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      Cell-Specific Transcriptome Analysis Shows That Adult Pillar and Deiters' Cells Express Genes Encoding Machinery for Specializations of Cochlear Hair Cells

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          Abstract

          The mammalian auditory sensory epithelium, the organ of Corti, is composed of hair cells and supporting cells. Hair cells contain specializations in the apical, basolateral and synaptic membranes. These specializations mediate mechanotransduction, electrical and mechanical activities and synaptic transmission. Supporting cells maintain homeostasis of the ionic and chemical environment of the cochlea and contribute to the stiffness of the cochlear partition. While spontaneous proliferation and transdifferentiation of supporting cells are the source of the regenerative response to replace lost hair cells in lower vertebrates, supporting cells in adult mammals no longer retain that capability. An important first step to revealing the basic biological properties of supporting cells is to characterize their cell-type specific transcriptomes. Using RNA-seq, we examined the transcriptomes of 1,000 pillar and 1,000 Deiters' cells, as well as the two types of hair cells, individually collected from adult CBA/J mouse cochleae using a suction pipette technique. Our goal was to determine whether pillar and Deiters' cells, the commonly targeted cells for hair cell replacement, express the genes known for encoding machinery for hair cell specializations in the apical, basolateral, and synaptic membranes. We showed that both pillar and Deiters' cells express these genes, with pillar cells being more similar to hair cells than Deiters' cells. The fact that adult pillar and Deiters' cells express the genes cognate to hair cell specializations provides a strong molecular basis for targeting these cells for mammalian hair cell replacement after hair cells are lost due to damage.

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          Prestin is the motor protein of cochlear outer hair cells.

          The outer and inner hair cells of the mammalian cochlea perform different functions. In response to changes in membrane potential, the cylindrical outer hair cell rapidly alters its length and stiffness. These mechanical changes, driven by putative molecular motors, are assumed to produce amplification of vibrations in the cochlea that are transduced by inner hair cells. Here we have identified an abundant complementary DNA from a gene, designated Prestin, which is specifically expressed in outer hair cells. Regions of the encoded protein show moderate sequence similarity to pendrin and related sulphate/anion transport proteins. Voltage-induced shape changes can be elicited in cultured human kidney cells that express prestin. The mechanical response of outer hair cells to voltage change is accompanied by a 'gating current', which is manifested as nonlinear capacitance. We also demonstrate this nonlinear capacitance in transfected kidney cells. We conclude that prestin is the motor protein of the cochlear outer hair cell.
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            Principal component analysis for clustering gene expression data.

            There is a great need to develop analytical methodology to analyze and to exploit the information contained in gene expression data. Because of the large number of genes and the complexity of biological networks, clustering is a useful exploratory technique for analysis of gene expression data. Other classical techniques, such as principal component analysis (PCA), have also been applied to analyze gene expression data. Using different data analysis techniques and different clustering algorithms to analyze the same data set can lead to very different conclusions. Our goal is to study the effectiveness of principal components (PCs) in capturing cluster structure. Specifically, using both real and synthetic gene expression data sets, we compared the quality of clusters obtained from the original data to the quality of clusters obtained after projecting onto subsets of the principal component axes. Our empirical study showed that clustering with the PCs instead of the original variables does not necessarily improve, and often degrades, cluster quality. In particular, the first few PCs (which contain most of the variation in the data) do not necessarily capture most of the cluster structure. We also showed that clustering with PCs has different impact on different algorithms and different similarity metrics. Overall, we would not recommend PCA before clustering except in special circumstances.
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              Auditory hair cell replacement and hearing improvement by Atoh1 gene therapy in deaf mammals.

              In the mammalian auditory system, sensory cell loss resulting from aging, ototoxic drugs, infections, overstimulation and other causes is irreversible and leads to permanent sensorineural hearing loss. To restore hearing, it is necessary to generate new functional hair cells. One potential way to regenerate hair cells is to induce a phenotypic transdifferentiation of nonsensory cells that remain in the deaf cochlea. Here we report that Atoh1, a gene also known as Math1 encoding a basic helix-loop-helix transcription factor and key regulator of hair cell development, induces regeneration of hair cells and substantially improves hearing thresholds in the mature deaf inner ear after delivery to nonsensory cells through adenovectors. This is the first demonstration of cellular and functional repair in the organ of Corti of a mature deaf mammal. The data suggest a new therapeutic approach based on expressing crucial developmental genes for cellular and functional restoration in the damaged auditory epithelium and other sensory systems.
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                Author and article information

                Contributors
                Journal
                Front Mol Neurosci
                Front Mol Neurosci
                Front. Mol. Neurosci.
                Frontiers in Molecular Neuroscience
                Frontiers Media S.A.
                1662-5099
                01 October 2018
                2018
                : 11
                : 356
                Affiliations
                [1] 1Department of Biomedical Sciences, Creighton University School of Medicine , Omaha, NE, United States
                [2] 2Chongqing Academy of Animal Science , Chongqing, China
                [3] 3Department of Otorhinolaryngology-Head and Neck Surgery, Beijing Tonren Hospital, Capital Medical University , Beijing, China
                [4] 4Department of Otolaryngology-Head and Neck Surgery, University of Nebraska Medical Center , Omaha, NE, United States
                Author notes

                Edited by: Albert Quintana, Autonomous University of Barcelona, Spain

                Reviewed by: Agnieszka J. Szczepek, Charité Universitätsmedizin Berlin, Germany; Sergi Simó, University of California, Davis, United States

                *Correspondence: David Z. Z. He hed@ 123456creighton.edu
                Article
                10.3389/fnmol.2018.00356
                6174830
                30327589
                e5c69ff1-75d3-4c60-a1e9-a678fbb074d9
                Copyright © 2018 Liu, Chen, Giffen, Stringham, Li, Judge, Beisel and He.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 24 May 2018
                : 11 September 2018
                Page count
                Figures: 9, Tables: 1, Equations: 0, References: 64, Pages: 17, Words: 10439
                Funding
                Funded by: National Institute on Deafness and Other Communication Disorders 10.13039/100000055
                Award ID: R01 DC016807
                Funded by: National Natural Science Foundation of China 10.13039/501100001809
                Award ID: 81600798
                Award ID: 81770996
                Award ID: 31771376
                Categories
                Neuroscience
                Original Research

                Neurosciences
                hair cell,pillar cell,deiters' cell,transcriptome,rna-seq,mouse
                Neurosciences
                hair cell, pillar cell, deiters' cell, transcriptome, rna-seq, mouse

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