Mechanisms of Probiotics in Alcoholic Liver Disease

 描述（由申请人提供）：重度酒精性肝病（ALD）具有较高的发病率和死亡率。最近的研究表明，益生菌逆转酒精诱导的肝脏脂肪变性和炎症，并改善动物模型和患者的肝酶。我们的实验室表明，益生菌菌株，鼠李糖乳杆菌Gorbach-Goldin（LGG）的管理，显着改善酒精治疗小鼠的肝酶和组织学。然而，LGG的有益作用机制，无论是活菌形式还是培养上清液形式，都不清楚。我们的总体假设是，LGG增加肠道cathelicidin相关的抗菌肽（CRAMP），肠三叶因子（ITF）的表达和肠道短链脂肪酸（SCFA），导致改善肠道微生物群的稳态，肠道屏障的完整性和肝脏脂肪代谢，这些可能是LGG补充剂减轻酒精诱导的肝损伤的机制。我们评估LGG作用的机制有4个具体目标：目标1将确定CRAMP在酒精介导的肠道微生物群变化中的作用，以及CRAMP在实验性ALD中LGG有益作用中的作用。我们将使用ALD的CRAMP敲除小鼠模型，使用宏基因组方法确定CRAMP和LGG在酒精诱导的肠道微生物群变化中的作用。目的2将确定LGG在促进ITF表达中的作用，并评估ITF是否积极调节肠紧密连接作为改善实验性ALD肠屏障完整性的机制。我们将使用分泌ITF的杯状细胞系来评估LGG在ITF产生中的作用以及ITF的作用。 在上皮细胞系中调节肠紧密连接。我们还将研究口服重组人ITF减轻酒精诱导的肠屏障和肝损伤的策略。目的3探讨丁酸在LGG对实验性ALD大鼠小肠紧密连接的保护作用中的作用及机制。代谢组学方法将用于分析肠腔中酒精和LGG干预诱导的SCFA变化。我们将探索丁酸在调控occludin表达中的潜在机制，包括肠组蛋白脱乙酰酶（HDAC）、miR 122 a和体外和体内HIF-1a介导的信号传导。目的4评价和比较LGG上清液（LGG）、活LGG（vLGG）和热灭活LGG（hiLGG）在小鼠模型中预防/治疗ALD的有效性。将在脂肪变性、脂肪性肝炎和纤维化的ALD的三种实验模型中测试这三种LGG制剂在ALD的预防/治疗中的有效性。该研究将对开发基于益生菌的预防和治疗ALD的新治疗策略产生重大影响。

 DESCRIPTION (provided by applicant): Severe alcoholic liver disease (ALD) has a high morbidity and mortality. Recent studies demonstrated that probiotics reversed alcohol-induced hepatic steatosis and inflammation, and improved liver enzymes in animal models and in patients. Our laboratory showed that administration of a probiotic strain, Lactobacillus rhamnosus Gorbach-Goldin (LGG), significantly improved liver enzymes and histology in alcohol treated mice. However, the beneficial mechanisms of action of LGG, either in viable bacteria form or in culture supernatant form, are not clear. Our overall hypothesis is that LGG increases intestinal cathelicidin-related antimicrobial peptide (CRAMP), intestinal trefoil factor (ITF) expression and intestinal short chain fatty acids (SCFAs), leading to improved gut microbiota homeostasis, intestinal barrier integrity and hepatic fat metabolism, and these are likely the mechanisms by which LGG supplementation attenuates alcohol induced liver injury. We evaluate mechanisms of LGG action with 4 specific aims: Aim 1 will determine the role of CRAMP in the alcohol-mediated changes in gut microbiota and the role of CRAMP in the beneficial effects of LGG in experimental ALD. We will use CRAMP knockout mouse model of ALD to determine the role of CRAMP and LGG in alcohol-induced changes in gut microbiota using a metagenomic approach. Aim 2 will determine the role of LGG in promoting ITF expression and evaluate whether ITF positively modulates intestinal tight junctions as a mechanism for improved intestinal barrier integrity in experimental ALD. We will use an ITF-secreting goblet cell line to evaluate the role of LGG in the production of ITF and the role of ITF in the modulation of intestinal tight junctions in an epithelial cell line. We will also examine th strategy that oral administration of recombinant human ITF attenuates alcohol-induced intestinal barrier and liver injury. Aim 3 will determine the role and mechanisms of butyric acid in the beneficial effects of LGG on intestinal tight junctions in experimental ALD. A metabolomics approach will be used to analyze the changes in SCFAs induced by alcohol and LGG intervention in the intestinal lumen. We will explore the potential mechanisms underlying the role of butyric acid in the regulation of occludin expression involving intestinal histone deacetylases (HDACs), miR122a, and HIF--mediated signaling in vitro and in vivo. Aim 4 will evaluate and compare the effectiveness of LGG supernatant (LGGs) with viable LGG (vLGG) and heat-inactivated LGG (hiLGG) in the prevention/therapy of ALD in mouse models. These three LGG preparations will be tested for the effectiveness in the prevention/therapy of ALD in three experimental models of ALD of steatosis, steatohepatitis and fibrosis. This study will have a major impact on the development of probiotic-based new therapeutic strategy for the prevention and treatment of ALD.