930 Regulation of Hepatic Stellate Cell Activation by MicroRNA Let-7 Family During Alcoholic Liver Injury

Abstract

Background: The intestinal mucus layer protects the epithelium against noxious agents and pathogenic bacteria present in the gastrointestinal tract. It is composed of mucins, predominantly mucin-2 (Muc2), secreted by goblet cells of the intestine. Experimental alcoholic liver disease is dependent on the translocation of bacterial products across the intestinal barrier into the systemic circulation, which induces an inflammatory response in the liver and contributes to steatohepatitis. The aim of our study was to investigate the impact of the intestinal mucus layer and in particular Muc2 on alcoholic liver disease. Methods and Results: We used the Tsukamoto-French mouse model which involves continuous intragastric feeding of isocaloric diet (n=4-5) or alcohol for 1 week in Muc2-/(n=9) and wildtype (WT) mice (n=10). Muc2 was abundantly expressed in the small and large intestine of WT mice, but undetectable in the liver. The intestinal mucus layer was considerably thinner in Muc2 deficient mice as shown by PAS staining. Alcohol feeding did not result in a compensatory upregulation of other intestinal mucins in Muc2-/mice. After alcohol feeding mice deficient in Muc2 were protected from steatohepatitis as evidenced by significantly lower ALT levels and hepatic triglyceride concentrations. In addition, hepatic oxidative stress was significantly reduced in Muc2-/mice following intragastric alcohol feeding as shown by immunohistochemistry for 4-hydroxynonenal (4-HNE) and by TBARS assay. There was no significant difference in plasma alcohol levels or hepatic alcohol metabolizing enzymes alcohol dehydrogenase 1 (ADH1) and cytochrome p450 enzyme 2E1 (Cyp2E1) in alcohol fed Muc2-/as compared to WT mice. Most notably, Muc2-/mice had significantly lower systemic plasma LPS levels after alcohol feeding. In contrast to WT mice, Muc2-/mice did not exhibit intestinal bacterial overgrowth, but a higher amount of intestinal probiotic Lactobacillus after alcohol administration as shown by qPCR for 16S rRNA and Lactobacillus spp. The antimicrobial proteins Reg3b and Reg3g were found to be expressed at significantly higher levels in the proximal small intestine in isocaloric diet and alcohol fed Muc2-/mice relative to WT mice as assessed by qPCR and Western blotting. As Reg3b and Reg3g are bactericidal c-type lectins, an increase in their expression might contribute to the observed reduction of the bacterial burden and suppression of bacterial overgrowth in the intestine. Conclusion: Intestinal mucin-2 deficiency protects from alcoholic steatohepatitis. We suggest a pathway that involves higher expression of enteric antimicrobial molecules which suppresses alcohol-associated intestinal bacterial overgrowth. Subsequently, lower amounts of bacterial products such as endotoxin translocate into the systemic circulation and cause less alcoholic liver disease.