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.

Public Health Relevance

Excessive alcohol drinking is a leading cause of chronic liver disease worldwide, which results in a spectrum of liver disorders that range from fatty liver to cirrhosis. The goals of this project are to investigate the effects of probiotic supplementation on alcohol-induced liver injury and the mechanisms related to hypoxia-inducible factor. This study will have a major impact on the development of new therapeutic targets for the prevention and treatment of ALD.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21AA020848-02
Application #
8597993
Study Section
Health Services Research Review Subcommittee (AA)
Program Officer
Radaeva, Svetlana
Project Start
2012-12-10
Project End
2014-11-30
Budget Start
2013-12-01
Budget End
2014-11-30
Support Year
2
Fiscal Year
2014
Total Cost
$160,313
Indirect Cost
$53,438
Name
University of Louisville
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
057588857
City
Louisville
State
KY
Country
United States
Zip Code
40292
Shao, Tuo; Zhao, Cuiqing; Li, Fengyuan et al. (2018) Intestinal HIF-1? deletion exacerbates alcoholic liver disease by inducing intestinal dysbiosis and barrier dysfunction. J Hepatol 69:886-895
Wu, Guicheng; Liu, Yanlong; Liu, Yunhuan et al. (2018) FGF 21 deficiency slows gastric emptying and reduces initial blood alcohol concentration in mice exposed to acute alcohol in fasting state. Biochem Biophys Res Commun 497:46-50
Yuan, Fuqiang; Chen, Xiaopan; Liu, Jie et al. (2017) Up-regulation of Siah1 by ethanol triggers apoptosis in neural crest cells through p38 MAPK-mediated activation of p53 signaling pathway. Arch Toxicol 91:775-784
Li, Fengyuan; Duan, Kangmin; Wang, Cuiling et al. (2016) Probiotics and Alcoholic Liver Disease: Treatment and Potential Mechanisms. Gastroenterol Res Pract 2016:5491465
Liu, Yanlong; Zhao, Cuiqing; Xiao, Jian et al. (2016) Fibroblast growth factor 21 deficiency exacerbates chronic alcohol-induced hepatic steatosis and injury. Sci Rep 6:31026
Chen, Xiaopan; Liu, Jie; Feng, Wen-ke et al. (2015) MiR-125b protects against ethanol-induced apoptosis in neural crest cells and mouse embryos by targeting Bak 1 and PUMA. Exp Neurol 271:104-11
Shi, Xue; Wei, Xiaoli; Yin, Xinmin et al. (2015) Hepatic and fecal metabolomic analysis of the effects of Lactobacillus rhamnosus GG on alcoholic fatty liver disease in mice. J Proteome Res 14:1174-82
Zhao, Cuiqing; Liu, Yanlong; Xiao, Jian et al. (2015) FGF21 mediates alcohol-induced adipose tissue lipolysis by activation of systemic release of catecholamine in mice. J Lipid Res 56:1481-91
Zhang, Min; Wang, Cuiling; Wang, Chunhong et al. (2015) Enhanced AMPK phosphorylation contributes to the beneficial effects of Lactobacillus rhamnosus GG supernatant on chronic-alcohol-induced fatty liver disease. J Nutr Biochem 26:337-44
Zhao, Haiyang; Zhao, Cuiqing; Dong, Yuanyuan et al. (2015) Inhibition of miR122a by Lactobacillus rhamnosus GG culture supernatant increases intestinal occludin expression and protects mice from alcoholic liver disease. Toxicol Lett 234:194-200

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