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Microbial metabolite butyrate facilitates M2 macrophage polarization and function.

Ji J, Shu D, Zheng M, Wang J, Luo C, Wang Y, Guo F, Zou X, Lv X, Li Y, Liu T, Qu H - Sci Rep (2016)

Bottom Line: Metabolites from intestinal microbes modulate the mucosal immune system by regulating the polarization and expansion of T cells.Butyrate attenuated intestinal inflammation in mice with dextran sulfate sodium (DSS)-induced colitis, with a significant increase in colonic expression of the M2 macrophage-associated protein, Arg1.Collectively, our study indicated that commensal microbe-derived butyrate is a novel activator of STAT6-mediated transcription through H3K9 acetylation driving M2 macrophage polarization, and delineated new insights into the immune interplay underlying inflammatory bowel disease.

View Article: PubMed Central - PubMed

Affiliation: Institute of Animal Science, Guangdong Academy of Agricultural Sciences, 1 Dafeng 1st Street, Wushan, Tianhe District, Guangzhou 510640, China.

ABSTRACT
Metabolites from intestinal microbes modulate the mucosal immune system by regulating the polarization and expansion of T cells. Whether the microbial metabolites influence macrophage polarization, however, is poorly understood. Here, we show that the large bowel microbial fermentation product, butyrate, facilitates M2 macrophage polarization, in vitro and in vivo. The supernatant from butyrate-treated M2 macrophage increased the migration and enhanced the wound closure rate of MLE-12 cells. Butyrate attenuated intestinal inflammation in mice with dextran sulfate sodium (DSS)-induced colitis, with a significant increase in colonic expression of the M2 macrophage-associated protein, Arg1. M2 macrophage treated with butyrate, had increased activation of the H3K9/STAT6 signaling pathway, suggesting a mechanism for butyrate facilitated M2 macrophage polarization. Collectively, our study indicated that commensal microbe-derived butyrate is a novel activator of STAT6-mediated transcription through H3K9 acetylation driving M2 macrophage polarization, and delineated new insights into the immune interplay underlying inflammatory bowel disease.

No MeSH data available.


Related in: MedlinePlus

Butyrate increases the expression of genes typical of M2 macrophage.(a) M0-BMDMs were directly exposed in series concentrations of butyrate for 24 h. Expression of Arg1, Fizz1, Ym1, and MR was measured by quantitative RT-PCR. (b) The relative quantity of Arg1, Fizz1, Ym1, and MR mRNA transcribed by nonpolarized (M0-BMDMs, gray bars) or M2-BMDM after 3, 6, 12, and 24 h with IL-4 in the absence (white bars) or presence (black bars) of 50 μg/mL butyrate. (c) Arg1 protein levels in the absence or presence of 50 μg/mL butyrate in M2-BMDMs after 24 h treatment. Data are mean ± SD for at least three independent experiments. Comparisons between means used t tests (*P < 0.05, **P < 0.01, ***P < 0.001).
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f1: Butyrate increases the expression of genes typical of M2 macrophage.(a) M0-BMDMs were directly exposed in series concentrations of butyrate for 24 h. Expression of Arg1, Fizz1, Ym1, and MR was measured by quantitative RT-PCR. (b) The relative quantity of Arg1, Fizz1, Ym1, and MR mRNA transcribed by nonpolarized (M0-BMDMs, gray bars) or M2-BMDM after 3, 6, 12, and 24 h with IL-4 in the absence (white bars) or presence (black bars) of 50 μg/mL butyrate. (c) Arg1 protein levels in the absence or presence of 50 μg/mL butyrate in M2-BMDMs after 24 h treatment. Data are mean ± SD for at least three independent experiments. Comparisons between means used t tests (*P < 0.05, **P < 0.01, ***P < 0.001).

Mentions: The cells were morphologically typical of bone marrow-derived macrophages (BMDMs) (Supplementary Fig. 1a) and 99.9% were F4/80+ as determined by flow cytometry (Supplementary Fig. 1b,c). Flow cytometry and Cell Counting Kit-8 assays showed no changes in BMDMs’ viability following butyrate treatment (Supplementary Fig. 2). To investigate the effect of butyrate on polarization of M2 macrophage, we first examined the hallmarks of M2 macrophage expression in BMDMs directly treated with butyrate. The results showed that treatment of M0 macrophages with butyrate did not stimulate the upregulation of M2 markers (Arg1, Fizz1, Ym1 and MR) (Supplementary Fig. 3). The data suggested that butyrate did not itself drive polarization of M0 macrophages towards the M2 phenotype. To determine whether butyrate facilitated IL-4–driven M2 polarization, M0 macrophages were activated by IL-4 in the presence of various concentrations (1 to 200 μg/ml) of butyrate. The relative expression of genes encoding Arg1, Fizz1, and Ym1, but not MR, was considerably higher in M2-BMDMs exposed to 50 μg/ml butyrate for 24 h (Fig. 1a). Meanwhile, stimulation with butyrate did not alter the surface expression of costimulatory molecules, CD80 or CD86, on M2-BMDMs (Supplementary Fig. 4). Butyrate clearly increased the expression of M2 macrophage marker genes in M2-BMDMs and 50 μg/mL was then used to determine its influence the progress of polarization, again triggered by IL-4. Compared to its absence, exposure of M2-BMDMs to butyrate significantly enhanced expression of Arg1, Fizz1, and Ym1 after 12 h treatment with IL-4 and the effect on Arg1 was most striking (Fig. 1b). Western blot analysis of Arg1 protein content at 24 h was consistent with the enhanced gene expression (Fig. 1c). Collectively, these data indicated that butyrate enhanced the IL-4-induced expression of M2-associated markers.


Microbial metabolite butyrate facilitates M2 macrophage polarization and function.

Ji J, Shu D, Zheng M, Wang J, Luo C, Wang Y, Guo F, Zou X, Lv X, Li Y, Liu T, Qu H - Sci Rep (2016)

Butyrate increases the expression of genes typical of M2 macrophage.(a) M0-BMDMs were directly exposed in series concentrations of butyrate for 24 h. Expression of Arg1, Fizz1, Ym1, and MR was measured by quantitative RT-PCR. (b) The relative quantity of Arg1, Fizz1, Ym1, and MR mRNA transcribed by nonpolarized (M0-BMDMs, gray bars) or M2-BMDM after 3, 6, 12, and 24 h with IL-4 in the absence (white bars) or presence (black bars) of 50 μg/mL butyrate. (c) Arg1 protein levels in the absence or presence of 50 μg/mL butyrate in M2-BMDMs after 24 h treatment. Data are mean ± SD for at least three independent experiments. Comparisons between means used t tests (*P < 0.05, **P < 0.01, ***P < 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4837405&req=5

f1: Butyrate increases the expression of genes typical of M2 macrophage.(a) M0-BMDMs were directly exposed in series concentrations of butyrate for 24 h. Expression of Arg1, Fizz1, Ym1, and MR was measured by quantitative RT-PCR. (b) The relative quantity of Arg1, Fizz1, Ym1, and MR mRNA transcribed by nonpolarized (M0-BMDMs, gray bars) or M2-BMDM after 3, 6, 12, and 24 h with IL-4 in the absence (white bars) or presence (black bars) of 50 μg/mL butyrate. (c) Arg1 protein levels in the absence or presence of 50 μg/mL butyrate in M2-BMDMs after 24 h treatment. Data are mean ± SD for at least three independent experiments. Comparisons between means used t tests (*P < 0.05, **P < 0.01, ***P < 0.001).
Mentions: The cells were morphologically typical of bone marrow-derived macrophages (BMDMs) (Supplementary Fig. 1a) and 99.9% were F4/80+ as determined by flow cytometry (Supplementary Fig. 1b,c). Flow cytometry and Cell Counting Kit-8 assays showed no changes in BMDMs’ viability following butyrate treatment (Supplementary Fig. 2). To investigate the effect of butyrate on polarization of M2 macrophage, we first examined the hallmarks of M2 macrophage expression in BMDMs directly treated with butyrate. The results showed that treatment of M0 macrophages with butyrate did not stimulate the upregulation of M2 markers (Arg1, Fizz1, Ym1 and MR) (Supplementary Fig. 3). The data suggested that butyrate did not itself drive polarization of M0 macrophages towards the M2 phenotype. To determine whether butyrate facilitated IL-4–driven M2 polarization, M0 macrophages were activated by IL-4 in the presence of various concentrations (1 to 200 μg/ml) of butyrate. The relative expression of genes encoding Arg1, Fizz1, and Ym1, but not MR, was considerably higher in M2-BMDMs exposed to 50 μg/ml butyrate for 24 h (Fig. 1a). Meanwhile, stimulation with butyrate did not alter the surface expression of costimulatory molecules, CD80 or CD86, on M2-BMDMs (Supplementary Fig. 4). Butyrate clearly increased the expression of M2 macrophage marker genes in M2-BMDMs and 50 μg/mL was then used to determine its influence the progress of polarization, again triggered by IL-4. Compared to its absence, exposure of M2-BMDMs to butyrate significantly enhanced expression of Arg1, Fizz1, and Ym1 after 12 h treatment with IL-4 and the effect on Arg1 was most striking (Fig. 1b). Western blot analysis of Arg1 protein content at 24 h was consistent with the enhanced gene expression (Fig. 1c). Collectively, these data indicated that butyrate enhanced the IL-4-induced expression of M2-associated markers.

Bottom Line: Metabolites from intestinal microbes modulate the mucosal immune system by regulating the polarization and expansion of T cells.Butyrate attenuated intestinal inflammation in mice with dextran sulfate sodium (DSS)-induced colitis, with a significant increase in colonic expression of the M2 macrophage-associated protein, Arg1.Collectively, our study indicated that commensal microbe-derived butyrate is a novel activator of STAT6-mediated transcription through H3K9 acetylation driving M2 macrophage polarization, and delineated new insights into the immune interplay underlying inflammatory bowel disease.

View Article: PubMed Central - PubMed

Affiliation: Institute of Animal Science, Guangdong Academy of Agricultural Sciences, 1 Dafeng 1st Street, Wushan, Tianhe District, Guangzhou 510640, China.

ABSTRACT
Metabolites from intestinal microbes modulate the mucosal immune system by regulating the polarization and expansion of T cells. Whether the microbial metabolites influence macrophage polarization, however, is poorly understood. Here, we show that the large bowel microbial fermentation product, butyrate, facilitates M2 macrophage polarization, in vitro and in vivo. The supernatant from butyrate-treated M2 macrophage increased the migration and enhanced the wound closure rate of MLE-12 cells. Butyrate attenuated intestinal inflammation in mice with dextran sulfate sodium (DSS)-induced colitis, with a significant increase in colonic expression of the M2 macrophage-associated protein, Arg1. M2 macrophage treated with butyrate, had increased activation of the H3K9/STAT6 signaling pathway, suggesting a mechanism for butyrate facilitated M2 macrophage polarization. Collectively, our study indicated that commensal microbe-derived butyrate is a novel activator of STAT6-mediated transcription through H3K9 acetylation driving M2 macrophage polarization, and delineated new insights into the immune interplay underlying inflammatory bowel disease.

No MeSH data available.


Related in: MedlinePlus