Probiotics and HIF signaling in Alcoholic Liver Disease

描述（申请人提供）：酒精性肝病（ALD）的发病机制是多因素的。动物和人类研究表明，肠道来源的内毒素是导致ALD的关键因素。内毒素来源于革兰氏阴性菌的细胞壁，由于肠道屏障功能受到严格调节，通常仅以微量穿透肠道上皮。然而，内毒素渗漏在病理条件下增加，如酗酒。肠上皮的屏障功能是由细胞旁顶端连接复合物（包括位于上皮细胞顶端的紧密连接和粘附连接）和黏液层提供的。黏液层的保护涉及粘蛋白和许多调节肽，如肠三叶因子（ITFs），它们在屏障的形成和稳定、肠道快速修复（'恢复）的增强、粘膜分化过程的调节和粘膜免疫反应的调节中发挥关键作用。屏障的形成和恢复也依赖于不间断的粘膜血流和持续的糖酵解。肠上皮细胞位于厌氧腔和高代谢固有层之间，形成陡峭的生理氧梯度。生理性和病理性缺氧诱导低氧诱导因子（hypoxia inducible factor， HIF） -1和-2，HIF -1和-2通过上调肠道ITF、粘蛋白和糖酵解酶等多种基因产物，代表了一种主要的适应性反应。我们的初步研究表明，慢性酒精喂养降低肠道HIF-2、ITF和紧密连接表达，导致内毒素血症增加，最终导致肝损伤。补充一种益生菌菌株，鼠李糖乳杆菌GG (LGG)，可以减轻这些有害影响和ALD。然而，其机制尚不清楚。我们的中心假设是，酒精喂养引起有利于氧化应激、炎症和线粒体损伤的变化，从而减少HIF反应信号和靶蛋白，导致肠道屏障功能破坏，增加内毒素血症和随后的ALD，而益生菌补充将通过增强HIF调节反应来减轻酒精诱导的肝损伤。为了验证这一假设，我们将携带以下具体目标：确定乙醇和益生菌对ALD中肠道HIF信号传导的影响及其机制。具体目标2。确定HIF信号是否需要益生菌补充剂的保护作用

DESCRIPTION (provided by applicant): The pathogenesis of alcoholic liver disease (ALD) is multi-factorial. Animal and human studies suggest that gut-derived endotoxin is a critical factor contributing to ALD. Endotoxins are derived from the cell wall of Gram-negative bacteria, and normally penetrate the gut epithelium in only trace amounts due to tightly regulated intestinal barrier function. However, endotoxin leakiness is increased under pathological conditions, such as alcohol abuse. The barrier function of the intestinal epithelium is provided by paracellular apical junction complexes, including tight junctions and adherens junctions located at the apical end of epithelial cells, and by mucus layers. Protection of the mucus layer involves mucins and a number of regulatory peptides, such as intestinal trefoil factors (ITFs), which play critical rols in formation and stabilization of the barrier, enhancement of rapid intestinal repair ('restitution), modulation of mucosal differentiation processes, and modulation of the mucosal immune response. Formation and restitution of the barrier is also dependent on uninterrupted mucosal blood flow and ongoing glycolysis. Intestinal epithelial cells are positioned between an anaerobic lumen and a highly metabolic lamina propria forming a steep physiologic oxygen gradient. Physiological and pathological hypoxia induces hypoxia-inducible factor (HIF) -1 and -2, which represent a major adaptation response by upregulating a variety of gene products such as intestinal ITF, mucins and glycolytic enzymes. Our preliminary studies showed that chronic alcohol feeding decreased intestinal HIF-2, ITF and tight junction expression leading to increased endotoxemia and ultimately liver injury. Supplementation of a probiotic strain, lactobacillus rhamnosus GG (LGG), attenuates those deleterious effects and ALD. However, the mechanisms are unknown. Our central hypothesis is that alcohol feeding causes a change favoring oxidative stress, inflammation and mitochondrial damage, which reduces HIF responsive signaling and the targeted proteins, leading to disruption of gut barrier function, increased endotoxemia and subsequent ALD, and that probiotic supplementation will attenuate alcohol-induced liver injury by potentiating HIF regulated responses. To test this hypothesis, we will carry our following specific aims: Specific Aim 1. Determine the effects of ethanol and probiotics on intestinal HIF signaling and mechanisms in ALD. Specific Aim 2. Determine whether HIF signaling is required for the protective effect(s) of probiotic supplementation against alcohol-induced liver injury. We will use intestine-specific HIF-1¿ and HIF-2¿ knockout mice to evaluate and compare the role of HIF-1¿ and HIF-2¿ in ALD. We will then use prolyl-4 hydroxylase-1 PH.D.1 knockout mice for a gain-of-function strategy to determine whether an increase HIF attenuates alcohol-induced mucus layer dysfunction in ALD. This study will have a major impact on the development of new therapeutic targets for the prevention and treatment of ALD.