Short-Chain Fatty Acids Promote Intracellular Bactericidal Activity in Head Kidney Macrophages From Turbot (Scophthalmus maximus L.) via Hypoxia Inducible Factor-1?
ABSTRACT: Short-chain fatty acids (SCFAs) are mainly produced by microbiota through the fermentation of carbohydrates in the intestine. Acetate, propionate, and butyrate are the most abundant SCFA metabolites and have been shown to be important in the maintenance of host health. In this study, head kidney macrophages (HKMs) were isolated and cultured from turbots. We found that the antibacterial activity of HKMs was increased after these cells were incubated with sodium butyrate, sodium propionate or sodium acetate. Interestingly, our results showed that all three SCFAs enhanced the expression of hypoxia inducible factor-1 ? (HIF-1?) in HKMs, and further study confirmed that butyrate augmented the oxygen consumption of these cells. Moreover, HIF-1? inhibition diminished the butyrate-promoted intracellular bacterial killing activity of macrophages, and SCFAs also raised the gene expression and activity of lysozymes in HKMs via HIF-1? signaling. In addition, our results suggested that butyrate induced HIF-1? expression and the bactericidal activity of HKMs through histone deacetylase inhibition, while G protein-coupled receptors did not contribute to this effect. Finally, we demonstrated that butyrate induced a similar response in the murine macrophage cell line RAW264.7. In conclusion, our results demonstrated that SCFAs promoted HIF-1? expression via histone deacetylase inhibition, leading to the enhanced production of antibacterial effectors and increased bacterial killing of macrophages.
Project description:<h4>Background</h4>Eosinophilic esophagitis (EoE) is a food allergen driven disease that is accompanied by interleukin (IL) 13 overexpression and esophageal barrier dysfunction allowing transepithelial food allergen permeation. Nutraceuticals, such as short-chain fatty acids (SCFAs) that restore barrier function and increase immune fitness may be a promising tool in the management of EoE. Here, we investigated the effects of the SCFAs acetate, propionate, and butyrate on an IL-13-compromised human esophageal epithelial barrier, including the mechanisms involved.<h4>Methods</h4>An air-liquid interface culture model of differentiated human EPC2-hTERT (EPC2) was used to study whether SCFAs could restore barrier function after IL-13-induced impairment. Esophageal epithelial barrier function was monitored by transepithelial electrical resistance (TEER) and FITC-dextran paracellular flux, and was further examined by qPCR and immunohistochemical analysis. G protein-coupled receptor (GPR) GPR41, GPR43, GPR109a, or histone deacetylase (HDAC) (ant)agonists were used to assess mechanisms of action of SCFAs.<h4>Results</h4>IL-13 stimulation decreased TEER and increased FITC flux, which was counteracted by butyrate and propionate, but not acetate treatment. Barrier proteins FLG and DSG1 mRNA expression was upregulated following butyrate and propionate treatment, whereas expression of eosinophil chemoattractant CCL26 and protease CAPN14 was downregulated. Similarly, butyrate and propionate restored FLG and DSG1 protein expression. Similar effects were observed with an HDAC antagonist but not with GPR agonists.<h4>Conclusion</h4>Nutraceuticals butyrate and propionate restore the barrier function of esophageal epithelial cells after an inflammatory insult and may be of therapeutic benefit in the management of EoE.
Project description:Recent investigations in neuroscience implicate the role of microbial-derived metabolites, such as short-chain fatty acids (SCFAs) in brain health and disease. The SCFAs acetate, propionate and butyrate have pleiotropic effects within the nervous system. They are crucial for the maturation of the brain's innate immune cells, the microglia, and modulate other glial cells through the aryl-hydrocarbon receptor. Investigations in preclinical and clinical models find that SCFAs exert neuroprotective and antidepressant affects, while also modulating the stress response and satiety<i>.</i> However, many investigations thus far have not assessed the impact of sex on SCFA activity. Our novel investigation tested the impact of physiologically relevant doses of SCFAs on male and female primary cortical astrocytes. We find that butyrate (0-25 μM) correlates with increased <i>Bdnf</i> and <i>Pgc1-α</i> expression, implicating histone-deacetylase inhibitor pathways. Intriguingly, this effect is only seen in females. We also find that acetate (0-1500 μM) correlates with increased <i>Ahr</i> and <i>Gfap</i> expression in males only, suggesting immune modulatory pathways. In males, propionate (0-35 μM) correlates with increased <i>Il-22</i> expression, further suggesting immunomodulatory actions. These findings show a novel sex-dependent impact of acetate and butyrate, but not propionate on astrocyte gene expression.
Project description:Butyrate is a major short-chain fatty acid (SCFA) produced by microbial fermentation of dietary fiber in the gastrointestinal tract. Butyrate is also a well-known broad-spectrum histone deacetylase (HDAC) inhibitor. Butyrate has been reported to improve energy metabolism in rodents, which is associated with its beneficial effects on skeletal muscle, brown fat tissue and pancreatic ?-cells. The present study investigated the direct effect of butyrate on hepatic gluconeogenesis in mouse primary hepatocytes and the underlying mechanism. Isolated mouse primary hepatocytes were incubated with sodium butyrate, other HDAC inhibitors and other SCFAs. Hepatic glucose production was measured and gluconeogenic gene expression was detected by polymerase chain reaction analysis. The phosphorylation of cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) was assessed by western blot analysis. The results revealed that sodium butyrate dose-dependently increased hepatic glucose production and gluconeogenic gene expression in isolated mouse primary hepatocytes. Trichostatin A, a potent broad-spectrum HDAC inhibitor, had the opposite effect. Similar to sodium butyrate, propionate, which is another SCFA, promoted hepatic glucose production and gluconeogenic gene expression in the presence or absence of gluconeogenic substrates, which were further enhanced by cAMP. Furthermore, sodium butyrate also increased the accumulation of intracellular ATP and induced the phosphorylation of CREB in mouse hepatocytes. In conclusion, the present study suggested that butyrate stimulates hepatic gluconeogenesis and induces gluconeogenic gene expression as a substrate and cAMP/CREB signaling activator.
Project description:Short-chain fatty acids (SCFAs), such as acetate, butyrate, and propionate, modulate immune responses in the gut. However, the effect of SCFAs on mucosal vaccine-induced immune cell migration is poorly understood. Here, we investigated whether SCFAs modulate chemokine expression induced by the killed whole-cell oral cholera vaccine, Shanchol™, in human intestinal epithelial cells. Shanchol™ induced expression of CCL2, CCL5, CCL20, and CXCL10 at the mRNA level, but not at the protein level. Interestingly, CCL20 secretion was substantially increased by co-stimulation with Shanchol™ and butyrate, while neither acetate nor propionate showed such effect. Enhanced CCL20 secretion was associated with GPR109A activation, and histone deacetylase (HDAC) inhibition. In addition, co-treatment with Shanchol™ and butyrate synergistically increased the secretion of adenosine triphosphate (ATP). Moreover, CCL20 secretion was decreased by inhibiting the extracellular ATP receptor P2X7. However, neither inflammasomes nor caspases were involved in CCL20 production. The culture supernatant of cells treated with Shanchol™ and butyrate augmented human immature dendritic cell migration. Collectively, these results suggest that butyrate enhances Shanchol™-induced CCL20 production in human intestinal epithelial cells via HDAC inhibition and ATP-P2X7 signaling by activating GPR109A. These effects potentially enhance the mucosal immune responses in the gut induced by this oral cholera vaccine.
Project description:Short-chain fatty acids (SCFAs) are crucial gut microbial metabolites that play a major role in the occurrence and development of hepatic fibrosis (HF). However, the effect of SCFAs on hepatic stellate cells (HSCs), the major pro-fibrogenic cells, is yet undefined. In this study, the effects of three major SCFAs (acetate, propionate, and butyrate) were assessed on the activation of HSCs. LX2 cells were activated with TGF-β1 and treated with sodium acetate (NaA), sodium propionate (NaP), or sodium butyrate (NaB). SCFA treatment significantly reduced the protein levels of α-SMA and the phosphorylation of Smad2 and decreased the mRNA expression of Acta2/Col1a1/Fn in cells compared to the TGF-β1 treatment. Among the three SCFAs, NaA revealed the best efficacy at alleviating TGF-β1-induced LX2 cell activation. Additionally, acetate accumulated in the cells, and G protein-coupled receptor (GPR) 43 silencing did not have any impact on the inhibition of LX2 cell activation by NaA. These findings indicated that NaA enters into the cells to inhibit LX2 cell activation independent of GPR43. The results of phosphokinase array kit and Western blot indicated that NaA increased the AMP-activated protein kinase (AMPK) activation and reduced the phosphorylation of c-Jun in cultured LX2 cells, and siRNA-peroxisome proliferator-activated receptor (PPAR) -γ abolished the inhibitory effects of NaA against TGF-β1-induced LX2 cell activation. In conclusion, this study showed that NaA inhibited LX2 cell activation by activating the AMPK/PPARγ and blocking the c-Jun signaling pathways. Thus, SCFAs might represent a novel and viable approach for alleviating HF.
Project description:Short-chain fatty acids (SCFAs) butyrate and propionate are metabolites from dietary fiber's fermentation by gut microbiota that can affect differentiation or functions of T cells, macrophages and dendritic cells. We show here that at low doses these SCFAs directly impact B cell intrinsic functions to moderately enhance class-switch DNA recombination (CSR), while decreasing at higher doses over a broad physiological range, AID and Blimp1 expression, CSR, somatic hypermutation and plasma cell differentiation. In human and mouse B cells, butyrate and propionate decrease B cell Aicda and Prdm1 by upregulating select miRNAs that target Aicda and Prdm1 mRNA-3'UTRs through inhibition of histone deacetylation (HDAC) of those miRNA host genes. By acting as HDAC inhibitors, not as energy substrates or through GPR-engagement signaling in these B cell-intrinsic processes, these SCFAs impair intestinal and systemic T-dependent and T-independent antibody responses. Their epigenetic impact on B cells extends to inhibition of autoantibody production and autoimmunity in mouse lupus models.
Project description:Short-chain fatty acids (SCFAs) acetate, propionate, and butyrate are produced in large quantities by the gut microbiome and contribute to a wide array of physiological processes. While the underlying mechanisms are largely unknown, many effects of SCFAs have been traced to changes in the cell’s epigenetic state. Here, we systematically investigate how SCFAs alter the epigenome. Using quantitative proteomics of histone modification states, we identified rapid and sustained increases in histone acetylation after the addition of butyrate or propionate, but not acetate. While decades of prior observations would suggest that hyperacetylation induced by SCFAs are due to inhibition of histone deacetylases (HDACs), we found that propionate and butyrate instead activate the acetyltransferase p300. Propionate and butyrate are rapidly converted to the corresponding acyl-CoAs which are then used by p300 to catalyze auto-acylation of the autoinhibitory loop, activating the enzyme for histone/protein acetylation. This data challenges the long-held belief that SCFAs mainly regulate chromatin by inhibiting HDACs, and instead reveals a previously unknown mechanism of HAT activation that can explain how an influx of low levels of SCFAs alters global chromatin states.
Project description:Aryl hydrocarbon receptor (AhR) ligands are important for gastrointestinal health and play a role in gut inflammation and the induction of T regulatory cells, and the short chain fatty acids (SCFAs) butyrate, propionate and acetate also induce similar protective responses. Initial studies with butyrate demonstrated that this compound significantly increased expression of Ah-responsive genes such as Cyp1a1/CYP1A1 in YAMC mouse colonocytes and Caco-2 human colon cancer cell lines. Butyrate synergistically enhanced AhR ligand-induced Cyp1a1/CYP1A1 in these cells with comparable enhancement being observed for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and also microbiota-derived AhR ligands tryptamine, indole and 1,4-dihydroxy-2-naphthoic acid (DHNA). The effects of butyrate on enhancing induction of Cyp1b1/CYP1B1, AhR repressor (Ahrr/AhRR) and TCDD-inducible poly(ADP-ribose)polymerase (Tiparp/TiPARP) by AhR ligands were gene- and cell context-dependent with the Caco-2 cells being the most responsive cell line. Like butyrate and propionate, the prototypical hydroxyamic acid-derived histone deacetylase (HDAC) inhibitors Panobinostat and Vorinostat also enhanced AhR ligand-mediated induction and this was accompanied by enhanced histone acetylation. Acetate also enhanced basal and ligand-inducible Ah responsiveness and histone acetylation, demonstrating that acetate was an HDAC inhibitor. These results demonstrate SCFA-AhR ligand interactions in YAMC and Caco-2 cells where SCFAs synergistically enhance basal and ligand-induced expression of AhR-responsive genes.
Project description:Short-chain fatty acids (SCFAs), particularly butyrate, are known to suppress inflammation, and regulate the gut bacterial ecology. However, little is known about propionate. We report here that propionate infusion in the caecum dramatically affected the structure of colonic microbiota of pigs based on 16s rRNA high-throughput sequencing. Sixteen pig models were perfused with saline or sodium propionate by a fistula in the caecum. At d 28, all pigs were slaughtered for analysing bacterial metabolites, colonic microbiota and the expression of genes related to inflammation. The results showed that caecal infusion of sodium propionate increased the concentration of propionate and decreased the butyrate concentration in colonic content. For biogenic amines, the tyramine concentration was increased, while the concentration of cadaverine was decreased by infusion of sodium propionate. Furthermore, at the level of phylum, propionate increased the abundance of Bacteroidetes and reduced the abundance of Firmicutes. Prevotella and Bacteroides counts were increased, while Turicibacter abundance was decreased at the level of genus. Real-time qPCR showed that the expression of NF-?B and IL-18 was upregulated by propionate infusion, whereas no significant differences were observed for the expression of other genes related to inflammatory processes. Taken together, these results provide a new evidence for the role of short-chain fatty acid propionate on the composition of microbial community and inflammatory cytokines.
Project description:Acetate, propionate and butyrate are the main short-chain fatty acids (SCFAs) that arise from the fermentation of fibers by the colonic microbiota. While many studies focus on the regulatory role of SCFAs, their quantitative role as a catabolic or anabolic substrate for the host has received relatively little attention. To investigate this aspect, we infused conscious mice with physiological quantities of stable isotopes [1-13C]acetate, [2-13C]propionate or [2,4-13C2]butyrate directly into the cecum, which is the natural production site in mice, and analyzed their interconversion by the microbiota as well as their metabolism by the host. Cecal interconversion - pointing to microbial cross-feeding - was high between acetate and butyrate, low between butyrate and propionate and almost absent between acetate and propionate. As much as 62% of infused propionate was used in whole-body glucose production, in line with its role as gluconeogenic substrate. Conversely, glucose synthesis from propionate accounted for 69% of total glucose production. The synthesis of palmitate and cholesterol in the liver was high from cecal acetate (2.8% and 0.7%, respectively) and butyrate (2.7% and 0.9%, respectively) as substrates, but low or absent from propionate (0.6% and 0.0%, respectively). Label incorporation due to chain elongation of stearate was approximately 8-fold higher than de novo synthesis of stearate. Microarray data suggested that SCFAs exert only a mild regulatory effect on the expression of genes involved in hepatic metabolic pathways during the 6h infusion period. Altogether, gut-derived acetate, propionate and butyrate play important roles as substrates for glucose, cholesterol and lipid metabolism. Mice were infused in cecum with stably-labelled isotopes of the three main short chain fatty acids or control solution. After 6 hrs, livers were removed and pooled RNA samples were subjected to gene expression profiling.