RESUMO
Mechanosensory hair cells of the inner ears and lateral line of vertebrates display heightened vulnerability to environmental insult, with damage resulting in hearing and balance disorders. An important example is hair cell loss due to exposure to toxic agents including therapeutic drugs such as the aminoglycoside antibiotics such as neomycin and gentamicin and antineoplastic agents. We describe two distinct cellular pathways for aminoglycoside-induced hair cell death in zebrafish lateral line hair cells. Neomycin exposure results in death from acute exposure with most cells dying within 1 hour of exposure. By contrast, exposure to gentamicin results primarily in delayed hair cell death, taking up to 24 hours for maximal effect. Washout experiments demonstrate that delayed death does not require continuous exposure, demonstrating two mechanisms where downstream responses differ in their timing. Acute damage is associated with mitochondrial calcium fluxes and can be alleviated by the mitochondrially-targeted antioxidant mitoTEMPO, while delayed death is independent of these factors. Conversely delayed death is associated with lysosomal accumulation and is reduced by altering endolysosomal function, while acute death is not sensitive to lysosomal manipulations. These experiments reveal the complexity of responses of hair cells to closely related compounds, suggesting that intervention focusing on early events rather than specific death pathways may be a successful therapeutic strategy.
RESUMO
Hearing and balance deficits have been reported during and following treatment with the antimalarial drug chloroquine. However, experimental work examining the direct actions of chloroquine on mechanoreceptive hair cells in common experimental models is lacking. This study examines the effects of chloroquine on hair cells using two common experimental models: the zebrafish lateral line and neonatal mouse cochlear cultures. Zebrafish larvae were exposed to varying concentrations of chloroquine phosphate or hydroxychloroquine for 1 h or 24 h, and hair cells assessed by antibody staining. A significant, dose-dependent reduction in the number of surviving hair cells was seen across conditions for both exposure periods. Hydroxychloroquine showed similar toxicity. In mouse cochlear cultures, chloroquine damage was specific to outer hair cells in tissue from the cochlear basal turn, consistent with susceptibility to other ototoxic agents. These findings suggest a need for future studies employing hearing and balance monitoring during exposure to chloroquine and related compounds, particularly with interest in these compounds as therapeutics against viral infections including coronavirus.
Assuntos
Sobrevivência Celular/efeitos dos fármacos , Cloroquina/análogos & derivados , Células Ciliadas Auditivas/efeitos dos fármacos , Hidroxicloroquina/toxicidade , Sistema da Linha Lateral/efeitos dos fármacos , Animais , Antivirais/toxicidade , Células Cultivadas , Cloroquina/toxicidade , Células Ciliadas Auditivas/citologia , Larva/efeitos dos fármacos , Camundongos , Modelos Animais , Ototoxicidade , Peixe-ZebraRESUMO
Aminoglycoside (AG) antibiotics are widely used to prevent life-threatening infections, and cisplatin is used in the treatment of various cancers, but both are ototoxic and result in loss of sensory hair cells from the inner ear. ORC-13661 is a new drug that was derived from PROTO-1, a compound first identified as protective in a large-scale screen utilizing hair cells in the lateral line organs of zebrafish larvae. Here, we demonstrate, in zebrafish larvae and in mouse cochlear cultures, that ORC-13661 provides robust protection of hair cells against both ototoxins, the AGs and cisplatin. ORC-13661 also prevents both hearing loss in a dose-dependent manner in rats treated with amikacin and the loading of neomycin-Texas Red into lateral line hair cells. In addition, patch-clamp recordings in mouse cochlear cultures reveal that ORC-13661 is a high-affinity permeant blocker of the mechanoelectrical transducer (MET) channel in outer hair cells, suggesting that it may reduce the toxicity of AGs by directly competing for entry at the level of the MET channel and of cisplatin by a MET-dependent mechanism. ORC-13661 is therefore a promising and versatile protectant that reversibly blocks the hair cell MET channel and operates across multiple species and toxins.
Assuntos
Antibacterianos/toxicidade , Antineoplásicos/toxicidade , Células Ciliadas Auditivas/efeitos dos fármacos , Ototoxicidade/prevenção & controle , Substâncias Protetoras/farmacologia , Tiofenos/farmacologia , Ureia/análogos & derivados , Amicacina/toxicidade , Aminoglicosídeos/toxicidade , Animais , Técnicas de Cultura de Células , Células Cultivadas , Cisplatino/toxicidade , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Células Ciliadas Auditivas/metabolismo , Humanos , Microscopia Intravital , Canais Iônicos/antagonistas & inibidores , Canais Iônicos/metabolismo , Masculino , Mecanotransdução Celular/efeitos dos fármacos , Camundongos , Ototoxicidade/etiologia , Técnicas de Patch-Clamp , Substâncias Protetoras/uso terapêutico , Ratos , Tiofenos/uso terapêutico , Imagem com Lapso de Tempo , Ureia/farmacologia , Ureia/uso terapêutico , Peixe-ZebraRESUMO
Growth of the craniofacial skeleton is a complex process controlled by both genetic and epigenetic factors, perturbations of which can lead to varying degrees of dysmorphology. Mouse models that recapitulate clinical craniofacial phenotypes are instrumental in studying the morphogenetic progression of diseases as well as uncovering their genetic and molecular bases. Commonly encountered phenotypes in these models include defects in the cranial base synchondroses, calvarial sutures, mandible or the midface, or any combination thereof, with the concurrent presence of altered overall craniofacial growth. However, the literature lacks an adequate normative timeline of developmental events and growth trends that shape the mouse craniofacial skeleton. In this report, we analyzed the postnatal craniofacial ontogeny (from postnatal day 7 [P7] through to P112) of male mice from the most widely used inbred mouse strain, C57BL/6J, using high-resolution microcomputed tomography (µCT) in combination with classic morphometric approaches. We also evaluated cranial base synchondroses at the histological level, and compared it to µCT-generated data to assess the timing and pattern of closure of these structures. Our data underscore the complex and unique growth patterns of individual bones and cranial regions and highlight the need to include younger animals in studies aimed at analyzing craniofacial growth processes. Furthermore, these data serve as a reference standard for future quantitative work.
RESUMO
Microtia is a term used to describe a wide array of phenotypic presentations of the outer ear. Although the majority of the cases are isolated in nature, much of our understanding of the causes of microtia has been driven by the identification of genes underlying syndromic forms where the anomaly co-presents with various other craniofacial and extra-craniofacial structural defects. In this review we discuss recent findings in mice deficient in Hoxa2, a key regulator of branchial arch patterning, which has necessitated a revision to the canonical model of pinna morphogenesis. The revised model will likely impact current classification schemes for microtia and, as we argue in this review, the interpretation of the developmental basis for various auricular malformations. In addition, we highlight recent studies in other mammalian species that are providing the first clues as to possible causes of at least some isolated anomalies and thus should now accelerate the search for the more elusive genetic contributions to the many isolated and non-syndromic cases of microtia. These findings, together with the application of new genome-level sequencing technologies and more thorough quantitative assessment of available mutant mouse resources, promise an exciting future for genetic studies in microtia.