Corneal Expression and Function of Slurp1

描述（申请人提供）：虽然分泌的Ly6/尿激酶型纤溶酶原激活物受体（uPAR）相关蛋白-1 （Slurp1）是角膜中表达最丰富的蛋白之一，并分泌到泪膜中，但其在眼表中的生物学功能尚不清楚。在本项目中，我们验证了Slurp1是抑制眼表炎症的关键免疫调节分子的中心假设，并在促炎条件下下调。这一假设得到了我们令人兴奋的初步数据的支持，这些数据表明，当角膜首次暴露于环境中时，Slurp1的表达(i)在小鼠眼睑打开时增加，（ii）在细菌脂多糖（LPS）注射或1型单纯疱疹病毒（HSV-1）感染并发中性粒细胞浸润的24小时内消失，（iii）在炎症的klf4条件null （Klf4CN）角膜中减少，以及（iv）被促炎的白细胞介素IL-4和IL-13抑制。更重要的是，(a) Slurp1的过表达阻碍了腺病毒性角膜炎中性粒细胞内流，(b) Slurp1结合uPAR配体uPA，表明Slurp1通过竞争性结合uPAR配体抑制炎症。我们将通过追求三个具体目标来检验我们的假设。在Aim 1中，我们将建立Slurp1的角膜功能。我们将通过监测Slurp1在几种不同实验模型中的表达来确定Slurp1的表达降低是否是角膜炎症的共同主题。通过利用分子方法降低或增加小鼠角膜Slurp1的表达，我们将分别确定这些变化是增加还是减轻炎症。在目标2中，我们将研究Slurp1功能的分子机制。我们将确定Slurp1是否与uPAR竞争结合其配体uPA、玻璃体连接蛋白和整合素，从而减少中性粒细胞迁移。在Aim 3中，我们将探究健康角膜和炎症角膜中Slurp1表达调控的分子事件。我们将确定kruppel样转录因子-4 （Klf4）是否控制睁眼时Slurp1的表达增加，以及在促炎条件下，不同刺激下产生的活性氧是否抑制Slurp1。该项目基于Slurp1是一种分子开关，在正常水平下抑制慢性炎症，在促炎条件下下调，这一新颖的想法，将产生与眼表健康和疾病相关的有价值的新信息。它创新地利用了我们独特的小鼠模型，以扩大基因调控和先天免疫反应之间的界面知识，并建立了解释眼表炎症调节的新范式。预期的结果将为眼表炎症性疾病的治疗干预确定一个新的目标。

DESCRIPTION (provided by applicant): Though the secreted Ly6/urokinase type plasminogen activator receptor (uPAR) related protein-1 (Slurp1) is one of the most abundantly expressed proteins in the cornea and is secreted into the tear film, its biological function in the ocular surface is unknown. In this project, we test the central hypothesis that Slurp1 is a key immunomodulatory molecule that suppresses inflammation at the ocular surface and is downregulated in pro-inflammatory conditions. This hypothesis is supported by our exciting preliminary data demonstrating that Slurp1 expression is (i) increased upon mouse eyelid opening when the cornea is first exposed to the environment, (ii) abrogated within 24h of bacterial lipopolysaccharide (LPS) injection or Herpes Simplex Virus type-1 (HSV-1) infection concurrent with neutrophil infiltration, (iii) decreased in the inflamed Klf4-conditional null (Klf4CN) corneas, and (iv) suppressed by pro-inflammatory interleukins IL-4 and IL-13. More importantly, (a) over-expression of Slurp1 hindered the neutrophil influx in adenoviral keratitis and (b) Slurp1 bound the uPAR ligand uPA, suggesting that Slurp1 inhibits inflammation by competitive binding of UPAR ligands. We will test our hypothesis by pursuing three Specific Aims. In Aim 1, we will establish the corneal function of Slurp1. We will determine whether reduced expression of Slurp1 is a common theme in corneal inflammation by monitoring its expression in several different experimental models. By utilizing molecular approaches to decrease or increase mouse corneal expression of Slurp1, we will determine whether these changes augment or mitigate inflammation respectively. In Aim 2, we will examine the molecular mechanisms underlying Slurp1 functions. We will determine whether Slurp1 competes with uPAR for binding to its ligands uPA, vitronectin, and integrins, resulting in decreased neutrophil migration. In Aim 3, we will interrogate the molecular events governing regulation of Slurp1 expression in healthy and inflamed corneas. We will determine if Kruppel-like transcription factor-4 (Klf4) governs the increased expression of Slurp1 upon eyelid opening, and reactive oxygen species produced in response to different stimuli suppress Slurp1 in pro-inflammatory conditions. This project, based on the novel idea that Slurp1 is a molecular switch that suppresses chronic inflammation when present in normal levels and is downregulated in pro-inflammatory conditions will generate valuable new information related to ocular surface health and disease. It makes innovative use of our unique mouse models to expand knowledge of the interface between gene regulation and innate immune response and establishes a new paradigm explaining regulation of inflammation at the ocular surface. The anticipated outcomes will define a novel target for therapeutic intervention in managing inflammatory disorders of the ocular surface.