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

描述(申请人提供)：在美国，酒精性肝病(ALD)影响着1000多万人，占肝硬变相关死亡的48%。目前，治疗酒精性肝硬变和癌症的唯一有效选择是肝移植。因此，开发新的治疗策略势在必行。酒精性内毒素血症是酒精性肝病的重要致病因素。越来越多的数据表明，过量的乙醇摄入通过两个主要机制诱导内毒素血症：1)刺激细菌过度生长；2)破坏肠粘膜屏障功能。我们的初步研究表明，乳铁蛋白(LF)可抑制酒精诱导的肠道高通透性。我们将检验乳铁蛋白可以维持肠道微生物区系平衡和保持肠道完整性的假说，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，并减轻ALD。建议研究的具体目的是：(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轻ALD。将雌性C57BL/6J小鼠分为几组，分别给予：(1)Liber-DeCarli对照液体饲料；(2)含酒精液体饲料；(3)LF(50 mg/kg)；我们将研究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；以及b)LF对肠道屏障功能的保护作用是由于LF诱导肠上皮细胞生长/迁移和/或抑制上皮细胞对内毒素刺激的炎症反应。(目标2)。确定LF是否能减轻酒精引起的肠道菌群失调。这一目标将描述和比较4组小鼠的肠道微生物群，其中I)Liber-DeCarli对照液体饮食，II)含酒精液体饮食，III)LF，IV)酒精+LF。我们将研究a)酒精治疗是否增加促炎肠道微生物(例如，革兰氏阴性内毒素产生菌)的患病率和/或丰度，并减少抗炎微生物(例如，乳杆菌、双歧杆菌)的出现；以及b)口服LF可恢复正常的肠道微生物群。(目标3)。确定在酒精和酒精加LF给药后不同的微生物和宿主基因表达网络。将对微生物和宿主转录本进行分析，以确定酒精和/或LF如何改变肠道粘膜中的代谢网络。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的代谢网络和2)表明肠道屏障功能障碍的宿主肠道的基因表达谱。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.