Abstract

Background: The gut-liver interaction has emerged as a critical component in alcohol-associated liver disease (ALD) pathogenesis. The central mediators are the gut luminal antigens, especially endotoxins that translocate to the liver. This occurs because of (i) compromised gut barrier integrity due to epithelial tight junction (TJ) disruption; and (ii) qualitative/quantitative gut microbiota changes and increased production of pathogenic antigens.

We have previously shown that ethanol consumption alters the liver methionine metabolic pathway causing a rise in intracellular S-adenosylhomocysteine (SAH) levels. While the levels of the key methyl donor, S-adenosylmethionine (SAM), is unchanged, the lowering of the SAM:SAH ratio impairs several methylation reactions leading to the generation of hallmark features of early alcohol-associated liver injury. We have further shown that treatment with betaine preserves the liver methylation potential to thereby prevent alcohol-induced liver damage.

This study was undertaken to examine the effect of alcohol on intestinal methionine metabolic pathway and explore whether any functional detriment of these alterations could be prevented by betaine treatment.

Methods: Adult male C57Bl/6 mice were fed the Lieber DeCarli control or ethanol diet for 6 weeks. At the end of the feeding regimen, blood, liver, cecal content and ileal segments were removed and analyzed.

Results: We observed that while SAM levels were maintained in the ileal mucosa of the ethanol-fed mice, there was a significant rise in SAH levels which decreased the SAM:SAH ratio by ~2-fold (p < 0.02) compared with controls. All these changes were like those previously reported in the livers of ethanol-fed mice.

Concomitant with the alterations in the crucial components of ileal methionine metabolic pathway that controls the cellular methylation potential, we observed disorganized localization of key members (occludin, claudin-1) of the multiprotein TJ complex by confocal microscopic analysis. Western blotting of the detergent insoluble fraction confirmed their loss in the ileal membrane fraction (where TJs are localized) of ethanol-fed mice compared with controls. The ethanol-induced TJ disruption was accompanied by systemic endotoxemia and ~2-, 8- and 3-fold enhanced hepatic expression of the respective mRNAs encoding the pro-inflammatory cytokines, TNFα, CCL2 and IL1β, respectively. Betaine supplementation not only prevented ethanol-induced TJ disruption and microbiota changes, but also mitigated systemic endotoxemia and liver inflammation.

Conclusion: Taken together, our results indicate that alcohol-induced alterations in the intestinal epithelial methionine metabolic pathway and the resulting impairments in methylation reaction promotes gut leakiness and progressive liver injury. Betaine by preventing intestinal methylation defects and microbiota changes prevents the development of ALD.