Arginase, Nitric oxide, and the Defective Immune Response to Helicobacter pylori

精氨酸酶、一氧化氮和对幽门螺杆菌的免疫反应缺陷

• 批准号: 7406308
• 负责人: Nuruddeen D Lewis
• 金额: $ 4.1万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-03-14 至 2012-03-13
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7406308
• 关键词: Acute; Address; American; Antibiotics; Apoptosis; Arginine; Bacteria; Biological; Biomedical Research; Carcinogens; Cell Line; Chronic; Chronic Gastritis; Class; Clinical; Enzymes; Figs - dietary; Gastric Adenocarcinoma; Gastric Tissue; Generations; Goals; Gram-Negative Bacteria; Helicobacter Infections; Helicobacter pylori; Histology; Host Defense; Hour; Human; Immune response; In Vitro; Individual; Infection; Inflammation; Laboratories; Life; Mammals; Metabolism; Modeling; Mucosal Immune Responses; Mus; Nitric Oxide; Nitric Oxide Synthase; Ornithine; Ornithine Decarboxylase; Pathway interactions; Patients; Peptic Ulcer; Pharmaceutical Preparations; Polyamines; Population; Production; Proteins; Public Health; Putrescine; Research; Research Personnel; Research Proposals; Risk Factors; Role; Screening procedure; Small Interfering RNA; Spermidine; Spermine; Stomach; System; Time; Training; Translations; Wild Type Mouse; World Health Organization; arginase; bactericide; burden of illness; career; desire; human NOS2A protein; in vivo; inhibitor/antagonist; insight; killings; macrophage; oxidation; pathogen; protein expression; response; small hairpin RNA

DESCRIPTION (provided by applicant): The overall objective of this proposal is to provide a training vehicle that will allow the applicant to be prepared for a successful career as an independent investigator in biomedical research. The research is focused on the bacterial pathogen Helicobacter pylori, which persists in the human stomach for the life of the host despite a vigorous innate and adaptive mucosal immune response. The long-term goal is to elucidate the role of the enzyme arginase in inducible nitric oxide (NO) synthase (iNOS)-dependent host defense against H. pylori. The hypothesis of this proposal is that L-arginine (L-Arg) metabolism by arginase II is pathogenic in H. pylori infection by restricting iNOS-dependent host defense and by contributing to the inflammation by generation of polyamines. The specific aims are: Aim 1, in vitro: To demonstrate that L-Arg metabolism by arginase II has important biological effects in the macrophage response to H. pylori. We propose that arginase II effectively competes with iNOS for L-Arg and inhibits iNOS translation and NO synthesis by decreasing the intracellular concentration of L-Arg. Using RAW 264.7 murine macrophages treated with arginase inhibitors and arginase II-specific siRNA/shRNA, and gastric macrophages isolated from arginase ll-/- and wild-type mice, we will assess: A.) i. iNOS expression, protein translation, and activity; and ii. killing of H. pylori; and B.) apoptosis and polyamine levels. Aim 2, in vivo: To determine the biological importance of L-Arg metabolism by arginase II in H. pylori infection. We will use two models of H. pylori infection, a chronic model in which mice are challenged with H. pylori for four months, and an acute model in which mice are challenged for 48 hours. Using both systems, wild-type mice will be treated with arginase inhibitors, and arginase ll-/- and wild-type mice will be compared. At these time points, the following will be assessed: A.) H. pylori colonization, histology, and iNOS expression in gastric tissues; B.) iNOS expression, NO generation, and apoptosis in isolated gastric macrophages. It is expected that these studies will provide new insights into the immunopathogenesis of H. pylori infection. Relevance to public health: H. pylori infects approximately 50% of the world's population and 20-40% of Americans. It causes chronic gastritis in all individuals and is the major cause of gastric adenocarcinoma in the world and the primary cause of peptic ulcer disease that is not drug-induced. Because 10-20% of all of those infected will present with clinical manifestations, the disease burden is enormous. Not all patients can be successfully treated with antibiotics, and there is no screening strategy in place; therefore, greater understanding of the reasons for the failed host immune response, as is being pursued in this study, are greatly needed.

描述（由申请人提供）：该提案的总体目标是提供一种培训工具，使申请人能够为作为生物医学研究独立研究者的成功职业生涯做好准备。该研究的重点是细菌病原体幽门螺杆菌，尽管宿主有强烈的先天性和适应性粘膜免疫反应，但它在宿主的一生中一直存在于人的胃中。长期目标是阐明精氨酸酶在诱导型一氧化氮（NO）合酶（iNOS）依赖性宿主防御幽门螺杆菌中的作用。该提议的假设是精氨酸酶 II 的 L-精氨酸 (L-Arg) 代谢通过限制 iNOS 依赖性宿主防御并通过产生多胺促进炎症而在幽门螺杆菌感染中致病。具体目标是： 目标 1，体外：证明精氨酸酶 II 代谢 L-Arg 在巨噬细胞对幽门螺杆菌的反应中具有重要的生物学效应。我们认为精氨酸酶 II 有效地与 iNOS 竞争 L-Arg，并通过降低细胞内 L-Arg 浓度来抑制 iNOS 翻译和 NO 合成。使用用精氨酸酶抑制剂和精氨酸酶 II 特异性 siRNA/shRNA 处理的 RAW 264.7 鼠巨噬细胞，以及从精氨酸酶 II-/- 和野生型小鼠中分离的胃巨噬细胞，我们将评估： A.) i． iNOS 表达、蛋白质翻译和活性； ii.杀死幽门螺杆菌； B.) 细胞凋亡和多胺水平。目标 2，体内：确定精氨酸酶 II 在幽门螺杆菌感染中 L-Arg 代谢的生物学重要性。我们将使用两种幽门螺杆菌感染模型，一种是小鼠受到幽门螺杆菌攻击四个月的慢性模型，另一种是小鼠受到幽门螺杆菌攻击48小时的急性模型。使用这两个系统，用精氨酸酶抑制剂治疗野生型小鼠，并对精氨酸酶II-/-和野生型小鼠进行比较。在这些时间点，将评估以下内容：A.) 胃组织中幽门螺杆菌定植、组织学和 iNOS 表达； B.) 分离的胃巨噬细胞中 iNOS 表达、NO 生成和凋亡。预计这些研究将为幽门螺杆菌感染的免疫发病机制提供新的见解。与公共卫生的相关性：幽门螺杆菌感染了世界上大约 50% 的人口和 20-40% 的美国人。它会引起所有个体的慢性胃炎，是世界上胃腺癌的主要原因，也是非药物引起的消化性溃疡病的主要原因。由于10-20%的感染者会出现临床表现，疾病负担是巨大的。并非所有患者都能成功地接受抗生素治疗，并且没有适当的筛查策略；因此，正如本研究所追求的那样，迫切需要更好地了解宿主免疫反应失败的原因。