Hearing loss is a prevalent disorder that affects people of all ages. On top of the existing hearing aids and cochlear implants, there is a growing effort to regenerate functional tissues and restore hearing. However, studying and evaluating these regenerative medicine approaches in a big animal model (e.g. pigs) whose anatomy, physiology, and organ size are similar to a human is challenging. In big animal models, the cochlea is bulky, intricate, and veiled in a dense and craggy otic capsule. These facts complicate 3D microscopic analysis that is vital in the cochlea, where structure-function relation is time and again manifested. To allow 3D imaging of an intact cochlea of newborn and juvenile pigs with a volume up to ∼ 250 mm³, we adapted the BoneClear tissue clearing technique, which renders the bone transparent. The transparent cochleae were then imaged with cellular resolution and in a timely fashion, which prevented bubble formation and tissue degradation, using an adaptive custom-built light-sheet fluorescence microscope. The adaptive light-sheet microscope compensated for deflections of the illumination beam by changing the angles of the beam and translating the detection objective while acquiring images. Using this combination of techniques, macroscopic and microscopic properties of the cochlea were extracted, including the density of hair cells, frequency maps, and lower frequency limits. Consequently, the proposed platform could support the growing effort to regenerate cochlear tissues and assist with basic research to advance cures for hearing impairments.

中文翻译：

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使用组织透明和定制光片显微镜对完整猪耳蜗进行三维成像


听力损失是一种影响所有年龄段人群的普遍疾病。除了现有的助听器和人工耳蜗之外，人们还在不断努力再生功能组织和恢复听力。然而，在解剖学、生理学和器官大小与人类相似的大型动物模型（例如猪）中研究和评估这些再生医学方法具有挑战性。在大型动物模型中，耳蜗体积庞大、错综复杂，并覆盖在致密且崎岖不平的耳囊中。这些事实使 3D 显微分析变得复杂，而该分析对于耳蜗至关重要，结构与功能关系在耳蜗中不断显现。为了对体积高达 ∼ 250 mm ³的新生猪和幼猪的完整耳蜗进行 3D 成像，我们采用了 BoneClear 组织透明技术，使骨骼透明。然后使用自适应定制光片荧光显微镜对透明耳蜗进行细胞分辨率及时成像，防止气泡形成和组织降解。自适应光片显微镜通过在采集图像时改变光束角度和平移检测物镜来补偿照明光束的偏转。利用这种技术组合，提取了耳蜗的宏观和微观特性，包括毛细胞的密度、频率图和频率下限。因此，拟议的平台可以支持日益增长的耳蜗组织再生努力，并协助基础研究以推进听力障碍的治疗。