Department of Biomedicine, and Clinic for Otolaryngology, Head and Neck Surgery, Hospital Basel, University of Basel, Basel, 4031, Switzerland.
Clinic for Otolaryngology, Head and Neck Surgery, Kantonsspital Graubünden, Chur, 7000, Switzerland.
Sci Rep. 2020 Feb 27;10(1):3609. doi: 10.1038/s41598-020-60262-5.
Acid-base homeostasis is critical for normal growth, development, and hearing function. The sodium-hydrogen exchanger 6 (NHE6), a protein mainly expressed in early and recycling endosomes, plays an important role in regulating organellar pH. Mutations in NHE6 cause complex, slowly progressive neurodegeneration. Little is known about NHE6 function in the mouse cochlea. Here, we found that all NHE isoforms were expressed in wild-type (WT) mouse cochlea. Nhe6 knockout (KO) mice showed significant hearing loss compared to WT littermates. Immunohistochemistry in WT mouse cochlea showed that Nhe6 was localized in the organ of Corti (OC), spiral ganglion (SG), stria vascularis (SV), and afferent nerve fibres. The middle and the inner ears of WT and Nhe6 KO mice were not different morphologically. Given the putative role of NHE6 in early endosomal function, we examined Rab GTPase expression in early and late endosomes. We found no change in Rab5, significantly lower Rab7, and higher Rab11 levels in the Nhe6 KO OC, compared to WT littermates. Because Rabs mediate TrkB endosomal signalling, we evaluated TrkB phosphorylation in the OCs of both strains. Nhe6 KO mice showed significant reductions in TrkB and Akt phosphorylation in the OC. In addition, we examined genes used as markers of SG type I (Slc17a7, Calb1, Pou4f1, Cal2) and type II neurons (Prph, Plk5, Cacna1g). We found that all marker gene expression levels were significantly elevated in the SG of Nhe6 KO mice, compared to WT littermates. Anti-neurofilament factor staining showed axon loss in the cochlear nerves of Nhe6 KO mice compared to WT mice. These findings indicated that BDNF/TrkB signalling was disrupted in the OC of Nhe6 KO mice, probably due to TrkB reduction, caused by over acidification in the absence of NHE6. Thus, our findings demonstrated that NHEs play important roles in normal hearing in the mammalian cochlea.
酸碱平衡对正常生长、发育和听力功能至关重要。钠离子-氢交换体 6(NHE6),一种主要在早期和回收型内体中表达的蛋白,在调节细胞器 pH 值方面发挥重要作用。NHE6 突变导致复杂的、进行性缓慢的神经退行性疾病。目前对于 NHE6 在小鼠耳蜗中的功能知之甚少。在这里,我们发现所有 NHE 同工型都在野生型(WT)小鼠耳蜗中表达。与 WT 同窝仔鼠相比,Nhe6 敲除(KO)小鼠表现出明显的听力损失。WT 小鼠耳蜗中的免疫组织化学显示 Nhe6 定位于柯蒂氏器(OC)、螺旋神经节(SG)、血管纹(SV)和传入神经纤维。WT 和 Nhe6 KO 小鼠的中耳和内耳在形态上没有差异。鉴于 NHE6 在早期内体功能中的假定作用,我们检查了早期和晚期内体中 Rab GTPase 的表达。与 WT 同窝仔鼠相比,我们发现 Nhe6 KO OC 中的 Rab5 没有变化,Rab7 显著降低,Rab11 水平升高。因为 Rab 介导 TrkB 内体信号转导,我们评估了两种品系 OC 中的 TrkB 磷酸化。Nhe6 KO 小鼠的 OC 中 TrkB 和 Akt 磷酸化显著减少。此外,我们还检查了作为 SG Ⅰ型(Slc17a7、Calb1、Pou4f1、Cal2)和Ⅱ型神经元(Prph、Plk5、Cacna1g)标志物的基因。与 WT 同窝仔鼠相比,我们发现 Nhe6 KO 小鼠 SG 中的所有标记基因表达水平均显著升高。抗神经丝因子染色显示 Nhe6 KO 小鼠的耳蜗神经中有轴突丢失,而 WT 小鼠则没有。这些发现表明 BDNF/TrkB 信号在 Nhe6 KO 小鼠的 OC 中被破坏,可能是由于 NHE6 缺失导致 TrkB 减少,从而导致过度酸化。因此,我们的研究结果表明,NHEs 在哺乳动物耳蜗的正常听力中发挥重要作用。