Replacement of the cochlear sensory epithelium using stem cell-derived inner ear organoids

摘要 感音神经性听力损失在退伍军人中非常普遍，但目前没有生物治疗方法 从而能够完全恢复耳蜗和听觉。干细胞治疗听力恢复 伟大的承诺。从多能干细胞产生内耳类器官的新进展和我们的研究 使耳蜗适应接受外来细胞的开创性努力已经铺平了前进的道路。然而，在这方面， 在将这些方法结合起来方面仍然存在几个主要障碍。首先，缺乏一个连贯的 因此，我们需要一种策略来识别正确的供体细胞，并根据耳蜗损伤的严重程度对其进行调整。在这 我们专注于干细胞治疗严重受损的耳蜗，其中极端或慢性 损伤已经使Corti器官减少为基质细胞的扁平上皮，迄今为止已经证明这种上皮对 再生的基因方法。这种方法可能是唯一可行的战略， 长期听力损失的老年退伍军人的感觉上皮。开发内耳类器官的方案 从多能干细胞产生潜在的供体细胞所有沿着耳发育轨迹。值得注意的是， 我们已经开发了在这些培养物中从非耳细胞中分离耳囊泡祖细胞的方法，但是 囊泡成熟为感觉上皮细胞需要复杂的商业细胞外基质， 体外高效分化。在这个提议中，我们检验了多能性的otic祖细胞将是 这需要替换严重受损的耳蜗中的毛细胞和支持细胞。提案测试 移植供体细胞在分化的各个阶段的影响，并检查的作用， 细胞外基质促进供体细胞存活和成熟。成功完成这些研究将 为严重听力损失的可行生物治疗开辟了新途径。

ABSTRACT Sensorineural hearing loss is highly prevalent in military Veterans, but there is currently no biological treatment enabling full restoration of the cochlea and hearing perception. Hearing restoration by stem cell therapy holds great promise. New advances in the generation of inner ear organoids from pluripotent stem cells and our pioneering efforts to condition the cochlea for receiving foreign cells have paved the way forward. However, there remain several major obstacles to combining these approaches. Foremost, there is a lack of a coherent strategy for identifying the right donor cells and tailoring these to the severity of injury in the cochlea. In this proposal, we are focused on stem cell therapy for the severely damaged cochlea, where extreme or chronic injury has reduced the organ of Corti to a flat epithelium of stromal cells that have so far proven refractory to genetic approaches to regeneration. This approach may be the only viable strategy for restoring the full sensory epithelium in older Veterans with long-term hearing loss. Protocols for developing inner ear organoids from pluripotent stem cells produce potential donor cells all along the otic developmental trajectory. Notably, we have developed methods to isolate otic vesicle progenitors from non-otic cells in these cultures, but maturation of the vesicles into sensory epithelia requires a complex, commercial extracellular matrix for efficient differentiation in vitro. In this proposal, we test the hypothesis that multipotent otic progenitors will be required to replace both hair cells and supporting cells in the severely damaged cochlea. The proposal tests the impact of implanting donor cells at various stages of differentiation and examines the role of the extracellular matrix in promoting donor cell survival and maturation. Successful completion of these studies will open new paths toward a viable biological treatment for severe hearing loss.