Cellular therapy for the inner ear

内耳细胞疗法

• 批准号: 8984514
• 负责人: Albert Edge
• 金额: $ 49.24万
• 依托单位: MASSACHUSETTS EYE AND EAR INFIRMARY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-12 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8984514
• 关键词: Acoustic Nerve; Acoustics; Action Potentials; Adult; Afferent Neurons; Animals; Auditory; Auditory Brainstem Responses; Auditory system; Behavior assessment; Behavioral; Bilateral; Biological Models; Biological Preservation; Brain; CNS processing; Cell Death; Cell Therapy; Cells; Chemicals; Cochlea; Complex; Denervation; Detection; Discrimination; Doxycycline; Ear; Excision; Functional disorder; Genes; Genetic; Glutamates; Goals; Grant; Hair Cells; Hearing; Immunohistochemistry; In Situ; In Vitro; Inferior Colliculus; Labyrinth; Lead; Measurement; Measures; Midbrain structure; Modeling; Morphology; Mus; Natural regeneration; Nerve; Nerve Degeneration; Nerve Regeneration; Nervous system structure; Neuronal Differentiation; Neurons; Noise; Organ of Corti; Ouabain; Outer Hair Cells; Perception; Peripheral; Process; Reading; Recovery; Recovery of Function; Role; Route; Schwann Cells; Sensorineural Hearing Loss; Signal Transduction; Small Interfering RNA; Stem cell transplant; Stem cells; Stimulus; Synapses; System; Testing; Therapeutic; Time; Tissues; Toxic effect; Up-Regulation; Work; axon growth; axonal guidance; base; behavior measurement; behavior test; deafness; embryonic stem cell; ganglion cell; hearing impairment; improved; in vivo; in vivo Model; inhibitor/antagonist; kainate; mouse model; neogenin; nerve stem cell; nerve supply; neural circuit; neurogenesis; neuron loss; neurotrophic factor; novel; novel strategies; otoacoustic emission; patch clamp; postsynaptic; progenitor; promoter; public health relevance; receptor; reconstitution; reinnervation; relating to nervous system; repaired; research study; response; restoration; sound; spiral ganglion; success; synaptic function; synaptogenesis; transdifferentiation

 DESCRIPTION (provided by applicant): Degeneration of neural circuitry, either through synaptic loss or neural cell death, is an important cause of nervous system dysfunction, and afferent auditory nerve loss leading to deafness is a manifestation of these processes occurring in the cochlea. Our previous work has shown that neurons from transplanted stem cells can reinnervate hair cells of the organ of Corti after chemical de-afferentation, suggesting that this neural circuit can be reconstituted. In these studies we ask questions about repair of both types of neural degeneration: first, we investigate whether reinnervation and synaptogenesis with hair cells can be increased by manipulating axonal guidance and neurotrophic factors in in vitro and in vivo models of synapse damage; second, we study the effect of progenitor cell transdifferentiation to neurons and the regeneration of this circuit in a model of complete afferent neuron loss. Measurements of synaptic and neural repair include electrophysiological and histological analysis of new synapses in vitro and immunohistochemical as well as peripheral, central, and behavioral assessments of auditory function in vivo. Understanding the mechanisms underlying re-formation of neural connections to hair cells in the adult ear is important to therapeutic approaches for the treatment of neural dysfunction that causes hearing loss. The Specific Aims comprise three inter-related experiments to probe key variables likely to influence the success of peripheral and central reinnervation of the auditory system. In Aim 1 we assess the effect of inhibitors of axonal guidance as well as promoters of axonal growth on the reinnervation of hair cells in cochlear models with loss of the afferent synapse but preservation of ganglion cells. In the in vitro system (afferent synapses lost due to kainate administration) we use electrophysiological measurements to assess synaptic function: firing of action potentials and excitatory postsynaptic currents by the neurons after stimulation of hair cells, and inhibition of this synaptic function by pharmacological blockers of glutamatergic synapses. In the in vivo system (noise damage causing afferent synapse loss) metrics include synaptic analysis as well as auditory brainstem response and distortion product otoacoustic emissions. Assessments are based on the morphology and number of synapses formed with inner and outer hair cells determined immunohistochemically. In Aim 2 we define the roles and optimal expression levels of Sox2 as well as Ngn1 for Schwann cell to neuronal conversion. We have isolated progenitor cells that can be converted to neurons from spiral ganglion, and we have shown that these cells arise from Schwann cells. We test the role of Sox2 and Ngn1 in this conversion. We assess conversion of Schwann cells to neurons in vivo using ouabain to produce a model of auditory nerve damage, and we stimulate the conversion of endogenous cells to spiral ganglion cells by modulating expression of the same genes. In Aim 3 we record the spiking activity from single neurons in the auditory midbrain to sound stimuli delivered to the reinnervated and untreated ears. We compare markers of neural recovery to behavioral measures of improved sound discrimination.

 描述（由申请人提供）：通过突触丧失或神经细胞死亡导致的神经回路退化是神经系统功能障碍的重要原因，导致耳聋的传入听觉神经丧失是这些过程在耳蜗中发生的表现。我们以前的工作表明，来自移植干细胞的神经元可以在化学去传入后重新支配Corti器官的毛细胞，这表明这种神经回路可以重建。在这些研究中，我们问的问题，这两种类型的神经退行性变的修复：首先，我们调查是否可以通过操纵轴突的指导和神经营养因子在体外和体内模型的突触损伤的毛细胞的神经再支配和突触发生增加;第二，我们研究祖细胞转分化为神经元和再生的影响，在模型中的传入神经元的完全损失，这个电路。突触和神经修复的测量包括体外新突触的电生理学和组织学分析，以及体内听觉功能的外周、中枢和行为评估。了解成年人耳毛细胞神经连接重建的机制对于治疗导致听力损失的神经功能障碍的治疗方法非常重要。具体目标包括三个相互关联的实验，以探讨可能影响听觉系统外周和中枢神经再支配成功的关键变量。在目的1中，我们评估了轴突导向抑制剂和轴突生长促进剂对耳蜗模型中毛细胞神经再支配的影响，其中耳蜗模型中传入突触丢失但神经节细胞保留。在体外系统中（由于红藻氨酸盐给药导致传入突触丢失）， 使用电生理学测量来评估突触功能：刺激毛细胞后神经元的动作电位和兴奋性突触后电流的放电，以及抑制 这种突触功能的药理学阻断剂的突触。在体内系统中（噪声损伤导致传入突触损失），指标包括突触分析以及听觉脑干反应和畸变产物耳声发射。评估是基于化学测定的与内毛细胞和外毛细胞形成的突触的形态和数量。在目的2中，我们定义了Sox 2和Ngn 1在雪旺细胞向神经元转化中的作用和最佳表达水平。我们已经从螺旋神经节中分离出了可以转化为神经元的祖细胞，我们已经证明这些细胞来自雪旺细胞。我们测试的作用，Sox 2和Ngn 1在这种转换。我们使用哇巴因评估体内雪旺细胞向神经元的转化以产生听神经损伤模型，并且我们通过调节相同基因的表达来刺激内源性细胞向螺旋神经节细胞的转化。在目标3中，我们记录了听觉中脑中单个神经元对传递到再神经支配和未处理的耳朵的声音刺激的尖峰活动。我们比较了神经恢复的标志物，以改善声音辨别的行为措施。