Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells

An Erratum to this article was published on 12 February 2014

Abstract

Gut commensal microbes shape the mucosal immune system by regulating the differentiation and expansion of several types of T cell1,2,3,4,5. Clostridia, a dominant class of commensal microbe, can induce colonic regulatory T (Treg) cells, which have a central role in the suppression of inflammatory and allergic responses3. However, the molecular mechanisms by which commensal microbes induce colonic Treg cells have been unclear. Here we show that a large bowel microbial fermentation product, butyrate, induces the differentiation of colonic Treg cells in mice. A comparative NMR-based metabolome analysis suggests that the luminal concentrations of short-chain fatty acids positively correlates with the number of Treg cells in the colon. Among short-chain fatty acids, butyrate induced the differentiation of Treg cells in vitro and in vivo, and ameliorated the development of colitis induced by adoptive transfer of CD4+CD45RBhi T cells in Rag1−/− mice. Treatment of naive T cells under the Treg-cell-polarizing conditions with butyrate enhanced histone H3 acetylation in the promoter and conserved non-coding sequence regions of the Foxp3 locus, suggesting a possible mechanism for how microbial-derived butyrate regulates the differentiation of Treg cells. Our findings provide new insight into the mechanisms by which host–microbe interactions establish immunological homeostasis in the gut.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Gut microbial metabolism is essential for the induction of colonic Treg cells.
Figure 2: Butyrate induces the differentiation of Treg cells in the colonic lamina propria.
Figure 3: Chromatin modification at the Foxp3 locus by butyrate.
Figure 4: Butyrate ameliorates T-cell-dependent experimental colitis.

Accession codes

Accessions

DDBJ/GenBank/EMBL

Gene Expression Omnibus

Data deposits

The microarray and ChIP-seq analysis data have been deposited at the Gene Expression Omnibus (GEO) under accession number GSE49655. The microbiome analysis data have been deposited at the DDBJ database (http://getentry.ddbj.nig.ac.jp/) under accession number DRA001105.

References

  1. 1

    Chung, H. et al. Gut immune maturation depends on colonization with a host-specific microbiota. Cell 149, 1578–1593 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  2. 2

    Ivanov, I. I. et al. Induction of intestinal Th17 cells by segmented filamentous bacteria. Cell 139, 485–498 (2009)

    CAS  PubMed  PubMed Central  Google Scholar 

  3. 3

    Atarashi, K. et al. Induction of colonic regulatory T cells by indigenous clostridium species. Science 331, 337–341 (2011)

    ADS  CAS  PubMed  Google Scholar 

  4. 4

    Geuking, M. B. et al. Intestinal bacterial colonization induces mutualistic regulatory T cell responses. Immunity 34, 794–806 (2011)

    CAS  PubMed  Google Scholar 

  5. 5

    Round, J. L. & Mazmanian, S. K. Inducible Foxp3+ regulatory T-cell development by a commensal bacterium of the intestinal microbiota. Proc. Natl Acad. Sci. USA 107, 12204–12209 (2010)

    ADS  CAS  PubMed  Google Scholar 

  6. 6

    Itoh, K. & Mitsuoka, T. Characterization of clostridia isolated from faeces of limited flora mice and their effect on caecal size when associated with germ-free mice. Lab. Anim. 19, 111–118 (1985)

    CAS  PubMed  Google Scholar 

  7. 7

    Thornton, A. M. et al. Expression of Helios, an Ikaros transcription factor family member, differentiates thymic-derived from peripherally induced Foxp3+ T regulatory cells. J. Immunol. 184, 3433–3441 (2010)

    CAS  PubMed  PubMed Central  Google Scholar 

  8. 8

    Yadav, M. et al. Neuropilin-1 distinguishes natural and inducible regulatory T cells among regulatory T cell subsets in vivo . J. Exp. Med. 209, 1713–1722 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  9. 9

    Weiss, J. M. et al. Neuropilin 1 is expressed on thymus-derived natural regulatory T cells, but not mucosa-generated induced Foxp3+ T reg cells. J. Exp. Med. 209, 1723–1742 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  10. 10

    Annison, G., Illman, R. J. & Topping, D. L. Acetylated, propionylated or butyrylated starches raise large bowel short-chain fatty acids preferentially when fed to rats. J. Nutr. 133, 3523–3528 (2003)

    CAS  PubMed  Google Scholar 

  11. 11

    Rubtsov, Y. P. et al. Regulatory T cell-derived interleukin-10 limits inflammation at environmental interfaces. Immunity 28, 546–558 (2008)

    CAS  PubMed  Google Scholar 

  12. 12

    Rakoff-Nahoum, S., Paglino, J., Eslami-Varzaneh, F., Edberg, S. & Medzhitov, R. Recognition of commensal microflora by Toll-like receptors is required for intestinal homeostasis. Cell 118, 229–241 (2004)

    CAS  PubMed  Google Scholar 

  13. 13

    Mazmanian, S. K., Round, J. L. & Kasper, D. L. A microbial symbiosis factor prevents intestinal inflammatory disease. Nature 453, 620–625 (2008)

    ADS  CAS  PubMed  Google Scholar 

  14. 14

    Candido, E. P., Reeves, R. & Davie, J. R. Sodium butyrate inhibits histone deacetylation in cultured cells. Cell 14, 105–113 (1978)

    CAS  PubMed  Google Scholar 

  15. 15

    Davie, J. R. Inhibition of histone deacetylase activity by butyrate. J. Nutr. 133, 2485S–2493S (2003)

    CAS  PubMed  Google Scholar 

  16. 16

    de Zoeten, E. F., Wang, L., Sai, H., Dillmann, W. H. & Hancock, W. W. Inhibition of HDAC9 increases T regulatory cell function and prevents colitis in mice. Gastroenterology 138, 583–594 (2010)

    CAS  PubMed  Google Scholar 

  17. 17

    Tao, R. et al. Deacetylase inhibition promotes the generation and function of regulatory T cells. Nature Med. 13, 1299–1307 (2007)

    CAS  PubMed  Google Scholar 

  18. 18

    Josefowicz, S. Z., Lu, L.-F. & Rudensky, A. Y. Regulatory T cells: mechanisms of differentiation and function. Annu. Rev. Immunol. 30, 531–564 (2012)

    CAS  PubMed  PubMed Central  Google Scholar 

  19. 19

    Zheng, Y. et al. Role of conserved non-coding DNA elements in the Foxp3 gene in regulatory T-cell fate. Nature 463, 808–812 (2010)

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

  20. 20

    Ruan, Q. et al. Development of Foxp3+ regulatory T cells is driven by the c-Rel enhanceosome. Immunity 31, 932–940 (2009)

    CAS  PubMed  PubMed Central  Google Scholar 

  21. 21

    Powrie, F., Leach, M. W. M., Mauze, S. S., Caddle, L. B. L. & Coffman, R. L. R. Phenotypically distinct subsets of CD4+ T cells induce or protect from chronic intestinal inflammation in C. B-17 scid mice. Int. Immunol. 5, 1461–1471 (1993)

    CAS  PubMed  Google Scholar 

  22. 22

    Maslowski, K. M. et al. Regulation of inflammatory responses by gut microbiota and chemoattractant receptor GPR43. Nature 461, 1282–1286 (2009)

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

  23. 23

    Brown, A. J. et al. The orphan G protein-coupled receptors GPR41 and GPR43 are activated by propionate and other short chain carboxylic acids. J. Biol. Chem. 278, 11312–11319 (2003)

    CAS  PubMed  Google Scholar 

  24. 24

    Inan, M. S. et al. The luminal short-chain fatty acid butyrate modulates NF-κB activity in a human colonic epithelial cell line. Gastroenterology 118, 724–734 (2000)

    CAS  PubMed  Google Scholar 

  25. 25

    Thibault, R. et al. Down-regulation of the monocarboxylate transporter 1 is involved in butyrate deficiency during intestinal inflammation. Gastroenterology 133, 1916–1927 (2007)

    CAS  PubMed  Google Scholar 

  26. 26

    Frank, D. N. et al. Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases. Proc. Natl Acad. Sci. USA 104, 13780–13785 (2007)

    ADS  CAS  Google Scholar 

  27. 27

    Scheppach, W. et al. Effect of butyrate enemas on the colonic mucosa in distal ulcerative colitis. Gastroenterology 103, 51–56 (1992)

    CAS  PubMed  Google Scholar 

  28. 28

    Harig, J. M., Soergel, K. H., Komorowski, R. A. & Wood, C. M. Treatment of diversion colitis with short-chain-fatty acid irrigation. N. Engl. J. Med. 320, 23–28 (1989)

    CAS  PubMed  Google Scholar 

  29. 29

    Miyao, T. et al. Plasticity of Foxp3+ T cells reflects promiscuous Foxp3 expression in conventional T cells but not reprogramming of regulatory T cells. Immunity 36, 262–275 (2012)

    CAS  PubMed  Google Scholar 

  30. 30

    Yamaguchi, T. et al. Control of immune responses by antigen-specific regulatory T cells expressing the folate receptor. Immunity 27, 145–159 (2007)

    CAS  PubMed  PubMed Central  Google Scholar 

  31. 31

    Weigmann, B. et al. Isolation and subsequent analysis of murine lamina propria mononuclear cells from colonic tissue. Nature Protocols 2, 2307–2311 (2007)

    CAS  PubMed  Google Scholar 

  32. 32

    Date, Y. et al. New monitoring approach for metabolic dynamics in microbial ecosystems using stable-isotope-labeling technologies. J. Biosci. Bioeng. 110, 87–93 (2010)

    CAS  PubMed  Google Scholar 

  33. 33

    Bouskra, D. et al. Lymphoid tissue genesis induced by commensals through NOD1 regulates intestinal homeostasis. Nature 456, 507–510 (2008)

    ADS  CAS  PubMed  Google Scholar 

  34. 34

    Kim, S. W. et al. Robustness of gut microbiota of healthy adults in response to probiotic intervention revealed by high-throughput pyrosequencing. DNA Res. 20, 241–253 (2013)

    CAS  PubMed  PubMed Central  Google Scholar 

  35. 35

    Reyes, A. et al. Viruses in the faecal microbiota of monozygotic twins and their mothers. Nature 466, 334–338 (2010)

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

  36. 36

    Larkin, M. A. et al. Clustal W and Clustal X version 2.0. Bioinformatics 23, 2947–2948 (2007)

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37

    Letunic, I. & Bork, P. Interactive Tree Of Life (iTOL): an online tool for phylogenetic tree display and annotation. Bioinformatics 23, 127–128 (2006)

    PubMed  Google Scholar 

  38. 38

    Fukuda, S. et al. Bifidobacteria can protect from enteropathogenic infection through production of acetate. Nature 469, 543–547 (2011)

    ADS  CAS  PubMed  Google Scholar 

  39. 39

    Fukuda, S. et al. Evaluation and characterization of bacterial metabolic dynamics with a novel profiling technique, real-time metabolotyping. PLoS ONE 4, e4893 (2009)

    ADS  PubMed  PubMed Central  Google Scholar 

  40. 40

    Kruger, N. J., Troncoso-Ponce, M. A. & Ratcliffe, R. G. 1H NMR metabolite fingerprinting and metabolomic analysis of perchloric acid extracts from plant tissues. Nature Protocols 3, 1001–1012 (2008)

    CAS  PubMed  Google Scholar 

  41. 41

    Wiklund, S. et al. Visualization of GC/TOF-MS-based metabolomics data for identification of biochemically interesting compounds using OPLS class models. Anal. Chem. 80, 115–122 (2008)

    CAS  PubMed  Google Scholar 

  42. 42

    Kikuchi, J., Shinozaki, K. & Hirayama, T. Stable isotope labeling of Arabidopsis thaliana for an NMR-based metabolomics approach. Plant Cell Physiol. 45, 1099–1104 (2004)

    CAS  PubMed  Google Scholar 

  43. 43

    Tian, C. et al. Top-down phenomics of Arabidopsis thaliana: metabolic profiling by one- and two-dimensional nuclear magnetic resonance spectroscopy and transcriptome analysis of albino mutants. J. Biol. Chem. 282, 18532–18541 (2007)

    CAS  PubMed  Google Scholar 

  44. 44

    Sekiyama, Y., Chikayama, E. & Kikuchi, J. Profiling polar and semipolar plant metabolites throughout extraction processes using a combined solution-state and high-resolution magic angle spinning NMR approach. Anal. Chem. 82, 1643–1652 (2010)

    CAS  PubMed  Google Scholar 

  45. 45

    Akiyama, K. et al. PRIMe: a Web site that assembles tools for metabolomics and transcriptomics. In Silico Biol. 8, 339–345 (2008)

    CAS  PubMed  Google Scholar 

  46. 46

    Chikayama, E. et al. Statistical indices for simultaneous large-scale metabolite detections for a single NMR spectrum. Anal. Chem. 82, 1653–1658 (2010)

    CAS  PubMed  Google Scholar 

  47. 47

    Sannasiddappa, T. H., Costabile, A., Gibson, G. R. & Clarke, S. R. The influence of Staphylococcus aureus on gut microbial ecology in an in vitro continuous culture human colonic model system. PLoS ONE 6, e23227 (2011)

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

  48. 48

    Morita, T. et al. Resistant proteins alter cecal short-chain fatty acid profiles in rats fed high amylose cornstarch. J. Nutr. 128, 1156–1164 (1998)

    CAS  PubMed  Google Scholar 

  49. 49

    Obata, Y. et al. Epithelial cell-intrinsic Notch signaling plays an essential role in the maintenance of gut immune homeostasis. J. Immunol. 188, 2427–2436 (2012)

    CAS  PubMed  Google Scholar 

  50. 50

    Furusawa, Y. et al. DNA double-strand breaks induced by cavitational mechanical effects of ultrasound in cancer cell lines. PLoS ONE 7, e29012 (2012)

    ADS  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We would like to thank P. Carninci, Y. Shinkai and M. Yoshida for discussion; Y. Chiba and S. Yamada for technical support; H. Sugahara for technical advice; and P. D. Burrows for critical reading and editing of the manuscript. This work was supported in part by grants from Japanese Ministry of Education, Culture, Sports, Science and Technology (24117524 to S.F.; 21022049 to K.Ha.; 20113003 to H.O.), The Japan Society for the Promotion of Science (24890293 to Y.F.; 252667 to Y.O.; 24380072 and 24658129 to S.F.; 22689017 to K.Ha.; 21390155 to H.O.), The Japan Science and Technology Agency (K.Ha., K.A. and K.Ho.), RIKEN President’s Special Research Grant (H.O.), RIKEN RCAI Young Chief Investigator program (K.Ha.), the Institute for Fermentation, Osaka (S.F.), the Mishima Kaiun Memorial Foundation (S.F.), The Takeda Science Foundation (S.F. and H.O.), The Mitsubishi Foundation (H.O.), and The Uehara Memorial Foundation (S.F. and K.Ha.).

Author information

Affiliations

Authors

Contributions

S.F., K.Ha., D.L.T., T.M., K.Ho. and H.O. conceived the study; K.Ha. and S.F. designed the experiments and wrote the manuscript with Y.Fur., Y.O. and H.O.; Y. Fur. and Y.O conducted a large part of experiments together with S.F., G.N., D.T., C.U., K.K., T.K., M.Ta., E.M. and K.Ha; S.F, S.O. and K.Ha. prepared germ-free, CRB-associated and gnotobiotic mice. K.A. and K.Ho. were involved in data discussion. S.F., Y.N., C.U. and J.K. performed metabolome analysis. S.F., T.K., S.M. and M.To. performed microbiome analysis. T.A.E. performed bioinformatic analyses. S.Hi. and T.M. performed HPLC analysis. S.F. and N.N.F. performed GC–MS analysis. Y.Fuj. performed histological analysis. T.L., J.M.C., D.L.T. and S.Ho. provided essential materials and contributed to the design of experiments. Y.Fur. and H.K. contributed to the ChIP assay. H.O. directed the study and took primary responsibility for editing the manuscript.

Corresponding authors

Correspondence to Shinji Fukuda or Koji Hase or Hiroshi Ohno.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-22 and Supplementary Table 1. (PDF 2523 kb)

PowerPoint slides

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Furusawa, Y., Obata, Y., Fukuda, S. et al. Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells. Nature 504, 446–450 (2013). https://doi.org/10.1038/nature12721

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing