Developing novel stem cell-based approaches to treat hearing loss

该项目的长期目标是开发一种基于干细胞的方法来再生受损的心脏功能。 听觉细胞和突触连接。我们之前的应用重点是使用干细胞再生毛细胞 基于细胞的表观遗传学方法。这项更新建议的主要目标是确定分子 小鼠毛细胞正常发育过程中突触形成的关键机制及新型干细胞的研制 基于细胞的方法重新支配小鼠的听觉毛细胞。毛细胞再生已取得显著进展 进步。与毛细胞再生相比，螺旋神经节神经元(SGN)的再生已有报道，但 结果相对有限。我们已经确定了一种循序渐进的方法来引导小鼠胚胎干细胞(ESCs) 分化为具有SGN特征的干细胞来源的SGN样神经元(ESN)。到目前为止，只有几个 有报道称，干细胞来源的神经元可以与小鼠毛细胞形成神经接触。然而，无论是 这些神经接触是真正的SGN-毛细胞突触尚不清楚。这些神经的功能 在突触水平上的接触还没有确定。再生的关键分子机制 对毛细胞进行功能再支配的神经元仍然不清楚。因此，迫切需要使用eSNs来 重新支配毛细胞，并确定对重新支配至关重要的分子机制。埃弗林B家族蛋白 与EphB受体结合，EphB受体是一类跨膜受体酪氨酸激酶。EPhinB信号转导参与其中 在各种发育项目中，包括突触发生。先前的研究表明，EPhinB1 信号转导对于兴奋性突触的诱导至关重要。我们的初步数据表明EPhinB信号转导的作用 在ESN-毛细胞再神经支配中。根据之前和我们的初步数据，我们假设EphinB 信号可能在调节神经元与感觉毛细胞形成功能性突触的过程中发挥关键作用。至 检验这一假设，提出了三个互补的具体目标。目标1将确定神经元的作用 EfNB1在SGN毛细胞发育过程中突触的形成。目标2将确定分子机制 ESN-毛细胞神经再支配的结果。AIMS 1和2的结果将指导毛细胞突触再生的设计 研究。在目标3中，我们将确定EPhinB1在多大程度上调节ESN毛细胞的再神经支配。 这一提议的完成将确定干细胞来源的神经元 恢复感觉毛细胞的神经。确定Efnb1在毛细胞再神经再生中的作用将指导突触 再生研究，将转化为治疗听力损失患者的临床试验。因此， 这项工作的成果将为探索基于干细胞的多学科方法开辟新的途径 再生毛细胞突触，恢复退伍军人和平民患者的听力功能。

The long-term aim of this project is to develop a stem cell-based approach to regenerate the function of damaged auditory cells and synaptic connections. Our previous application focused on hair cell regeneration using stem cell-based epigenetic approaches. The major objectives of this renewal proposal are to determine the molecular mechanism critical for mouse hair cell synapse formation during normal development and develop a novel stem cell-based approach to reinnervate mouse auditory hair cells. Hair cell regeneration has obtained significant progress. Compared to hair cell regeneration, spiral ganglion neuron (SGN) regeneration has been reported but with relatively limited results. We have identified a stepwise method to guide mouse embryonic stem cells (ESCs) to differentiate into ESC-derived SGN-like neurons (ESNs) that showed SGN features. Up to date, only a few reports show that stem cell-derived neurons can form neural contacts with mouse hair cells. However, whether these neural contacts are bona fide SGN-hair cell synapses remains unclear. The function of these neural contacts has not been determined at the synaptic level. The molecular mechanism critical for regenerated neurons to functionally reinnervate hair cells remains obscure. Therefore, there is a critical need to use ESNs to reinnervate hair cells and determine the molecular mechanism critical for reinnervation. EphrinB family proteins bind to the EphB receptors, a family of transmembrane receptor tyrosine kinases. EphrinB signaling is involved in a variety of developmental programs, including synaptogenesis. Previous studies show that EphrinB1 signaling is critical for excitatory synaptic induction. Our preliminary data suggest the role of EphrinB signaling in ESN-hair cell reinnervation. Based on previous and our preliminary data, we hypothesize that EphrinB signaling may play a critical role in regulating neurons to form functional synapses with sensory hair cells. To test this hypothesis, three complementary specific aims are proposed. Aim 1 will determine the role of neuronal Efnb1 in SGN-hair cell synapse formation during development. Aim 2 will determine the molecular mechanism of ESN-hair cell reinnervation. Results of Aims 1 and 2 will guide the design of hair cell synapse regeneration research. In Aim 3, we will determine the extent to which EphrinB1 regulates ESN-hair cell reinnervation. Completion of this proposal will determine the molecular mechanisms critical for stem cell-derived neurons to reinnervate sensory hair cells. Identifying the role of Efnb1 in hair cell reinnervation will guide the synapse regeneration research, which will be translated into clinical trials to treat hearing loss patients. Therefore, the outcomes of this work will open new avenues to explore a stem cell-based multidisciplinary approach to regenerate hair cell synapses and restore the hearing function of Veteran and civilian patients.