IMMUNOBIOLOGY OF A 30 KDA PROTEIN OF HAEMOPHILUS DUCREYI

本项目旨在了解在感染的初始阶段，嗜血杆菌与人类中性粒细胞（PMN）和宿主上皮细胞的分子相互作用。宿主细胞对病原体的附着和内化通常是感染过程的第一步。在体外，H. ducreyi在缺乏血清调节素的情况下被PMN和上皮细胞结合并内化，这表明配体（粘连素）/受体相互作用(s)。在感染过程的早期进行免疫干预可以改变或预防细胞附着和可能的疾病。pmn是在软下腺病变开始时募集的第一批细胞，因此可能是抵抗感染的重要防御。这些研究将增强我们对附着于上皮细胞和PMN在感染过程中的作用的整体理解，PMN杀死杜克雷伊嗜血杆菌的能力，并可能导致新的疫苗策略。有五个具体目标。在第一个特异性目标中，将构建一个30 kDa外膜蛋白的等基因突变体。如以下章节所述，30kda蛋白与细胞粘附素和其他细菌中介导血清抗性的蛋白质有关。第二个特定目标是确定这种蛋白质的功能。我们将使用突变体/亲本比较来确定其在PMN、上皮细胞和细胞基质蛋白粘附中的可能作用。我们还将进行30kda突变体/亲本对血清和吞噬杀伤的抗性比较。在第三个特定目标中，我们将在温度依赖性感染兔模型和人杜克雷伊氏杆菌感染实验模型中测试杜克雷伊氏杆菌30kda蛋白的等基因突变体。第四个具体目标涉及某些与疫苗有关的问题。我们将确定在软下疳感染期间是否会引发30kda蛋白的抗体，以记录在体内的表达。我们将测试纯化重组表达的30kda保护动物免受侵袭性感染的能力。在Specific Aim 5中，我们将鉴定除30kda蛋白外的PMN和细胞粘附素。

This project seeks to understand the molecular interactions of Haemophilus decreyi with human neutrophils (PMN) and host epithelial cells during the initial stages of infection. Attachment to, and internalization of, pathogens by host cells is often the first step in the infectious process. In vitro, H. ducreyi is bound and internalized by PMN in the absence of serum opsonins and by epithelial cells suggesting a ligand (adhesin)/receptor interaction(s). Immunological intervention early in the infectious process could modify or prevent cellular attachment and possibly disease. PMNs are the first cells recruited at the onset of chancroidal lesion and therefore might be an important defense against infection. These studies will enhance our overall understanding of the role of attachment to epithelial cells and PMN in the infectious process, the ability of PMN to kill H. ducreyi, and perhaps, lead to new vaccine strategies. There are five Specific Aims. In the first Specific Aim, an isogenic mutant of the 30 kDa outer membrane protein will be constructed. As described in detail in the following sections, the 30 kDa protein is related to cellular adhesins and proteins which mediate serum resistance in other bacteria. The second Specific aim is to determine the function of this protein. We will use mutant/parent comparisons to determine its possible role in adherence to PMN, epithelial cells, and cellular matrix proteins,. We will also perform 30 kDa mutant/parent comparisons for resistance to serum and phagocytic killing. In the third Specific aim, we will test an isogenic mutant of the H. ducreyi 30 kDa protein in the temperature dependent rabbit model of infection and in the human experimental model of H. ducreyi infection. The fourth Specific aim addresses certain vaccine related issues. We will determine if antibodies to the 30 kDa protein are elicited during chancroid infection to document in vivo expression. We will test the ability of purified recombinantly expressed 30 kDa to protect animals from a challenge infection. In Specific Aim 5 we will identify PMN and cellular adhesins other than the 30 kDa protein.