Effect of Lactoferrin on Alcohol-Induced Dysbiosis and Gut Barrier Dysfunction

描述（由申请人提供）：在美国，酒精性肝病（ALD）影响超过1000万人，占肝病相关死亡的48%。目前，酒精性肝硬化和癌症的唯一有效治疗选择是肝移植。因此，开发新的治疗策略势在必行。酒精诱导的内毒素血症是导致ALD的关键因素。累积的数据表明，过量的乙醇摄入通过两种主要机制诱导内毒素血症：1）刺激细菌过度生长和2）破坏肠道粘膜屏障功能障碍。我们的初步研究表明，乳铁蛋白（LF）抑制酒精诱导的肠道通透性过高。我们将研究乳铁蛋白可以维持肠道微生物群的平衡并保持肠道的完整性的假设，尽管饮酒，因此，乳铁蛋白可以防止内毒素转移到门静脉，减少内毒素血症，并减弱ALD。所提出的研究的具体目的是：（目的1）确定LF是否通过减少酒精诱导的肠漏和内毒素血症来减弱ALD。将雌性C57 Bl/6 J小鼠分成几组，并用以下食物处理：i）Liber-DeCarli对照流质饮食，ii）含酒精的流质饮食，iii）LF（50 mg/kg），iv）酒精加各种剂量的LF。我们将检查a）口服LF是否阻止内毒素从肠道转移到门静脉并减少酒精诱导的肝脏炎症和损伤;和B）LF对肠道屏障功能的保护作用是否归因于LF诱导的肠道上皮细胞生长/迁移和/或抑制上皮细胞对LPS刺激的炎症反应。(Aim 2）。确定LF是否减弱酒精诱导的肠道生态失调。该目的将分析和比较用i）Liber-DeCarli对照流质饮食、ii）含酒精流质饮食、iii）LF、iv）酒精加LF处理的4组小鼠中的肠道微生物组。我们将研究a）酒精治疗是否增加促炎性肠道微生物（例如，产生革兰氏阴性内毒素的细菌）并减少抗炎微生物（例如，乳酸杆菌、双歧杆菌）;和B）口服给予LF恢复正常的肠道微生物组。(Aim 3）。确定微生物和宿主基因表达网络，不同的酒精和酒精加LF管理。将对微生物和宿主转录组进行分析，以确定酒精和/或LF如何改变肠粘膜中的代谢网络。我们将确定a）口服酒精是否诱导1）产生酒精代谢致病产物的肠道微生物代谢网络和2）指示肠道屏障功能障碍的宿主肠道中的基因表达谱。我们还将研究口服LF是否能改善酒精诱导的微生物和宿主肠道基因表达谱的变化。

DESCRIPTION (provided by applicant): In the U.S., alcohol liver disease (ALD) affects more than 10 million people and accounts for 48% of liver cirrhosis-associated deaths. Currently, the only effective treatment option for alcohol-induced liver cirrhosis and cancer is liver transplantation. Therefore, it is imperative to develop new therapeutic strategies. Alcohol-induced endotoxemia is a critical factor in causing ALD. Accumulating data demonstrate that excess ethanol intake induces endotoxemia through two main mechanisms: 1) stimulation of bacterial overgrowth and 2) disruption of gut mucosal barrier dysfunction. Our preliminary studies demonstrate that lactoferrin (LF) suppresses alcohol-induced gut hyper-permeability. We will examine the hypotheses that lactoferrin can maintain the balance of gut microbiota and preserve the integrity of the intestines despite alcohol consumption, and as a result, lactoferrin prevents the transfer of endotoxin to the portal vein, reduces endotoxemia, and attenuates ALD. The Specific Aims of the proposed studies are: (Aim 1) Determine whether LF attenuates ALD by reducing alcohol-induced gut leakage and endotoxemia. Female C57Bl/6J mice will be divided into several groups and treated with: i) Liber-DeCarli control liquid diet, ii) alcohol-containing liquid diet, iii) LF (50 mg/kg), iv) alcohol plus various doses of LF. We will examine whether a) oral administration of LF prevents endotoxin transfer from the gut to the portal vein and decreases alcohol-induced hepatic inflammation and injury; and b) the protective effects of LF on gut barrier function are due to LF-induced gut epithelial cell growth/migration and/or suppression of the inflammatory response of epithelial cells to LPS stimulation. (Aim 2). Determine whether LF attenuates alcohol-induced enteric dysbiosis. This aim will profile and compare enteric microbiomes in 4 groups of mice treated with i) Liber-DeCarli control liquid diet, ii) alcohol-containing liquid diet, iii) LF, iv) alcohol plus LF. We will investigate whether a) alohol treatment increases the prevalence and/or abundance of pro-inflammatory enteric microbes (e.g., Gram-negative endotoxin producing bacteria) and reduces the occurrence of anti-inflammatory microbes (e.g., lactobacilli, bifidobacteria); and b) oral administration of LF restores the normal enteric microbiome. (Aim 3). Identify microbial and host gene-expression networks that differ following alcohol and alcohol plus LF administration. Microbial and host transcriptomes will be profiled in order to determine how alcohol and/or LF alter the metabolic networks operating in the enteric mucosa. We will determine whether a) oral administration of alcohol induces gene expression profiles in 1) metabolic networks of enteric microbes that produce pathogenic products of alcohol metabolism and 2) host intestines that are indicative of gut barrier dysfunction. We will also examine whether oral administration of LF ameliorates alcohol-induced changes in microbial and host enteric gene expression profiles.