Single cell RNA-Seq reveals Fgf signaling dynamics during sensory hair cell regeneration
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ABSTRACT:
ORGANISM(S): Danio rerio
PROVIDER: GSE123241 | GEO | 2018/12/03
REPOSITORIES: GEO
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Project description:Deafness due to the terminal loss of inner ear hair cells is one of the most common sensory diseases. However, non-mammalian animals (e.g. birds, amphibian and fish) regenerate damaged hair cells. In order to better understand the reasons underpinning such regeneration disparities in vertebrates, we set out to define the changes in gene expression associated with the regeneration of hair cells in the zebrafish lateral line at high resolution. We performed RNA-Seq analyses on regenerating support cells purified by fluorescence activated cell sorting (FACS). The zebrafish lateral line provides an experimentally accessible system to define the complex signaling events triggered by injury and regeneration, because these cells can be acutely killed by exposure to neomycin, after which they regenerate rapidly. Lateral line hair cells are located in the center of a mechanosensory organ known as the neuromast and are surrounded by inner support cells and an outer ring of mantle cells. Tg(sqET20) larvae express GFP strongly in mantle cells and to a lesser degree in inner support cells. We isolated GFP positive and GFP negative cells from 5 days post fertilization (dpf) Tg(sqET20) larvae at 1, 3 and 5 hours post neomycin treatment, as well as from a non-treated control. Transgenic zebrafish Tg(sqET20) larvae at 5 days post fertilization were exposed to neomycin, dissociated, and FACS sorted into GFP positive and GFP negative populations at 1, 3, and 5 hours following treatment, along with a mock treated 1 hr control. The experiment was performed in triplicate, for a total of 24 samples.
2014-03-26 | E-GEOD-56176 | biostudies-arrayexpress
Project description:Deafness due to the terminal loss of inner ear hair cells is one of the most common sensory diseases. However, non-mammalian animals (e.g. birds, amphibian and fish) regenerate damaged hair cells. In order to better understand the reasons underpinning such regeneration disparities in vertebrates, we set out to define the changes in gene expression associated with the regeneration of hair cells in the zebrafish lateral line at high resolution. We performed RNA-Seq analyses on regenerating support cells purified by fluorescence activated cell sorting (FACS). The zebrafish lateral line provides an experimentally accessible system to define the complex signaling events triggered by injury and regeneration, because these cells can be acutely killed by exposure to neomycin, after which they regenerate rapidly. Lateral line hair cells are located in the center of a mechanosensory organ known as the neuromast and are surrounded by inner support cells and an outer ring of mantle cells. Tg(sqET20) larvae express GFP strongly in mantle cells and to a lesser degree in inner support cells. We isolated GFP positive and GFP negative cells from 5 days post fertilization (dpf) Tg(sqET20) larvae at 1, 3 and 5 hours post neomycin treatment, as well as from a non-treated control.
2014-03-26 | GSE56176 | GEO
Project description:Mammalian inner ear and fish lateral line sensory hair cells depend on fluid motion to transduce environmental signals and elicit a response. In mammals, actively maintained ionic homeostasis of the cochlear and vestibular fluid (endolymph) is essential for hair cell function and numerous mammalian hearing and vestibular disorders arise from disrupted endolymph ion homeostasis. Lateral line hair cells, however, are openly exposed to the aqueous environment with fluctuating ionic composition. How sensory transduction in the lateral line is maintained during environmental changes of ionic composition is not fully understood. Using lineage labeling, in vivo time lapse imaging and scRNA-seq, we discovered highly motile skin-derived cells that invade mature mechanosensory organs of the zebrafish lateral line and differentiate into Neuromast-associated (Nm) ionocytes. Furthermore, the invasive behavior is adaptive as it is triggered by drastic fluctuations in environmental stimuli. Our findings challenge the notion of an entirely placodally-derived lateral line and identify Nm ionocytes as likely regulators of mechanosensory hair cell function possibly by modulating the ionic microenvironment. The discovery of lateral line ionocytes provides an experimentally accessible in vivo system to study cell invasion and migration, as well as the physiological adaptation of vertebrate organs to changing environmental conditions.
2021-03-11 | GSE168695 | GEO
Project description:A major cause of human deafness and vestibular dysfunction is permanent loss of the mechanosensory hair cells of the inner ear. In non-mammalian vertebrates such as zebrafish, regeneration of missing hair cells can occur throughout life. While a comparative approach has the potential to reveal the basis of such differential regenerative ability, the degree to which the inner ears of fish and mammals share common hair and supporting cell types remains unresolved. Here we perform single-cell RNA sequencing of the zebrafish inner ear at embryonic through adult stages to catalog the diversity of hair and non-sensory supporting cells. We identify a putative progenitor population for hair and supporting cells, as well as distinct hair and supporting cell types in the maculae versus cristae. The hair and supporting cell types differ from those described for the lateral line, a distributed mechanosensory organ in zebrafish in which most studies of hair cell regeneration have been conducted. In the maculae, we identify two subtypes of hair cells that share gene expression with mammalian striolar or extrastriolar hair cells. In situ hybridization reveals that these hair cell subtypes occupy distinct spatial domains within the two major macular organs, the utricle and saccule, consistent with the reported distinct electrophysiological properties of hair cells within these domains. These findings suggest that primitive specialization of spatially distinct striolar and extrastriolar hair cells likely arose in the last common ancestor of fish and mammals. The similarities of inner ear cell type composition between fish and mammals also support using zebrafish as a relevant model for understanding inner ear-specific hair cell function and regeneration.
2023-01-04 | GSE211728 | GEO
Project description:Hearing loss is most commonly caused by the destruction of mechanosensory hair cells in the ear. This condition is usually permanent: Despite the presence of putative hair-cell progenitors in the cochlea, hair cells are not naturally replenished in adult mammals. Unlike those of the mammalian ear, the progenitor cells of nonmammalian vertebrates can regenerate hair cells through- out life. The basis of this difference remains largely unexplored but may lie in molecular dissimilarities that affect how progenitors respond to hair-cell death. We analyzed gene expression in hair-cell progenitors of the lateral-line system. We developed a transgenic line of zebrafish called alpl:mCherry that expresses a red fluorescent protein in the presumptive hair-cell progenitors known as mantle cells. Fluorescence-activated cell sorting from the skins of transgenic larvae, followed by microarray-based expression analysis, revealed a constellation of transcripts that are specifically enriched in these cells versus hair cells and non-fluorescent skin cells. Gene expression analysis after hair-cell ablation uncovered a cohort of genes that are differentially regulated early in regeneration, suggesting possible roles in the response of progen- itors to hair-cell death. These results provide a resource for studying hair-cell regeneration and the biology of sensory progenitor cells.
2014-04-01 | GSE55853 | GEO
Project description:Single cell RNA-seq of SVZ lateral ventricle walls from non-irradiated mice brains and 5 days post-irradiation 4Gy-irradiated mice brains (according to the model SVZ regeneration after a moderate dose of irradiation that we previously described (Daynac et al. 2013). We used this model of SVZ reconstitution to gain insights on the annotation and and temporal ordering of neural progenitors clusters.
2023-03-27 | E-MTAB-12495 | biostudies-arrayexpress
Project description:Sensorineural hearing loss is included in the most common disabilities, and often caused by loss of sensory hair cells in the cochlea. Hair cell regeneration has long been a main target for developing novel therapeutics for sensorineural hearing loss. In the mammalian cochlea, hair cell regeneration occurs in very limited situations, while auditory epithelia of non-mammalians retain the capacity for hair cell regeneration. In the avian basilar papilla, an auditory sensory epithelium, supporting cells, which are sources for regenerated hair cells, are usually quiescent, while hair cell loss induces both direct transdifferentiation of supporting cells and mitotic division of supporting cells. In the present study, we aimed to establish an explant culture model for hair cell regeneration in chick basilar papillae, and validated usefulness of our model to investigate the initial phase of hair cell regeneration. Histological assessments demonstrated that hair cell regeneration via direct transdifferentiation of supporting cells occurred in our model. Labeling assay using 5-ethynyl-2'-deoxyuridine (EdU) revealed the occurrence of mitotic division of supporting cells in the specific location in basilar papillae, while no EdU labeling was identified in newly generated HCs. RNA sequencing indicated alterations in known signaling pathways associated with hair cell regeneration, which is consistent with previous findings. In addition, unbiased analyses of RNA sequencing data indicated novel genes and signaling pathways that could be related to the ignition of supporting cell activation in chick basilar papillae. These results indicate the advantages of our model using explant cultures of chick basilar papillae for exploring molecular mechanisms for hair cell regeneration. Further studies such as single-cell RNA sequencing will allow us to capture the spatiotemporal information by using our explant culture model.
2020-10-23 | GSE154375 | GEO
Project description:The inner ear utilizes sensory hair cells as mechano-electric transducers for sensing sound and balance. In mammals, these hair cells lack the capacity for regeneration. Unlike mammals, hair cells from non-mammalian vertebrates, such as birds, can be regenerated throughout the life of the organism making them a useful model for studying inner ear genetics pathways. The zinc finger transcription factor GATA3 is required for inner ear development and mutations cause sensory neural deafness in humans. In the avian cochlea GATA3 is expressed throughout the sensory epithelia; however, expression is limited to the striola of the utricle. The striola corresponds to an abrupt change in morphologically distinct hair cell types and a 180° shift in hair cell orientation. We used 3 complimentary approaches to identify potential downstream targets of GATA3 in the avian utricle. Specifically we used microarray expression profiling of GATA3 knockdown by siRNA and GATA3 over-expression treatments as well as direct comparisons of GATA3 expressing cells from the striola and non GATA3 expressing cells from the extra-striola. To identify genes that are co-expressed with GATA3 at the striola reversal zone, we compared gene expression in cells micro-dissected from the sensory epithelia of the chick utricle striola to cells from the surrounding extra-striola. There are 2 biological samples and experiments include technical replicates as well as dye-switches for a total of 8 microarrays.
2009-12-24 | E-GEOD-14783 | biostudies-arrayexpress
Project description:Hearing loss is most commonly caused by the destruction of mechanosensory hair cells in the ear. This condition is usually permanent: Despite the presence of putative hair-cell progenitors in the cochlea, hair cells are not naturally replenished in adult mammals. Unlike those of the mammalian ear, the progenitor cells of nonmammalian vertebrates can regenerate hair cells through- out life. The basis of this difference remains largely unexplored but may lie in molecular dissimilarities that affect how progenitors respond to hair-cell death. We analyzed gene expression in hair-cell progenitors of the lateral-line system. We developed a transgenic line of zebrafish called alpl:mCherry that expresses a red fluorescent protein in the presumptive hair-cell progenitors known as mantle cells. Fluorescence-activated cell sorting from the skins of transgenic larvae, followed by microarray-based expression analysis, revealed a constellation of transcripts that are specifically enriched in these cells versus hair cells and non-fluorescent skin cells. Gene expression analysis after hair-cell ablation uncovered a cohort of genes that are differentially regulated early in regeneration, suggesting possible roles in the response of progen- itors to hair-cell death. These results provide a resource for studying hair-cell regeneration and the biology of sensory progenitor cells. Two sets of analyses were performed. The first compared baseline expression levels in four sorted cell types from two different transgenic lines of zebrafish, alpl:mCherry;pou4f3:GFP and alpl:mCherry:ET20. alpl:mCherry;pou4f3:GFP larvae express GFP in hair cells and mCherry in mantle cells. alpl:mCherry:ET20 larvae express GFP in most mantle cells and mChery in all mantle cells. The four cell types compared were GFP+ hair cells, mCherry+ mantle cells, mCherry+/GFP+ mantle cells from alpl:mCherry:ET20 larvae, and non-fluorescent (NF) cells. Two hair-cell, five mCherry+ mantle cell, four mCherry+/GFP+ mantle cell, and six NF cell samples were analyzed. This excludes a few samples that were discardced based on failure to cluster by principal component analysis. A second analysis, separately imported, compared gene expression in mCherry+ mantle cells and NF cells at four time points following chemical ablation of hair cells with copper sulfate. These were one, three, five, and eleven hours after treatment (hpCu). Untreated samples served as controls. For mCherry+ cells, five untreated and four each of 1 hpCu, 3 hpCu, 5 hpCu and 11 hpCu were analyzed. For NF cells, six untreated, seven 1 hpCu, and four each of 3 hpCu, 5 hpCu and 11 hpCu samples were analyzed. Untreated mCherry+ and NF cells were shared between both analyses.
2014-04-01 | E-GEOD-55853 | biostudies-arrayexpress
Project description:Latent transdifferentiation potential exists in supporting cells at neonatal stage in the organ of Corti in mouse, but this plasticity is lost rapidly during the first week of postnatal maturation, leading to the failure of sensory hair cell regeneration. To investigate the molecular mechanism underlying the loss of transdifferentiation potential, we compared epigenetic changes in E17.5, P1 and P6 supporting cells. In E17.5 and P1 supporting cells, hair cell gene enhancers are kept in a primed state (H3K4me1+ H3K27ac-) which can be readily activated during transdifferentiation induced by Notch blocking. As supporting cells mature, a substantial amount of hair cell gene enhancers are decommissioned by H3K4me1, leading to the loss of transdifferentiation potential. In contrast, hair cell gene enhancers retain their commissioned epigenetic status in mature utricular supporting cells, sustaining the transidifferentiation potential in utricular supporting cells in mature animals. Our findings reveal the epigenetic mechanisms underlying the failure of sensory hair cell regeneration in mammalian cochlea.
2021-07-09 | GSE150010 | GEO