Bile Acids and Gut Microbiome in the Pathogenesis of Inflammation in Cirrhosis

描述(由申请人提供)： 酒精性肝病和肝硬变是美国发病率和死亡率的主要原因之一，在退伍军人健康系统中也是如此。然而，尽管有几个预防和治疗酒精滥用的计划，但很大一部分受影响的退伍军人继续饮酒，即使是在肝硬变的情况下。这可能会导致大量的负担，这种负担源于不受控制的炎症环境和细菌易位导致的内毒素血症，细菌移位导致感染、多器官衰竭和死亡。这些患者是戒酒药物治疗的糟糕候选者，往往没有依从性，并且由于持续饮酒而无法接受肝脏移植。因此，预防这种无节制的炎症和细菌易位是至关重要的；然而，这些背后的确切机制尚不清楚，这是我们知识中的一个主要差距。胆汁酸(BAS)和肠道微生物区系是肠道环境的关键成分，它们的相互作用影响细菌移位和炎症。我们小组一直致力于利用独立于培养的分析来描述微生物区系的作用，以及炎症和BAs对它们的调节，特别是在肝硬变中破坏膜稳定的继发性BAs。我们还发现，经常饮酒的肝硬化症患者粪便继发性碱性磷酸酶和肠道炎性细胞因子的表达显著增加，这些细胞因子可能介导这些患者的肠道损伤并加剧炎症。因此，在肠道微生物区系改变或生物失调的背景下，研究继发性BAS在肠道和全身炎症的发病机制中的作用，并将其与禁酒后的酒精性肝硬变和伴有非酒精性肝病的肝硬变进行比较，对于阐明肠道环境在肝病进展中的作用至关重要。我们的中心假设：肝硬变患者，特别是继续饮酒的酒精性肝硬变患者， 由于与肠道微生物区系改变相关的次级胆汁酸增加，导致肠道和全身炎症加剧。这一中心假设将通过以下两个具体目标进行检验：具体目标1：利用系统生物学方法，确定持续酗酒的肝硬变患者与戒酒和非酒精性肝硬变患者相比，次级胆汁酸对全身和肠道炎症、肠道微生物群和内毒素血症的影响。我们将用系统生物学的方法在100名受试者中研究BAS与十二指肠、回肠、结肠和粪便来源的微生物群与年龄和肝硬变严重程度匹配的积极饮酒的肝硬化组与禁酒性酒精性肝硬化组、伴NAFLD的肝硬化组和健康对照组的全身和肠壁炎症的相互作用。具体目的2：明确胆汁酸和肠道微生物区系在无菌小鼠肠道和全身炎症、内毒素血症和细菌移位的发病机制中的作用。我们将利用无菌小鼠来阐明胆汁酸和微生物区系对内毒素血症、炎症和细菌易位的个体贡献。将使用6组无菌小鼠：1)不干预，2)单独喂养脱氧胆酸，3)以健康人粪便定植，4)酒精性肝硬变粪便定植。各组在定植后将进行肠道和全身炎症、内毒素血症、BA谱(血清、胆汁、肝脏、粪便、肠道内容物)和微生物群变化(第3-4组)的评估，并进行组间比较。这些研究将提供一个新的综合平台，利用前沿的翻译和系统生物学方法研究次级胆汁酸和肠道微生物区系在所有肝硬变患者肠道和全身炎症发展中的作用。我们希望这项提案的结果将增加我们对重点治疗方法的开发的洞察力，使这一未得到充分服务的退伍军人群体受益。

DESCRIPTION (provided by applicant): Alcoholic liver disease and cirrhosis are one of the leading causes of morbidity and mortality in the US, as well as in the VA Health System. However, despite several programs to prevent and treat alcohol abuse, a significant proportion of affected Veterans continue to drink, even in the setting of cirrhosis. This can lead to a substantial burden that stems from an unchecked inflammatory milieu and endotoxemia due to bacterial translocation that precipitates infections, multi-organ failure and death. These patients are poor candidates for abstinence pharmacological therapy, are often non-adherent and cannot be offered liver transplant due to their continued drinking. Therefore prevention of this unchecked inflammation and bacterial translocation is critical; however, the exact mechanisms behind these are unclear, which is a major gap in our knowledge. Bile acids (BAs) and gut microbiota are key components of the intestinal milieu, whose interaction impacts bacterial translocation and inflammation. Our group has focused on delineating the role of microbiota, using culture- independent analyses, with inflammation and their modulation by BAs, especially the membrane-destabilizing secondary BAs in cirrhosis. We also discovered that actively drinking cirrhotics have a highly significant increase in fecal secondary BAs and intestinal inflammatory cytokine expression that may mediate intestinal injury and potentiate inflammation in these patients. Therefore, the study of the role of secondary BAs in the pathogenesis of intestinal and systemic inflammation in the context of altered gut microbiota or dysbiosis, in actively drinking cirrhotics compared to abstinent alcoholic cirrhotics and cirrhotics with non-alcoholic liver disease is critical in delineating the role of the intestinal milieu in the progression of liver disease. Our central hypothesis: Patients with cirrhosis, especially alcoholic cirrhotic patients who continue to drink, develop enhanced intestinal and systemic inflammation due to an increase in secondary bile acids associated with altered gut microbiota. This central hypothesis will be tested using these two specific aims: Specific aim 1: To define the effect of secondary bile acids on systemic and gut inflammation, gut microbiome and endotoxemia in cirrhotic patients with continued alcohol abuse compared to abstinent alcoholic cirrhotics and non-alcoholic cirrhotics using a systems biology approach. We will study the interaction of BAs with microbiome of the duodenal, ileal, colonic and fecal origin with systemic and intestinal wall inflammation in age- and cirrhosis-severity matched actively drinking cirrhotics compared to abstinent alcoholic cirrhotics, cirrhotic with NAFLD and healthy controls using a systems biology approach in 100 subjects. Specific Aim 2: To define the role of bile acids and gut microbiota in the pathogenesis of the increased intestinal and systemic inflammation, endotoxemia and bacterial translocation in a germ-free mouse model after colonization with stool from alcoholic cirrhotic patients. We will be utilizing germ-free mice to elucidate the individual contributions of bile acids and microbiota on endotoxemia, inflammation and bacterial translocation. Six groups of germ-free mice will be used: 1) without intervention 2) fed deoxycholic acid alone 3) colonized with healthy human stool 4) colonized with alcoholic cirrhotic stool. Each group will be evaluated for intestinal and systemic inflammation, endotoxemia, BA profile (serum, gallbladder, liver, fecal, intestinal contents) and microbiome changes (Groups 3-4) will be performed after colonization and compared between groups. The studies will provide a novel integrative platform to study the role of secondary bile acids and gut microbiota in the development of intestinal and systemic inflammation in all cirrhotic patients using cutting-edge translational and systems biology approaches. We expect the results of this proposal to increase our insight into the development of focused therapeutic approaches that benefit this underserved population of Veterans.