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Browsing by Author "Holt, Jeffrey R."
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Item Functional development of mechanosensitive hair cells in stem cell-derived organoids parallels native vestibular hair cells(Nature Publishing Group, 2016-05-24) Liu, Xiao-Ping; Koehler, Karl R.; Mikosz, Andrew M.; Hashino, Eri; Holt, Jeffrey R.; Department of Otolaryngology—Head & Neck Surgery, IU School of MedicineInner ear sensory epithelia contain mechanosensitive hair cells that transmit information to the brain through innervation with bipolar neurons. Mammalian hair cells do not regenerate and are limited in number. Here we investigate the potential to generate mechanosensitive hair cells from mouse embryonic stem cells in a three-dimensional (3D) culture system. The system faithfully recapitulates mouse inner ear induction followed by self-guided development into organoids that morphologically resemble inner ear vestibular organs. We find that organoid hair cells acquire mechanosensitivity equivalent to functionally mature hair cells in postnatal mice. The organoid hair cells also progress through a similar dynamic developmental pattern of ion channel expression, reminiscent of two subtypes of native vestibular hair cells. We conclude that our 3D culture system can generate large numbers of fully functional sensory cells which could be used to investigate mechanisms of inner ear development and disease as well as regenerative mechanisms for inner ear repair.Item Generation of inner ear organoids with functional hair cells from human pluripotent stem cells(Nature Publishing group, 2017-06) Koehler, Karl R.; Nie, Jing; Longworth-Mills, Emma; Liu, Xiao-Ping; Lee, Jiyoon; Holt, Jeffrey R.; Hashino, Eri; Otolaryngology -- Head and Neck Surgery, School of MedicineHuman inner ear tissue derived from pluripotent stem cells could provide a powerful platform for drug discovery or a source of sound- or motion-sensing cells for patients with hearing loss or balance dysfunction. Here we report a method for differentiating human pluripotent stem cells to inner ear organoids that harbor functional hair cells. Using a three-dimensional culture system, we modulate TGF, BMP, FGF, and Wnt signaling to generate multiple otic vesicle–like structures from a single stem-cell aggregate. Over two months, the vesicles develop into inner ear organoids with sensory epithelia that are innervated by sensory neurons. Additionally, using CRISPR/Cas9, we generate an ATOH1-2A-eGFP cell line to detect hair cell induction and demonstrate that derived hair cells exhibit electrophysiological properties similar to those of native sensory hair cells. Our culture system will be useful for elucidating mechanisms of human inner ear development and testing potential inner ear therapies.