ACANTHAMOEBA KERATITIS BIOLOGY, IMMUNOLOGY and THERAPY

描述（申请人提供）：棘阿米巴角膜炎是由致病性自由生活的阿米巴虫引起的一种危及视力的角膜疾病。棘阿米巴角膜炎的致病级联涉及一系列过程，包括：(1)滋养体通过凝集素-糖蛋白相互作用与角膜上皮细胞结合，(2)产生破坏角膜上皮细胞和基质细胞的细胞病变因子，(3)产生促进滋养体通过基底膜和基质侵入和渗透的蛋白水解酶，(4)降解构成角膜基质的I型和IV型胶原的胶原降解酶，(5)角膜膜金属蛋白酶的激活；(6)神经会阴炎的诱导。间质溶解是棘阿米巴角膜炎的主要致盲并发症。确定基质病发病机制中涉及的因素将是重要的。在目前的应用中，我们正在使用不同的策略来攻击致病级联中的关键步骤，以减轻正在进行的角膜炎和保护角膜组织。因此，治疗方式被设计为在生物体侵入角膜基质后抑制角膜组织的侵入和破坏。该项目的具体目标是：1)确定棘阿米巴纤溶酶原激活物（aPA）在棘阿米巴角膜炎发病机制中的作用；2)分析甘露糖诱导的棘阿米巴蛋白（133 -kDa）的胶原溶解活性；3)确定甘露糖诱导的棘阿米巴蛋白（133-kDa）是否激活基质金属蛋白酶4)确定甘露糖诱导的棘阿米巴蛋白（133-kDa）和棘阿米巴纤溶酶原激活物（aPA）作为免疫原诱导免疫和减轻棘阿米巴感染发病机制的可行性。5)克隆133-kDa蛋白并分析其片段是否适合作为亚单位疫苗，以诱导对棘阿米巴感染的更好保护。这些研究利用了棘阿米巴角膜炎的中国仓鼠模型，这与人类的对应物相似。棘阿米巴角膜炎的持续存在和耐药菌株的出现强调了这一努力的重要性。该项目的长期目标是开发一种抗疾病疫苗，作为治疗棘阿米巴角膜炎的辅助治疗手段。

DESCRIPTION (provided by applicant): Acanthamoeba keratitis is a sight-threatening corneal disease caused by pathogenic free-living amoebae. The pathogenic cascade of Acanthamoeba keratitis involves a series of processes that include: (1) binding of the trophozoites to the corneal epithelial cells via lectin-glycoprotein interactions, (2) generation of cytopathic factors that destroy the corneal epithelium and stromal cells, (3) production of proteolytic enzymes that facilitate the invasion and penetration of trophozoites through the basement membrane and stroma, (4) elaboration of collagenolytic enzymes that degrade types I and IV collagens, which constitute the corneal matrix, (5) activation of corneal membrane metalloproteinases and, (6) induction of perineuritis. Stromal dissolution is a major blinding complication of Acanthamoeba keratitis. It will be important to determine what factors are involved in pathogenesis of stromal disease. In the present application, we are using different strategies to attack crucial steps in the pathogenic cascade in an effort to mitigate ongoing keratitis and preserve corneal tissues. Accordingly, therapeutic modalities are designed to inhibit invasion and destruction of corneal tissues after the organisms have invaded corneal matrix. The specific aims for this project are: 1) determine the role of Acanthamoeba plasminogen activator (aPA) in the pathogenesis of Acanthamoeba keratitis, 2) analyze the collagenolytic activity of the mannose-induced Acanthamoeba protein (133 -kDa), 3 ) determine if the mannose-induced Acanthamoeba protein (133-kDa) activate matrix metalloproteinases 4) determine feasibility of using the mannose-induced protein (133-kDa) and Acanthamoeba plasminogen activator (aPA) as immunogens for inducing immunity and mitigating the pathogenesis of Acanthamoeba infections. 5) Clone the 133-kDa protein and analyze the fragment that might be suitable for use as a subunit vaccine to induce better protection against Acanthamoeba infections. These studies utilize the Chinese hamster model of Acanthamoeba keratitis, that is similar to the human counterpart. Continued presence of Acanthamoeba keratitis and emergence of the drug resistant strains is underscoring the significance of this endeavor. The long-range goal of this project is to develop an anti-disease vaccine as a therapeutic adjunct for the management of Acanthamoeba keratitis.