Improved TMC1 gene therapy restores hearing and balance in mice with genetic inner ear disorders.
Ontology highlight
ABSTRACT:
SUBMITTER: Nist-Lund CA
PROVIDER: S-EPMC6342993 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The diagnostics of inner ear diseases are primarily functional, but there is a growing interest in inner ear biomarkers. The present scoping review aimed to elucidate gaps in the literature regarding the definition, classification system, and an overview of the potential uses of inner ear biomarkers. Relevant biomarkers were categorized, and their possible benefits were evaluated. The databases OVID Medline, EMBASE, EBSCO COINAHL, CA PLUS, WOS BIOSIS, WOS Core Collection, Proquest Dissertations, Theses Global, PROSPERO, Cochrane Library, and BASE were searched using the keywords "biomarker" and "inner ear". Of the initially identified 1502 studies, 34 met the inclusion criteria. The identified biomarkers were classified into diagnostic, prognostic, therapeutic, and pathognomonic; many were detected only in the inner ear or temporal bone. The inner-ear-specific biomarkers detected in peripheral blood included otolin-1, prestin, and matrilin-1. Various serum antibodies correlated with inner ear diseases (e.g., anti-type II collagen, antinuclear antibodies, antibodies against cytomegalovirus). Further studies are advised to elucidate the clinical significance and diagnostic or prognostic usage of peripheral biomarkers for inner ear disorders, filling in the literature gaps with biomarkers pertinent to the otology clinical practice and integrating functional and molecular biomarkers. These may be the building blocks toward a well-structured guideline for diagnosing and managing some audio-vestibular disorders.
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Project description:Usher syndrome is a syndromic form of hereditary hearing impairment that includes sensorineural hearing loss and delayed-onset retinitis pigmentosa (RP). Type 1 Usher syndrome (USH1) is characterized by congenital profound sensorineural hearing impairment and vestibular areflexia, with adolescent-onset RP. Systemic treatment with antisense oligonucleotides (ASOs) targeting the human USH1C c.216G>A splicing mutation in a knockin mouse model of USH1 restores hearing and balance. Herein, we explore the effect of delivering ASOs locally to the ear to treat hearing and vestibular dysfunction associated with Usher syndrome. Three localized delivery strategies were investigated in USH1C mice: inner ear injection, trans-tympanic membrane injection, and topical tympanic membrane application. We demonstrate, for the first time, that ASOs delivered directly to the ear correct Ush1c expression in inner ear tissue, improve cochlear hair cell transduction currents, restore vestibular afferent irregularity, spontaneous firing rate, and sensitivity to head rotation, and successfully recover hearing thresholds and balance behaviors in USH1C mice. We conclude that local delivery of ASOs to the middle and inner ear reach hair cells and can rescue both hearing and balance. These results also demonstrate the therapeutic potential of ASOs to treat hearing and balance deficits associated with Usher syndrome and other ear diseases.
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Project description:Recent work has demonstrated that transmembrane channel-like 1 protein (TMC1) is an essential component of the sensory transduction complex in hair cells of the inner ear. A closely related homolog, TMC2, is expressed transiently in the neonatal mouse cochlea and can enable sensory transduction in Tmc1-null mice during the first postnatal week. Both TMC1 and TMC2 are expressed at adult stages in mouse vestibular hair cells. The extent to which TMC1 and TMC2 can substitute for each other is unknown. Several biophysical differences between TMC1 and TMC2 suggest these proteins perform similar but not identical functions. To investigate these differences, and whether TMC2 can substitute for TMC1 in mature hair cells, we generated a knock-in mouse model allowing Cre-inducible expression of Tmc2. We assayed for changes in hair cell sensory transduction and auditory and vestibular function in Tmc2 knockin mice (Tm[Tmc2]) in the presence or absence of endogenous Tmc1, Tmc2 or both. Our results show that expression of Tm[TMC2] restores sensory transduction in vestibular hair cells and transiently in cochlear hair cells in the absence of TMC1. The cellular rescue leads to recovery of balance but not auditory function. We conclude that TMC1 provides some additional necessary function, not provided by TMC2.
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Project description:Purpose: To identify orthologous genes in Xenopus that are implicated in deafness and vestibular disorders in humans and to compare RNA-Seq and microarray to determine the advantages of each technology for Xenopus transcriptomic analysis. Methods: Inner ear RNA from X. laevis stages 56-58 was isolated with the Qiagen® RNeasy® Mini Kit. RNA quality was assessed using the Agilent 2100 Bioanalyzer. The RNA was sent to the National Center for Genome Resources, for Illumina-Solexa sequencing using the genome analyzer II and to MIT for microarray processing usign the Affymetrix GeneChip® X. laevis Genome 2.0 Array. The OMIM database was mined for identification of genes causing deafness and vestibular disorder in humans. Human sequences for each of the deafness and vestibular disorders genes were downloaded from Ensembl. Blast-2.2.27+ was used to mapped the human sequences against X. tropicalis predicted proteins from JGI or to Xenopus consensus sequences downloaded from Affymetrix. The alignment of the human sequences to the predicted proteins resulted in the identification of the protein designation number. The protein designation number was used to obtain the sacaffold number and coordinates from JGI genome browser. The scaffold number and coordinates were input into Alpheus sequence variant detection pipeline to obtain the number of reads associated with each gene. The reads were log2 transformed for comparison to the microarray. In the case of the alignment of the human sequences to the Xenopus consensus sequences resulted in the PSID. The PSID was used to obtain the GCRMA intensity value. The GCRMA value and the number of reads were compared to determine which technology detected more genes. Results: Using Affymetrix GeneChip® X. laevis Genome 2.0 Arrays and Illumina-Solexa sequencing methods, we determined that the transcriptional profile of the Xenopus inner ear comprises hundreds of genes that are orthologous to OMIM® genes implicated in deafness and vestibular disorders in humans. Analysis of genes that mapped to both technologies showed that RNA-Seq detected more of both deafness and vestibular disorder than the Microarray. RNA-Seq and microarray detected 108 genes but RNA-Seq detected 48 genes that were below threshold criteria for microarray. While the microarray detected 11 genes that were below the expression criteria for RNA-Seq and 23 genes were below the threshold criteria for both technologies. Further analysis of the 108 genes detected by both technologies showed a minor correlation between the two technologies. Conclusions: As part of this study we identified candidate scaffold regions of the Xenopus tropicalis genome that can be used for gene mutagenesis. Furthermore, results from this study showed that the two technologies can complement each other and that a combination of both approaches can provide a more complete analysis of gene expression than either method alone. Inner ear RNA from X. laevis larval stages 56-58 was isolated and shipped to the National Center for Genome Resources, for Illumina-Solexa sequencing or to the Massachusetts Institute of Technology BioMicro Center for microarray analysis with the Affymetrix GeneChip® X. laevis Genome 2.0 Array. RNA-Sequencing was completed using the Illumina-Solexa platform for sequencing by synthesis. Short-insert paired end (SIPE) libraries were prepared from total RNA according to Illuminaâ??s mRNA-Seq Sample Prep Protocol v2.0 (Illumina, San Diego, CA, USA). The resultant double-stranded cDNA concentration was measured on a NanoDrop spectrophotometer, and size and purity were determined on the 2100 Bioanalyzer using a DNA 1000 Nano kit. The cDNA libraries were cluster amplified on Illumina flowcells, sequenced on the GAII Sequencer as 36-cycle single-end reads, and processed using Illumina software v1.0. Illumina reads were aligned to the X. tropicalis genome using the algorithm for genomic mapping and alignment program (GMAP) and Alpheus® Sequence Variant Detection System v3.1.
2016-02-28 | E-GEOD-69701 | biostudies-arrayexpress
Project description:Auditory and vestibular disorders are prevalent sensory disabilities caused by genetic and environmental (noise, trauma, chemicals) factors that often damage mechanosensory hair cells of the inner ear. Development of treatments for inner ear disorders of hearing and balance relies on the use of animal models such as fish, amphibians, reptiles, birds, and non-human mammals. Here, we aimed to augment the utility of the genus Xenopus for uncovering genetic mechanisms essential for the maintenance of inner ear structure and function.Using Affymetrix GeneChip(®) X. laevis Genome 2.0 Arrays and Illumina-Solexa sequencing methods, we determined that the transcriptional profile of the Xenopus laevis inner ear comprises hundreds of genes that are orthologous to OMIM(®) genes implicated in deafness and vestibular disorders in humans. Analysis of genes that mapped to both technologies demonstrated that, with our methods, a combination of microarray and RNA-Seq detected expression of more genes than either platform alone.As part of this study we identified candidate scaffold regions of the Xenopus tropicalis genome that can be used to investigate hearing and balance using genetic and informatics procedures that are available through the National Xenopus Resource (NXR), and the open access data repository, Xenbase. The results and approaches presented here expand the viability of Xenopus as an animal model for inner ear research.
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Project description:Purpose: To identify orthologous genes in Xenopus that are implicated in deafness and vestibular disorders in humans and to compare RNA-Seq and microarray to determine the advantages of each technology for Xenopus transcriptomic analysis. Methods: Inner ear RNA from X. laevis stages 56-58 was isolated with the Qiagen® RNeasy® Mini Kit. RNA quality was assessed using the Agilent 2100 Bioanalyzer. The RNA was sent to the National Center for Genome Resources, for Illumina-Solexa sequencing using the genome analyzer II and to MIT for microarray processing usign the Affymetrix GeneChip® X. laevis Genome 2.0 Array. The OMIM database was mined for identification of genes causing deafness and vestibular disorder in humans. Human sequences for each of the deafness and vestibular disorders genes were downloaded from Ensembl. Blast-2.2.27+ was used to mapped the human sequences against X. tropicalis predicted proteins from JGI or to Xenopus consensus sequences downloaded from Affymetrix. The alignment of the human sequences to the predicted proteins resulted in the identification of the protein designation number. The protein designation number was used to obtain the sacaffold number and coordinates from JGI genome browser. The scaffold number and coordinates were input into Alpheus sequence variant detection pipeline to obtain the number of reads associated with each gene. The reads were log2 transformed for comparison to the microarray. In the case of the alignment of the human sequences to the Xenopus consensus sequences resulted in the PSID. The PSID was used to obtain the GCRMA intensity value. The GCRMA value and the number of reads were compared to determine which technology detected more genes. Results: Using Affymetrix GeneChip® X. laevis Genome 2.0 Arrays and Illumina-Solexa sequencing methods, we determined that the transcriptional profile of the Xenopus inner ear comprises hundreds of genes that are orthologous to OMIM® genes implicated in deafness and vestibular disorders in humans. Analysis of genes that mapped to both technologies showed that RNA-Seq detected more of both deafness and vestibular disorder than the Microarray. RNA-Seq and microarray detected 108 genes but RNA-Seq detected 48 genes that were below threshold criteria for microarray. While the microarray detected 11 genes that were below the expression criteria for RNA-Seq and 23 genes were below the threshold criteria for both technologies. Further analysis of the 108 genes detected by both technologies showed a minor correlation between the two technologies. Conclusions: As part of this study we identified candidate scaffold regions of the Xenopus tropicalis genome that can be used for gene mutagenesis. Furthermore, results from this study showed that the two technologies can complement each other and that a combination of both approaches can provide a more complete analysis of gene expression than either method alone.
2016-02-28 | GSE69701 | GEO
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Project description:The proteins that form the permeation pathway of mechanosensory transduction channels in inner-ear hair cells have not been definitively identified. Genetic, anatomical, and physiological evidence support a role for transmembrane channel-like protein (TMC) 1 in hair cell sensory transduction, yet the molecular function of TMC proteins remains unclear. Here, we provide biochemical evidence suggesting TMC1 assembles as a dimer, along with structural and sequence analyses suggesting similarity to dimeric TMEM16 channels. To identify the pore region of TMC1, we used cysteine mutagenesis and expressed mutant TMC1 in hair cells of Tmc1/2-null mice. Cysteine-modification reagents rapidly and irreversibly altered permeation properties of mechanosensory transduction. We propose that TMC1 is structurally similar to TMEM16 channels and includes ten transmembrane domains with four domains, S4-S7, that line the channel pore. The data provide compelling evidence that TMC1 is a pore-forming component of sensory transduction channels in auditory and vestibular hair cells.
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