From intestinal dysbiosis to alcohol-associated liver disease

Article information

Clin Mol Hepatol. 2020;26(4):595-605

Publication date (electronic) : 2020 September 11

doi : https://doi.org/10.3350/cmh.2020.0086

Beatriz Garcia Mendes ¹^,², Bernd Schnabl^,²

¹Department of Clinical Analysis, Federal University of Santa Catarina, Florianopolis, SC, Brazil

²Department of Medicine, University of California San Diego, La Jolla, CA, USA

Corresponding author : Bernd Schnabl Department of Medicine, University of California San Diego, MC0063, 9500 Gilman Drive, La Jolla, CA 92093, USA Tel: +1-858-822-5311, Fax: +1-858-822-5370 E-mail: beschnabl@ucsd.edu

Editor: Sang Gyune Kim, Soonchunhyang University College of Medicine, Korea

Received 2020 April 29; Revised 2020 July 8; Accepted 2020 July 27.

Abstract

Alcohol-associated intestinal dysbiosis and bacterial overgrowth can lead to a dysregulation of tryptophan metabolism and lower production of indoles. Several of these indole derivatives are aryl hydrocarbon receptor ligands that, in turn, are involved in antimicrobial defense via induction of interleukin-22 (IL-22). IL-22 increases the expression of intestinal regenerating islet-derived 3 (Reg3) lectins, which maintain low bacterial colonization of the inner mucus layer and reduce bacterial translocation to the liver. Chronic alcohol consumption is associated with reduced intestinal expression of Reg3β and Reg3γ, increased numbers of mucosa-associated bacteria and bacterial translocation. Translocated microbial products and viable bacteria reach the liver and activate the innate immune system. Release of inflammatory molecules promotes inflammation, contributes to hepatocyte death and results in a fibrotic response. This review summarizes the mechanisms by which chronic alcohol intake changes the gut microbiota and contributes to alcohol-associated liver disease by changing microbial-derived metabolites.

Keywords: Dysbiosis; Tryptophan; Aryl hydrocarbon receptor; Interleukin-22; Alcohol-associated liver disease

INTRODUCTION

Alcohol consumption is one of the major causes of chronic liver disease in Western countries. In the United States, according to the 2018 National Survey on Drug Use and Health, 14.4 million adults ages 18 and older (5.8% of this age group) had alcohol use disorder, including 9.2 million men (7.6% of men in this age group) and 5.3 million women (4.1% of women in this age group). In 2018, of the 83,517 liver disease deaths among individuals ages 12 and older, 47.8% involved alcohol [1].

The global burden of alcohol-associated liver disease is immense and comprises relatively mild and reversible alcohol-associated hepatic steatosis (fatty liver) to fibrosis and cirrhosis, and alcoholic hepatitis [2,3]. Besides liver diseases, alcohol use is linked to multiple and chronic diseases, including increased risk of cancers [4]; cardiovascular disease [5]; pancreatitis [6]; disruption in the circadian clock [7]; and impaired immune function increasing the susceptibility to bacterial and viral infections [8]. The susceptibility of patients with alcohol use disorder to develop alcohol-associated liver disease is variable indicating that, although alcohol is necessary, it is not enough to cause progressive organ dysfunction [9,10]. Consequently, factors other than the toxicity of alcohol are involved in generating health complications, one of which may be alcohol-induced changes in intestinal microbiota composition and/or function [11]. Other risk factors for progressive alcohol-associated liver disease is the amount of consumed alcohol (>1 drink/day for women, >2 drinks per day for men), drinking pattern (drinking without meal, binge drinking), genetic factors, female gender, smoking, increased body mass index and concomitant chronic liver diseases [12,13].

This review summarizes the mechanisms by which chronic alcohol intake changes the intestinal microbiota and contributes to alcohol-associated liver disease.

INTESTINAL DYSBIOSIS

The intestinal microbiota is the community of microorganisms (bacteria, archaea, fungi and viruses) that reside in the gut [14]. The human gut microbiota houses more than 10 different bacterial phyla, and there is a balance between commensal and pathogenic microbes under homeostatic conditions [15]. Dysbiosis occurs when disease or environmental factors disrupt this microbial balance contributing to the manifestation or continuation of a given disease that cannot be attributed to a single bacterial species [16,17]. Alcohol use is associated with enteric dysbiosis and intestinal bacterial overgrowth in both preclinical models and patients with alcohol abuse [18-20].

Alcohol-associated changes in the enteric microbiota are required for the development of the liver disease because intestinal decontamination with non-absorbable antibiotics (polymyxin B and neomycin) prevents alcohol-associated intestinal bacterial overgrowth and dysbiosis in mice. Importantly, reducing the intestinal bacterial burden suppressed subclinical intestinal inflammation after chronic alcohol feeding, stabilized the gut barrier and reduced ethanol-induced steatohepatitis in mice [21,22]. In ethanolfeed rats, the same antibiotic cocktail prevented liver injury and reduced the hepatic pathology score (including steatosis, inflammation, and necrosis) [23].

Interestingly, Lactobacillus was strongly suppressed and almost absent in mice fed intragastric ethanol for 3 weeks as compared with control (isocaloric) fed animals [19]. However, treatment with prebiotic fructooligosaccharides – a stimulator of beneficial bacteria growth such as lactobacilli and bifidobacteria – improved ethanol-induced steatohepatitis by inducing gene and protein expression of the bactericidal c-type lectins regenerating islet derived 3 gamma (Reg3γ) and by reducing intestinal bacterial overgrowth [19].

In humans, chronic alcohol consumption alters the composition of mucosa-associated microbiota with a lower abundance of Bacteroidetes and a higher abundance of Proteobacteria in a subset of alcoholic patients with and without liver disease compared with healthy controls [20]. While phylum Bacteroidetes is involved in carbohydrates fermentation leading to short-chain fatty acid production (mainly acetate and propionate) [24,25], Proteobacteria are known to produce lipopolysaccharides (LPS), a potent activator of the toll-like receptor (TLR)-4 [26]. The fecal microbiota of patients with alcohol use disorder and alcohol-associated liver disease was characterized by quantitative and qualitative alterations, with a reduction of bacterial diversity, reduction of Akkermansia and increase of Bacteroides. Akkermansia muciniphila produces shortchain fatty acids such as acetic acid from mucin and supplies energy to goblet cells, improving the intestinal barrier function [27]. Moreover, several reports indicate its effects on glucose and lipid metabolism, and that certain food ingredients such as polyphenols may increase its abundance in the gut [28]. In alcohol-dependent patients with high intestinal permeability, the level of Faecalibacterium prausnitzii, a bacterial species known for its antiinflammatory properties, was decreased. Conversely, those patients had higher plasma interleukin (IL)-8 levels, an inflammatory cytokine [29]. Intestinal dysbiosis has been associated with the severity of alcohol dependence and cirrhosis, and deteriorating dysbiosis is associated with cirrhosis progression [30]. Severe alcoholic hepatitis was associated with higher fecal proportions of Bifidobacteria, Streptococcus, Enterobacteria [30,31], and Enterococcus [30], and fewer proportions of Atopobium [30]. For example, differences in fecal microbiota composition were observed in patients with alcohol use disorder and alcoholic hepatitis as compared with non-alcoholic subjects [31]. In patients with alcoholic hepatitis, 5.59% of fecal bacteria were Enterococcus spp., compared with almost none in controls (0.023%) [31]. Fecal samples from patients with alcoholic hepatitis had about 2,700-fold more Enterococcus faecalis (E. faecalis) than non-alcoholic controls [31]. The exotoxin cytolysin, secreted by E. faecalis, was discovered to exert a deleterious effect on ethanol-induced liver disease in mice [31]. The presence of cytolysin-positive (cytolytic) E. faecalis correlates with liver disease severity and mortality in patients with alcoholic hepatitis [31], but no correlation was found in patients with non-alcoholic fatty liver disease [32].

Bacterial infections are a serious complication of cirrhosis, as they can lead to decompensation, multiple organ failure, and/or death. Because bacterial translocation from the gut lumen to extraintestinal sites causes infections, prevention of infections is mostly based on the use of orally administered, poorly absorbed antibiotics (known as selective intestinal decontamination) [33,34]. Several antibiotics were tested and/or used for this purpose, such as polymyxin, neomycin, gentamycin, colistin, paromomycin, and trimethoprim/sulfamethoxazole [35,36]. Currently, norfloxacin and rifaximin are the forms of selective intestinal decontamination for which there is the most evidence in cirrhosis [37-39]. However, the major drawback of routine antibiotic prophylaxis is the emergence of multidrug-resistant organisms [34]. Thus, novel therapies have been proposed based on their ability to modify the altered intestinal microbiota such as probiotics and prebiotics [19,40-45], fecal microbiota transplantation [46,47], phage therapy [31], among others. Evidence for their use in clinical practice is limited and all require further studies.

CHANGES IN MICROBIAL METABOLITES

Changes in the gut microbiota affect the host immune system by altering tryptophan metabolism [48]. Furthermore, endogenous tryptophan metabolites synthesized by the host (kynurenines, serotonin, and melatonin), and bacterial metabolites (indole, indole derivatives, skatole, and tryptamine) play an important role in regulating intestinal and systemic immune homeostasis [49]. Detailed pathways of tryptophan metabolism were recently reviewed by Hendrikx and Schnabl [50].

Several bacterial species convert tryptophan into indole and indole derivatives mainly via the enzyme tryptophanase, which is expressed in many gram-negative, as well as gram-positive bacterial species including Escherichia coli, Clostridium spp. and Bacteroides spp. [51]. Diverse indole derivatives, such as indole-3-aldehyde, indole-3-acetic acid, indole-3-propionic acid, indole-3-acetaldehyde, and indole acrylic acid bind and activate the aryl hydrocarbon receptor (AhR) [48-50]. AhR is a cytosolic ligand-activated transcription factor that is important in xenobiotic metabolism and serves as a regulator of immunity and inflammation, which involves modulating adaptive immunity and gut barrier function [48,49]. Activated AhR acts as anti-inflammatory signaling pathway that regulates the development of intraepithelial lymphocytes and innate lymphoid cells, which are important in the defense against invading pathogens and maintenance of intestinal homeostasis [52]. Moreover, AhR has been implicated in antimicrobial defense via induction of IL-22 expression by group 3 innate lymphoid cells (ILC3) [53,54]. IL-22 further regulates the microbial composition and enhances antimicrobial defense via the induction of antimicrobial proteins [52].

Dietary tryptophan supplementation altered intestinal microbial composition and diversity, improved intestinal mucosal barrier function, activated AhR signaling, and downregulated expression of inflammatory cytokines in the large intestine of weaned piglets [55]. Moreover, the metabolite indole-3-acetic acid, produced by Bacteroides spp. and Clostridium spp. [51], modulated inflammatory responses of hepatocytes and macrophages attenuating release of pro-inflammatory cytokines and cytokine-induced lipogenesis [56]. In a mouse model of ethanol-induced liver disease, ethanol-associated dysbiosis reduced intestinal levels of indole-3-acetic acid and activation of the AhR, which resulted in a decreased expression of IL-22 in the intestine and reduced expression of Reg3γ. Oral supplementation of indole-3-acetic acid protected mice from ethanol-induced steatohepatitis by inducing intestinal expression of IL-22 and Reg3γ, which prevented bacterial translocation to the liver [22]. The prevention of liver damage by non-absorbable antibiotics was associated with restored expression of IL-22 mRNA in lamina propria cells and IL-23-driven production of IL-22 by ILC3 [22], most likely by increasing intestinal levels of indole-3-lactic acid [57] and/or possibly other indole derivatives. Indole-3-lactic acid, produced by Bifidobacterium [58] and Lactobacillus spp. [57,59], have been reportedly involved in inducing immunoregulatory T cells [57] and in suppressing inflammatory T cells [60]. Indole was also shown to prevent LPS-mediated detrimental effects in the liver by downregulation of inflammatory mediators [61]. Metabolomic analysis revealed that fecal and serum levels of tryptophan were decreased in alcoholic hepatitis patients when compared with controls [62]. In line with the decrease of fecal levels of tryptophan-derived metabolites, indole-3-acetic acid, indole-3-propionic acid, and indole-3-lactic acid were also significantly reduced in alcoholic hepatitis patients [22,62].

The metabolic syndrome is associated with reduced capacity of the microbiota to metabolize tryptophan into derivatives that can activate AhR. Fecal samples of individuals with metabolic syndrome contain low levels of tryptophan-based metabolites and have reduced AhR activity. AhR ligand deficiency was also observed in mice fed a high-fat diet [63]. Treatment with either an AhR agonist or a Lactobacillus strain, to compensate for the impaired microbiota-derived AhR ligand signaling, reduces glucose intolerance and liver steatosis in animal models [63]. Depletion of indole-3-acetic acid in the liver and cecum was observed in mice fed a high-fat diet compared with those fed a regular diet [56]. This study also demonstrated that indole-3-acetic acid attenuates the release of inflammatory cytokines that induce the liver synthesis of free fatty acids, which in turn stimulate macrophages. Moreover, indole-3-acetic acid alleviates lipogenesis mediated by cytokine and free fatty acids via its direct action on hepatocytes in an AhR-dependent manner [56]. The treatment with indole-3-acetic acid attenuates high-fat diet-induced NAFLD in mice, as evidenced by amelioration of insulin resistance, lipid metabolism, oxidative stress, and inflammation [64].

In summary, fatty liver disease-associated dysbiosis results in changes of bacterial-derived metabolites, such as tryptophan metabolites. Indole derivatives are ligands for AhR and are important in immunoregulation and host defense.

IL-22

As aforementioned, gut microbiota can stimulate IL-22 expression via the production of tryptophan metabolites. IL-22, a member of the IL-10 family of cytokines, is produced by ILC3 upon stimulation and other immune cells such as Th17, Th22, natural killer cells, and γδ T cells [65]. Specific myeloid cells may also produce IL-22 under certain circumstances such as mouse macrophage-derived IL-22 produced in response to ethanol-induced cell death [66].

IL-22 acts via a transmembrane receptor complex that consists of two different subunits, IL-22R1 and IL-10R2 [67]. In hepatocytes, the biological effect of IL-22 is mediated by activation of the signal transducer and activator of transcription 3 (STAT3) and subsequent induction of anti-apoptotic and proliferative genes [68]. IL-22 is unusual among most interleukins because it does not directly regulate the function of immune cells. Rather, IL-22 targets cells at outer-body barriers, such as the skin and tissues of the digestive and respiratory systems, as well as cells of the pancreas, liver, kidney, and joints [67], controlling bacterial infection, homeostasis, and tissue repair. Thus, IL-22 is produced by immune cells and targets epithelial cells in several organs, including the intestine and the liver.

Numerous in vitro and in vivo studies have shown that IL-22 has many benefits to the liver, such as preventing hepatitis [69], stimulating liver regeneration [70], improving fatty liver [71,72], and alcoholic liver disease [68,73], and alleviating liver fibrosis [74]. In a mouse model of ethanol-induced liver disease, intestinal IL-22 has the beneficial effect of reducing bacterial translocation in the intestine by increasing the expression of Reg3γ. Bacteria engineered to produce IL-22 in the intestine (without increasing systemic IL-22) ameliorate experimental ethanol-induced steatohepatitis via induction of Reg3γ [22]. Besides, IL-22 treatment may effectively inhibit bacterial infection [75] and ameliorate kidney injury [76], two deleterious conditions that are often associated with severe alcoholic hepatitis and contribute to death of patients. More importantly, IL-22 is a promising drug for the treatment of alcoholic hepatitis because of its hepatoprotective and antifibrotic effects and relatively few know side effects during short-term use [76-78].

In addition to the direct effects on liver cells, IL-22 is a key cytokine that links intestinal immune activation to epithelial cell repair and barrier protection following damage [79]. Intestinal epithelial cells express the IL-22 receptor complex that binds IL-22 resulting in the induction of antimicrobial peptides and regenerative pathways that collectively aid in limiting bacterial invasion while promoting epithelial proliferation, wound healing, and repair [65,80,81].

ANTIMICROBIAL PEPTIDES

Chemical barriers consist of antimicrobial peptides, including defensins, lysozymes, secretory phospholipase A2, and C-type lectins, mainly involved in the segregation of gut bacteria and intestinal epithelial cells in the intestine [82]. Among those antimicrobial peptides, C-type lectins regenerating islet-derived 3 beta (Reg3β) and Reg3γ are abundantly expressed in intestinal epithelial cells and Paneth cells of the small intestine and act to maintain the inner mucus layer devoid of bacterial colonization [50,82,83]. They are upregulated upon bacterial colonization of the gut and during intestinal infection and inflammation, thereby contributing to the spatial segregation of intestinal bacteria and the epithelium [84,85]. Reg3γ is bactericidal against gram-positive bacteria by binding to peptidoglycan on the bacterial cell surface while Reg3β binds directly to LPS protecting mice against gram-negative bacteria [86-89]. Reg3β and Reg3γ also influence crypt regeneration, epithelial cell proliferation, and protect intestinal stem cells and Paneth cells from undergoing apoptosis during tissue damage [87,90,91].

Chronic alcohol consumption was associated with lower intestinal levels of Reg3β and Reg3γ in the small intestine [19,91]. Treatment with prebiotics partially restored Reg3γ protein levels, reduced bacterial overgrowth and ameliorated ethanol-induced steatohepatitis [19]. Intestinal deficiency in Reg3β and Reg3γ increases numbers of mucosa-associated bacteria, enhances bacterial translocation and promotes the progression of ethanol-induced fatty liver disease toward steatohepatitis, whereas intestine-specific overexpression of Reg3γ protects mice against it [88]. Duodenal biopsies from alcohol-dependent patients showed lower levels of Reg3γ [19] and an increased number of bacteria covering small intestinal mucosa surfaces than duodenal biopsies from non-alcoholic individuals [88]. Further, deficiency of mucin-2, the most abundantly secreted mucin in the small and large intestine, is associated with higher Reg3β and Reg3γ expression by Paneth cells and enterocytes [84], protecting mice against ethanol-induced liver disease [92]. These data indicate that antimicrobial defense plays an important role in preventing bacterial translocation and protects against alcohol-associated liver disease development [50,93,94].

BACTERIAL TRANSLOCATION

Ethanol is associated with increased intestinal permeability, defined as the passage of microbial products (including endotoxin, peptidoglycan, bacterial DNA, lipopeptides, and β-glucan) from the intestinal lumen to the mesenteric lymph nodes, and extraintestinal sites [14,95]. This increased paracellular transport across disrupted tight junctions between the enterocytes commonly occurs in patients with advanced alcohol-associated liver cirrhosis [10,96] or alcoholic hepatitis [97]. The degree of liver injury correlates with endotoxemia in patients with cirrhosis and is higher in alcoholic cirrhosis compared with other etiologies [98]. Furthermore, in patients with liver cirrhosis, endotoxemia has been associated with hepatic failure, encephalopathy, and death [99].

Results in patients with early (pre-cirrhotic) stages of alcohol-associated liver disease are mixed. Elevations in plasma LPS have been observed during the early stages of alcohol-associated liver disease [100]. However, not all heavily drinking subjects (~50%) presented with increased intestinal permeability. Indeed, patients with dysbiosis had higher intestinal permeability while patients without microbial alterations did not, despite heavy chronic alcohol consumption. Three weeks of alcohol cessation reversed changes in intestinal permeability [29,94]. This raises the possibility that only a subset of heavy drinking patients has increased intestinal permeability.

Increased intestinal permeability is a common feature in preclinical models of ethanol-induced liver disease [101]. In addition, microbial derived products appear to play an important role for ethanol-induced liver disease in mice. Mice with genetic deletions in the LPS signaling pathway are resistant to alcohol-induced liver damage [102]. Despite endotoxemia occurring in preclinical mouse models of ethanol-induced liver disease, LPS alone is not enough to result in liver injury and steatosis. We have demonstrated in three independent mouse studies that blocking translocation of viable bacteria to the liver is sufficient to reduce ethanol-induced liver disease despite the same systemic LPS level [31,88,103].

Ethanol impairs the expression of intestinal antimicrobial proteins, which induces a quantitative increase of bacteria in the mucus and epithelial cell layer. This facilitates translocation of viable bacteria from the gut lumen to mesenteric lymph nodes and the liver by mechanisms that are poorly understood and deserve future investigation. The most common bacteria involved in translocation of viable bacteria are derived from the family of Enterobacteriaceae (Escherichia coli, Klebsiella, etc.), Enterococcus and Streptococcus, while anaerobic microorganisms are rarely responsible [104]. In mice, chronic ethanol administration changes bacterial alpha diversity in the ileum, largely driven by an increase in gramnegative bacteria. Moreover, gram-negative Prevotella not only increased in the mucus layer of the ileum but also in liver samples suggesting that translocation of viable bacteria to the liver might be associated with microbiota changes in the distal gastrointestinal tract [101]. Gastric acid suppression by proton pump inhibitor increases intestinal Enterococcus and its translocation to the liver, exacerbating ethanol-induced liver disease both in mice and humans [103]. E. faecalis was detectable in the liver of mice given cytolytic and non-cytolytic E. faecalis and fed an ethanol diet, but not when mice were fed a control diet indicating that ethanol-induced changes in the gut barrier are necessary for the translocation of cytolytic E. faecalis. Transplantation of feces from cytolysin-positive patients increased translocation of cytolytic E. faecalis to the liver after ethanol administration. Bacteriophages that target cytolytic E. faecalis reduced translocation of cytolysin to the liver and abolished ethanol-induced liver disease in humanized mice [31]. These examples illustrate that translocation of viable bacteria from the gut lumen is sufficient and necessary for ethanol-induced liver disease in mice.

ALCOHOL-ASSOCIATED LIVER DISEASE

When microbial products or viable bacteria reach the liver, activation of the TLRs and the cytosolic nucleotide-binding oligomerization domain-like receptors present in both parenchymal and non-parenchymal cells occur [10,77]. LPS stimulates the innate immune response through the binding to TLR4 and its co-receptor cluster of differentiation 14 (CD14), activating, in turn, NF-κB and IL-6/STAT3 signaling in Kupffer cells, macrophages, and hepatic stellate cells [105]. TLR4 signaling is required for liver steatosis, inflammation, and a fibrotic response after chronic alcohol intake [106]. Mice with mutant TLR4, deficiency of its cellular co-receptor CD14 [107] or with inactivation of Kupffer cells are protected from ethanol-induced liver injury [102,108]. Additionally, alcohol induces LPS binding protein, TLR4 and CD14 expression thus enhancing responsiveness to endotoxin [108,109]. TLR4 activation in Kupffer cells stimulates the production of cytokines (IL-1β and tumor necrosis factor [TNF]-α), chemokines, reactive oxygen species, and leukotrienes which leads to T lymphocyte and neutrophil recruitment, hepatic stellate activation, and collagen production [106,110]. Additionally, patients with alcohol dependence have elevated plasma peptidoglycans level and increased mRNA expression of IL-1β, IL-8, and IL-18 in peripheral blood mononuclear cells. Induction of these cytokines was likely related to increased expression and activation of TLR2 receptors as well as activation of the transcription factor AP-1 and the NLRP3 inflammasome, contributing to inflammation in the liver and progression of the disease [29].

Hepatic non-immune cells, such as endothelial cells and hepatic stellate cells, also respond to bacterial products through TLRs releasing inflammatory cytokines and chemokines including IL-1, IL-6, and TNF-α as well as profibrogenic cytokines including transforming growth factor (TGF)-β1 [110]. TGF-β1, a key activator of hepatic stellate cells, can upregulate the synthesis of some extracellular matrix proteins and the cellular receptors of several matrix proteins to further promote hepatocyte injury and death in the liver [111]. Ethanol-induced oxidative stress in the liver and ethanol metabolites (acetaldehyde and adducts such as malondialdehyde or 4-hydroxynonenal) also sensitize hepatic stellate cells to activation by LPS, which results in liver injury and fibrosis after the combination of chronic alcohol feeding and LPS [112].

We have recently demonstrated that cytolysin secreted by E. faecalis can directly kill hepatocyte and promote ethanol-induced liver disease in mice [31]. All these processes together can ultimately culminate in hepatic injury and systemic inflammation contributing further to immune disarray, predisposing individuals to complications such as infections and hepatic decompensation (Fig. 1) [113,114].

Figure 1.

The mechanism by which chronic alcohol intake changes the intestinal microbiota and contributes to alcohol-associated liver disease. Alcohol abuse is accompanied by intestinal dysbiosis and bacterial overgrowth. Certain gut bacteria can metabolize tryptophan into indole derivatives that can bind and activate the AhR. Activated AhR induces the expression of IL-22 in lamina propria immune cells, which stimulates mucosal defense via the production of antimicrobial peptides, such as Reg3β and Reg3γ, in intestinal epithelial cells and Paneth cells. Intestinal deficiency of Reg3β and Reg3γ increases the numbers of mucosa-associated bacteria and facilitates bacterial translocation through the portal vein. In the liver, viable bacteria and microbial products induce hepatic inflammation, hepatocyte death and fibrotic responses. Reg3γ, regenerating islet-derived 3 gamma; Reg3β, regenerating islet-derived 3 beta; IL-22, interleukin-22; AhR, aryl hydrocarbon receptor.

CONCLUSIONS

We have considerably increased our knowledge about how chronic alcohol intake changes the intestinal microbiota and contributes to disease progression. Gut dysbiosis and bacterial translocation result in immune activation and hepatic injury. Therefore, restoration of intestinal eubiosis can be used as prevention and therapeutic approach. This could be achieved by manipulating of gut microbiota through dietary changes, probiotic, prebiotic or symbiotic therapy. A promising therapeutic approach is the modification of the tryptophan-aryl hydrocarbon receptor pathway to increase the intestinal expression of IL-22 and antimicrobial proteins like Reg3γ and Reg3β. Genetically engineered bacteria producing specific factors to restore this pathway, might have fewer side effects than systemic administration. A small clinical trial showed promising results using an IL-22 agonist in patients with alcoholic hepatitis. Finally, editing the microbiota with bacteriophages is another therapeutic modality that could be explored to reduce the progression of alcohol-associated liver disease. Future clinical trials are required to test these possibilities.

Notes

Authors’ contribution

BGM drafted the manuscript; BS edited and revised the manuscript; both approved the manuscript’s final version.

Conflicts of Interest: B.S. has been consulting for Ferring Research Institute, Intercept Pharmaceuticals, HOST Therabiomics and Patara Pharmaceuticals. B.S.’s institution UC San Diego has received grant support from BiomX, NGM Biopharmaceuticals, CymaBay Therapeutics, Synlogic Operating Company and Axial Biotherapeutics.

Acknowledgements

This study was supported in part by NIH grants R01 AA020703, R01 AA24726, U01 AA026939, and services provided by P30 DK120515 and P50 AA011999.

Abbreviations

AhR

aryl hydrocarbon receptor

CD14

cluster of differentiation 14

E. faecalis

Enterococcus faecalis

interleukin

ILC3

group 3 innate lymphoid cells

LPS

lipopolysaccharides

Reg3β

regenerating islet-derived 3 beta

Reg3γ

regenerating islet-derived 3 gamma

STAT3

signal transducer and activator of transcription 3

TGF

transforming growth factor

TLRs

toll-like receptors

TNF

tumor necrosis factor

References

1. National Institute on Alcohol Abuse and Alcoholism (NIAAA). Alcohol Facts and Statistics. NIAAA web site, <https://www.niaaa.nih.gov/publications/brochures-and-fact-sheets/alcohol-facts-and-statistics>. Accessed 21 Mar 2020.

2. Teschke R. Alcoholic liver disease: current mechanistic aspects with focus on their clinical relevance. Biomedicines 2019;7:68.

3. Rehm J, Samokhvalov AV, Shield KD. Global burden of alcoholic liver diseases. J Hepatol 2013;59:160–168.

4. Bagnardi V, Rota M, Botteri E, Tramacere I, Islami F, Fedirko V, et al. Alcohol consumption and site-specific cancer risk: a comprehensive dose-response meta-analysis. Br J Cancer 2015;112:580–593.

5. Piano MR. Alcohol’s effects on the cardiovascular system. Alcohol Res 2017;38:219–241.

6. Yadav D, Lowenfels AB. The epidemiology of pancreatitis and pancreatic cancer. Gastroenterology 2013;144:1252–1261.

7. Udoh US, Valcin JA, Swain TM, Filiano AN, Gamble KL, Young ME, et al. Genetic deletion of the circadian clock transcription factor BMAL1 and chronic alcohol consumption differentially alter hepatic glycogen in mice. Am J Physiol Gastrointest Liver Physiol 2017;314:G431–G447.

8. Barr T, Helms C, Grant K, Messaoudi I. Opposing effects of alcohol on the immune system. Prog Neuropsychopharmacol Biol Psychiatry 2016;65:242–251.

9. Bluemel S, Williams B, Knight R, Schnabl B. Precision medicine in alcoholic and nonalcoholic fatty liver disease via modulating the gut microbiota. Am J Physiol Gastrointest Liver Physiol 2016;311:G1018–G1036.

10. Brenner DA, Paik YH, Schnabl B. Role of gut microbiota in liver disease. J Clin Gastroenterol 2015;49 Suppl 1(0 1):S25–S27.

11. Engen PA, Green SJ, Voigt RM, Forsyth CB, Keshavarzian A. The gastrointestinal microbiome: alcohol effects on the composition of intestinal microbiota. Alcohol Res 2015;37:223–236.

12. Seitz HK, Bataller R, Cortez-Pinto H, Gao B, Gual A, Lackner C, et al. Alcoholic liver disease. Nat Rev Dis Primers 2018;4:16.

13. Avila MA, Dufour JF, Gerbes AL, Zoulim F, Bataller R, Burra P, et al. Recent advances in alcohol-related liver disease (ALD): summary of a gut round table meeting. Gut 2020;69:764–780.

14. Sarin SK, Pande A, Schnabl B. Microbiome as a therapeutic target in alcohol-related liver disease. J Hepatol 2019;70:260–272.

15. Bishehsari F, Magno E, Swanson G, Desai V, Voigt RM, Forsyth CB, et al. Alcohol and gut-derived inflammation. Alcohol Res 2017;38:163–171.

16. Sommer F, Anderson JM, Bharti R, Raes J, Rosenstiel P. The resilience of the intestinal microbiota influences health and disease. Nat Rev Microbiol 2017;15:630–638.

17. Canesso MCC, Lacerda NL, Ferreira CM, Gonçalves JL, Almeida D, Gamba C, et al. Comparing the effects of acute alcohol consumption in germ-free and conventional mice: the role of the gut microbiota. BMC Microbiol 2014;14:240.

18. Hartmann P, Chu H, Duan Y, Schnabl B. Gut microbiota in liver disease: too much is harmful, nothing at all is not helpful either. Am J Physiol Gastrointest Liver Physiol 2019;316:G563–G573.

19. Yan AW, Fouts DE, Brandl J, Stärkel P, Torralba M, Schott E, et al. Enteric dysbiosis associated with a mouse model of alcoholic liver disease. Hepatology 2011;53:96–105.

20. Mutlu EA, Gillevet PM, Rangwala H, Sikaroodi M, Naqvi A, Engen PA, et al. Colonic microbiome is altered in alcoholism. Am J Physiol Gastrointest Liver Physiol 2012;302:G966–G978.

21. Chen P, Miyamoto Y, Mazagova M, Lee KC, Eckmann L, Schnabl B. Microbiota protects mice against acute alcohol-induced liver injury. Alcohol Clin Exp Res 2015;39:2313–2323.

22. Hendrikx T, Duan Y, Wang Y, Oh JH, Alexander LM, Huang W, et al. Bacteria engineered to produce IL-22 in intestine induce expression of REG3G to reduce ethanol-induced liver disease in mice. Gut 2019;68:1504–1515.

23. Adachi Y, Moore LE, Bradford BU, Gao W, Thurman RG. Antibiotics prevent liver injury in rats following long-term exposure to ethanol. Gastroenterology 1995;108:218–224.

24. Rios-Covian D, Salazar N, Gueimonde M, de Los Reyes-Gavilan CG. Shaping the metabolism of intestinal Bacteroides population through diet to improve human health. Front Microbiol 2017;8:376.

25. den Besten G, van Eunen K, Groen AK, Venema K, Reijngoud DJ, Bakker BM. The role of short-chain fatty acids in the interplay between diet, gut microbiota, and host energy metabolism. J Lipid Res 2013;54:2325–2340.

26. d’Hennezel E, Abubucker S, Murphy LO, Cullen TW. Total lipopolysaccharide from the human gut microbiome silences toll-like receptor signaling. mSystems 2017;2:e00046–17.

27. Naito Y, Uchiyama K, Takagi T. A next-generation beneficial microbe: Akkermansia muciniphila. J Clin Biochem Nutr 2018;63:33–35.

28. Xu Y, Wang N, Tan HY, Li S, Zhang C, Feng Y. Function of Akkermansia muciniphila in obesity: interactions with lipid metabolism, immune response and gut systems. Front Microbiol 2020;11:219.

29. Leclercq S, De Saeger C, Delzenne N, de Timary P, Stärkel P. Role of inflammatory pathways, blood mononuclear cells, and gutderived bacterial products in alcohol dependence. Biol Psychiatry 2014;76:725–733.

30. Llopis M, Cassard AM, Wrzosek L, Boschat L, Bruneau A, Ferrere G, et al. Intestinal microbiota contributes to individual susceptibility to alcoholic liver disease. Gut 2016;65:830.

31. Duan Y, Llorente C, Lang S, Brandl K, Chu H, Jiang L, et al. Bacteriophage targeting of gut bacterium attenuates alcoholic liver disease. Nature 2019;575:505–511.

32. Lang S, Demir M, Duan Y, Martin A, Schnabl B. Cytolysin-positive Enterococcus faecalis is not increased in patients with non-alcoholic steatohepatitis. Liver International 2020;40:860–865.

33. Garcia-Tsao G. Prophylactic antibiotics in cirrhosis: are they promoting or preventing infections? Clin Liver Dis (Hoboken) 2019;14:98–102.

34. Yan K, Garcia-Tsao G. Novel prevention strategies for bacterial infections in cirrhosis. Expert Opin Pharmacother 2016;17:689–701.

35. Fernández J, Tandon P, Mensa J, Garcia-Tsao G. Antibiotic prophylaxis in cirrhosis: good and bad. Hepatology 2016;63:2019–2031.

36. Bode C, Schäfer C, Fukui H, Bode JC. Effect of treatment with paromomycin on endotoxemia in patients with alcoholic liver disease--a double-blind, placebo-controlled trial. Alcohol Clin Exp Res 1997;21:1367–1373.

37. Soni H, Kumar-M P, Sharma V, Bellam BL, Mishra S, Mahendru D, et al. Antibiotics for prophylaxis of spontaneous bacterial peritonitis: systematic review & Bayesian network meta-analysis. Hepatol Int 2020;14:399–413.

38. Kaji K, Saikawa S, Takaya H, Fujinaga Y, Furukawa M, Kitagawa K, et al. Rifaximin alleviates endotoxemia with decreased serum levels of soluble CD163 and mannose receptor and partial modification of gut microbiota in cirrhotic patients. Antibiotics (Basel) 2020;9:145.

39. Zapater P, Caño R, Llanos L, Ruiz-Alcaraz AJ, Pascual S, Barquero C, et al. Norfloxacin modulates the inflammatory response and directly affects neutrophils in patients with decompensated cirrhosis. Gastroenterology 2009;137:1669–1679. e1.

40. Horvath A, Durdevic M, Leber B, di Vora K, Rainer F, Krones E, et al. Changes in the intestinal microbiome during a multispecies probiotic intervention in compensated cirrhosis. Nutrients 2020;12:1874.

41. Macnaughtan J, Figorilli F, García-López E, Lu H, Jones H, Sawhney R, et al. A double-blind, randomized placebo-controlled trial of probiotic Lactobacillus casei shirota in stable cirrhotic patients. Nutrients 2020;12:1651.

42. Vidot H, Cvejic E, Finegan LJ, Shores EA, Bowen DG, Strasser SI, et al. Supplementation with synbiotics and/or branched chain amino acids in hepatic encephalopathy: a pilot randomised placebo-controlled clinical study. Nutrients 2019;11:1810.

43. Xu J, Ma R, Chen LF, Zhao LJ, Chen K, Zhang RB. Effects of probiotic therapy on hepatic encephalopathy in patients with liver cirrhosis: an updated meta-analysis of six randomized controlled trials. Hepatobiliary Pancreat Dis Int 2014;13:354–360.

44. Pande C, Kumar A, Sarin SK. Addition of probiotics to norfloxacin does not improve efficacy in the prevention of spontaneous bacterial peritonitis: a double-blind placebo-controlled randomizedcontrolled trial. Eur J Gastroenterol Hepatol 2012;24:831–839.

45. Stadlbauer V, Mookerjee RP, Hodges S, Wright GA, Davies NA, Jalan R. Effect of probiotic treatment on deranged neutrophil function and cytokine responses in patients with compensated alcoholic cirrhosis. J Hepatol 2008;48:945–951.

46. Shasthry SM. Fecal microbiota transplantation in alcohol related liver diseases. Clin Mol Hepatol 2020;26:294–301.

47. Bajaj JS, Fagan A, Gavis EA, Kassam Z, Sikaroodi M, Gillevet PM. Long-term outcomes of fecal microbiota transplantation in patients with cirrhosis. Gastroenterology 2019;156:1921–1923. e3.

48. Agus A, Planchais J, Sokol H. Gut microbiota regulation of tryptophan metabolism in health and disease. Cell Host Microbe 2018;23:716–724.

49. Gao J, Xu K, Liu H, Liu G, Bai M, Peng C, et al. Impact of the gut microbiota on intestinal immunity mediated by tryptophan metabolism. Front Cell Infect Microbiol 2018;8:13.

50. Hendrikx T, Schnabl B. Indoles: metabolites produced by intestinal bacteria capable of controlling liver disease manifestation. J Intern Med 2019;286:32–40.

51. Roager HM, Licht TR. Microbial tryptophan catabolites in health and disease. Nat Commun 2018;9:3294.

52. Lamas B, Natividad JM, Sokol H. Aryl hydrocarbon receptor and intestinal immunity. Mucosal Immunol 2018;11:1024–1038.

53. Sun M, Ma N, He T, Johnston LJ, Ma X. Tryptophan (Trp) modulates gut homeostasis via aryl hydrocarbon receptor (AhR). Crit Rev Food Sci Nutr 2020;60:1760–1768.

54. Rankin LC, Girard-Madoux MJ, Seillet C, Mielke LA, Kerdiles Y, Fenis A, et al. Complementarity and redundancy of IL-22-producing innate lymphoid cells. Nat Immunol 2016;17:179–186.

55. Liang H, Dai Z, Liu N, Ji Y, Chen J, Zhang Y, et al. Dietary L-tryptophan modulates the structural and functional composition of the intestinal microbiome in weaned piglets. Front Microbiol 2018;9:1736.

56. Krishnan S, Ding Y, Saedi N, Choi M, Sridharan GV, Sherr DH, et al. Gut microbiota-derived tryptophan metabolites modulate inflammatory response in hepatocytes and macrophages. Cell Rep 2018;23:1099–1111.

57. Cervantes-Barragan L, Chai JN, Tianero MD, Di Luccia B, Ahern PP, Merriman J, et al. Lactobacillus reuteri induces gut intraepithelial CD4+ CD8αα+ T cells. Science 2017;357:806–810.

58. Sakurai T, Odamaki T, Xiao JZ. Production of indole-3-lactic acid by bifidobacterium strains isolated fromhuman infants. Microorganisms 2019;7:340.

59. Zelante T, Iannitti RG, Cunha C, De Luca A, Giovannini G, Pieraccini G, et al. Tryptophan catabolites from microbiota engage aryl hydrocarbon receptor and balance mucosal reactivity via interleukin-22. Immunity 2013;39:372–385.

60. Wilck N, Matus MG, Kearney SM, Olesen SW, Forslund K, Bartolomaeus H, et al. Salt-responsive gut commensal modulates TH17 axis and disease. Nature 2017;551:585–589.

61. Beaumont M, Neyrinck AM, Olivares M, Rodriguez J, de Rocca Serra A, Roumain M, et al. The gut microbiota metabolite indole alleviates liver inflammation in mice. FASEB J 2018;32:6681–6693.

62. Gao B, Duan Y, Lang S, Barupal D, Wu TC, Valdiviez L, et al. Functional microbiomics reveals alterations of the gut microbiome and host co‐metabolism in patients with alcoholic hepatitis. Hepatol Commun 2020;4:1168–1182.

63. Natividad JM, Agus A, Planchais J, Lamas B, Jarry AC, Martin R, et al. Impaired aryl hydrocarbon receptor ligand production by the gut microbiota is a key factor in metabolic syndrome. Cell Metab 2018;28:737–749. e4.

64. Ji Y, Gao Y, Chen H, Yin Y, Zhang W. Indole-3-acetic acid alleviates nonalcoholic fatty liver disease in mice via attenuation of hepatic lipogenesis, and oxidative and inflammatory stress. Nutrients 2019;11:2062.

65. Ngo VL, Abo H, Maxim E, Harusato A, Geem D, Medina-Contreras O, et al. A cytokine network involving IL-36γ, IL-23, and IL-22 promotes antimicrobial defense and recovery from intestinal barrier damage. Proc Natl Acad Sci U S A 2018;115:E5076–E5085.

66. Liu Y, Verma VK, Malhi H, Gores GJ, Kamath PS, Sanyal A, et al. Lipopolysaccharide downregulates macrophage-derived IL-22 to modulate alcohol-induced hepatocyte cell death. Am J Physiol Cell Physiol 2017;313:C305–C313.

67. Sabat R, Ouyang W, Wolk K. Therapeutic opportunities of the IL-22-IL-22R1 system. Nat Rev Drug Discov 2014;13:21–38.

68. Ki SH, Park O, Zheng M, Morales-Ibanez O, Kolls JK, Bataller R, et al. Interleukin-22 treatment ameliorates alcoholic liver injury in a murine model of chronic-binge ethanol feeding: Role of signal transducer and activator of transcription 3. Hepatology 2010;52:1291–1300.

69. Radaeva S, Sun R, Pan HN, Hong F, Gao B. Interleukin 22 (IL-22) plays a protective role in T cell-mediated murine hepatitis: IL-22 is a survival factor for hepatocytes via STAT3 activation. Hepatology 2004;39:1332–1342.

70. Zhou H, Xie G, Mao Y, Zhou K, Ren R, Zhao Q, et al. Enhanced regeneration and hepatoprotective effects of Interleukin 22 fusion protein on a predamaged liver undergoing partial hepatectomy. J Immunol Res 2018;2018:5241526.

71. Hartmann P, Seebauer CT, Mazagova M, Horvath A, Wang L, Llorente C, et al. Deficiency of intestinal mucin-2 protects mice from diet-induced fatty liver disease and obesity. Am J Physiol Gastrointest Liver Physiol 2016;310:G310–G322.

72. Brandl K, Schnabl B. Is intestinal inflammation linking dysbiosis to gut barrier dysfunction during liver disease? Expert Rev Gastroenterol Hepatol 2015;9:1069–1076.

73. Xing WW, Zou MJ, Liu S, Xu T, Wang JX, Xu DG. Interleukin-22 protects against acute alcohol-induced hepatotoxicity in mice. Biosci Biotechnol Biochem 2011;75:1290–1294.

74. Hu BL, Shi C, Lei RE, Lu DH, Luo W, Qin SY, et al. Interleukin-22 ameliorates liver fibrosis through miR-200a/beta-catenin. Sci Rep 2016;6:36436.

75. Eidenschenk C, Rutz S, Liesenfeld O, Ouyang W. Role of IL-22 in microbial host defense. In : Fillatreau S, O’Garra A, eds. Interleukin-10 in Health and Disease Berlin: Springer; 2014;:213–236.

76. Xu MJ, Feng D, Wang H, Guan Y, Yan X, Gao B. IL-22 ameliorates renal ischemia-reperfusion injury by targeting proximal tubule epithelium. J Am Soc Nephrol 2014;25:967–977.

77. Arab JP, Sehrawat TS, Simonetto DA, Verma VK, Feng D, Tang T, et al. An open-label, dose-escalation study to assess the safety and efficacy of IL-22 agonist F-652 in patients with alcohol-associated hepatitis. Hepatology 2020;72:441–453.

78. Gao B, Shah VH. Combination therapy: new hope for alcoholic hepatitis? Clin Res Hepatol Gastroenterol 2015;39 Suppl 1(Suppl 1):S7–S11.

79. Zindl CL, Lai JF, Lee YK, Maynard CL, Harbour SN, Ouyang W, et al. IL-22-producing neutrophils contribute to antimicrobial defense and restitution of colonic epithelial integrity during colitis. Proc Natl Acad Sci U S A 2013;110:12768–12773.

80. Hammer AM, Morris NL, Cannon AR, Khan OM, Gagnon RC, Movtchan NV, et al. Interleukin-22 prevents microbial dysbiosis and promotes intestinal barrier regeneration following acute injury. Shock 2017;48:657–665.

81. Sugimoto K, Ogawa A, Mizoguchi E, Shimomura Y, Andoh A, Bhan AK, et al. IL-22 ameliorates intestinal inflammation in a mouse model of ulcerative colitis. J Clin Invest 2008;118:534–544.

82. Okumura R, Takeda K. Maintenance of intestinal homeostasis by mucosal barriers. Inflamm Regen 2018;38:5.

83. Shin JH, Seeley RJ. Reg3 proteins as gut hormones? Endocrinology 2019;160:1506–1514.

84. Burger-van Paassen N, Loonen LM, Witte-Bouma J, Korteland-van Male AM, de Bruijn AC, van der Sluis M, et al. Mucin muc2 deficiency and weaning influences the expression of the innate defense genes reg3β, reg3γ and angiogenin-4. PLoS One 2012;7e38798.

85. Vaishnava S, Yamamoto M, Severson KM, Ruhn KA, Yu X, Koren O, et al. The antibacterial lectin RegIIIγ promotes the spatial segregation of microbiota and host in the intestine. Science 2011;334:255–258.

86. Brandl K, Plitas G, Schnabl B, DeMatteo RP, Pamer EG. MyD88-mediated signals induce the bactericidal lectin RegIIIγ and protect mice against intestinal Listeria monocytogenes infection. J Exp Med 2007;204:1891–1900.

87. van Ampting MT, Loonen LM, Schonewille AJ, Konings I, Vink C, Iovanna J, et al. Intestinally secreted c-type lectin Reg3b attenuates Salmonellosis but not Listeriosis in mice. Infect Immun 2012;80:1115–1120.

88. Wang L, Fouts DE, Stärkel P, Hartmann P, Chen P, Llorente C, et al. Intestinal REG3 lectins protect against alcoholic steatohepatitis by reducing mucosa-associated microbiota and preventing bacterial translocation. Cell Host Microbe 2016;19:227–239.

89. Miki T, Holst O, Hardt WD. The bactericidal activity of the C-type lectin RegIIIβ against gram-negative bacteria involves binding to lipid A. J Biol Chem 2012;287:34844–34855.

90. Zhao D, Kim YH, Jeong S, Greenson JK, Chaudhry MS, Hoepting M, et al. Survival signal REG3α prevents crypt apoptosis to control acute gastrointestinal graft-versus-host disease. J Clin Invest 2018;128:4970–4979.

91. Hanash AM, Dudakov JA, Hua G, O’Connor MH, Young LF, Singer NV, et al. Interleukin-22 protects intestinal stem cells from immune-mediated tissue damage and regulates sensitivity to graft versus host disease. Immunity 2012;37:339–350.

92. Hartmann P, Chen P, Wang HJ, Wang L, McCole DF, Brandl K, et al. Deficiency of intestinal mucin-2 ameliorates experimental alcoholic liver disease in mice. Hepatology 2013;58:108–119.

93. Chu H, Williams B, Schnabl B. Gut microbiota, fatty liver disease, and hepatocellular carcinoma. Liver Res 2018;2:43–51.

94. Stärkel P, Schnabl B. Bidirectional Communication between liver and gut during alcoholic liver disease. Semin Liver Dis 2016;36:331–339.

95. Stärkel P, Leclercq S, de Timary P, Schnabl B. Intestinal dysbiosis and permeability: the yin and yang in alcohol dependence and alcoholic liver disease. Clin Sci (Lond) 2018;132:199–212.

96. Ponziani FR, Zocco MA, Cerrito L, Gasbarrini A, Pompili M. Bacterial translocation in patients with liver cirrhosis: physiology, clinical consequences, and practical implications. Expert Rev Gastroenterol Hepatol 2018;12:641–656.

97. Lang S, Duan Y, Liu J, Torralba MG, Kuelbs C, Ventura-Cots M, et al. Intestinal fungal dysbiosis and systemic immune response to fungi in patients with alcoholic hepatitis. Hepatology 2020;71:522–538.

98. Bajaj JS, Heuman DM, Hylemon PB, Sanyal AJ, White MB, Monteith P, et al. Altered profile of human gut microbiome is associated with cirrhosis and its complications. J Hepatol 2014;60:940–947.

99. Fukui H. Gut-liver axis in liver cirrhosis: how to manage leaky gut and endotoxemia. World J Hepatol 2015;7:425–442.

100. Parlesak A, Schäfer C, Schütz T, Bode JC, Bode C. Increased intestinal permeability to macromolecules and endotoxemia in patients with chronic alcohol abuse in different stages of alcohol-induced liver disease. J Hepatol 2000;32:742–747.

101. Bluemel S, Wang L, Kuelbs C, Moncera K, Torralba M, Singh H, et al. Intestinal and hepatic microbiota changes associated with chronic ethanol administration in mice. Gut Microbes 2020;11:265–275.

102. Hritz I, Mandrekar P, Velayudham A, Catalano D, Dolganiuc A, Kodys K, et al. The critical role of toll-like receptor (TLR) 4 in alcoholic liver disease is independent of the common TLR adapter MyD88. Hepatology 2008;48:1224–1231.

103. Llorente C, Jepsen P, Inamine T, Wang L, Bluemel S, Wang HJ, et al. Gastric acid suppression promotes alcoholic liver disease by inducing overgrowth of intestinal Enterococcus. Nat Commun 2017;8:837.

104. Alexopoulou A, Agiasotelli D, Vasilieva LE, Dourakis SP. Bacterial translocation markers in liver cirrhosis. Ann Gastroenterol 2017;30:486–497.

105. Meroni M, Longo M, Dongiovanni P. Alcohol or gut microbiota: who is the guilty? Int J Mol Sci 2019;20:4568.

106. Inokuchi S, Tsukamoto H, Park E, Liu ZX, Brenner DA, Seki E. Toll-like receptor 4 mediates alcohol-induced steatohepatitis through bone marrow-derived and endogenous liver cells in mice. Alcohol Clin Exp Res 2011;35:1509–1518.

107. Hartmann P, Seebauer CT, Schnabl B. Alcoholic liver disease: the gut microbiome and liver cross talk. Alcohol Clin Exp Res 2015;39:763–775.

108. Uesugi T, Froh M, Arteel GE, Bradford BU, Wheeler MD, Gäbele E, et al. Role of lipopolysaccharide-binding protein in early alcohol-induced liver injury in mice. J Immunol 2002;168:2963–2969.

109. Schäfer C, Parlesak A, Schütt C, Bode JC, Bode C. Concentrations of lipopolysaccharide-binding protein, bactericidal/permeability-increasing protein, soluble CD14 and plasma lipids in relation to endotoxaemia in patients with alcoholic liver disease. Alcohol Alcohol 2002;37:81–86.

110. Fukui H. Role of gut dysbiosis in liver diseases: what have we learned so far? Diseases 2019;7:58.

111. Liu M, Xu Y, Han X, Yin L, Xu L, Qi Y, et al. Dioscin alleviates alcoholic liver fibrosis by attenuating hepatic stellate cell activation via the TLR4/MyD88/NF-κB signaling pathway. Sci Rep 2015;5:18038.

112. Schaffert CS, Duryee MJ, Hunter CD, Hamilton BC 3rd, DeVeney AL, Huerter MM, et al. Alcohol metabolites and lipopolysaccharide: roles in the development and/or progression of alcoholic liver disease. World J Gastroenterol 2009;15:1209–1218.

113. Kumar R, Goh BG, Kam JW, Chang PE, Tan CK. Comparisons between non-alcoholic steatohepatitis and alcohol-related hepatocellular carcinoma. Clin Mol Hepatol 2020;26:196–208.

114. Arroyo V, Moreau R, Kamath PS, Jalan R, Ginès P, Nevens F, et al. Acute-on-chronic liver failure in cirrhosis. Nat Rev Dis Primers 2016;2:16041.

Article information Continued

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.