Bioelectricity in Gut Epithelium Drives Pathogenic Bacterial Targeting

项目摘要 我们的肠道含有大约100万亿个肠道细菌，它们共同促进营养吸收， 成熟的免疫系统，以及发挥核心作用，保护宿主免受肠道细菌 感染.然而，许多肠道细菌病原体已经发展出在肠道中定殖的策略， 粘膜并引起疾病。临床上重要的肠道细菌，如沙门氏菌、志贺氏菌、耶尔森氏菌和 致病性大肠大肠杆菌，是一个主要的公共卫生问题，由于他们的致病能力，造成严重的 具有潜在致命后果的肠道和肠外疾病及其传播难易程度 通过受污染的食物和水。这些细菌已经发展出共同的策略， 并侵入相对少量的卵泡相关上皮（FAE）细胞，称为微折叠（M）细胞， 引起炎症。极低剂量的污染，有时只有几种病原体， 引起严重肠炎和/或播散性感染。人们仍然不太清楚为什么这么少的细菌 病原体通常被数百万（如果不是数十亿）的肠道微生物包围， 他们的目标入口-FAE的低丰度M细胞。 在此之前，我们已经证明了气管粘液中存在内源性生物电场 恒河猴的上皮细胞，并首次检测到沙门氏菌感染产生的电 场（IGEF）在小鼠盲肠FAE。这些生物电信号在胚胎发育过程中起着关键作用。 发育，组织再生和伤口愈合，以及在播散性感染，因为我们证明 在我们最近的工作中。通过施加模拟IGEF的电场，我们已经证明了Escherosal E。杆菌 迁移到阳极和致病性沙门氏菌迁移到阴极，排他地和同时地。 在这个探索性的R21中，我们提出了一种新的致病细菌生物电控制机制， 面向.我们的中心假设是，在FAE周围存在一个活跃的上皮细胞"电池"， 被细菌病原体本质上用于侵入性靶向。我们将测试我们的假设，通过 以下具体目的：1）在空间上定义和表征肠上皮的生物电活动。使用 先进的电生理学技术，我们将测量和操纵离子电流 FAE和周围绒毛上皮的密度、跨上皮电位和跨膜电位， 离体小鼠盲肠模型。成功完成将建立第一个肠道生物电剖面 上皮2)探讨细菌侵袭性靶向作用中肠上皮细胞生物电的机制。我们 工作假设是肠道病原体利用局部生物电来战略性地靶向FAE， 细菌表面的电特性。这将在基因上进行测试，并在体外和体外实验中得到证实。 vivo.了解生物电特性如何引导病原体进入M细胞可以为未来提供信息 用于预防/治疗胃肠道感染和炎症的药物方法。

Project Summary Our gut contains about 100 trillion commensal bacteria that collectively contribute to nutrient absorption and maturation of the immune system, as well as play a central role in protecting the host from enteric bacterial infections. However, many enteric bacterial pathogens have developed strategies to colonize the intestinal mucosa and cause diseases. Clinically significant enteric bacteria, such as Salmonella, Shigella, Yersinia, and pathogenic E. coli, are a major public health concern due to their pathogenic capacities to cause severe diarrheal and extraintestinal diseases with potentially fatal consequences, and their ease of transmission through contaminated food and water. These bacteria have developed common strategies to specifically target and invade a relatively small number of follicle-associated epithelial (FAE) cells known as Microfold (M) cells to induce inflammation. Contamination with extremely low doses, sometimes with only a few pathogens, can cause severe enteritis and/or disseminated infections. It remains poorly understood how so few bacterial pathogens, which are typically surrounded by millions (if not billions) of commensal microbes, find a way to their targeted portal of entry—the low abundance M cells of the FAE. Previously, we have demonstrated the existence of endogenous bioelectric fields in the tracheal mucus epithelium of the rhesus monkey and, and for the first time, detected Salmonella infection-generated electric fields (IGEF) in mouse cecum FAE. These bioelectrical signals play critical roles during embryonic development, tissue regeneration and wound healing, as well as in disseminated infections as we demonstrate in our most recent work. By applying electric fields mimicking IGEF we have shown that commensal E. coli migrate to the anode and pathogenic Salmonella migrate to the cathode, exclusively and simultaneously. In this exploratory R21, we propose a novel mechanism of bioelectrical control in pathogenic bacterial targeting. Our central hypothesis is that an active epithelial “battery” exists around the FAE, which is intrinsically exploited by bacterial pathogens for invasive targeting. We will test our hypothesis through the following specific aims: 1) Spatially define and characterize bioelectrical activities at gut epithelia. Using advanced electrophysiological techniques, we will measure and pharmacologically manipulate ionic current density, trans-epithelial potential, and transmembrane potential in FAE and surrounding villus epithelium in an ex vivo mouse cecum model. Successful completion will establish the first bioelectricity profile of intestinal epithelium. 2) Dissect the mechanisms of bioelectricity at gut epithelia in bacterial invasive targeting. Our working hypothesis is that enteric pathogens utilize local bioelectricity to strategically target the FAE depending on the surface electrical properties of the bacteria. This will be tested genetically and affirmed in vitro and ex vivo. Understanding how the bioelectric properties guide pathogen entry into M cells could inform future pharmaceutical approaches to prevent/treat gastrointestinal infection and inflammation.