Mechanisms of otoprotection: Impact of Survivin-NO-signaling on microcirculation and hearing preservation after cochlear implantation

尽管微创电极植入、保留结构的手术技术和术中糖皮质激素的应用，但残留听力的患者植入人工耳蜗术后仍有可能导致部分或完全丧失残余听力。其潜在的分子机制仍不完全清楚，尽管这是发展耳保护策略的关键。在我们之前的工作中，我们令人信服地表明，微循环障碍以及亚硝化应激诱导的细胞保护机制干扰参与了内耳损伤。一氧化氮合酶(NO)可调节细胞保护的凋亡抑制蛋白Survivin在螺旋神经节和耳蜗侧壁上的表达，是Corti器功能微循环的宿主。在以往研究的基础上，我们假设HCI损伤后，Survivin-NO轴对微循环和听力功能的影响和潜在的信号通路，以及它们最终与耳保护或耳毒性的关系，都还没有被研究过。我们假设HCI影响耳蜗壁，导致微循环障碍，以及eNOS/iNOS驱动的NO诱导，调节Survivin的表达。这些应激状态最终可能导致听力功能受损。因此，需要研究微循环和Survivin-NO信号在HCI中的作用。项目的目标是：(1)监测微循环和听力功能；(2)测量HCI诱导的内皮细胞激活和NO诱导；(3)剖析与HCI/NO相关的Survivin信号通路的影响；(4)靶向抑制Survivin与HCI相关性听力损害的相关性；(5)HCI相关效应对听力参数、微循环、内皮细胞激活、NO诱导以及Survivin相关耳保护蛋白表达的统计相关性分析；(6)与局部糖皮质激素治疗相比，局部NO抑制或诱导以及全身吸入NO对Survivin-NO信号的调节作为一种新的耳保护策略在HCI中是一种新的耳保护策略。我们将使用我们建立的动物模型--正常听力豚鼠--来分析耳蜗侧壁加基础电极插入的情况，以及条件Survivin-敲除小鼠模型。实验包括听力测量、体内荧光显微镜、Survivin靶向脂质体纳米载体的应用、免疫组织化学和激光捕获-显微解剖。该项目不仅将揭示关键的HCI相关耳保护机制，而且可能为临床上新的耳保护策略开辟翻译前景。

In spite of minimal-invasive electrode insertion, structure-preserving surgical techniques and intraoperative glucocorticoid application, cochlea implantation in patients with hearing remnants (hCI) may still result in partial or complete loss of residual hearing. The underlying molecular mechanisms are still incompletely understood, though key for the development of otoprotective strategies.In our previous work, we convincingly showed that microcirculatory impairment as well as nitrosative stress-induced interference with cytoprotective mechanisms are involved in inner ear damage. Nitric oxide (NO) was found to modulate the expression of the cytoprotective inhibitor-of-apoptosis-protein Survivin in the spiral ganglion as well as in the cochlear lateral wall, hosting the functional microcirculation of the Organ of Corti. The impact and underlying signaling pathways of the Survivin-NO-axis on microcirculation and hearing function following hCI insertion trauma as well as their ultimate otoprotective or ototoxic relevance have not yet been investigated.Based on our previous studies, we hypothesize that hCI affects the lateral cochlear wall, resulting in microcirculatory disturbance as well as to an eNOS/iNOS-driven NO induction, modulating the expression of Survivin. These stress conditions may ultimately contribute to impairment of hearing function. Hence, the role of microcirculation and Survivin-NO-signaling for hCI shall here be investigated. Project aims are: (1) monitoring microcirculation and hearing function; (2) measurement of hCI-induced endothelial cell activation and NO induction; (3) dissecting the impact of hCI-/NO-associated Survivin-modulating signaling cascades (4) relevance of targeted survivin inhibition for hCI-associated hearing impairment; (5) statistical correlation analysis of hCI-associated effects on parameters of hearing, microcirculation, endothelial cell activation, NO induction as well as on the expression of Survivin-associated otoprotective proteins; (6) modulation of Survivin-NO-signaling using local NO-inhibition or -induction as well as systemic inhalative NO application as a novel otoprotection strategy in hCI compared with local glucocorticoid treatment.We will use our established animal model, the normal hearing guinea pig, for analysis of the lateral cochlear wall complemented by basal electrode insertion as well as a conditional survivin-knock out mouse model. Experiments involve audiometry, in-vivo-fluorescence microscopy, application of Survivin-targeting liposomal nanocarriers, immunohistochemistry, and laser capture-microdissection.The project will not only uncover fundamental crucial hCI-associated otoprotective mechanisms but may also open translational vistas for novel otoprotection strategies in the clinics.