Supplementation of Saturated Long-Chain Fatty Acids Maintains Intestinal Eubiosis and Reduces Ethanol-induced Liver Injury in Mice

Abstract

Background & Aims Alcoholic liver disease is a leading cause of mortality. Chronic alcohol consumption is accompanied by intestinal dysbiosis, and development of alcoholic liver disease requires gut-derived bacterial products. However, little is known about how alterations to the microbiome contribute to pathogenesis of alcoholic liver disease. Methods We used the Tsukamoto-French mouse model which involves continuous intragastric feeding of isocaloric diet or alcohol for 3 weeks. Bacterial DNA from the cecum was extracted for deep metagenomic sequencing. Targeted metabolomics assessed concentrations of saturated fatty acids in cecal contents. To maintain intestinal metabolic homeostasis, diets of ethanol-fed and control mice were supplemented with saturated long-chain fatty acids (LCFA). Bacterial genes involved in fatty acid biosynthesis, amounts of lactobacilli, and saturated LCFA were measured in fecal samples of non-alcoholic individuals and patients with active alcohol abuse. Results Analyses of intestinal contents from mice revealed alcohol-associated changes to the intestinal metagenome and metabolome, characterized by reduced synthesis of saturated LCFA. Maintaining intestinal levels of saturated fatty acids in mice resulted in eubiosis, stabilized the intestinal gut barrier and reduced ethanol-induced liver injury. Saturated LCFA are metabolized by commensal Lactobacillus and promote their growth. Proportions of bacterial genes involved in fatty acid biosynthesis were lower in feces from patients with active alcohol abuse than controls. Total levels of LCFA correlated with those of lactobacilli in fecal samples from patients with active alcohol abuse but not in controls. Conclusion In humans and mice, alcohol causes intestinal dysbiosis, reducing the capacity of the microbiome to synthesize saturated LCFA and the proportion of Lactobacillus species. Dietary approaches to restore levels of saturated fatty acids in the intestine might reduce ethanol-induced liver injury in patients with alcoholic liver disease.

Intestinal fungi contribute to development of alcoholic liver disease

“…Chronic ethanol administration induced bacterial dysbiosis in mice given vehicle (4,21,22) (Supplemental Figure 2E). Antifungal treatment did not produce major changes in the bacterial composition in isocaloric or ethanol diet-fed mice by comparison of bacterial 16S rRNA data sets (Supplemental Figure 2E).…”

“…Gene expression results are expressed relative to the levels of control (WT) mice or cells. DNA was isolated from feces as previously described (4,21,42). Published microbial primer sequences for 16S rRNA gene (43) and fungal-specific 18S rDNA gene were used (44).…”

“…Baseline features of this cohort are provided in Supplemental Table 2. Stool samples were collected, and DNA was extracted from fecal samples (21). Written informed consent was signed by each participant after the nature and possible consequences of the studies were explained.…”

Identification of cytochrome CYP2E1 as critical mediator of synergistic effects of alcohol and cellular lipid accumulation in hepatocytes in vitro

“…Moreover, we found that alcohol and oleate combination induced significantly more pro-inflammatory activity in hepatocytes than either of the two stimuli alone. Previous studies have found that effects of free fatty acids on hepatocytes may dependent on their degree of saturation [42,43]. In the present study we mainly applied the unsaturated fatty acid oleate to induce hepatocellular lipid accumulation because we have previously shown that the oleate induced in vitro steatosis causes cellular alterations similar to those found in hepatic tissue of NAFLD-patients [19,20].…”

Biomarker discovery of liver diseases using mass spectrometry-based metabolomics.

“…Although the underlying mechanisms are still unclear, recent studies showed that saturated fatty acids were metabolized by commensal Lactobacilli to promote their expansion [184]. Fatty acids can be synthesized by bacteria in the host in addition to being derived from the fat in the diet.…”