Characterizing the Inflammatory Role of the Short-Chain Fatty Acid Butyrate in Macrophages

Abstract

Inflammatory bowel disease (IBD), which includes Crohn's disease and ulcerative colitis, is characterized by chronic inflammation of the gastrointestinal tract. Although the cause of IBD is not uncovered yet, its sharp rise in incidence in the Western world highlights the importance of unraveling its pathogenesis. It is proven that the increasing number of IBD cases is accompanied by a decreased consumption of indigestible starch and fiber. In the human gastrointestinal tract, dietary fiber is fermented by commensal bacteria into short-chain fatty acids (SCFAs), including acetate, propionate and butyrate. <br/> SCFAs were shown to induce anti-inflammatory phenotypes in immune cells and are critical for maintaining intestinal homeostasis. However, once dysbiosis in the gut develops, fiber-rich foods and particularly butyrate can exacerbate IBD pathogenesis. It is also known that mutations of the major anti-inflammatory cytokine IL-10 or its receptor result in early onset IBD highlighting the importance of immune dysregulation on disease development. To date, there is no specific curative treatment available for IBD. <br/> Therefore, the study aims to investigate the impact of the SCFA butyrate on the immune response of macrophages under inflammatory conditions to gain insights into the pathogenesis being relevant for potential novel therapeutic approaches. <br/> To mimic the leaky gut microenvironment under pro-inflammatory conditions during IBD in vitro, primary human macrophages are exposed to lipopolysaccharide both in the presence and absence of butyrate. Small interfering RNA-mediated knockdown experiments of histone deacetylase enzymes and exposure of macrophages to histone deacetylase inhibitors with varying specificities are performed to depict the operating principle of butyrate. Protein expression of various cytokines is assessed by homogenous time-resolved fluorescence and gene expression is determined using reverse transcription quantitative real-time PCR. For investigating the genome-wide effect of butyrate, chromatin immunoprecipitation sequencing is performed. <br/> The findings indicate that the SCFA butyrate is a potent inhibitor of histone deacetylase class I enzymes under inflammatory conditions. Moreover, butyrate can inhibit the release of the anti-inflammatory cytokine IL-10 in a toll-like receptor 4-dependent manner. The butyrate's blockage of IL-10 results in an inhibited downstream phosphorylation of Signal transducer and activator of transcription 3. <br/> Next, this inhibitory effect of butyrate can be mimicked by histone deacetylase inhibitors with varying specificities and an increase of acetylation of histone 3 lysine 27 is observed after butyrate administration in primary human macrophages emphasizing the feature of histone deacetylase inhibition of butyrate. Subsequently, we demonstrated a differential regulation of cytokine expression and inflammatory response in macrophages due to epigenetic reprogramming by butyrate. <br/> These findings elucidate the potential reasons behind the detrimental effects of butyrate treatment during the acute inflammatory phase of IBD, suggesting its potential use in remission and as maintenance therapy. Future studies should seek to provide a deeper understanding of the interaction between butyrate, histone deacetylase inhibitors, and the immune system in the context of IBD, especially focusing on the context-dependent effects of butyrate.