Abstract and Introduction
Background Alcohol abuse represents the most common cause of liver disease in the Western countries. Pre-clinical and clinical studies showed that alcohol consumption affects amount and composition of gut microbiota. Moreover, gut flora plays an important role in the pathogenesis of alcoholic liver injury.
Aim To review the relationship between alcohol administration and changes on gut microbiota, its involvement in the pathogenesis of alcoholic liver disease, and how gut microbiota modulation could be a target for the treatment of alcoholic liver disease.
Methods Articles were identified using the PubMed database with the search terms 'Alcohol', 'Gut Microbiota', 'Alcoholic liver disease', 'Probiotic', 'Prebiotic', 'Symbiotic' and 'Antibiotic'. English-language articles were screened for relevance. Full review of publications for the relevant studies was conducted, including additional publications that were identified from individual article reference lists.
Results Alcohol abuse induces changes in the composition of gut microbiota, although the exact mechanism for this alteration is not well known. The translocation of bacterial products into the portal blood appears to play a key role in alcohol-induced liver damage. Several studies show that the modulation of gut microbiota seem to be a promising strategy to reduce alcohol-induced liver injury.
Conclusions Further studies are needed to better understand the relationship between alcohol administration and changes in gut microbiota, and its involvement in alcoholic liver disease. Moreover larger studies are needed to confirm the preliminary results on the therapeutic effects of gut microbiota modulation.
Alcohol abuse represents a risk factor for a number of diseases and a considerable contributor to the global burden of disease. Harmful alcohol use is recognised to be the most common cause of alcoholic liver disease (ALD) and liver cirrhosis in the Western countries. The progression of alcoholic liver disease is influenced by several factors (i.e. duration of alcohol abuse, drinking pattern and alcohol amount, nutritional status, gender, ethnicity, iron overload, co-existing metabolic syndrome, chronic hepatitis B and C virus co-infections and polymorphisms of genes involved in alcohol metabolism). Among these, recent studies have suggested that gut flora could play an important role in the pathophysiology of alcoholic liver injury and other alcohol-related diseases.[4,5]
It is well known that alcohol leads to mucosal damage, quantitative and qualitative alterations of gut flora (i.e. small intestinal bacterial overgrowth and dysbiosis), and increased gut permeability, resulting in the translocation of endotoxins and other bacterial products into portal blood flow. Bacterial products stimulate the release of pro-inflammatory mediators such as reactive oxygen species (ROS), leukotrienes, chemokines and cytokines (e.g. TNF-α and IL-1β) exerting injurious effects to cells resulting in inflammatory infiltration and fibrosis in the liver, and possibly in other organs.
The aim of this review was to discuss the effects of chronic alcohol consumption on gut microbiota composition and the relationship between gut bacterial-derived products and ALD. The modulation of gut flora as a target for the treatment of ALD and for other alcohol-related diseases will also be discussed.
Alcohol-induced Changes on Gut Microbiota
The gut of normal human subjects is a perfect habitat for various types of bacteria. The term 'gut microbiota' refers to a complex mixture of diverse microbes present in the gastrointestinal lumen. It consists of approximately 1014 microbial cells, which is 10 times the number of somatic cells in the human body.[9,10] Their collective genome, named 'microbiome', contains at least 100 times as many genes as our own genome. About 98% of the intestinal microbiota belongs to one of the four bacterial phyla, namely Firmicutes, Bacteroidetes, Proteobacteria and Actinobacteria,[12,13] with little representation of other bacterial phyla. Some of these bacteria, such as Lactobacilli and Bifidobacteria (>85% of total bacteria), live in a commensalism state, while others, such as Clostridium and Fusobacterium, could potentially become pathogens. Fungi, protozoa, archaea and viral particles are also represented in microbiota. These microorganisms start colonising the gut after birth, with a succession of different populations until a stable microbiota has been established.
The exact role of microbiota is still largely unknown. Although some functions have already been demonstrated (i.e. catabolism of dietary fibre, maintenance of barrier functions, vitamin synthesis, drug and toxin metabolism and behavioural conditioning),[12,16] this number is still growing. In particular, it seems that gut microbiota could be involved in inhibiting the growth of pathogenic bacteria, in stimulating the production of total and pathogen-specific mucosal IgA, in the production of nutrients for mucosal cells, development and modulation of immune system and immunological tolerance.[12,17] Thus, gut microbiota seems to be involved in a bi-directional interaction with host, acting as a 'super-organism' in the maintenance of human homeostasis. Moreover, several diseases seem to be associated to the alteration of this equilibrium.[12,14]
Bacterial species and concentration vary along gastrointestinal tract. There are inter-individual variations related to age, dietary habits, geographical origin, stress and lifestyle and intra-individual variations related to periods of illness, medications or dietary changes.[16,18] On this connection, both pre-clinical and clinical studies showed a relationship between alcohol administration and modification of gut microbiota.
Alteration of Gut Microbiota in Pre-clinical Studies
Recent advances in the understanding of the effects of alcohol administration on the amount and composition of gut microbiota have emerged from animal studies (Table 1). Mutlu and colleagues studied the composition of gut microbiota in two groups of rats after a period of 10 weeks intragastric administration of alcohol or dextrose. Ileal and colonic mucosal microbiota composition was analysed with Length Heterogeneity PCR (LH-PCR) fingerprinting technique. A significant alteration of colonic microbiota (dysbiosis) was found in alcohol-fed rats with respect to dextrose-fed rats. Moreover alcohol-induced colonic dysbiosis was prevented by the administration of lactobacillus GG or oats. Using an interesting design based on the genetic analysis of gut microbiome, Yan and colleagues investigated the changes in the intestinal microbial community in a group of alcohol-fed rats compared to controls after 3 weeks of alcohol consumption. They observed a bacterial overgrowth in the proximal small intestine, dysbiosis and suppression of Reg3b and Reg3g anti-microbial molecules. The reduction in anti-microbial proteins Reg3b and Reg3g following chronic alcohol exposure might contribute to the enteric dysbiosis, observed after alcohol consumption. The same authors observed a relative abundance of Bacteroidetes and Verrucomicrobia in alcohol-fed mice compared with a relative predominance of Firmicutes in control mice. Moreover, an overgrowth of Akkermansia muciniphila was observed in alcohol-fed mice. Since this microbe is able to degrade mucin,[21,22] some authors hypothesised that it could promote bacterial translocation (one of the factors involved in alcohol liver injury). Moreover, Yan and colleagues in the same study showed that the population of Lactobacilli was depleted in alcohol-treated mice. This last evidence could explain the beneficial effect of probiotics in the prevention of increased gut permeability, endotoxinemia and liver injury reported in alcohol-fed mice. In contrast with previous data a study based on metagenomic analysis of the gut microbiome showed a decline of both Bacteriodetes and Firmicutes phyla in alcohol-fed mice, with an increase in the Gram-negative Proteobacteria and Gram-positive Actinobacteria phyla. The bacterial genera that showed the biggest expansion were the Gram-negative Alcaligenes and Gram-positive Corynebacterium.
Recently, Campos Canesso and co-workers compared germ-free mice with conventional mice in an acute alcohol intake model (administration of ethanol in their drinking water for 7 days, with a higher dose of alcohol administered on day 7). Alcohol consumption produced intestinal bacterial overgrowth and dysbiosis in conventional mice with a prevalence of Enterobacteriaceae. Moreover, germ-free mice showed a reduced liver pathology (lipid content) after alcohol administration compared with conventional mice. Interestingly, the administration of intestinal contents from conventional mice to germ-free mice induced inflammation in the small intestine and the liver. Finally, the administration of fibres reduced gut permeability and protected conventional mice from liver injury after acute alcohol consumption.
Alteration of Gut Microbiota in Human Studies
Few studies investigated the relationship between alcohol abuse and changes in human gut microbiota (Table 2). Bode and colleagues evaluated types and numbers of bacteria examined in aspirates from the jejunum of alcoholic and hospitalised control patients. An increased number of microorganisms was found in chronic alcoholic patients, suggesting that bacterial overgrowth might contribute to functional and/or morphological abnormalities of the small intestine, commonly found in alcoholic patients. In a subsequent study, the same authors evaluated, by using hydrogen breath test, small intestine bacterial overgrowth (SIBO) in chronic alcoholics (n = 45) and in subjects without history of alcohol abuse (n = 60). A significant higher prevalence of SIBO in alcoholics with respect to controls was found. No differences were found in the prevalence of SIBO in alcoholic patients with liver cirrhosis compared to alcoholic patients without liver cirrhosis. These results have been confirmed by Hauge and colleagues in alcoholic patients admitted to hospital for detoxification who underwent upper gastrointestinal endoscopy with gastric and duodenal biopsies for bacteriological culture. A high prevalence of bacterial overgrowth (Gram-positive aerobic cocci) in the upper gastrointestinal tract was found in alcoholic patients. According to the authors, bacterial overgrowth could explain, at least in part, the gastrointestinal symptoms frequently complained by alcoholic patients (i.e. diarrhoea, nausea and abdominal pain). The same group of researchers have already demonstrated that the prevalence of Helicobacter pylori infection did not differ between alcoholic patients and controls. This result was confirmed by Buzás in a larger sample study
Casafont Morencos and colleagues compared 89 patients with alcoholic cirrhosis and 40 healthy subjects to assess the prevalence of intestinal bacterial overgrowth by H2-glucose breath test. Intestinal bacterial overgrowth was documented approximately in one-third of patients with alcoholic cirrhosis while in none of the healthy subjects. Moreover the prevalence of intestinal bacterial overgrowth was significantly higher in decompensated patients. The prevalence of spontaneous bacterial peritonitis was significantly higher in patients who had intestinal bacterial overgrowth than in patients who did not. In these patients, bacterial overgrowth may be a condition pre-disposing to the infection of the ascitic fluid. Some authors speculated that bacterial overgrowth in the upper small intestine might contribute to mucosal damage and affect the absorption of macro- and micronutrients in alcoholic patients.
In a recent retrospective chart review, Gabbard and colleagues confirmed a higher prevalence of SIBO in patients with moderate alcohol consumption with respect to teetotaler patients, using H2- lactulose breath test.
Besides quantitative changes of gut microbiota (SIBO), chronic alcohol consumption has been also associated to qualitative microbiota changes, such as dysbiosis, a condition in which the symbiotic relationship between gut microbioma and host is lost.
In a recent study, Mutlu and colleagues evaluated the mucosa-associated colonic microbiome of 48 alcoholic patients, with and without liver disease, compared to 18 healthy subjects. Colonic biopsy samples were analysed for microbiota composition using LH-PCR fingerprinting and multitag pyrosequencing. Confirming pre-clinical data an alteration in function and composition of colonic microbiome was found in alcoholic patients. In particular, alcoholic patients with dysbiosis showed a reduction in Bacteroidetes and an increase in Proteobacteria. Moreover there was no correlation between the duration of sobriety and the presence of dysbiosis, suggesting that the effects of chronic alcohol consumption are not temporary but rather long-lasting.
The higher abundances of Proteobacteria is in line with Chen and colleagues who also pointed out an increased prevalence of Fusobacteria in HBV- and alcohol-related cirrhotic patients. On the contrary, Prevotellaceae seem to be more represented in alcohol-related cirrhosis than in HBV-related cirrhosis and controls, probably due to ethanol metabolism in human gut. In contrast to the pre-clinical data by Yan in experimental fed alcohol mice, clinical studies by Mutlu and Chen showed a reduction in intestinal Bacteroides in alcoholic patients.[34,35] This difference could be explained by the increased population's heterogeneity and different study methods. However, further studies are needed to confirm these preliminary results.
Bhonchal and colleagues confirmed the presence of qualitative and quantitative alterations of small intestinal microflora in chronic alcoholics with ALD using microbiological analysis of duodenal (D2) biopsies obtained during upper gastrointestinal endoscopy. Although conducted with other methods, this study is in line with previous studies showing bacterial overgrowth, both of Gram-positive cocci and Gram-negative bacilli, in alcoholic patients with ALD.
Based on these available studies, it is difficult to draft a definitive position on the effects of alcohol on gut microbiota. The major limitations are related to the small sample size of the studies, heterogeneity of analytical methods, differences in alcohol abuse parameters and in the degree of ALD. However, it seems that alcohol consumption and abuse is able to produce quantitative and qualitative alterations of gut micriobiota. To date, there are ongoing studies about metagenomics, transcriptomics and metabolomics in alcoholic patients. These studies will help to explain microbe-microbe and microbe-host interactions, and may suggest new strategies to manipulate commensal microflora.
Mechanisms of Alcohol-induced Changes on Gut Microbiota
The reason for bacterial overgrowth and dysbiosis in alcoholic patients is not completely known. Since alcohol can reduce gastrointestinal motility, this has been proposed as the key mechanism for faecal stasis and bacterial overgrowth, resulting in a higher number of luminal bacteria.[6,38] Alcohol-induced suppression of innate immune response (i.e. Reg family proteins suppression) and adaptive immune response (GALT lymphoid cells depletion) represent another proposed mechanism.[39,40] These data suggested that alcohol consumption could suppress the Th I-type cellular immune response leading to increased susceptibility to oral infections. Kavanaugh and colleagues showed a substantial loss of both T-cells and dendritic cells in intestine of rats exposed both to alcohol and burn injury compared with intestine of rats receiving either burn or sham injury. Pre-clinical studies also showed that alcohol administration in mice is able to suppress bactericidal protein expression, regenerating islet derived (Reg)-3b and Reg 3g. The treatment with prebiotics partially restored Reg3g protein levels, while it mitigated bacterial overgrowth.[20,42] Finally, it is not yet well known if alcohol exerts a direct effect on the intestinal microbiota.
Aliment Pharmacol Ther. 2015;41(10):917-927. © 2015 Blackwell Publishing