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Local immune response to food antigens drives meal-induced abdominal pain

Abstract

Up to 20% of people worldwide develop gastrointestinal symptoms following a meal1, leading to decreased quality of life, substantial morbidity and high medical costs. Although the interest of both the scientific and lay communities in this issue has increased markedly in recent years, with the worldwide introduction of gluten-free and other diets, the underlying mechanisms of food-induced abdominal complaints remain largely unknown. Here we show that a bacterial infection and bacterial toxins can trigger an immune response that leads to the production of dietary-antigen-specific IgE antibodies in mice, which are limited to the intestine. Following subsequent oral ingestion of the respective dietary antigen, an IgE- and mast-cell-dependent mechanism induced increased visceral pain. This aberrant pain signalling resulted from histamine receptor H1-mediated sensitization of visceral afferents. Moreover, injection of food antigens (gluten, wheat, soy and milk) into the rectosigmoid mucosa of patients with irritable bowel syndrome induced local oedema and mast cell activation. Our results identify and characterize a peripheral mechanism that underlies food-induced abdominal pain, thereby creating new possibilities for the treatment of irritable bowel syndrome and related abdominal pain disorders.

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Fig. 1: OVA-specific immune response and VHS in post-infectious mice.
Fig. 2: Mast cells have a key role in the development of OVA-induced VHS.
Fig. 3: H1R-mediated nociceptive neuron sensitization drives OVA-induced VHS.
Fig. 4: Intramucosal injection of food antigens induces an immediate mucosal response in patients with IBS.

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Data availability

The data for 16S rRNA gene sequences are available from the European Nucleotide Archive (https://www.ebi.ac.uk/ena/) under accession number PRJEB41204. All other relevant data generated and analysed during the current study are available upon reasonable request to the corresponding author. Source data are provided with this paper.

References

  1. Schäfer, T. et al. Epidemiology of food allergy/food intolerance in adults: associations with other manifestations of atopy. Allergy 56, 1172–1179 (2001).

    Article  PubMed  Google Scholar 

  2. Honda, K. & Littman, D. R. The microbiota in adaptive immune homeostasis and disease. Nature 535, 75–84 (2016).

    Article  ADS  CAS  PubMed  Google Scholar 

  3. Mowat, A. M. I. Anatomical basis of tolerance and immunity to intestinal antigens. Nat. Rev. Immunol. 3, 331–341 (2003).

    Article  CAS  PubMed  Google Scholar 

  4. Bouziat, R. et al. Reovirus infection triggers inflammatory responses to dietary antigens and development of celiac disease. Science 356, 44–50 (2017).

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  5. Spiller, R. & Garsed, K. Postinfectious irritable bowel syndrome. Gastroenterology 136, 1979–1988 (2009).

    Article  PubMed  Google Scholar 

  6. Card, T. et al. Post-infectious IBS: defining its clinical features and prognosis using an internet-based survey. United European Gastroenterol. J. 6, 1245–1253 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  7. Ford, A. C., Lacy, B. E. & Talley, N. J. Irritable bowel syndrome. N. Engl. J. Med. 376, 2566–2578 (2017).

    Article  CAS  PubMed  Google Scholar 

  8. Barbara, G. et al. Mast cell-dependent excitation of visceral-nociceptive sensory neurons in irritable bowel syndrome. Gastroenterology 132, 26–37 (2007).

    Article  CAS  PubMed  Google Scholar 

  9. Seiffart, V. et al. IL10-deficiency in CD4+ T cells exacerbates the IFNγ and IL17 response during bacteria induced colitis. Cell. Physiol. Biochem. 36, 1259–1273 (2015).

    Article  CAS  PubMed  Google Scholar 

  10. Mondelaers, S. U. et al. Effect of genetic background and postinfectious stress on visceral sensitivity in Citrobacter rodentium-infected mice. Neurogastroenterol. Motil. 28, 647–658 (2016).

    Article  CAS  PubMed  Google Scholar 

  11. Lee, J. W. et al. Subjects with diarrhea-predominant IBS have increased rectal permeability responsive to tryptase. Dig. Dis. Sci. 55, 2922–2928 (2010).

    Article  CAS  PubMed  Google Scholar 

  12. Wu, Y. L., Stubbington, M. J. T., Daly, M., Teichmann, S. A. & Rada, C. Intrinsic transcriptional heterogeneity in B cells controls early class switching to IgE. J. Exp. Med. 214, 183–196 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Kassinen, A. et al. The fecal microbiota of irritable bowel syndrome patients differs significantly from that of healthy subjects. Gastroenterology 133, 24–33 (2007).

    Article  CAS  PubMed  Google Scholar 

  14. Wouters, M. M. et al. Histamine receptor H1-mediated sensitization of TRPV1 mediates visceral hypersensitivity and symptoms in patients with irritable bowel syndrome. Gastroenterology 150, 875–887.e9 (2016).

    Article  CAS  PubMed  Google Scholar 

  15. Hughes, P. A. et al. Post-inflammatory colonic afferent sensitisation: different subtypes, different pathways and different time courses. Gut 58, 1333–1341 (2009).

    Article  CAS  PubMed  Google Scholar 

  16. Liu, J. N. et al. The prevalence of serum specific IgE to superantigens in asthma and allergic rhinitis patients. Allergy Asthma Immunol. Res. 6, 263–266 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Forbes-Blom, E., Camberis, M., Prout, M., Tang, S. C. & Le Gros, G. Staphylococcal-derived superantigen enhances peanut induced Th2 responses in the skin. Clin. Exp. Allergy 42, 305–314 (2012).

    Article  CAS  PubMed  Google Scholar 

  18. Proft, T. & Fraser, J. D. Bacterial superantigens. Clin. Exp. Immunol. 133, 299–306 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Jones, A. L., Curran-Everett, D. & Leung, D. Y. M. Food allergy is associated with Staphylococcus aureus colonization in children with atopic dermatitis. J. Allergy Clin. Immunol. 137, 1247–1248.e3 (2016).

    Article  PubMed  Google Scholar 

  20. Rolland-Fourcade, C. et al. Epithelial expression and function of trypsin-3 in irritable bowel syndrome. Gut 66, 1767–1778 (2017).

    Article  CAS  PubMed  Google Scholar 

  21. Barbara, G. et al. Activated mast cells in proximity to colonic nerves correlate with abdominal pain in irritable bowel syndrome. Gastroenterology 126, 693–702 (2004).

    Article  PubMed  Google Scholar 

  22. De Schryver, E. et al. Local immunoglobulin e in the nasal mucosa: clinical implications. Allergy Asthma Immunol. Res. 7, 321–331 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  23. Tobin, M. C. et al. Atopic irritable bowel syndrome: a novel subgroup of irritable bowel syndrome with allergic manifestations. Ann. Allergy Asthma Immunol. 100, 49–53 (2008).

    Article  PubMed  Google Scholar 

  24. Jones, M. P., Walker, M. M., Ford, A. C. & Talley, N. J. The overlap of atopy and functional gastrointestinal disorders among 23,471 patients in primary care. Aliment. Pharmacol. Ther. 40, 382–391 (2014).

    Article  CAS  PubMed  Google Scholar 

  25. Halpin, S. J. & Ford, A. C. Prevalence of symptoms meeting criteria for irritable bowel syndrome in inflammatory bowel disease: systematic review and meta-analysis. Am. J. Gastroenterol. 107, 1474–1482 (2012).

    Article  PubMed  Google Scholar 

  26. Pearson, J. S., Niven, R. M., Meng, J., Atarodi, S. & Whorwell, P. J. Immunoglobulin E in irritable bowel syndrome: another target for treatment? A case report and literature review. Therap. Adv. Gastroenterol. 8, 270–277 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Gilfillan, A. M. & Rivera, J. The tyrosine kinase network regulating mast cell activation. Immunol. Rev. 228, 149–169 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Longstreth, G. F. et al. Functional bowel disorders. Gastroenterology 130, 1480–1491 (2006).

    Article  PubMed  Google Scholar 

  29. Bengtsson, M., Persson, J., Sjölund, K. & Ohlsson, B. Further validation of the visual analogue scale for irritable bowel syndrome after use in clinical practice. Gastroenterol. Nurs. 36, 188–198 (2013).

    Article  PubMed  Google Scholar 

  30. Feyerabend, T. B. et al. Cre-mediated cell ablation contests mast cell contribution in models of antibody- and T cell-mediated autoimmunity. Immunity 35, 832–844 (2011).

    Article  CAS  PubMed  Google Scholar 

  31. Oettgen, H. C. et al. Active anaphylaxis in IgE-deficient mice. Nature 370, 367–370 (1994).

    Article  ADS  CAS  PubMed  Google Scholar 

  32. Reikvam, D. H. et al. Depletion of murine intestinal microbiota: effects on gut mucosa and epithelial gene expression. PLoS ONE 6, e17996 (2011).

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  33. Moritoki, Y. et al. B-cell depletion with anti-CD20 ameliorates autoimmune cholangitis but exacerbates colitis in transforming growth factor-β receptor II dominant negative mice. Hepatology 50, 1893–1903 (2009).

    Article  CAS  PubMed  Google Scholar 

  34. Kim, M. S. & Kim, T. S. IgA+ plasma cells in murine intestinal lamina propria as a positive regulator of Treg differentiation. J. Leukoc. Biol. 95, 461–469 (2014).

    Article  PubMed  Google Scholar 

  35. Huang, Z.-L. et al. Altered sleep-wake characteristics and lack of arousal response to H3 receptor antagonist in histamine H1 receptor knockout mice. Proc. Natl Acad. Sci. USA 103, 4687–4692 (2006).

    Article  ADS  CAS  PubMed  PubMed Central  Google Scholar 

  36. Lauffer, A. et al. Subacute stress and chronic stress interact to decrease intestinal barrier function in rats. Stress 19, 225–234 (2016).

    Article  CAS  PubMed  Google Scholar 

  37. Hildebrand, F., Tadeo, R., Voigt, A. Y., Bork, P. & Raes, J. LotuS: an efficient and user-friendly OTU processing pipeline. Microbiome 2, 30 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  38. Callahan, B. J. et al. DADA2: high-resolution sample inference from Illumina amplicon data. Nat. Methods 13, 581–583 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Gloor, G. B., Wu, J. R., Pawlowsky-Glahn, V. & Egozcue, J. J. It’s all relative: analyzing microbiome data as compositions. Ann. Epidemiol. 26, 322–329 (2016).

    Article  PubMed  Google Scholar 

  40. Palarea-Albaladejo, J. & Martín-Fernández, J. A. ZCompositions – R package for multivariate imputation of left-censored data under a compositional approach. Chemom. Intell. Lab. Syst. 143, 85–96 (2015).

    Article  CAS  Google Scholar 

  41. Oksanen, J. et al. The vegan package. Community Ecology Package https://cran.r-project.org/web/packages/vegan/index.html (2008).

  42. Reber, L. L. et al. Imaging protective mast cells in living mice during severe contact hypersensitivity. JCI Insight 2, e92900 (2017).

    Article  PubMed Central  Google Scholar 

  43. Joulia, R. et al. Mast cells form antibody-dependent degranulatory synapse for dedicated secretion and defence. Nat. Commun. 6, 6174 (2015).

    Article  ADS  CAS  PubMed  Google Scholar 

  44. McCloy, R. A. et al. Partial inhibition of Cdk1 in G 2 phase overrides the SAC and decouples mitotic events. Cell Cycle 13, 1400–1412 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Pfaffl, M. W. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 29, e45 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Brierley, S. M., Jones, R. C. W., III, Gebhart, G. F. & Blackshaw, L. A. Splanchnic and pelvic mechanosensory afferents signal different qualities of colonic stimuli in mice. Gastroenterology 127, 166–178 (2004).

    Article  PubMed  Google Scholar 

  47. Ibeakanma, C. & Vanner, S. TNFα is a key mediator of the pronociceptive effects of mucosal supernatant from human ulcerative colitis on colonic DRG neurons. Gut 59, 612–621 (2010).

    Article  CAS  PubMed  Google Scholar 

  48. Stewart, T., Beyak, M. J. & Vanner, S. Ileitis modulates potassium and sodium currents in guinea pig dorsal root ganglia sensory neurons. J. Physiol. (Lond.) 552, 797–807 (2003).

    Article  CAS  Google Scholar 

  49. Moore, B. A., Stewart, T. M. R., Hill, C. & Vanner, S. J. TNBS ileitis evokes hyperexcitability and changes in ionic membrane properties of nociceptive DRG neurons. Am. J. Physiol. Gastrointest. Liver Physiol. 282, G1045–G1051 (2002).

    Article  CAS  Google Scholar 

  50. Ibeakanma, C. et al. Brain-gut interactions increase peripheral nociceptive signaling in mice with postinfectious irritable bowel syndrome. Gastroenterology 141, 2098–2108.e5 (2011).

    Article  CAS  PubMed  Google Scholar 

  51. Denadai-Souza, A. et al. Functional proteomic profiling of secreted serine proteases in health and inflammatory bowel disease. Sci. Rep. 8, 7834 (2018).

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  52. Denadai-Souza, A. et al. Effect of tryptase inhibition on joint inflammation: a pharmacological and lentivirus-mediated gene transfer study. Arthritis Res. Ther. 19, 124 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  53. van Dongen, J. J. et al. Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia 17, 2257–2317 (2003).

    Article  PubMed  Google Scholar 

  54. Roskin, K. M. et al. IgH sequences in common variable immune deficiency reveal altered B cell development and selection. Sci. Transl. Med. 7, 302ra135 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  55. Warners, M. J. et al. Abnormal responses to local esophageal food allergen injections in adult patients with eosinophilic esophagitis. Gastroenterology 154, 57–60.e2 (2018).

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

We thank I. Croux and R. De Keyser for technical assistance; L. Beullens, A. W. Essilfie, H. Willekens and H. Volkaerts for nursing assistance in the studies involving human subjects; M. Rasulova for assistance with ELISpot plate read-out; and the Cell and Tissue Imaging Cluster for the use of the NIKON inverted TillVision microscope, epifluorescence microscope BX 41 Olympus and Zeiss LSM880—Airyscan—and Zeiss LSM 780—SP Mai Tai HP DS—confocal microscopes (Cell and Tissue Imaging Cluster, CIC, supported by Hercules AKUL/15/37_GOH1816N and FWO G.0929.15 to P. Vanden Berghe). Flow cytometry was done at the FACS Core facility (KU Leuven) and we thank P. Andrée Penttila and P. Kumar for technical assistance. We thank the National Reference Centre (NRC) for S. aureus of Université Libre de Bruxelles (ULB) for SAgs-positive S. aureus strains; and Genentech for the IgG2a anti-CD20 antibody and its isotype control. M.V.F., M.F.V. and D.B. were supported by FWO PhD fellowships (1110019N, 11C2219N and 1127415N, respectively). C. Breynaert is supported by the Clinical Research Fund, University Hospitals, Leuven. J.A.-L., M.M.W., P.J., N.S. and Y.A.A. were supported by FWO postdoctoral fellowships (12X9820N, 1248513N, 12R5219N, 12V3619N and 12H8220N, respectively). During this project J.A.-L. was also previously supported by an FWO PhD fellowship (11Y2116N). T.B.F. and H.-R.R. were supported by SFB-TRR156-A07. S.D.B. is supported by NIH grant NIAID R01 AI125567 and an endowment from the Crown Family Foundation. G.E.B. was funded by a KU Leuven university grant (Global Opportunities for Associations GOA 14.011, C14/18/086) and an FWO grant (G0A9516N).

Author information

Authors and Affiliations

Authors

Contributions

J.A.-L., M.V.F., A.D.-S., M.M.W. and G.E.B. planned and designed the experiments. J.A.-L. and M.V.F. performed mouse infections and pharmacological treatments and carried out in vivo animal protocols. J.A.-L., M.V.F., I.A. and M.C.-E. performed immunoglobulin ELISAs. J.A.-L., M.C.-E. and S.U.M. performed diarrhoea-development experiments. J.A.-L. recorded VMRs and compliance, except for pyrilamine treatment and in Hrh1−/− mice, which were performed by M.V.F. E.P. performed afferent nerve recordings. J.A.-L. performed colonic permeability assessments, assisted by R.F. J. Si and J.R. performed analysis of murine microbiota. M.V.F. and S.T. performed histopathology assessments in mouse samples. N.S. performed cytokine quantification by cytometric bead array. J.A.-L., I.A. and N.F. performed RNA extraction and quantitative PCR experiments. M.F.V., M.V.F., I.A., N.F. and G.B. performed FACS experiments, assisted by G.M., S.I.M., C. Bosteels and B.N.L. ELISPOT assay was performed by L.D. and A.D.-S. C.L.-L. and J.J.-P. performed patch-clamp experiments, assisted by D.E.R. J.A.-L. performed Ca2+ experiments, assisted by Y.A.A. and K.T. J.A.-L. performed bacterial DNA extraction from faecal samples. P.J. and M.F.V. performed immunofluorescence staining of human samples and total corrected cell fluorescence analysis. P.J. and M.C.-E. performed and analysed immunofluorescence staining of mouse samples and mast cell live imaging analysis. R.B., G.E.B., K.V.B. and D.B. performed food antigen injections and collection of biopsies. J.A.-L. and D.B. performed the analysis of the reactions to food antigens. C. Breynaert and R.S. performed the allergy work-up in humans. A.D.-S. designed, performed and analysed protease activity assays. A.D.-S. and M.N. evaluated histamine presence in mouse and human samples, assisted by D.C. and P.A. IgH sequence analysis of IgE+ clones was performed by R.A.H., assisted by S.D.B. T.B.F. and H.-R.R. provided Cpa3Cre/+ mice, and J. Strid provided Igh7−/− mice. S.H. and F.A.R. provided experimental support. J.A.-L., M.V.F., M.F.V., A.D.-S., D.E.R., S.J.V., M.M.W. and G.E.B. reviewed data. J.A.-L., A.D.-S. and G.E.B. wrote and revised the manuscript. All other authors corrected and approved the final version of the manuscript. M.F.V. and P.J. contributed equally to this manuscript.

Corresponding author

Correspondence to Guy E. Boeckxstaens.

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Competing interests

The authors declare no competing interests.

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Extended data figures and tables

Extended Data Fig. 1 Extended analysis of the OVA-specific immune response and VHS in post-infectious mice.

a, b, diarrhoea development quantification by (a) water content in faeces and (b) whole-gut transit time upon gavage of carmine red dye in OVA/sham + OVA, OVA/infected + OVA (n = 10/group) mice. c, quantification of OVA-specific IgE in intestinal homogenates of OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA mice (n = 9, 10, 10 and 9, respectively) at 7 weeks post-infection. d, ear-swelling after intradermal injection of OVA in OVA/sham + OVA (n = 6), OVA/infected + OVA (n = 10), at 7 weeks post-infection, and OVA-allergy mice (n = 6, 10 and 6, respectively). e, f, VMR to colorectal distention in (e) OVA/sham + OVA, OVA/infected + saline, saline/infected + OVA and OVA/infected + OVA mice (n = 12, 11, 11 and 13, respectively) and (f) tracing of a electromyographic response to 20-μL-, 40-μL-, 60-μL- and 80-μL-volume colorectal distention in a OVA/infected + OVA mouse at baseline and 7 weeks post-infection. g, VMR to colorectal distention in OVA/infected + OVA (n = 7) and OVA/infected = Saline (n = 7 and 5, respectively) mice at baseline (BL), 7 weeks post-infection (PI) and after 1, 2, 3 and 4 weeks after stopping oral OVA or saline re-exposure, respectively. h, colonic permeability in BALB/c mice expressed as passage of fluorescein sodium (left) and transepithelial resistance (right) in OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA mice (n = 8, 11, 12 and 8, respectively) at 7 weeks post-infection. i, scheme illustrating the post-infectious protocol with prior OVA tolerization. j, VMR to colorectal distention in mice OVA-tolerized (high-dose) + saline/infected + OVA, OVA-tolerized (high-dose) + OVA/infected + OVA, OVA-tolerized (low-dose) + saline/infected + OVA and OVA-tolerized (low-dose) + OVA/infected + OVA (n = 6, 9, 6 and 8, respectively). k, l, VMR to colorectal distention in mice OVA/infected repeatedly gavaged with BSA compared to OVA (n = 6 and 10, respectively). m, scheme illustrating the post-infectious protocol in mice treated with anti-IgE antibody. n, VMR to colorectal distention in OVA/infected + OVA mice treated with anti-IgE antibody or control antibody (n = 8/group). o, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial resistance (right) of (l) VHS mice treated with anti-IgE antibody or control antibody (n = 8/group). p, VMR to colorectal distention in OVA/infected + OVA mice with WT or Igh7−/− background (n = 10/group). q, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial resistance (right) of OVA/infected + OVA mice with Igh7−/− background or WT mice (n = 10/group) at 7 weeks post-infection. r, scheme illustrating the protocol in mice that received monoclonal OVA-specific IgE antibody. s, VMR to colorectal distention in naïve mice treated with monoclonal OVA-specific IgE antibody or monoclonal Dinitrophenyl (DNP) antibody (n = 7 and 6, respectively). Two-tailed Mann–Whitney test in a and b for every time point, and in o and q (left); two-tailed t-test for q (right). Kruskal–Wallis test (Dunn’s multiple-comparisons test) in c and d. One-way ANOVA (Sidak’s multiple-comparisons test) in h. Two-way repeated ANOVA (Sidak’s multiple-comparisons test) in e, g, j–l, n, p and s. Data shown as Box-and-whiskers (centre line, median; box, 25th-75th percentiles; whiskers, 10th-90th percentiles) in a, b and d and median ± IQR in c, e, g, h, j–l, n-q and s. VMR, visceromotor response; BL, baseline; w, week; d; day.

Source data

Extended Data Fig. 2 Changes in microbiota composition are not likely to be associated with the development in VHS.

a, changes in the gut microbiome composition over time studied by ordination (PCA) plot before the infection, 10 days after infection (R2 = 0.11, p-value = 0.02), and after OVA gavage (R2 = 0.18, p-value = 0.002) between OVA/infected and saline/infected (n = 8/group) mice. The numbers in brackets indicate the variation explained by each axis (PCA: principal component analysis). b–d, drivers of variation in the murine intestinal community after OVA re-exposure. b, Comparison of mean abundance of taxa between OVA- and saline-infected mice. Bacterial abundances were computed using centred log-ratio transformation (CLR; two-sided Mann–Whitney test, FDR <0.2). c, ordination biplot of PCA analysis showing the significant contributors in the community variation. Arrows indicate the relative importance of each genus (permutation-based P < 0.05). d, abundance of genus Robinsoniella, Eisenbergiella, Senegalimassilia, Murimonas, Clostridium_XVIII and Stomatobaculum by group and VHS (two-sided Mann–Whitney test, FDR >0.1). e, scheme illustrating the post-infectious protocol in mice treated with antibiotics. f, g, VMR to colorectal distention in VHS (OVA/infected + OVA) mice treated with antibiotics or control, and compared to OVA/infected + saline mice (n = 10, 7 and 6, respectively) (g depicts the VMM response at 7 weeks post-infection). In d, VHS threshold was established based on the 95th percentile of visceromotor responses at baseline (AUC >4.8). There was no significant association between VHS and bacterial abundance in any group (two-tailed Mann–Whitney test). X-axis indicates absence (0) and presence (1) of VHS. Two-way ANOVA 9(Sidak’s multiple-comparisons test) in f and g. Data shown as box-and-whiskers ± 1.5 IQR in d and as median ± IQR in f and g. VMR, visceromotor response; BL, baseline; w, week.

Source data

Extended Data Fig. 3 Analysis of ‘low grade’ inflammation in colonic samples of post-infectious mice.

a, Representative H&E-stained colonic sections from OVA/sham + OVA, OVA/infected + saline, saline/infected + OVA and OVA/infected + OVA at 7 weeks post-infection. b, thickness of the colonic muscularis (left), length of the crypts (middle) and distance between crypts (right), measured in OVA/sham + OVA, saline/infected + OVA (n = 6), OVA/infected + saline and OVA/infected + OVA mice (n = 8, 6, 8 and 8, respectively) at 7 weeks post-infection. c–k, concentration of (c) IFN-γ, (d) TNF, (e) IL-6, (f) KC (murine IL-8 homologue), (g) MCP-1, (h) IL-17A, (i) IL-13, (j) IL-4 and (k) IL-10 assessed in colonic supernatant and (l) IL-13 and (m) IL-5 in colon-draining lymph nodes of OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA mice (n = 8, 11, 12 and 8, respectively, for c–k; n = 7, 7, 9 and 9, respectively for l and m). n–q, immune cell quantification along the intestinal tract: (n and p) gating strategy; (o) quantification of population of CD11b+Ly6G+SiglecF- neutrophils, CD11b+SiglecF+ eosinophils, CD117+FcRεI+ mast cells, CD3+ T cells, CD3+CD4+ T cells, CD3+CD4+Foxp3+ T cells and (q) Helios-Nrp1loFOXP3+ pTregs in the colon and small intestine in OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA mice (n = 7, 6, 7 and 7, respectively, for o (colon); 9, 8, 9 and 10, respectively, for o (small intestine); 9, 9, 8 and 10, respectively, for q (Helios-Nrp1loFOXP3+ pTregs in colon); 9, 9, 8 and 10, respectively, for q (Helios-Nrp1loFOXP3+ pTregs in small intestine);. One-way ANOVA (Sidak’s multiple-comparisons test) in b–m, o and q. Data shown median ± IQR in b–m, o and q.

Source data

Extended Data Fig. 4 Extended analysis of mast cell activation as key players in the development of OVA-induced VHS in post-infectious mice.

a–c, avidin-fluorescence intensity over time (expressed as TCCF) in MC from OVA/sham + OVA, saline/infected + OVA OVA/infected + saline and OVA/infected + OVA mice (n = 222 [7 mice], 145 [6 mice], 202 [8 mice] and 239 [9 mice], respectively), (a) shown as percentage at times = 0, 7, 14, 21 and 28 min, (b) micrographs and (c) shown at time = 28 min as median per mouse. In b, arrows point to representative MC showing degranulation (yellow, 28 min) compared to baseline (white, 0 min). d, representative image of double-staining using DAPI and avidin in fixed colonic tissue from healthy mice (n = 3). e, Histamine quantification in supernatant collected from OVA/sham + OVA, saline/infected + OVA, OVA/infected + saline and OVA/infected + OVA (n = 10/group) at 7 weeks post-infection. f, VMR to colorectal distention in OVA/infected + OVA mice (d) treated with doxantrazole or vehicle (n = 14 and 11, respectively). g, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of OVA/infected + OVA mice treated with doxantrazole or vehicle (n = 8/group) at 8 weeks post-infection. h, quantification of population of CD117+FcRεI+ mast cells in intestinal lamina propria from Cpa3Cre/+ and WT mice (n = 3/group). i, VMR to colorectal distention in OVA/infected + OVA mice with Cpa3Cre/+ background or WT littermates (n = 13 and 12, respectively). j, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) in OVA/infected + OVA mice with Cpa3Cre/+ background or WT littermates (n = 12/group) at 9 weeks post-infection. k, scheme illustrating the post-infectious protocol in mice treated with anti-CD20 antibody and bortezomib. l, anti-CD20 (5D2 clone) antibody and bortezomib depleted CD19+ Ly6K+ plasma cells (left) and B220+ IgM+ B cells (right) from the colon of mice compared to control antibody/vehicle and naïve mice (n = 2, 1 and 4, respectively) and (m) gating strategy used to validate B cell and plasma cell depletion. n, quantification of OVA-specific IgE in colon samples homogenates from OVA/infected + OVA mice treated with anti-CD20 antibody and bortezomib or control antibody and vehicle (n = 12 and 9, respectively). o, VMR to colorectal distention in OVA/infected + OVA mice treated with anti-CD20 antibody/bortezomib or control antibody/vehicle (n = 3 and 4, respectively). Mixed-effects model (Dunnett’s multiple-comparisons test) in a and e. One-way ANOVA (Sidak’s multiple-comparisons test) in c. two-way repeated measures ANOVA (Sidak’s multiple-comparisons test) in f, (I) and o. Two-tailed Mann–Whitney in g, h, j and n. Data are shown as median ± IQR in a, e–j, l, n and o, and box-and-whiskers (centre line, median; box, 25th and 75th percentiles; whiskers, 10th and 90th percentiles) in c. VMR, visceromotor response; BL, baseline; w, week.

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Extended Data Fig. 5 H1R mediates OVA-induced VHS in post-infectious mice.

a, representative tracing and Fura-2 ratiometric fields of DRG neurons (TRPV1+ neuron [right] indicated by arrowhead). b, c, VMR to colorectal distention in OVA/infected + OVA mice (a) treated with vehicle and pyrilamine (n = 9 and 8, respectively) and (b) with WT and Hrh1−/− background (n = 13 and 9, respectively). Two-way repeated measures ANOVA (Sidak’s multiple-comparisons test) in b and c. Data are shown as median ± IQR.

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Extended Data Fig. 6 SEB induces MC-mediated OVA-induced VHS upon comparable to C. rodentium infection.

a, scheme illustrating the SEB protocol. b, VMR to colorectal distention in saline/SEB + OVA, OVA/SEB + OVA and OVA (low dose) + OVA/SEB + OVA mice (n = 10, 14 and 4, respectively). c, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of saline/SEB + OVA and OVA/SEB + OVA (n = 8 and 7, respectively) mice. d, heat-map of the gene expression of inflammatory genes and mast-cell-related genes in OVA/saline + OVA (a), saline/SEB + OVA (b) and OVA/SEB + OVA mice (c) (n = 21, 7 and 7, respectively; except for Tryptase α/β-1: n = 7/group). e, VMR to colorectal distention in SEB-VHS mice treated with doxantrazole or vehicle (n = 8 and 9, respectively). f, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of SEB-VHS mice treated with doxantrazole or vehicle (n = 8 and 7, respectively). g, VMR to colorectal distention in OVA/SEB + OVA mice with Cpa3Cre/+ background or WT littermates (n = 5 and 6, respectively). h, colonic permeability expressed as passage of fluorescein sodium (left) and transepithelial electrical resistance (right) of OVA/SEB + OVA mice with Cpa3Cre/+ background or WT littermates (n = 5 and 6, respectively). i, quantification of OVA-specific IgE in colon homogenates of saline/SEB + OVA and OVA/SEB + OVA mice (n = 8 and 7, respectively). j, ear-swelling after intradermal injection of OVA in saline/SEB + OVA, OVA/SEB + OVA and OVA-allergy mice (n = 9, 7 and 6, respectively) mice. k, % of HV and IBS patients (n = 64 and 84, respectively) positive for S. aureus (left) and SAg-encoding S. aureus (right) in faecal samples. Numbers below the bars represent the ratio of positive and total HV and patients. Two-way repeated ANOVA (Sidak’s multiple-comparisons test) in b, e and g. Two-tailed unpaired t-test in c and f. Two-tailed Mann–Whitney test in h and i. Kruskal–Wallis test (Dunn’s multiple-comparisons test) in d and j. Two-sided Fisher’s exact test in k. Data shown as median ± IQR in b, e–i and box-and-whiskers (centre line, median; box, 25th-75th percentiles; whiskers, 10th-90th percentiles) in j. VMR, visceromotor response; BL, baseline; w, week. In d, Tpsab1, Il4, Il6 and Il10 gene expression differences were statistically significant in OVA/SEB + OVA vs. OVA/saline + OVA (adjusted P = 0.0005, 0.0004, 0.0163 and 0.0039, respectively).

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Extended Data Fig. 7 Extended analysis of mast cells and IgE in human samples.

a, measurement of trypsin-like activity in supernatants of rectal biopsies from HV and IBS patients (n = 13 and 48, respectively) in basal conditions. b, c, number of (a) MC (CD117+) and (b) IgE+-MC (CD117+ IgE+) quantified in mucosal rectal biopsies from HV and IBS patients (n = 15 and 22, respectively). d, quantification of CD117+ IgE+ cells (MC) at ≤ 5 μm from nerve fibres (β tubulin III+ cells) in HV and IBS patients (n = 15 and 17, respectively). e, correlation between the IgE TCCF and max mucosal diameter after food-antigen injection in IBS patients (n = 7). f, number of IgE+ clones was quantified in mucosal rectal biopsies cDNA from HV and IBS patients (n = 10 and 20, respectively) using deep sequencing. Two-tailed Mann–Whitney test in a–d and f. two-tailed Pearson’s correlation in e. Data are shown as median ± IQR in a–d, and as box-and-whiskers (centre line, median; box, 25th and 75th percentiles; whiskers, 10th and 90th percentiles) in f. HPF = high power field.

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Extended Data Fig. 8 Graphical representation of the mechanism proposed: local immune response to dietary antigens triggered by bacterial infection leads to food-induced abdominal pain.

Bacterial infection (or bacterial toxins, SEB) can trigger break of oral tolerance to food antigens leading to food-induced VHS upon food-antigen re-exposure. OVA-specific IgE antibodies bind to and sensitizes tissue-resident mast cells, which are activated upon re-exposure to OVA during feeding and release mediators that sensitize afferent neurons via H1R-mediated pathway. Components of this figure were created using Servier Medical Art templates, which are licensed under a Creative Commons Attribution 3.0 Unported License; https://smart.servier.com.

Extended Data Table 1 Colonic compliance (volume-pressure relationship) evaluated in animals subjected to VMR recordings
Extended Data Table 2 Demographic data and characteristics of IBS patients participating in food-antigen injection protocol

Supplementary information

Supplementary Table 1

A list of primer sequences for gene detection by RT-qPCR.

Reporting Summary

Video 1

Injection of food antigens in the rectosigmoid mucosa. Representative injection of food-antigen solutions in the rectosigmoid mucosa of participants.

Video 2

Mucosal edema reaction to the positive-control histamine. Representative reaction 12-min after histamine injection.

Video 3

Mucosal edema reaction to food antigens in HV Representative reaction 12-min after food-antigen injection (milk) in HV.

Video 4

Mucosal edema reaction to food antigens in IBS patients. Representative reaction after 12-min food-antigen injection (milk) in IBS patients.

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Aguilera-Lizarraga, J., Florens, M.V., Viola, M.F. et al. Local immune response to food antigens drives meal-induced abdominal pain. Nature 590, 151–156 (2021). https://doi.org/10.1038/s41586-020-03118-2

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