ErbB2信号通路参与听神经病谱系障碍髓鞘生成的机制研究

Auditory nerve spectrum disorders (ANSD), which are sensorineural hearing disorders resulting from the pathological basis of demyelinating of the eighth cranial nerve, have had no definitely effective treatment in clinical practice to date. Peripheral nerve myelin sheath is mainly differentiated from Schwann cells (SCs). In the previous study, we found that Sox10 and ErbB2 gene were crucial to the survival of SCs, which affected the development of Spiral Ganglion Neurons (SGNs) and guide axon growth. Using abdominal injection of diphtheria toxin (DT) , we successfully established animal model of acute ANSD and analyzed its pathogenesis in the view of audiology and morphology. On the basis of the previous research, this project will further investigate: 1) use Sox10 and ErbB2 gene knockout mice and establish the SCs-SGNs myelinating co-culture system in vitro, in order to explore the molecular mechanisms of ErbB2 signaling pathway involved in the differentiation and maturation of SCs and the effects to the development of SGNs; 2) inject DT to establish the acute ANSD animal model, which will help us to verify that interference of ErbB2 signaling pathway may promote the proliferation of myelin sheath and the repair of axons in the ANSD mice. In our project, we will take the development and regeneration of myelin sheath as a breakthrough point, and comprehensively analyze the role of ErbB2 signaling pathway in the occurrence and progression of ANSD and the mechanisms of promoting neural repair from the molecule-cell-organization and the overall aspect. That will provide new orientations for the treatment of the disease.

听神经病谱系障碍（ANSD）是以第Ⅷ对颅神经脱髓鞘病变为病理基础的感音神经性听力障碍，目前尚无确切的治疗方法。外周神经髓鞘主要由Schwann细胞（SCs）分化发育而来。我们发现，Sox10和ErbB2基因决定了SCs的存活，它们可能促进神经元发育和引导轴突生长。我们用腹腔注射白喉毒素（DT）成功构建急性ANSD动物模型，并从听力学和形态学角度分析其发病机制。本项目在前期研究基础上拟：1）培育Sox10、ErbB2转基因小鼠和建立SCs-SGNs体外成髓鞘共培养体系，探讨ErbB2通路参与SCs分化的分子机制及对SGNs的影响；2）注射DT构建急性ANSD动物模型，探讨干预ErbB2信号通路对ANSD小鼠髓鞘增生和轴突损伤后修复的作用。本项目以髓鞘发育和再生作为切入点，从分子-细胞-组织-整体角度全面分析ErbB2信号通路在ANSD发生中的作用及神经修复机制，为治疗疾病提供新的方向。

研究听神经病谱系障碍(ANSD)的耳毒性动物模型多种多样。这种疾病的潜在机制尚不清楚。本研究用白喉毒素(DT)注射小鼠，观察小鼠听觉脑干反应(ABR)和正常耳声发射(OAEs)的电生理变化。Myosin7a和CtBP-2免疫染色显示毛细胞和带状突触的数量变化。透射电镜(TEM)观察了DT损伤后特殊神经通路的形态学变化。采用免疫化学、real - time PCR和western blot检测CD-9作为白喉毒素受体相关蛋白的表达情况。结果表明，DT组ABR阈值降低，耳声发射(OAEs)保持正常。DT诱导内毛细胞(ihc)的丢失具有时间和空间特异性，而外毛细胞存活。内耳带状突触的缺失在细胞死亡之前就已被发现。与第3天内内毛细胞数量保持稳定不同，螺旋神经节神经元(SGNs)及其髓鞘在21天内继续发生退行性变。我们发现CD9在ihc和听神经髓鞘中呈点状表达。DT治疗后CD9水平升高，而新生儿CD9水平明显降低。综上所述，适当给药可成功建立ANSD动物模型。注射DT后，IHC的缺陷可以在时间和空间特异性的基础上早期得到稳定。SGNs和髓鞘长期呈继发性损害。ihc中的带状突触缺失可能是细胞损伤的早期迹象。CD9的表达可能影响DT对小鼠听觉通路的耳毒性。