reversible gene knockdown. We have developed the transgenic mouse line Actin-tTS::Nkcc1 tetO/tetO for manipulatable expression of the cochlear K+ circulation protein, NKCC1. Nkcc1 transcription was blocked by the binding of a tetracycline-dependent transcriptional silencer to the tetracycline operator sequences inserted upstream of the Nkcc1 translation initiation site. Administration of the tetracycline derivative doxycycline reversibly regulated Nkcc1 knockdown. Progeny from pregnant/lactating mothers fed doxycycline-free chow from embryonic day 0 showed strong suppression of Nkcc1 expression (~90% downregulation) and Nkcc1 null phenotypes at postnatal day 35 (P35). P35 transgenic mice from mothers fed doxycycline-free chow starting at P0 (delivery) showed weaker suppression of Nkcc1 expression (~70% downregulation) and less hearing loss with mild cochlear structural changes. Treatment of these mice at P35 with doxycycline for 2 weeks reactivated Nkcc1 transcription to control levels and improved hearing level at high frequency; i.e., these doxycycline-treated mice exhibited partially reversible hearing loss. Thus, development of the Actin-tTS::Nkcc1 tetO/tetO transgenic mouse line provides a mouse model for the study of variable hearing loss through reversible knockdown of Nkcc1.