PATHOGENESIS OF ACANTHAMOEBA KERATITIS

棘阿米巴角膜炎是一种使人衰弱的角膜感染。目前，该病的诊断并不直截了当，治疗也有问题，包括每小时、24小时、局部应用一种药物组合，持续很长一段时间。尽管进行了积极的治疗，复发仍有可能发生。鉴于这种疾病的破坏性以及与治疗有关的问题，我们的目标是找到一种方法来识别处于危险中的个人，并为他们提供合理设计的策略，以防止感染。寄生虫对宿主细胞的粘附是感染发病的关键第一步。在之前的资助期间，我们证明棘阿米巴表达一种甘露糖受体，该受体介导阿米巴与角膜上皮细胞的粘附。为了了解甘露糖介导的变形虫与宿主细胞的粘附触发最终杀死宿主细胞的事件的机制，在Aim 1中，我们将克隆编码变形虫甘露糖受体的cDNA，并将确定甘露糖受体是否是具有调节信号转导事件潜力的跨膜蛋白。在Aim 2中，我们将确定是否口服免疫具有阿米巴甘露糖受体碳水化合物识别结构域的仓鼠导致泪液中抗体水平升高，如果是这样，免疫反应是否提供对感染的保护。我们最近的研究表明，在甘露糖介导的变形虫与宿主细胞的粘附之后，一种特定的蛋白酶P3被分泌到共培养培养基中。Aim 3中提出的研究是为了验证一个假设，即P3可能在涉及变形虫诱导的角膜上皮细胞溶解的接触依赖事件的级联反应中起重要作用。希望本研究将有助于更好地了解棘阿米巴角膜炎的分子基础，并最终提高预防该病的前景。此外，这项研究将有助于对感染的致病机制和细胞生物学的基本理解，因此，有利于未来研究人员寻求预防其他病原体引起的眼部感染。

Acanthamoeba keratitis is a debilitating infection of the cornea. At present, diagnosis of the disease is not straightforward and treatment is problematic, consisting of hourly, around the clock, topical application of a combination of drugs for an extended period of time. Despite the aggressive treatment, recrudescence may occur. In view of the devastating nature of the disease and the problems associated with the therapy, our goals are to find a means to identify individuals who are at risk and provide them with rationally designed strategies to protect against the infection. The adhesion of the parasite to the host cells is the critical first step in the pathogenesis of infection. During the previous funding period, we demonstrated that Acanthamoebae express a mannose receptor which mediates adhesion of the amoeba to corneal epithelial cells. For an understanding of the mechanism by which the mannose-mediated adhesion of the amoeba to host cells triggers events which ultimately kill the host cells, in Aim 1, we shall clone a cDNA encoding the amoeba mannose receptor and will establish whether the mannose receptor is a transmembrane protein with potential for modulating signal transduction events. In Aim 2, we shall determine whether oral immunization of hamsters with specifically the carbohydrate recognition domain of the amoeba mannose receptor leads to an elevated antibody level in their tears, and, if so, whether the immune response provides protection against the infection. We have recently shown that subsequent to the mannose-mediated adhesion of the amoeba to host cells, a specific proteinase, P3, is secreted into the co-culture media. Studies proposed in Aim 3 are to test a hypothesis that P3 may be an important player in the cascade of contact-dependent events involved in the amoeba-induced cytolysis of corneal epithelial cells. It is hoped that this study will contribute to a better understanding of the molecular basis of Acanthamoeba keratitis and will ultimately improve the prospects of preventing the disease. In addition, this study will contribute to the basic understanding of the pathogenic mechanisms and cell biology of infections in general and as such, benefit future investigators looking to prevent ocular infections caused by other pathogens.