ULCER-ASSOCIATED MAJOR ANTIGEN OF HELICOBACTER PYLORI

幽门螺杆菌溃疡相关主要抗原

• 批准号: 2458848
• 负责人: MURALI K TUMMURU
• 金额: $ 10.57万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-08-01 至 2000-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2458848
• 关键词: Helicobacter; SDS polyacrylamide gel electrophoresis; bacterial antigens; bacterial genetics; biopsy; cell adhesion; cytokine; duodenal ulcer; endoscopy; enzyme linked immunosorbent assay; humoral immunity; immunoglobulin A; inflammation; interleukin 1; interleukin 8; leukocyte activation disorder; microorganism culture; molecular pathology; peptic ulcer; plasmids; protein purification; site directed mutagenesis; virulence

Helicobacter pylori is a curved, microaerophilic, gram-negative bacterium that is the major cause of chronic gastritis in humans. While most infected persons remain asymptomatic, infection with this pathogen is a significant risk factor for the development of peptic ulcer disease and gastric carcinoma. The major objective of the proposed work is to characterize the role of the 131 kDa immunodominant antigen (CagA; cytotoxin-associated gene product) in the pathogenesis of H.pylori- associated duodenal ulcer disease. About 50-60% of H.pylori isolates produce the CagA antigen, and virtually all H. pylori-infected patients with duodenal ulceration develop a serologic response to CagA. Previous studies have shown that mucosal neutrophil infiltration and epithelial surface degeneration are significantly greater in patients with a local gastric immune response to CagA than in those without such a response. Previous studies have also shown that CagA-producing H. pylori strains induce significantly more interleukin-8 (lL-8) synthesis by gastric epithelial cells than do CagA-negative strains. In addition,recent studies have shown that patients infected with CagA+ strains induce a specific pattern of cytokine expression in gastric tissue involving IL-1 alpha and IL-1 beta. The gene encoding CagA has now been cloned, and recombinant CagA protein is recognized by sera from an H.pylori-infected patients. H.pylori can vary the size of CagA and the size variation is associated with deletion or insertion of repeat sequences. Sequence analysis of the region upstream from cagA, which is not present in CagA strains, identified three genes (cagB, cagC, and cagD ). The deduced CagC sequence was found to have significant homology with the Bordetella pertussis toxin secretion protein (PtIC). The hypothesis of this study is that the CagA antigen of H.pylori is associated with duodenal ulcer disease, and is an important virulence factor. The specific aims are 1) to study the relationship between CagA production and H.pylori-associated duodenal ulcer disease, 2) to determine the role of CagA in gastric inflammation, 3) to characterize the genetic differences between CagA+ and CagA H.pylori strains.To accomplish the first objective, native CagA from H.pylori strain 84-183 will be purified and its structural and biochemical characteristics determined. Next, mucosal immune responses to CagA will be compared in patients with gastritis alone and those with duodenal ulceration. Subsequently, H.pylori isolates from patients with gastritis alone and duodenal ulcer will be tested for CagA size and determine the relationship between the type of CagA expressed and clinical outcome. Isogenic pairs of CagA+ and CagA strains will be used to determine whether CagA plays a role in cytokine induction and neutrophil activation within gastric tissues. The cagA neighboring genes that are present exclusively in CagA+ strains will be characterized and disrupted by insertional mutagenesis, and the mutants will be tested for cytotoxin secretion and/or cytokine induction. Understanding the pathogenic role of CagA and its neighboring gene products in H.pylori infection may aid in the development of strategies to decrease the morbidity associated with complications of H.pylori infection.

幽门螺杆菌是一种弯曲的、微需氧的革兰氏阴性细菌 这是人类慢性胃炎的主要原因。虽然大多数 感染者仍然没有症状，感染该病原体是一种 消化性溃疡疾病发展的重要风险因素， 胃癌拟议工作的主要目标是 表征131 kDa免疫显性抗原（CagA; 细胞毒素相关基因产物）在幽门螺杆菌发病机制中的作用- 相关的十二指肠溃疡疾病。大约50-60%的幽门螺杆菌分离株 产生CagA抗原和几乎所有的H.幽门螺杆菌感染者 十二指肠溃疡患者对CagA产生血清学应答。先前 研究表明，粘膜中性粒细胞浸润和上皮细胞 表面变性在局部 胃对CagA的免疫应答比那些没有这种应答的人要高。 以前的研究也表明，产生CagA的H.幽门螺杆菌菌株 通过胃粘膜诱导显著更多白细胞介素-8（IL-8）合成 上皮细胞比CagA阴性菌株。此外，最近的研究 已经表明，感染CagA+菌株的患者诱导特异性 胃组织中涉及IL-1 α的细胞因子表达模式， IL-1 β。编码CagA的基因现已被克隆和重组， CagA蛋白被来自幽门螺杆菌感染患者的血清识别。 幽门螺杆菌可以改变CagA的大小，并且大小变化与CagA的大小相关。 具有重复序列的缺失或插入。 序列分析 cagA上游的区域，其不存在于CagA菌株中， 鉴定了三个基因（cagB、cagC和cagD）。推导的CagC序列 发现与百日咳杆菌毒素具有显著的同源性 分泌蛋白（PtIC）。本研究假设CagA 幽门螺杆菌抗原与十二指肠溃疡病有关，是一种 重要的毒力因子。具体目标是：1）研究 幽门螺杆菌相关性十二指肠溃疡与CagA的关系 溃疡病，2）确定CagA在胃炎症中的作用， 3)描述CagA+和CagA幽门螺杆菌之间的遗传差异 为了实现第一个目标，将来自幽门螺杆菌的天然CagA 将菌株84-183纯化，并对其结构和生化特性进行分析。 特性决定。接下来，将研究针对CagA的粘膜免疫应答。 比较单纯胃炎和十二指肠溃疡患者 溃疡随后，幽门螺杆菌分离自胃炎患者 单独和十二指肠溃疡将测试CagA大小，并确定 CagA表达类型与临床结局之间的关系。 CagA+和CagA菌株的同基因对将用于确定是否 CagA在细胞因子诱导和中性粒细胞活化中起作用， 胃组织cagA邻近基因只存在于 在CagA+菌株中，将通过插入 诱变，并将测试突变体的细胞毒素分泌和/或 细胞因子诱导了解CagA的致病作用及其 幽门螺杆菌感染的邻近基因产物可能有助于发展 减少与并发症相关的发病率的战略 幽门螺杆菌感染。