Pathogenesis of Helicobacter pylori infection

描述(由申请人提供)： 摘要幽门螺杆菌是一种定植于人体胃的革兰氏阴性细菌。幽门螺杆菌感染与患远端胃癌和消化性溃疡疾病的风险增加有关。在以前的研究中，我们已经对一种分泌型幽门螺杆菌毒素(VacA)进行了详细的分析。幽门螺杆菌基因组包含三个与VacA有远亲关系的基因，每个基因编码一个250 kDa的蛋白质。与VacA类似，这些VacA并列蛋白被预测为由V型(自动转运蛋白途径)分泌。与已经被非常详细地研究的VacA相反，到目前为止，对H.Pylori VacA并列的研究很少。假设：这一建议的假设如下：(1)VacA并列基因在体内具有重要功能；(2)VacA并列基因的表达受转录调控；(3)VacA并列基因由自身转运蛋白途径分泌；(4)VacA并列基因调控幽门螺杆菌与宿主细胞的相互作用。研究目的：本研究的长期目标是了解幽门螺杆菌在人胃粘膜中定植和存活的分子机制，了解幽门螺杆菌感染导致胃癌或消化性溃疡疾病发生的分子机制，并开发有效的预防胃癌和消化性溃疡疾病的策略。具体目标是：(1)分析幽门螺杆菌引起的胃病动物模型中的VacA并列蛋白；(2)分析幽门螺杆菌VacA并列蛋白的转录；(3)分析VacA并列蛋白的分泌和功能活性。方法：构建幽门螺杆菌等基因突变株，研究VacA基因在体内的作用。然后，我们将在幽门螺杆菌感染的动物模型中比较野生型和突变菌株在胃中定植的能力，并确定这些幽门螺杆菌蛋白是否调节胃炎、胃损伤或胃癌的发展。我们将分析在体内和体外编码VacA类似物的基因的转录，并阐明这些基因的转录调控机制。最后，我们将使用蛋白质组学和免疫学相结合的方法来分析VacA paralog的分泌和蛋白降解过程，并比较野生型和突变株与培养的胃上皮细胞的相互作用。 公共卫生相关性： 幽门螺杆菌感染在退伍军人中很常见，退伍军人有患幽门螺杆菌引起的疾病的风险，如胃癌和消化性溃疡。在前往世界各地旅行的过程中，退伍军人面临感染幽门螺杆菌菌株的风险，与美国发现的菌株相比，这种菌株患胃癌的风险相对更高。退伍军人管理局目前每年花费数百万美元用于抑酸药物疗法，以防止幽门螺杆菌引起的并发症。幽门螺杆菌在人体胃内定植并引起胃部疾病的分子机制的研究有望为预防和治疗幽门螺杆菌感染提供新的途径，这将对退伍军人的护理具有重要的临床意义。

DESCRIPTION (provided by applicant): ABSTRACT Helicobacter pylori is a Gram-negative bacterium that colonizes the human stomach. H. pylori infection is associated with an increased risk of cancer of the distal stomach and peptic ulcer disease. In previous studies, we have conducted detailed analyses of a secreted H. pylori toxin (VacA). The H. pylori genome contains three genes that are distantly related to vacA, and each encodes a protein >250 kDa in size. Similar to VacA, these VacA paralogs are predicted to be secreted by a type V (autotransporter pathway). In contrast to VacA, which has been studied in great detail, thus far there has been very little study of H. pylori VacA paralogs. Hypotheses: The hypotheses of this proposal are as follows: (i) VacA paralogs have important functions in vivo; (ii) the expression of VacA paralogs is transcriptionally regulated; (iii) VacA paralogs are secreted by an autotransporter pathway; and (iv) VacA paralogs modulate interactions of H. pylori with host cells. Study Objectives: The long-term goals of this work are to understand the molecular mechanisms that allow H. pylori to colonize and persist in the human gastric mucosa, to understand the molecular mechanisms by which H. pylori infection leads to the development of gastric cancer or peptic ulcer disease, and to develop effective strategies for the prevention of gastric cancer and peptic ulcer disease. The specific objectives are (i) to analyze VacA paralogs in animal models of H. pylori-induced gastric disease; (ii) to analyze the transcription of H. pylori vacA paralogs; and (iii) to analyze the secretion and functional activities of VacA paralogs. Methods: To investigate the role of VacA paralogs in vivo, we will construct isogenic H. pylori mutant strains. We will then compare the ability of wild-type and mutant strains to colonize the stomach in animal models of H. pylori infection, and we will determine whether these H. pylori proteins modulate the development of gastric inflammation, gastric injury or gastric cancer. We will analyze the transcription of genes encoding VacA paralogs both in vivo and in vitro, and we will elucidate the mechanisms by which the transcription of these genes is regulated. Finally, we will use a combination of proteomic and immunologic methods to analyze the secretion and proteolytic processing of VacA paralogs, and we will compare the interactions of wild-type and mutant strains with cultured gastric epithelial cells. PUBLIC HEALTH RELEVANCE: Helicobacter pylori infection is common among Veterans, and Veterans are at risk for development of H. pylori-induced diseases such as stomach cancer and peptic ulcers. During travel to various parts of the world, Veterans are at risk for acquiring H. pylori strains that confer a relatively higher risk of gastric cancer compared to strains found in the United States. The Veterans Administration currently spends millions of dollars each year for acid-suppressive drug regimens to prevent H. pylori-induced complications. Investigation of the molecular mechanisms by which H. pylori colonizes the human stomach and causes gastric diseases is likely to lead to the development of new approaches for the prevention and treatment of H. pylori infection, which would have important clinical implications for the care of Veterans.