HSPGs in Ocular Surface Diseases

描述（由申请人提供）：细菌性角膜炎是一种威胁视力的感染性疾病，与严重的眼部疾病相关。据估计，全世界每年的细菌性角膜炎发病率约为50万例，仅在美国就有3万例。眼表通常适合微生物病原体居住，因为眼表具有阻止微生物定植的解剖屏障、不断清除病原体的机械系统以及迅速建立有效根除入侵病原体的协调防御反应。当这些机制中的一种或几种被破坏，宿主-病原体相互作用的平衡转向有利于致病机制时，就会发生感染。因此，更好地了解宿主-病原体相互作用是一项重要的公共卫生优先事项，但导致微生物角膜炎发病的细菌和宿主机制尚不完全清楚。本应用程序的中心目标是确定syndecan- 1，一种主要的细胞表面硫酸肝素蛋白聚糖（HSPG），如何促进细菌性角膜炎的发病机制。Syndecan-1在角膜上皮中大量表达，但其在角膜中的功能尚不清楚。Syndecan-1在细胞表面作为受体，在细胞外环境中作为可溶性HSPGs，因为它的胞外外结构域在一定病理条件下可以脱落。初步研究表明，缺乏syndecan-1的小鼠可显著抵抗金黄色葡萄球菌的角膜感染，提示syndecan-1是促进金黄色葡萄球菌角膜炎的重要宿主因子。syndecan-1以这种方式发挥作用的潜在机制尚不清楚，但初步数据也表明，金黄色葡萄球菌诱导角膜上皮细胞中syndecan-1脱落，并利用syndecan-1外结构域的能力，以硫酸肝素（HS）依赖的方式对抗中性粒细胞介导的防御机制。基于这些数据，本研究将在《3 Specific Aims》中检验细菌颠覆HSPGs是细菌性角膜炎发病机制中的一个重要毒力机制的总体假设。目的1将定义在金黄色葡萄球菌角膜炎中如何诱导syndecan-1脱落。Aim 2将阐明促进金黄色葡萄球菌角膜炎的syndecan-1外结构域HS的结构特征，Aim 3将确定syndecan-1外结构域抑制中性粒细胞杀伤金黄色葡萄球菌的生物学机制。拟议的研究将使用新的和已建立的遗传、生化和细胞生物学方法，并结合细菌性角膜炎的体内模型。预计这些研究将揭示角膜上皮HSPGs先前未知的生物学功能，并增加我们对细菌性角膜炎发病机制核心的分子和细胞过程的理解。

DESCRIPTION (provided by applicant): Microbial keratitis is a sight-threatening infectious disease associated with significant ocular morbidity. By one estimate, the annual incidence of microbial keratitis is approximately 500,000 patients worldwide and 30,000 patients in the US alone. The ocular surface is generally inhabitable for microbial pathogens because of anatomical barriers that impede microbial colonization, mechanical systems that constantly clear pathogens, and coordinated defense responses that are rapidly mounted to effectively eradicate invading pathogens. Infection occurs when one or several of these mechanisms are compromised and the balance of host-pathogen interactions shifts to favor pathogenic mechanisms. Thus, a better understanding of host-pathogen interactions is a major public health priority, but the bacterial and host mechanisms that contribute to the pathogenesis of microbial keratitis are incompletely understood. The central goal of this application is to define how syndecan- 1, a major cell surface heparin sulfate proteoglycan (HSPG), promotes the pathogenesis of bacterial keratitis. Syndecan-1 is abundantly expressed in the corneal epithelium, but much remains unknown of its functions in the cornea. Syndecan-1 functions as receptors on the cell surface and also as soluble HSPGs in the extracellular environment because its extracellular ectodomain can be shed under certain pathological conditions. Preliminary studies showed that mice deficient in syndecan-1 significantly resist corneal infection by Staphylococcus aureus, suggesting that syndecan-1 is an important host factor that promotes S. aureus keratitis. The underlying mechanisms of how syndecan-1 functions in this manner are not understood, but preliminary data also suggested that S. aureus induces syndecan-1 shedding in corneal epithelial cells and exploits the capacity of syndecan-1 ectodomains to counteract neutrophil-mediated defense mechanisms in a heparin sulfate (HS)-dependent manner. Based on these data, this proposal will examine the overall hypothesis that bacterial subversion of HSPGs is an important virulence mechanism in the pathogenesis of bacterial keratitis in 3 Specific Aims. Aim 1 will define how syndecan-1 shedding is induced in S. aureus keratitis. Aim 2 will elucidate the structural features of syndecan-1 ectodomain HS that promote S. aureus keratitis, and Aim 3 will determine the biological mechanism of how syndecan-1 ectodomains inhibit S. aureus killing by neutrophils. The proposed studies will use both novel and established genetic, biochemical, and cell biological approaches in conjunction with in vivo models of bacterial keratitis. The proposed studies are anticipated to uncover previously unknown biological functions of corneal epithelial HSPGs and to increase our understanding of molecular and cellular processes that are central to the pathogenesis of bacterial keratitis.