胆红素通过酸敏感离子通道介导的耳蜗听神经毒性机制研究

Hyperbilirubinemia will damage the developing neuronal system in newborns. The accumulation of bilirubin in central nucleus will lead to acute or chronic dysfunction as in bilirubin encephalopathy or kernicterus. Hearing loss is one of the common symptoms and sequelae, and it could be caused by the damage to the auditory nuclei or to the peripheral auditory system, which is the cochlea. Hyperbilirubinemia could lead to oxidative stress, endoplasmic reticulum stress, and inflammation. Ischemia, inflammation, and hypoxia would active acid-sensing ion channels (ASICs), which would regulate apoptosis, cell differentiation, and other important processes. In our previous work, we demonstrate that spiral ganglion neurons (SGNs) were the most sensitive cell types to hyperbilirubinemia via organ of Corti and SGNs culture, and the neurotoxicity decreased when pH level increased in the medium. Based on these evidence, we hypothesized that hyperbilirubinemia would activate apoptosis in SGNs via the activation of ASICs, which lead to the cell death and auditory dysfunction. In this research, we will verify the neurotoxicity and the mechanism by regulating the acivity of ASICs in organ culture system as well as in ASIC knockout mice, , and provide the possibility to prevent or ameliorate the neurotoxicity in SGNs via regulation of the ASICs in future.

高胆红素血症给新生儿发育中的神经系统带来不同程度的损伤，当其选择性的沉积于特定的中枢核团时则会诱发多种急慢性功能障碍，引起胆红素脑病，危及患者健康。听力损伤是其常见症状及后遗症之一，其原因可为中枢性和外周性。高胆红素血症可引起氧化应激、内质网应激及炎症反应等。在缺血、缺氧以及炎症等状态下，可激活酸敏感离子通道，进而调节细胞凋亡、分化等生理过程。申请人前期预实验通过耳蜗柯蒂氏器及螺旋神经节培养中证实了胆红素对于耳蜗的细胞毒性作用首先发生于神经元，并且增加培养液pH值可减弱胆红素的神经毒性，因此，我们认为高胆红素血症可以通过激活螺旋神经节细胞的酸敏感离子通道进而激活凋亡通路，进而引起细胞凋亡及耳蜗功能变化导致听力损伤，本课题将通过调节酸敏感离子通道活性来验证其在胆红素外周毒性中的作用，并通过基因敲除小鼠进一步证实，为今后通过该通道的调节来改善、预防高胆红素血症带来的外周神经毒性提供理论基础。

高胆红素血症发生时，过多的胆红素选择性沉积于特定的核团会诱发多种急慢性功能障碍，给发育中的神经系统带来不同程度的损伤。听力损伤是其常见症状及后遗症之一，而高胆红素造成的听觉外周损害机制并不明了。高胆红素血症可引起氧化应激、内质网应激及炎症反应，在缺血、缺氧以及炎症等状态下，酸敏感离子通道可被激活，进而调节细胞凋亡、分化等生理过程。为探讨酸敏感离子通道在胆红素造成的外周神经毒性中的作用，我们建立了胆红素在耳蜗及螺旋神经元培养体系中的损伤模型，探究胆红素造成螺旋神经元损伤的细胞及分子机制，并使用酸敏感离子通道抑制剂进行干预，通过耳蜗及毛细胞的形态学及分子生物学层面发生的改变结合高通量的转录组水平测序分析，得到如下结论：在偏酸性环境下，酸敏感离子通道被激活，加重了胆红素造成的螺旋神经节损伤；使用抑制剂，可通过抑制Caspase-3介导的凋亡通路的活化减轻胆红素造成的神经损伤；在酸敏感离子通道抑制后，下游肿瘤坏死因子信号通路、NF-κB信号通路、细胞因子及受体作用通路等通路的改变参与了酸敏感离子通道抑制后神经元损伤减轻的保护机制。本课题明确了高胆红素通过酸敏感离子通道激活后引起螺旋神经元的细胞凋亡造成了外周神经毒性，并为进一步的预防及干预胆红素造成的听觉外周损伤毒性作用提供理论基础。