Acetate, the key modulator of inflammatory responses in acute alcoholic hepatitis

Abstract

Acute alcoholic hepatitis is characterized by disproportionate macrophage inflammatory cytokine responses to bacterial lipopolysaccharide. Lack of knowledge of the underlying mechanism has limited progress toward effective therapy. We postulated a novel mechanism by which ethanol increases histone acetylation, increasing proinflammatory gene transcription and cytokine synthesis. Cytokine responses to lipopolysaccharide in a human macrophage cell line cultured in 86 mM ethanol, 1 mM acetate, and normal media were measured by multiplex immunoassay. Changes in histone acetylation were determined by immunofluorescence microscopy and chromatin immunoprecipitation on presentation. The effect of ethanol and acetate on acetyl-coenzyme A (acetyl-coA) synthetases, which convert acetate to acetyl-coA, the substrate for histone acetylation, was determined by quantitative reverse-transcription polymerase chain reaction and immunoblotting. Knockdown of acetyl-coA synthetases by short hairpin RNA (shRNA) was used to determine their role in ethanol's enhancement of the inflammatory cytokine response. Ethanol-exposed macrophages developed enhanced interleukin 6 (IL6), IL8, and tumor necrosis factor alpha responses to lipopolysaccharide with time-dependent increases in histone acetylation that could be prevented by inhibition of ethanol metabolism. Chromatin immunoprecipitation confirmed increased histone acetylation at promoter regions of specific cytokine genes. The effect of ethanol was reproduced by incubation with acetate, the principal hepatic metabolite of ethanol, and both ethanol and acetate reduced histone deacetylase activity and up-regulated acetyl-coA synthetases. Knockdown of the acetyl-coA synthetases abrogated the effect of ethanol on cytokine production. Synthesis of metabolically available acetyl-coA from acetate is critical to the increased acetylation of proinflammatory gene histones and consequent enhancement of the inflammatory response in ethanol-exposed macrophages. This mechanism is a potential therapeutic target in acute alcoholic hepatitis.