MECHANISM OF PATHOGENIC ACTIVITY OF PEPTIDOGLYCAN

肽聚糖的致病活性机制

• 批准号: 3143359
• 负责人: Roman Dziarski
• 金额: $ 20.55万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-07-01 至 1996-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3143359
• 关键词: Escherichia coli; Staphylococcus aureus; affinity chromatography; antireceptor antibody; athymic mouse; carbohydrate receptor; hamsters; laboratory mouse; leukocyte activation /transformation; lipopolysaccharides; lymphocyte; macrophage; molecular pathology; monoclonal antibody; peptidoglycan; receptor binding; tissue /cell culture

Bacterial infections are still a major cause of morbidity and mortality, especially in immunocompromised individuals. Peptidoglycan and lipopolysaccharide can reproduce most major signs and symptoms of infections with Gram-positive and Gram-negative bacteria, yet their molecular mechanism of action on host cells is virtually unknown. This laboratory has recently discovered one dominant peptidoglycan-binding protein on mouse lymphocytes and macrophages, and demonstrated that this protein is identical with the recently discovered lipopolysaccharide receptor. The overall goals of these studies are to unravel the molecular mechanisms of action of these bacterial constituents on host cells and to develop new treatments to prevent pathologic effects of bacterial infections. As a first step towards these goals, this project will test the hypothesis that peptidoglycan and lipopolysaccharide act on host cells through one specific cell surface receptor. This project has six specific aims: (i) To isolate this peptidoglycan-lipopolysaccharide receptor protein from mouse, sheep, and cow lymphocytes and macrophages; (ii) To obtain hamster monoclonal antibodies to this receptor protein; (iii) To prove that these monoclonal antibodies indeed bind to one peptidoglycan-lipopolysaccharide receptor, by demonstrating binding of these antibodies to this receptor on cells and in an isolated form, and by showing inhibition of ligand binding by monoclonal antibodies and inhibition of monoclonal antibody binding by the receptor ligands; (iv) To obtain further evidence for the function of the peptidoglycan-lipopolysaccharide receptor protein as a cell activating receptor by demonstrating agonistic or antagonistic nature of anti-receptor monoclonal antibodies for activation of macrophages and B cells, and modulation of cell surface expression and function of this receptor by these antibodies; (v) To characterize the fine specificity of this receptor by determining the structural requirements of peptidoglycan and lipopolysaccharide for the binding to this receptor, by studying competitive inhibition of ligand binding by a series of natural and synthetic peptidoglycan and lipopolysaccharide partial structures and analogs; (vi) To obtain highly purified peptidoglycan-lipopolysaccharide receptor protein by affinity chromatography with immobilized anti-receptor monoclonal antibodies. These studies will: (i) verify the hypothesis of receptor-mediated activation of lymphocytes and macrophages by two most important bacterial cell wall constituents; (ii) provide new tools to study novel mechanisms of signal transduction in leukocyte activation by bacterial cell wall components; (iii) enable future development of reagents that can prevent or reverse septic shock and other pathologic effects of bacterial products; and (iv) result in a discovery of a potentially clinically useful marker for various leukocyte subpopulations and hematologic neoplasms.

细菌感染仍然是发病的主要原因 死亡率，尤其是免疫功能低下的个体。 肽聚糖和脂多糖可以再现大多数主要体征 以及革兰氏阳性和革兰氏阴性感染的症状 细菌，但它们对宿主细胞作用的分子机制是 几乎不为人所知。 该实验室最近发现了一种 小鼠淋巴细胞上的显性肽聚糖结合蛋白 巨噬细胞，并证明该蛋白质与 最近发现的脂多糖受体。 整体 这些研究的目标是揭示其分子机制 这些细菌成分对宿主细胞的作用并发展 预防细菌病理作用的新疗法 感染。 作为实现这些目标的第一步，该项目将 检验肽聚糖和脂多糖作用的假设 通过一种特定的细胞表面受体作用于宿主细胞。 这 项目有六个具体目标： (i) 隔离这一点 来自小鼠、绵羊的肽聚糖-脂多糖受体蛋白， 以及牛淋巴细胞和巨噬细胞； (ii) 获得仓鼠 该受体蛋白的单克隆抗体； (三) 证明 这些单克隆抗体确实与一个结合 肽聚糖-脂多糖受体，通过证明结合 这些针对细胞上和分离的受体的抗体 形式，并通过显示单克隆配体结合的抑制 抗体和单克隆抗体结合的抑制 受体配体； (iv) 获取该功能的进一步证据 肽聚糖-脂多糖受体蛋白作为细胞 通过表现出激动性或拮抗性来激活受体 抗受体单克隆抗体激活的性质 巨噬细胞和 B 细胞以及细胞表面表达的调节 以及这些抗体对该受体的功能； (v) 至 通过确定来表征该受体的精细特异性 肽聚糖和脂多糖的结构要求 通过研究竞争性抑制来与该受体结合 一系列天然和合成肽聚糖的配体结合 以及脂多糖部分结构和类似物； (六) 至 获得高纯度的肽聚糖-脂多糖受体 固定化抗受体亲和层析法检测蛋白质 单克隆抗体。 这些研究将：（i）验证 受体介导的淋巴细胞激活假说 巨噬细胞由两种最重要的细菌细胞壁成分组成； (ii) 提供新工具来研究新的信号机制 细菌细胞壁转导白细胞活化 成分; (iii) 促进未来试剂的开发 预防或逆转感染性休克和其他病理影响 细菌产品； (iv) 导致发现潜在的 各种白细胞亚群的临床有用标记物和 血液肿瘤。