RESUMO
In recent years, exome sequencing (ES) has shown great utility in the diagnoses of Mendelian disorders. However, after rigorous filtering, a typical ES analysis still involves the interpretation of hundreds of variants, which greatly hinders the rapid identification of causative genes. Since the interpretations of ES data require comprehensive clinical analyses, taking clinical expertise into consideration can speed the molecular diagnoses of Mendelian disorders. To leverage clinical expertise to prioritize candidate genes, we developed PhenoApt, a phenotype-driven gene prioritization tool that allows users to assign a customized weight to each phenotype, via a machine-learning algorithm. Using the ability to rank causative genes in top-10 lists as an evaluation metric, baseline analysis demonstrated that PhenoApt outperformed previous phenotype-driven gene prioritization tools by a relative increase of 22.7%-140.0% in three independent, real-world, multi-center cohorts (cohort 1, n = 185; cohort 2, n = 784; and cohort 3, n = 208). Additional trials showed that, by adding weights to clinical indications, which should be explained by the causative gene, PhenoApt performance was improved by a relative increase of 37.3% in cohort 2 (n = 471) and 21.4% in cohort 3 (n = 208). Moreover, PhenoApt could assign an intrinsic weight to each phenotype based on the likelihood of its being a Mendelian trait using term frequency-inverse document frequency techniques. When clinical indications were assigned with intrinsic weights, PhenoApt performance was improved by a relative increase of 23.7% in cohort 2 and 15.5% in cohort 3. For the integration of PhenoApt into clinical practice, we developed a user-friendly website and a command-line tool.
Assuntos
Doenças Genéticas Inatas/genética , Perda Auditiva Neurossensorial/genética , Deficiência Intelectual/genética , Aprendizado de Máquina , Microcefalia/genética , Nistagmo Congênito/genética , Escoliose/genética , Estudos de Coortes , Biologia Computacional , Bases de Dados Genéticas , Exoma , Doenças Genéticas Inatas/diagnóstico , Doenças Genéticas Inatas/patologia , Testes Genéticos , Genótipo , Perda Auditiva Neurossensorial/diagnóstico , Perda Auditiva Neurossensorial/patologia , Humanos , Deficiência Intelectual/diagnóstico , Deficiência Intelectual/patologia , Microcefalia/diagnóstico , Microcefalia/patologia , Nistagmo Congênito/diagnóstico , Nistagmo Congênito/patologia , Fenótipo , Escoliose/diagnóstico , Escoliose/patologia , Software , Sequenciamento do ExomaRESUMO
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.
Assuntos
Autofagia , Proteína DEAD-box 58 , Glaucoma , Pressão Intraocular , Malha Trabecular , Animais , Feminino , Humanos , Masculino , Camundongos , Doenças da Aorta , Autofagia/efeitos dos fármacos , Proteína DEAD-box 58/metabolismo , Proteína DEAD-box 58/genética , Hipoplasia do Esmalte Dentário , Glaucoma/patologia , Glaucoma/metabolismo , Glaucoma/genética , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Perda Auditiva Neurossensorial/metabolismo , Interferon beta/metabolismo , Pressão Intraocular/genética , Metacarpo/anormalidades , Camundongos Endogâmicos C57BL , Doenças Musculares , Mutação , Odontodisplasia , Atrofia Óptica/genética , Atrofia Óptica/metabolismo , Atrofia Óptica/patologia , Osteoporose , Linhagem , Receptores Imunológicos , Malha Trabecular/metabolismo , Malha Trabecular/efeitos dos fármacos , Calcificação VascularRESUMO
BACKGROUND: Plexins are large transmembrane receptors for the semaphorin family of signalling proteins. Semaphorin-plexin signalling controls cellular interactions that are critical during development as well as in adult life stages. Nine plexin genes have been identified in humans, but despite the apparent importance of plexins in development, only biallelic PLXND1 and PLXNA1 variants have so far been associated with Mendelian genetic disease. METHODS: Eight individuals from six families presented with a recessively inherited variable clinical condition, with core features of amelogenesis imperfecta (AI) and sensorineural hearing loss (SNHL), with variable intellectual disability. Probands were investigated by exome or genome sequencing. Common variants and those unlikely to affect function were excluded. Variants consistent with autosomal recessive inheritance were prioritised. Variant segregation analysis was performed by Sanger sequencing. RNA expression analysis was conducted in C57Bl6 mice. RESULTS: Rare biallelic pathogenic variants in plexin B2 (PLXNB2), a large transmembrane semaphorin receptor protein, were found to segregate with disease in all six families. The variants identified include missense, nonsense, splicing changes and a multiexon deletion. Plxnb2 expression was detected in differentiating ameloblasts. CONCLUSION: We identify rare biallelic pathogenic variants in PLXNB2 as a cause of a new autosomal recessive, phenotypically diverse syndrome with AI and SNHL as core features. Intellectual disability, ocular disease, ear developmental abnormalities and lymphoedema were also present in multiple cases. The variable syndromic human phenotype overlaps with that seen in Plxnb2 knockout mice, and, together with the rarity of human PLXNB2 variants, may explain why pathogenic variants in PLXNB2 have not been reported previously.
Assuntos
Amelogênese Imperfeita , Deficiência Intelectual , Linhagem , Humanos , Animais , Masculino , Feminino , Camundongos , Deficiência Intelectual/genética , Deficiência Intelectual/patologia , Amelogênese Imperfeita/genética , Amelogênese Imperfeita/patologia , Receptores de Superfície Celular/genética , Proteínas do Tecido Nervoso/genética , Alelos , Criança , Perda Auditiva/genética , Perda Auditiva/patologia , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Adulto , Mutação/genética , Adolescente , Pré-Escolar , FenótipoRESUMO
BACKGROUND: The stria vascularis (SV), located in the lateral wall of the cochlea, maintains cochlear fluid homeostasis and mechanoelectrical transduction (MET) activity required for sound wave conduction. The pathogenesis of a number of human inheritable deafness syndromes, age related hearing loss, drug-induced ototoxicity and noise-induced hearing loss results from the morphological changes and functional impairments in the development of the SV. In this study, we investigate the implications of intercellular communication within the SV in the pathogenesis of sensorineural hearing loss (SNHL). We aim to identify commonly regulated signaling pathways using publicly available single-cell transcriptomic sequencing (scRNA-seq) datasets. METHODS: We analyzed scRNA-seq data, which was derived from studying the cochlear SV in mice with SNHL compared to normal adult mice. After quality control and filtering, we obtained the major cellular components of the mouse cochlear SV and integrated the data. Using Seurat's FindAllMarkers and FindMarkers packages, we searched for novel conservative genes and differential genes. We employed KEGG and GSEA to identify molecular pathways that are commonly altered among different types of SNHL. We utilized pySCENIC to discover new specific regulatory factors in SV subpopulation cells. With the help of CellChat, we identified changes in subpopulation cells showing similar trends across different SNHL types and their alterations in intercellular communication pathways. RESULTS: Through the analysis of the integrated data, we discovered new conserved genes to SV specific cells and identified common downregulated pathways in three types of SNHL. The enriched genes for these pathways showing similar trends are primarily associated with the Electron Transport Chain, related to mitochondrial energy metabolism. Using the CellChat package, we further found that there are shared pathways in the incoming signaling of specific intermediate cells in SNHL, and these pathways have common upstream regulatory transcription factor of Nfe2l2. Combining the results from pySCENIC and CellChat, we predicted the transcription factor Nfe2l2 as an upstream regulatory factor for multiple shared cellular pathways in IC. Additionally, it serves as an upstream factor for several genes within the Electron Transport Chain. CONCLUSION: Our bioinformatics analysis has revealed that downregulation of the mitochondrial electron transport chain have been observed in various conditions of SNHL. E2f1, Esrrb, Runx1, Yy1, and Gata2 could serve as novel important common TFs regulating the electron transport chain. Adm has emerged as a potential new marker gene for intermediate cells, while Itgb5 and Tesc show promise as potential new marker genes for marginal cells in the SV. These findings offer a new perspective on SV lesions in SNHL and provide additional theoretical evidence for the same drug treatment and prevention of different pathologies of SNHL.
Assuntos
Perda Auditiva Neurossensorial , Estria Vascular , Adulto , Humanos , Animais , Camundongos , Estria Vascular/metabolismo , Estria Vascular/patologia , Análise da Expressão Gênica de Célula Única , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Cóclea , Fatores de Transcrição/metabolismoRESUMO
Identification of genes associated with nonsyndromic hearing loss is a crucial endeavor given the substantial number of individuals who remain without a diagnosis after even the most advanced genetic testing. PKHD1L1 was established as necessary for the formation of the cochlear hair-cell stereociliary coat and causes hearing loss in mice and zebrafish when mutated. We sought to determine if biallelic variants in PKHD1L1 also cause hearing loss in humans. Exome sequencing was performed on DNA of four families segregating autosomal recessive nonsyndromic sensorineural hearing loss. Compound heterozygous p.[(Gly129Ser)];p.[(Gly1314Val)] and p.[(Gly605Arg)];p[(Leu2818TyrfsTer5)], homozygous missense p.(His2479Gln) and nonsense p.(Arg3381Ter) variants were identified in PKHD1L1 that were predicted to be damaging using in silico pathogenicity prediction methods. In vitro functional analysis of two missense variants was performed using purified recombinant PKHD1L1 protein fragments. We then evaluated protein thermodynamic stability with and without the missense variants found in one of the families and performed a minigene splicing assay for another variant. In silico molecular modeling using AlphaFold2 and protein sequence alignment analysis were carried out to further explore potential variant effects on structure. In vitro functional assessment indicated that both engineered PKHD1L1 p.(Gly129Ser) and p.(Gly1314Val) mutant constructs significantly reduced the folding and structural stabilities of the expressed protein fragments, providing further evidence to support pathogenicity of these variants. Minigene assay of the c.1813G>A p.(Gly605Arg) variant, located at the boundary of exon 17, revealed exon skipping leading to an in-frame deletion of 48 amino acids. In silico molecular modeling exposed key structural features that might suggest PKHD1L1 protein destabilization. Multiple lines of evidence collectively associate PKHD1L1 with nonsyndromic mild-moderate to severe sensorineural hearing loss. PKHD1L1 testing in individuals with mild-moderate hearing loss may identify further affected families.
Assuntos
Surdez , Mutação de Sentido Incorreto , Linhagem , Receptores de Superfície Celular , Estereocílios , Animais , Feminino , Humanos , Masculino , Surdez/genética , Sequenciamento do Exoma , Genes Recessivos , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Modelos Moleculares , Receptores de Superfície Celular/genética , Estereocílios/metabolismo , Estereocílios/patologia , Estereocílios/genéticaRESUMO
Biallelic pathogenic variants in MAP3K20, which encodes a mitogen-activated protein kinase, are a rare cause of split-hand foot malformation (SHFM), hearing loss, and nail abnormalities or congenital myopathy. However, heterozygous variants in this gene have not been definitively associated with a phenotype. Here, we describe the phenotypic spectrum associated with heterozygous de novo variants in the linker region between the kinase domain and leucine zipper domain of MAP3K20. We report five individuals with diverse clinical features, including craniosynostosis, limb anomalies, sensorineural hearing loss, and ectodermal dysplasia-like phenotypes who have heterozygous de novo variants in this specific region of the gene. These individuals exhibit both shared and unique clinical manifestations, highlighting the complexity and variability of the disorder. We propose that the involvement of MAP3K20 in endothelial-mesenchymal transition provides a plausible etiology of these features. Together, these findings characterize a disorder that both expands the phenotypic spectrum associated with MAP3K20 and highlights the need for further studies on its role in early human development.
Assuntos
Craniossinostoses , Displasia Ectodérmica , Perda Auditiva Neurossensorial , Heterozigoto , Humanos , Displasia Ectodérmica/genética , Displasia Ectodérmica/patologia , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Masculino , Feminino , Craniossinostoses/genética , Fenótipo , Pré-Escolar , Deformidades Congênitas dos Membros/genética , Criança , Mutação , Lactente , MAP Quinase Quinase Quinases/genéticaRESUMO
V-ATPase is an ATP hydrolysis-driven proton pump involved in the acidification of intracellular organelles and systemic acid-base homeostasis through H+ secretion in the renal collecting ducts. V-ATPase dysfunction is associated with hereditary distal renal tubular acidosis (dRTA). ATP6V1B1 encodes the B1 subunit of V-ATPase that is integral to ATP hydrolysis and subsequent H+ transport. Patients with pathogenic ATP6V1B1 mutations often exhibit an early onset of sensorineural hearing loss. However, the mechanisms underlying this association remain unclear. We employed morpholino oligonucleotide-mediated knockdown and CRISPR/Cas9 gene editing to generate Atp6v1ba-deficient (atp6v1ba-/-) zebrafish as an ortholog model for ATP6V1B1. The atp6v1ba-/- zebrafish exhibited systemic acidosis and significantly smaller otoliths compared to wild-type siblings. Moreover, deficiency in Atp6v1ba led to degeneration of inner ear hair cells, with ultrastructural changes indicative of autophagy. Our findings indicate a critical role of ATP6V1B1 in regulating lysosomal pH and autophagy in hair cells, and the results provide insights into the pathophysiology of sensorineural hearing loss in dRTA. Furthermore, this study demonstrates that the atp6v1ba-/- zebrafish model is a valuable tool for further investigation into disease mechanisms and potential therapies for acidosis-related hearing impairment.
Assuntos
Acidose Tubular Renal , Acidose , Perda Auditiva Neurossensorial , Compostos Organometálicos , ATPases Vacuolares Próton-Translocadoras , Animais , Humanos , Peixe-Zebra/metabolismo , ATPases Vacuolares Próton-Translocadoras/genética , ATPases Vacuolares Próton-Translocadoras/metabolismo , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Mutação , Acidose Tubular Renal/genética , Células Ciliadas Auditivas/patologia , Concentração de Íons de Hidrogênio , Cabelo/metabolismo , Trifosfato de AdenosinaRESUMO
HDR syndrome is a rare disease characterized by hypoparathyroidism, deafness, and renal dysplasia. An autosomal dominant disease caused by heterozygous pathogenic GATA3 variants, the penetrance of each associated condition is variable. Literature reviews have provided some answers, but many questions remain, in particular what the relationship is between genotype and phenotype. The current study examines 28 patients with HDR syndrome combined with an exhaustive review of the literature. Some conditions such as hearing loss are almost always present, while others described as rare initially, do not seem to be so rare after all (genital malformations and basal ganglia calcifications). By modeling pathogenic GATA3 variants found in HDR syndrome, we found that missense variations appear to always be located in the same area (close to the two Zinc Finger domain). We describe new pathogenic GATA3 variants, of which some seem to always be associated with certain conditions. Many audiograms were studied to establish a typical audiometric profile associated with a phenotype in HDR. As mentioned in the literature, hearing function should always be assessed as early as possible and follow up of patients with HDR syndrome should include monitoring of parathyroid function and vesicoureteral reflux in order to prevent complications.
Assuntos
Fator de Transcrição GATA3 , Hipoparatireoidismo , Fenótipo , Humanos , Hipoparatireoidismo/genética , Hipoparatireoidismo/patologia , Fator de Transcrição GATA3/genética , Masculino , Feminino , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Estudos de Associação Genética , Nefrose/genética , Nefrose/patologia , Criança , Predisposição Genética para Doença , Mutação , Pré-Escolar , Estudos de CoortesRESUMO
MPZL2-related hearing loss is a rare form of autosomal recessive hearing loss characterized by progressive, mild sloping to severe sensorineural hearing loss. Thirty-five previously reported patients had biallelic truncating variants in MPZL2, with the exception of one patient with a missense variant of uncertain significance and a truncating variant. Here, we describe the clinical characteristics and genotypes of five patients from four families with confirmed MPZL2-related hearing loss. A rare missense likely pathogenic variant [NM_005797.4(MPZL2):c.280C>T,p.(Arg94Trp)] located in exon 3 was confirmed to be in trans with a recurrent pathogenic truncating variant that segregated with hearing loss in three of the patients from two unrelated families. This is the first recurrent likely pathogenic missense variant identified in MPZL2. Apparently milder or later-onset hearing loss associated with rare missense variants in MPZL2 indicates that some missense variants in this gene may cause a milder phenotype than that resulting from homozygous or compound heterozygous truncating variants. This study, along with the identification of truncating loss of function and missense MPZL2 variants in several diverse populations, suggests that MPZL2-related hearing loss may be more common than previously appreciated and demonstrates the need for MPZL2 inclusion in hearing loss testing panels.
Assuntos
Moléculas de Adesão Celular , Perda Auditiva Neurossensorial , Humanos , Moléculas de Adesão Celular/genética , Surdez/genética , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Mutação de Sentido Incorreto/genética , Linhagem , FenótipoRESUMO
Autosomal dominant sensorineural hearing loss (ADSNHL) is a genetically heterogeneous disorder caused by pathogenic variants in various genes, including MYH14. However, the interpretation of pathogenicity for MYH14 variants remains a challenge due to incomplete penetrance and the lack of functional studies and large families. In this study, we performed exome sequencing in six unrelated families with ADSNHL and identified five MYH14 variants, including three novel variants. Two of the novel variants, c.571G > C (p.Asp191His) and c.571G > A (p.Asp191Asn), were classified as likely pathogenic using ACMG and Hearing Loss Expert panel guidelines. In silico modeling demonstrated that these variants, along with p.Gly1794Arg, can alter protein stability and interactions among neighboring molecules. Our findings suggest that MYH14 causative variants may be more contributory and emphasize the importance of considering this gene in patients with nonsyndromic mainly post-lingual severe form of hearing loss. However, further functional studies are needed to confirm the pathogenicity of these variants.
Assuntos
Sequenciamento do Exoma , Perda Auditiva Neurossensorial , Cadeias Pesadas de Miosina , Miosina Tipo II , Linhagem , Humanos , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Feminino , Masculino , Cadeias Pesadas de Miosina/genética , Adulto , Mutação/genética , Predisposição Genética para Doença , Criança , Genes Dominantes , Pessoa de Meia-Idade , AdolescenteRESUMO
Aymé-Gripp syndrome (AYGRPS) is a multisystemic disorder caused by a subset of pathogenic variants in the MAF gene. Major clinical features include bilateral early cataracts, sensorineural hearing loss (SNHL), and a characteristic facial appearance along with variable neurodevelopmental delay. Pericarditis resulting in pericardial effusion of varying degree has been observed in a subset of affected individuals and could represent a severe feature in neonatal or infantile age. Here, we describe a syndromic infant with massive pericardial effusion and craniofacial features that oriented toward the suspicion of AYGRPS, which was subsequently confirmed by the molecular analysis of MAF. Pericardial effusion was first observed prenatally and documented to be recurrent, progressive, and severe in the first months of life, thus requiring pericardiocentesis and surgical procedures. In this report, we provide further delineation of the minor clinical characteristics, particularly focusing on cardiac features of AYGRPS. A dedicated cardiac surveillance of these findings may help reduce the morbidity and mortality of this rare condition.
Assuntos
Derrame Pericárdico , Feminino , Humanos , Recém-Nascido , Catarata/diagnóstico , Catarata/genética , Catarata/patologia , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/patologia , Perda Auditiva Neurossensorial/diagnóstico , Derrame Pericárdico/patologia , Derrame Pericárdico/diagnóstico , Fenótipo , Deficiências do Desenvolvimento/diagnóstico , Deficiências do Desenvolvimento/genética , Deficiências do Desenvolvimento/patologiaRESUMO
A heterozygous gain-of-function variant in the acyl-CoA oxidase 1 (ACOX1) gene, c.710A>G (p.Asn237Ser), is known to cause Mitchell syndrome, a very rare progressive disorder characterized by episodic demyelination, sensory polyneuropathy, and hearing loss. Only eight patients have been described so far. A single patient has been treated with intravenous immunoglobulin administration, indicating clinical improvement. In this study, we describe a 10-year-old girl carrying the identical mutation, who presented with progressive sensorineural deafness, visual abnormalities, skin ichthyosis, and gait ataxia from infantile age with progressive worsening and loss of walking ability by the age of 10 years. Antioxidant therapies and monthly intravenous immunoglobulin infusions showed excellent clinical results: after 1 year of treatment, the child is now able to walk, run, and jump. We emphasize the importance of early genetic diagnosis since an effective treatment is available for this rare condition.
Assuntos
Mutação com Ganho de Função , Criança , Feminino , Humanos , Doenças Desmielinizantes/genética , Doenças Desmielinizantes/tratamento farmacológico , Doenças Desmielinizantes/patologia , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/tratamento farmacológico , Perda Auditiva Neurossensorial/patologia , Imunoglobulinas Intravenosas/uso terapêutico , Imunomodulação/genética , Imunomodulação/efeitos dos fármacos , FenótipoRESUMO
PURPOSE: Radiographic review of pathologies that associate with third window syndrome. METHODS: Case series and literature review. RESULTS: Eight unique third window conditions are described and illustrated, including superior, lateral, and posterior semicircular canal dehiscence; carotid-cochlear, facial-cochlear, and internal auditory canal-cochlear dehiscence, labyrinthine erosion from endolymphatic sac tumor, and enlarged vestibular aqueduct. CONCLUSION: The present study highlights the characteristic imaging features and symptoms to differentiate third window pathologies for expedient diagnosis and management planning.
Assuntos
Perda Auditiva Neurossensorial , Doenças do Labirinto , Deiscência do Canal Semicircular , Aqueduto Vestibular , Humanos , Doenças do Labirinto/diagnóstico por imagem , Doenças do Labirinto/patologia , Perda Auditiva Neurossensorial/patologia , Aqueduto Vestibular/patologia , Cóclea/diagnóstico por imagem , Cóclea/patologia , Canais Semicirculares/diagnóstico por imagem , Canais Semicirculares/patologiaRESUMO
Hearing is essential for communication, and its loss can cause a serious disruption to one's social life. Hearing loss is also recognized as a major risk factor for dementia; therefore, addressing hearing loss is a pressing global issue. Sensorineural hearing loss, the predominant type of hearing loss, is mainly due to damage to the inner ear along with a variety of pathologies including ischemia, noise, trauma, aging, and ototoxic drugs. In addition to genetic factors, oxidative stress has been identified as a common mechanism underlying several cochlear pathologies. The cochlea, which plays a major role in auditory function, requires high-energy metabolism and is, therefore, highly susceptible to oxidative stress, particularly in the mitochondria. Based on these pathological findings, the potential of antioxidants for the treatment of hearing loss has been demonstrated in several animal studies. However, results from human studies are insufficient, and future clinical trials are required. This review discusses the relationship between sensorineural hearing loss and reactive oxidative species (ROS), with particular emphasis on age-related hearing loss, noise-induced hearing loss, and ischemia-reperfusion injury. Based on these mechanisms, the current status and future perspectives of ROS-targeted therapy for sensorineural hearing loss are described.
Assuntos
Perda Auditiva Neurossensorial , Estresse Oxidativo , Espécies Reativas de Oxigênio , Humanos , Perda Auditiva Neurossensorial/metabolismo , Perda Auditiva Neurossensorial/patologia , Animais , Espécies Reativas de Oxigênio/metabolismo , Antioxidantes/uso terapêutico , Antioxidantes/metabolismo , Cóclea/metabolismo , Cóclea/patologia , Perda Auditiva Provocada por Ruído/metabolismo , Perda Auditiva Provocada por Ruído/patologia , Perda Auditiva Provocada por Ruído/tratamento farmacológico , Traumatismo por Reperfusão/metabolismo , Mitocôndrias/metabolismoRESUMO
P2RX2 encodes the P2X2 receptor, which is an adenosine triphosphate (ATP) gated (purinoreceptor) ion channel. P2RX2 c. 178G > T (p.V60L) mutation was previously identified in two unrelated Chinese families, as the cause of human DFNA41, a form of dominant, early-onset and progressive sensorineural hearing loss. We generated and characterized a knock-in mouse model based on human p.V60L mutation that recapitulates the human phenotype. Heterozygous KI mice started to exhibit hearing loss at 21-day-old and progressed to deafness by 6-month-old. Vestibular dysfunction was also observed in mutant mice. Abnormal morphology of the inner hair cells and ribbon synapses was progressively observed in KI animals suggesting that P2rx2 plays a role in the membrane spatial location of the ribbon synapses. These results suggest that P2rx2 is essential for acoustic information transfer, which can be the molecular mechanism related to hearing loss.
Assuntos
Perda Auditiva Neurossensorial/genética , Receptores Purinérgicos P2X2/genética , Trifosfato de Adenosina/metabolismo , Animais , Modelos Animais de Doenças , Técnicas de Introdução de Genes , Células Ciliadas Auditivas Internas/patologia , Perda Auditiva Neurossensorial/patologia , Heterozigoto , Humanos , Camundongos , Mutação/genética , Linhagem , Fenótipo , Sinapses/genética , Sinapses/patologia , Doenças Vestibulares/genética , Doenças Vestibulares/patologiaRESUMO
Hearing loss is a frequent sensory impairment in humans and genetic factors account for an elevated fraction of the cases. We have investigated a large family of five generations, with 15 reported individuals presenting non-syndromic, sensorineural, bilateral and progressive hearing loss, segregating as an autosomal dominant condition. Linkage analysis, using SNP-array and selected microsatellites, identified a region of near 13 cM in chromosome 20 as the best candidate to harbour the causative mutation. After exome sequencing and filtering of variants, only one predicted deleterious variant in the NCOA3 gene (NM_181659, c.2810C > G; p.Ser937Cys) fit in with our linkage data. RT-PCR, immunostaining and in situ hybridization showed expression of ncoa3 in the inner ear of mice and zebrafish. We generated a stable homozygous zebrafish mutant line using the CRISPR/Cas9 system. ncoa3-/- did not display any major morphological abnormalities in the ear, however, anterior macular hair cells showed altered orientation. Surprisingly, chondrocytes forming the ear cartilage showed abnormal behaviour in ncoa3-/-, detaching from their location, invading the ear canal and blocking the cristae. Adult mutants displayed accumulation of denser material wrapping the otoliths of ncoa3-/- and increased bone mineral density. Altered zebrafish swimming behaviour corroborates a potential role of ncoa3 in hearing loss. In conclusion, we identified a potential candidate gene to explain hereditary hearing loss, and our functional analyses suggest subtle and abnormal skeletal behaviour as mechanisms involved in the pathogenesis of progressive sensory function impairment.
Assuntos
Surdez/genética , Predisposição Genética para Doença , Perda Auditiva Neurossensorial/genética , Coativador 3 de Receptor Nuclear/genética , Adulto , Animais , Surdez/patologia , Modelos Animais de Doenças , Orelha Interna/metabolismo , Orelha Interna/patologia , Exoma/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Células Ciliadas Auditivas/metabolismo , Células Ciliadas Auditivas/patologia , Perda Auditiva Neurossensorial/patologia , Humanos , Masculino , Camundongos , Linhagem , Sequenciamento do Exoma , Peixe-Zebra/genéticaRESUMO
Reactive oxygen species (ROS) produced by NADPH oxidases (Nox) contribute to the development of different types of sensorineural hearing loss (SNHL), a common impairment in humans with no established treatment. Although the essential role of Nox3 in otoconia biosynthesis and its possible involvement in hearing have been reported in rodents, immunohistological methods targeted at detecting Nox3 expression in inner ear cells reveal ambiguous results. Therefore, the mechanism underlying Nox3-dependent SNHL remains unclear and warrants further investigation. We generated Nox3-Cre knock-in mice, in which Nox3 was replaced with Cre recombinase (Cre). Using Nox3-Cre;tdTomato mice of either sex, in which tdTomato is expressed under the control of the Nox3 promoter, we determined Nox3-expressing regions and cell types in the inner ear. Nox3-expressing cells in the cochlea included various types of supporting cells, outer hair cells, inner hair cells, and spiral ganglion neurons. Nox3 expression increased with cisplatin, age, and noise insults. Moreover, increased Nox3 expression in supporting cells and outer hair cells, especially at the basal turn of the cochlea, played essential roles in ROS-related SNHL. The extent of Nox3 involvement in SNHL follows the following order: cisplatin-induced hearing loss > age-related hearing loss > noise-induced hearing loss. Here, on the basis of Nox3-Cre;tdTomato, which can be used as a reporter system (Nox3-Cre+/-;tdTomato+/+ and Nox3-Cre+/+;tdTomato+/+), and Nox3-KO (Nox3-Cre+/+;tdTomato+/+) mice, we demonstrate that Nox3 inhibition in the cochlea is a promising strategy for ROS-related SNHL, such as cisplatin-induced HL, age-related HL, and noise-induced HL.SIGNIFICANCE STATEMENT We found Nox3-expressing regions and cell types in the inner ear, especially in the cochlea, using Nox3-Cre;tdTomato mice, a reporter system generated in this study. Nox3 expression increased with cisplatin, age, and noise insults in specific cell types in the cochlea and resulted in the loss (apoptosis) of outer hair cells. Thus, Nox3 might serve as a molecular target for the development of therapeutics for sensorineural hearing loss, particularly cisplatin-induced, age-related, and noise-induced hearing loss.
Assuntos
Cóclea/metabolismo , Perda Auditiva Neurossensorial/metabolismo , NADPH Oxidases/metabolismo , Superóxidos/metabolismo , Envelhecimento/patologia , Animais , Cisplatino/toxicidade , Cóclea/patologia , Feminino , Técnicas de Introdução de Genes , Perda Auditiva Provocada por Ruído/metabolismo , Perda Auditiva Neurossensorial/etiologia , Perda Auditiva Neurossensorial/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ruído/efeitos adversosRESUMO
Mediator protein complex subunit 12 (Med12) is a core component of the basal transcriptional apparatus and plays a critical role in the development of many tissues. Mutations in Med12 are associated with X-linked intellectual disability syndromes and hearing loss; however, its role in nervous system function remains undefined. Here, we show that temporal conditional deletion of Med12 in astrocytes in the adult CNS results in region-specific alterations in astrocyte morphology. Surprisingly, behavioral studies revealed rapid hearing loss after adult deletion of Med12 that was confirmed by a complete abrogation of auditory brainstem responses. Cellular analysis of the cochlea revealed degeneration of the stria vascularis, in conjunction with disorganization of basal cells adjacent to the spiral ligament and downregulation of key cell adhesion proteins. Physiologic analysis revealed early changes in endocochlear potential, consistent with strial-specific defects. Together, our studies reveal that Med12 regulates auditory function in the adult by preserving the structural integrity of the stria vascularis.SIGNIFICANCE STATEMENT Mutations in Mediator protein complex subunit 12 (Med12) are associated with X-linked intellectual disability syndromes and hearing loss. Using temporal-conditional genetic approaches in CNS glia, we found that loss of Med12 results in severe hearing loss in adult animals through rapid degeneration of the stria vascularis. Our study describes the first animal model that recapitulates hearing loss identified in Med12-related disorders and provides a new system in which to examine the underlying cellular and molecular mechanisms of Med12 function in the adult nervous system.
Assuntos
Astrócitos/fisiologia , Perda Auditiva Neurossensorial/etiologia , Complexo Mediador/deficiência , Estria Vascular/patologia , Animais , Astrócitos/metabolismo , Astrócitos/ultraestrutura , Moléculas de Adesão Celular/metabolismo , Condicionamento Clássico/fisiologia , Potenciais Evocados Auditivos do Tronco Encefálico , Medo , Feminino , Reação de Congelamento Cataléptica , Técnicas de Inativação de Genes , Perda Auditiva Neurossensorial/patologia , Perda Auditiva Neurossensorial/fisiopatologia , Masculino , Complexo Mediador/fisiologia , Camundongos , Especificidade de Órgãos , Emissões Otoacústicas Espontâneas , Distribuição Aleatória , Reflexo de SobressaltoRESUMO
The multiligand receptors megalin (Lrp2) and cubilin (Cubn) and their endocytic adaptor protein Dab2 (Dab2) play essential roles in maintaining the integrity of the apical endocytic pathway of proximal tubule (PT) cells and have complex and poorly understood roles in the development of chronic kidney disease. Here, we used RNA-sequencing and CRISPR/Cas9 knockout (KO) technology in a well-differentiated cell culture model to identify PT-specific transcriptional changes that are directly consequent to the loss of megalin, cubilin, or Dab2 expression. KO of Lrp2 had the greatest transcriptional effect, and nearly all genes whose expression was affected in Cubn KO and Dab2 KO cells were also changed in Lrp2 KO cells. Pathway analysis and more granular inspection of the altered gene profiles suggested changes in pathways with immunomodulatory functions that might trigger the pathological changes observed in KO mice and patients with Donnai-Barrow syndrome. In addition, differences in transcription patterns between Lrp2 and Dab2 KO cells suggested the possibility that altered spatial signaling by aberrantly localized receptors contributes to transcriptional changes upon the disruption of PT endocytic function. A reduction in transcripts encoding sodium-glucose cotransporter isoform 2 was confirmed in Lrp2 KO mouse kidney lysates by quantitative PCR analysis. Our results highlight the role of megalin as a master regulator and coordinator of ion transport, metabolism, and endocytosis in the PT. Compared with the studies in animal models, this approach provides a means to identify PT-specific transcriptional changes that are directly consequent to the loss of these target genes.NEW & NOTEWORTHY Megalin and cubilin receptors together with their adaptor protein Dab2 represent major components of the endocytic machinery responsible for efficient uptake of filtered proteins by the proximal tubule (PT). Dab2 and megalin expression have been implicated as both positive and negative modulators of kidney disease. We used RNA sequencing to knock out CRISPR/Cas9 cubilin, megalin, and Dab2 in highly differentiated PT cells to identify PT-specific changes that are directly consequent to knockout of each component.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Proteína 9 Associada à CRISPR/genética , Sistemas CRISPR-Cas , Técnicas de Inativação de Genes , Túbulos Renais Proximais/metabolismo , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Receptores de Superfície Celular/metabolismo , Transcrição Gênica , Proteínas Adaptadoras de Transdução de Sinal/genética , Agenesia do Corpo Caloso/genética , Agenesia do Corpo Caloso/metabolismo , Agenesia do Corpo Caloso/patologia , Animais , Proteínas Reguladoras de Apoptose/genética , Células Cultivadas , Bases de Dados Genéticas , Redes Reguladoras de Genes , Perda Auditiva Neurossensorial/genética , Perda Auditiva Neurossensorial/metabolismo , Perda Auditiva Neurossensorial/patologia , Hérnias Diafragmáticas Congênitas/genética , Hérnias Diafragmáticas Congênitas/metabolismo , Hérnias Diafragmáticas Congênitas/patologia , Humanos , Túbulos Renais Proximais/patologia , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Masculino , Camundongos Knockout , Monodelphis , Miopia/genética , Miopia/metabolismo , Miopia/patologia , Proteinúria/genética , Proteinúria/metabolismo , Proteinúria/patologia , Receptores de Superfície Celular/genética , Erros Inatos do Transporte Tubular Renal/genética , Erros Inatos do Transporte Tubular Renal/metabolismo , Erros Inatos do Transporte Tubular Renal/patologiaRESUMO
Perception of sound is initiated by mechanically gated ion channels at the tips of stereocilia. Mature mammalian auditory hair cells require transmembrane channel-like 1 (TMC1) for mechanotransduction, and mutations of the cognate genetic sequences result in dominant or recessive heritable deafness forms in humans and mice. In contrast, zebrafish lateral line hair cells, which detect water motion, require Tmc2a and Tmc2b. Here, we use standard and multiplex genome editing in conjunction with functional and behavioral assays to determine the reliance of zebrafish hearing and vestibular organs on Tmc proteins. Surprisingly, our approach using multiple mutant alleles demonstrates that hearing in zebrafish is not dependent on Tmc1, nor is it fully dependent on Tmc2a and Tmc2b. Hearing however is absent in triple-mutant zebrafish that lack Tmc1, Tmc2a and Tmc2b. These outcomes reveal a striking resemblance of Tmc protein reliance in the vestibular sensory epithelia of mammals to the maculae of zebrafish. Moreover, our findings disclose a logic of Tmc use where hearing depends on a complement of Tmc proteins beyond those employed to sense water motion.