Significance of Corneal Cell Invasion By Bacteria

描述（由申请人提供）：角膜上皮对微生物的屏障功能通常被认为涉及表面粘蛋白、连接屏障和细胞外抗菌剂。 此外，我们发现角膜上皮细胞可以表现为“非专职吞噬细胞”，即它们可以内化细菌，然后抑制它们在酸化溶酶体内的活力。这种潜在的先天防御微生物的重要性仍有待确定，这是该项目的目标。 另一个目标是确定细菌如何逃避这种防御。铜绿假单胞菌是最常见和最具破坏性的角膜病原体之一。对于其他上皮衬里的身体部位，统计数据甚至更糟，其中铜绿假单胞菌感染经常导致死亡。虽然通常被称为细胞外病原体，但我们已经证明铜绿假单胞菌可以通过逃避酸化溶酶体并在质膜泡内建立复制小生境而在角膜（和气道）上皮细胞中茁壮成长。虽然溶酶体逃避和“泡龛”的形成都需要细菌III型分泌系统（T3SS），但这些能力是可分离的。溶酶体逃避依赖于T3SS效应子ExoS的ADP-核糖基化（ADPr）活性，而不是两个T3SS易位子之一的PopB。与此形成鲜明对比的是，铜绿假单胞菌需要ExoS和PopB来建立水泡小生境。 该项目的三个目标是：目标1。研究角膜上皮细胞杀死细胞内细菌的机制，特别是探索MyD88的作用，我们已经发现MyD88参与其中。目的2：确定铜绿假单胞菌如何使用ExoS逃避角膜上皮细胞中的酸化溶酶体。目标3.探索PopB和ExoS如何允许细胞内铜绿假单胞菌建立膜泡龛。 角膜上皮细胞的吞噬活性和细菌逃避这种防御都是新概念。了解导致这些非常不同结果的宿主-病原体相互作用，可能会导致管理感染的新策略。

DESCRIPTION (provided by applicant): Corneal epithelial barrier function against microbes is generally thought to involve surface mucins, junctional barriers, and extracellular antimicrobials. Additionally, we have found that corneal epithelial cells can behave as "non-professional phagocytes", i.e. they can internalize bacteria and then suppress their viability within acidified lysosomes. The importance of this potential innate defense against microbes remains to be determined and is a goal of this project. The other goal is to determine how bacteria evade this defense. Pseudomonas aeruginosa is among the most common and the most devastating of corneal pathogens. The statistics are even worse for other epithelial lined body sites, where P. aeruginosa infection very often results in mortality. While often referred to as an extracellular pathogen, we have shown P. aeruginosa can thrive in corneal (and airway) epithelial cells, by evading acidified lysosomes and establishing replicative niches inside plasma membrane blebs. While lysosome evasion and "bleb-niche" formation both require the bacterial type III secretion system (T3SS), these capacities are separable. Lysosome evasion depends on the ADP-ribosylation (ADPr) activity of the T3SS effector ExoS, but not PopB, one of two T3SS translocators. In stark contrast, both ExoS and PopB are needed for P. aeruginosa to establish bleb-niches. The three aims of this project are: Aim 1. To study mechanisms by which corneal epithelial cells kill intracellular bacteria; specifically exploring the role of MyD88, which we hae found is involved. Aim 2: To determine how P. aeruginosa uses ExoS to evade acidified lysosomes in corneal epithelial cells. Aim 3. To explore how PopB and ExoS then allow intracellular P. aeruginosa to establish membrane bleb niches. Phagocytic activity of corneal epithelial cells and bacterial evasion of this defense are both novel concepts. Understanding the host-pathogen interactions that result in these very different outcomes, could lead to new strategies for managing infection.