Connexin Function and Mechanisms of Cx26 Deficiency Induced Hearing Loss

摘要 缝隙连接基因Cx26(GJB2)突变是导致遗传性耳聋的主要原因，从深部到深部 出生时先天耳聋到儿童时期轻度迟发性听力损失。小鼠模型显示Cx26 缺乏可导致耳蜗性发育障碍、毛细胞变性、耳蜗内电位(EP) 耳蜗肌的主动放大功能下降。我们进一步发现，耳蜗肌的发育 先天性耳聋的主要原因是疾病，而不是毛细胞退化，而迟发性耳聋 听力损失与外毛细胞(OHC)电动能力降低有关，即使毛细胞没有 缝隙连接蛋白表达。我们还证明了K+再循环假说不是Cx26的耳聋机制 缺乏症。然而，Cx26缺乏导致这些病理变化的详细机制仍然存在。 不清楚。此外，人们对人类耳蜗的病理变化知之甚少。最近，基因治疗 Cx26病毒在耳蜗处表达后，未能恢复听力。主要原因是缺少必需品。 对Cx26在耳蜗中的作用和Cx26缺乏性耳聋机制的认识。 在本方案中，我们将继续研究Cx26的功能及其细胞和分子机制。 Cx26缺乏导致先天性耳聋和迟发性听力损失。Cx26缺乏导致 耳蜗性发育障碍，提示缝隙连接(GJ)通道为细胞间通讯 导管对耳蜗管的发育至关重要。许多因素，如启动子、转录因子和 MiRNAs，可以调节发育过程中的基因表达。然而，这些监管机构都不是可渗透的。 到GJ通道，但miRNAs除外。MiRNAs可以广泛地调节基因表达，并在 器官发育。MiRNAs缺陷可导致耳蜗性发育障碍。在这项研究中，我们将 首先测试Cx26缺乏是否会扰乱小鼠脑内miRNA的表达和细胞间通讯 先天性耳聋患者耳蜗骨发育的影响。其次，我们将定义Cx26如何 OHC电动功能缺陷导致迟发性听力损失，哪些患者是很好的候选对象 用于实施预防和治疗干预措施，因为他们早年听力正常。 最近，连接蛋白的非通道功能已经出现。除了形成GJ通道外，连接蛋白还可以 参与细胞骨架的形成。我们将测试Cx26缺乏是否会损害细胞骨架 在OHC的支持细胞中形成，从而改变OHC的负荷(膜张力)并下降 OHC电活动和主动耳蜗声放大。最后，我们将使用反向突变筛查 筛查非综合征患者颞骨中Cx26基因突变的方法 听力损失，其耳蜗的病理变化已被诊断，以确定突变诱导 人类耳蜗的病理变化。显然，这些研究将揭示详细的耳聋 Cx26缺乏的机制。这些信息对于开发有效的治疗方法是迫切需要的。

Summary Mutations of gap junction gene Cx26 (GJB2) cause the most of hereditary deafness, ranging from profound congenital deafness at birth to mild late-onset hearing loss in childhood. Mouse models show that Cx26 deficiency can induce cochlear developmental disorders, hair cell degeneration, endocochlear potential (EP) reduction, and active cochlear amplification declining. We further found that the cochlear developmental disorder rather than hair cell degeneration is a primary cause for congenital deafness, whereas late-onset hearing loss is associated with reduction of outer hair cell (OHC) electromotility even hair cells have no connexin expression. We also evidenced that K+-recycling hypothesis is not the deafness mechanism of Cx26 deficiency. However, detailed mechanisms of these pathological changes induced by Cx26 deficiency remain unclear. Moreover, little is known about pathological changes in the human cochlea. Recently, gene therapy with viral-expression of Cx26 in the cochlea was failed to restore hearing. The main reason is lack of required knowledge of Cx26 function in the cochlea and deafness mechanisms by Cx26 deficiency. In this proposal, we will continually investigate Cx26 function and cellular and molecular mechanisms of Cx26 deficiency-induced congenital deafness and late-onset hearing loss. Cx26 deficiency causes the cochlear developmental disorder, indicating that gap junction (GJ) channels as an intercellular communication conduit are crucial for the cochlear development. Many factors, such as promoters, transcription factors, and miRNAs, can regulate gene expressions during development. However, none of these regulators is permeable to GJ channels except miRNAs. miRNAs can regulate gene expression broadly and have a critical role in the organ development. Deficiency of miRNAs can cause cochlear developmental disorders. In this study, we will first test whether Cx26 deficiency can disrupt miRNA expression and intercellular communication in the cochlea to affect cochlear development in the congenital deafness. Secondly, we will define how Cx26 deficiency decline OHC electromotility leading to late-onset hearing loss, which patients are good candidates for administration of preventive and therapeutic interventions due to normal hearing in their earlier life. Recently, connexin non-channel function has been emerged. Besides forming GJ channels, connexins can participate in cell cytoskeleton formation. We will test whether Cx26 deficiency can impair cytoskeleton formation in the OHC’s supporting cells, thereby changing OHC-loading (membrane tension) and declining OHC electromotility and active cochlear amplification. Finally, we will use backward-mutation screening approach to screen Cx26 mutations in the archival human temporal bones from patients with nonsyndromic hearing loss, whose pathological changes in the cochlea have been diagnosed, to identify mutation-induced pathological changes in the human cochlea. Apparently, these studies will reveal detailed deafness mechanisms by Cx26 deficiency. Such information is urgently required for developing efficient therapies.