SIGNIFICANCE OF CORNEAL CELL INVASION BY BACTERIA

描述：（从抽象）广泛的长期目标，这是 研究的目的是了解细菌侵入角膜细胞的作用 在假单胞菌引起的细菌性感染性角膜炎的发病机制中 铜绿假单胞菌，并制定治疗和/或预防策略 这种疾病。 由于铜绿假单胞菌被认为是细胞外的 病原体，对这种疾病的发病机制的研究集中在 细菌粘附于角膜、细菌毒素和宿主 炎症反应。 现在已经证明铜绿假单胞菌 可以侵入角膜细胞;在角膜感染期间和各种感染中， 体外模型 细胞内细菌可以逃避宿主的防御， 抗生素，因此可以解释为什么铜绿假单胞菌角膜炎往往是 这一建议的第一个目的是， 为了测试铜绿假单胞菌对角膜细胞的侵袭是否有助于 感染性角膜炎的发展。 所提出的方法包括 鉴定抑制铜绿假单胞菌侵入角膜方法 细胞，然后使用这些方法来抑制侵袭， 监测对疾病发展的影响。 铜绿假单胞菌不能 感染完整健康角膜，且感染最常见是 与角膜损伤或接触透镜佩戴相关。 因此，两个 研究铜绿假单胞菌角膜的不同实验模型 感染;涉及磨损损伤的鼠模型， 角膜和角膜接触透镜过度磨损的兔子模型。 水平 细菌侵入细胞将由庆大霉素存活率决定 通过肉眼检查评估疾病的严重程度 的角膜。 接触透镜配戴是铜绿假单胞菌的主要原因 人类的角膜感染 第二个要检验的假设是， 在将铜绿假单胞菌引入眼睛后，接触透镜 磨损最终增加了细胞内细菌的总数， 角膜与不戴镜片的眼睛相比， 三种机制。 （1）环境条件 发生在接触透镜下，其改变了 角膜细胞也改变了这些细胞对P. 2）配戴隐形透镜增加铜绿假单胞菌的感染 细胞的侵入继发于增加的细菌粘附于这些细胞， 接触透镜的佩戴阻碍了 通过干扰细胞内铜绿假单胞菌从眼表 浅表上皮细胞脱落和/或细胞停滞 在接触透镜下面。 该假设将通过以下方式进行检验：1） 检测环境操作对铜绿假单胞菌的影响 进入培养的角膜上皮细胞和整个小鼠角膜， 体外; 2）比较铜绿假单胞菌侵入角膜浅层 从佩戴的人接触透镜上取下的上皮细胞和对照 受试者;和3）评估细胞内细菌是否停滞 角膜接触透镜配戴过程中兔眼内的细菌。 感染性角膜炎是 最严重的角膜接触透镜配戴并发症。 因为隐形眼镜是 最常佩戴在健康的眼睛中，即使这种情况的发生率很低， 严重的疾病是不可接受的。 使某些患者更 易受感染和新的预防和/或 可以通过拟议的研究确定感染的治疗。

DESCRIPTION: (from the abstract) The broad long term goal of this research is to understand the role of corneal cell invasion by bacteria in the pathogenesis of bacterial infectious keratitis due to Pseudomonas aeruginosa, and to develop strategies for treatment and/or prevention of this disease. Since P. aeruginosa is believed to be an extracellular pathogen, research into the pathogenesis of this disease has focused on bacterial adherence to the cornea, bacterial toxins, and the host inflammatory response. It has now been demonstrated that P. aeruginosa can invade corneal cells; during corneal infection and in various in vitro models. Intracellular bacteria can evade host defenses and some antibiotics, and could thus explain why P. aeruginosa keratitis is often persistent and difficult to treat. The first aim of this proposal is to test whether corneal cell invasion by P. aeruginosa contributes to the development of infectious keratitis. The proposed methods involve identification of means to inhibit P. aeruginosa invasion of corneal cells, and then use of these methods to inhibit invasion whilst monitoring the effect on development of disease. P. aeruginosa cannot infect an intact healthy cornea and infection is most commonly associated with either corneal injury or contact lens wear. Thus, two different experimental models for studying P. aeruginosa corneal infection will be utilized; a murine model involving abrasion injury to the cornea, and a rabbit model for contact lens overwear. Levels of bacterial invasion into cells will be determined by gentamicin survival assays, and the severity of disease assessed by macroscopic examination of the cornea. Contact lens wear is the leading cause of P. aeruginosa corneal infection in humans. The second hypothesis to be tested is that following the introduction of P. aeruginosa into the eye, contact lens wear eventually increases the total number of intracellular bacteria on the cornea in comparison to eyes that do not wear lenses, by one or more of three mechanisms. These are: 1) that environmental conditions occurring under a contact lens which alter the metabolic function of corneal cells also alter the susceptibility of these cells to P. aeruginosa invasion; 2) that contact lens wear increases P. aeruginosa invasion of cells secondarily to increased bacterial adherence to these cells; and 3) that contact lens wear impedes the clearance of intracellular P. aeruginosa from the ocular surface by interfering with superficial epithelial cell exfoliation and/or by stagnation of cells under the contact lens. This hypothesis will be tested by: 1) examining the effect of environmental manipulations on P. aeruginosa entry into cultured corneal epithelial cells and whole mouse cornea in vitro; 2) comparing P. aeruginosa invasion into superficial corneal epithelial cells removed from human contact lens wearing and control subjects; and 3) evaluating whether intracellular bacteria are stagnated in the rabbit eye during contact lens wear. Infectious keratitis is the most serious complication of contact lens wear. Since contact lenses are most frequently worn in healthy eyes, even a low incidence of this serious disease is unacceptable. Factors that make certain patients more susceptible to infection and new approaches for the prevention and/or treatment of infection could be identified by the proposed studies.