Neutrophil calprotectin and neutrophil extracellular traps (NETs) in corneal defense and damage during fungal keratitis.

 描述(申请人提供)：角膜感染和溃疡是全球范围内视力受损和失明的重要原因。曲霉和镰刀菌属的丝状真菌是这些角膜感染的主要原因，占发展中国家微生物角膜炎病例的50%以上，那里温暖的气候和大量的农业劳动力容易暴露孢子(分生孢子)和眼睛损伤。由于隐形眼镜的使用、局部皮质类固醇的使用和眼科手术的增加，它们在美国也是一个令人担忧的问题。美国疾病控制与预防中心调查了2005-2006年与特定镜片护理产品有关的镰刀菌角膜炎的暴发，并召回了该镜片溶液。环境中真菌分生孢子的普遍存在意味着这些感染是一个持续的问题。目前的抗真菌治疗受到严重副作用和许多真菌种类耐药性增加的限制和困扰。因此，有必要更好地了解真菌感染的病理生理学和更有效的治疗手段。我们的实验室建立了一种成熟的真菌性角膜炎小鼠模型，在该模型中，我们已经确定了中性粒细胞(PMN)在抗真菌防御中的重要性，部分原因是由于活性氧(ROS)的产生和营养铁络合。利用这一模型和分离的中性粒细胞，拟议的研究试图通过测试中性粒细胞S100A8和S100A9(二聚体：钙保护素)是通过锌螯合和随后抑制真菌抗氧化防御来实现最佳真菌控制所必需的假设，来扩大PMN在抗真菌免疫中的作用(目标1)。这项研究的第二个主要目标将是研究真菌性角膜炎期间角膜中性粒细胞胞外陷阱(NETS)的形成，并使用化学抑制剂和基因敲除小鼠来确定NETS和网络介导的钙保护素释放在抗真菌免疫中的必要性(目标2)。Net对宿主组织有潜在的损害，我们将确定这些结构是否会导致不必要的眼部炎症和损害，以及阻止净释放是否代表一种潜在的治疗策略。在这些研究的结论中，我们将更好地了解中性粒细胞如何控制真菌感染，以及我们如何将锌封存等策略用于治疗目的。我们还将确定一些中性粒细胞防御是否会导致角膜混浊和视力丧失，这将导致更有效地治疗真菌性角膜炎。

 DESCRIPTION (provided by applicant): Corneal infections and ulceration are significant causes of visual impairment and blindness worldwide. Filamentous fungi of the Aspergillus and Fusarium genera are a leading cause of these corneal infections, accounting for upwards of 50% of microbial keratitis cases in developing countries, where warm climates and a high volume of agricultural labor predisposes to spore (conidia) exposure and ocular trauma. They are also a concern in the United States due to increasing contact lens use, use of topical corticosteroids and ocular surgery. An outbreak of Fusarium keratitis associated with a particular lens care product in 2005-2006 was investigated by the CDC and resulted in recall of the lens solution. The ubiquitous presence of fungal conidia in the environment means that these infections are an ongoing concern. Current anti-fungal therapies are limited and plagued by severe side-effects and increasing resistance of many fungal species. Thus, there is a need for greater understanding of fungal infection pathophysiology and more effective therapeutic tools. Our lab has a well-established murine model of fungal keratitis in which we have identified the importance of neutrophils (PMNs) in anti-fungal defense, in part due to reactive oxygen species (ROS) production and nutrient iron chelation. Using this model and isolated neutrophils, proposed studies seek to broaden the role of PMNs in anti-fungal immunity by testing the hypothesis that neutrophil S100A8 and S100A9 (dimer: calprotectin) is necessary for optimal fungal control through zinc chelation and subsequent inhibition of fungal antioxidant defenses (Aim 1). The second major goal of the study will be to investigate the formation of neutrophil extracellular traps (NETs) in the cornea during fungal keratitis and use chemical inhibitors and knock-out mice to determine the necessity of NETs, and NET-mediated calprotectin release, in anti-fungal immunity (Aim 2). NETs are potentially damaging to host tissue and we will determine whether these structures lead to unnecessary ocular inflammation and damage, and whether blocking NET release represents a potential therapeutic strategy. At the conclusion of these studies we will better understand how neutrophils control fungal infection and how we can co-opt strategies such as zinc sequestration for therapeutic purposes. We will also determine whether some neutrophil defenses contribute to corneal opacification and vision loss which will lead to more effective management of fungal keratitis.