Neutrophil and Toll-Like Receptor 4 Contributions to Endotoxemia-Enhanced Gentamicin Ototoxicity

 描述(由申请人提供)：当严重的细菌感染(例如脑膜炎、心内膜炎、腹膜炎；通常包括菌血症)导致全身炎症反应综合征(SIRS)时，就会发生败血症。氨基糖苷类抗生素是为这些危及生命的感染保留的一种基本抗生素。然而，尽管氨基糖苷类化合物具有杀菌效果，但它们也会导致终生听力损失和前庭功能障碍，影响生活质量指标，如听力和语言技能、教育程度。最近的证据表明，实验诱导的SIRS与细菌脂多糖(LPS)一起增加了氨基糖苷类药物的耳蜗量，从而增强了这些药物的耳毒性副作用。这与耳蜗促炎分子(如细胞因子和趋化因子)在mRNA和蛋白质水平上的增加不谋而合。此外，患有TLR4(脂多糖的主要受体)功能障碍的小鼠耳蜗氨基糖苷类和促炎细胞因子水平降低。总而言之，这些先导数据表明，TLR4介导的耳蜗炎症是提高耳蜗氨基糖苷类和促炎细胞因子水平的原因。患有中性粒细胞减少症的患者，特别是新生儿和接受化疗的患者，特别容易发生败血症。在脓毒症中，中性粒细胞的主要作用是清除细菌，但在全身炎症反应综合征(SIRS)中，这也会在非感染组织中诱导中性粒细胞介导的并存血管损伤，导致与脓毒症相关的大部分发病率和死亡率。这一建议的总体假设是，SIRS通过先天免疫反应失调耳蜗血迷路屏障，增加耳蜗氨基糖苷类化合物的摄取，从而导致耳毒性。拟议的实验将(I)测量TLR4基因和中性粒细胞缺陷小鼠SIRS期间耳蜗炎标志物的水平；(Ii)表征SIRS期间以中性粒细胞为重点的耳蜗固有免疫反应；以及(Iii)表征耳蜗炎反应如何扰乱血迷路屏障生理学，增强氨基糖苷类药物诱导的耳毒性。这些数据将使我们能够确定免疫反应分子介体和中性粒细胞对SIRS增强的耳蜗氨基糖苷负荷的相对贡献。这项拟议研究的数据将与氨基糖苷类药物在脓毒症治疗中的临床应用相关，通过表征哪些因素会增加氨基糖苷类药物所致耳毒性的风险。对氨基糖苷类药物的适应症和禁忌症进行更强有力的循证定义，将降低与氨基糖苷类药物相关的终生耳毒性的患病率和程度。

 DESCRIPTION (provided by applicant): Sepsis occurs when serious bacterial infection (e.g. meningitis, endocarditis, peritonitis; usually including bacteremia) induces Systemic Inflammatory Response Syndrome (SIRS). Aminoglycosides are an essential class of antibiotics reserved for these life-threatening infections. Yet, despite their bactericidal efficacy, aminoglycosides can also induce life-long hearing loss and vestibular deficits, affecting quality of life indicators, e.g., listening and language skills, educational attainment. Recent evidence suggests that experimentally- induced SIRS, with bacterial lipopolysaccharide (LPS), enhances cochlear loading of aminoglycosides, potentiating the ototoxic side-effects of these drugs. This coincides with increased cochlear levels of pro- inflammatory molecules (such as cytokines and chemokines) at the mRNA and protein levels. Furthermore, cochlear levels of aminoglycosides and pro-inflammatory cytokines are reduced in mice with dysfunctional TLR4 (the primary receptor for LPS). Together, these pilot data suggest that TLR4-mediated inflammation in the cochlea is responsible for enhancing cochlear levels of aminoglycosides and pro-inflammatory cytokines. Patients with neutropenia, especially neonates and patients treated with chemotherapy, are exceptionally vulnerable to sepsis. During sepsis, the primary role of neutrophils is to eliminate bacteria, but during SIRS, this can also induce collateral neutrophil-mediated vascular injury in non-infected tissues, causing much of the morbidity and mortality associated with sepsis. The overall hypothesis of this proposal is that SIRS dysregulates the blood-labyrinth barrier of the cochlea via the innate immune response, increasing cochlear uptake of aminoglycosides and subsequent ototoxicity. The proposed experiments will (i) measure cochlear inflammatory marker levels during SIRS in TLR4- and neutrophil-deficient mice over time; (ii) characterize the cochlear innate immune response during SIRS with a focus on neutrophils; and (iii) Characterize how the cochlear inflammatory response perturbs blood-labyrinth barrier physiology potentiating aminoglycoside- induced ototoxicity. These data will allow us to determine the relative contributions of immune-response molecular mediators and neutrophils to SIRS-enhanced cochlear aminoglycoside loading. The data from the proposed studies will be relevant to clinical use of aminoglycosides in the management of sepsis, by characterizing which factors increase the risk of aminoglycoside-induced ototoxicity. Stronger, evidence-based definition of the indications and contraindications for aminoglycoside therapy will reduce the prevalence and degree of the life-long ototoxicity associated with aminoglycosides.