Authors
J. AGUILERA-LIZARRAGA (1), M. FLORENS (1), D. BALEMANS (1), S. THEOFANOUS (1), E. PERNA (1), H. RODEWALD (2), M. WOUTERS (1), G. BOECKXSTAENS (1) / [1] KU, Leuven, Belgium, Clinical and Experimental Medicine, [2] German Cancer Research Center (DKFZ), Heidelberg, Germany, Cellular Immunology
Introduction Superantigens trigger an aberrant immune response by a polyclonal activation of T-cells, and are mainly produced by Staphylococcus aureus and Streptococcus pyogenes. Previously, we showed that in mice, similar to an infection with Citrobacter rodentium, Staphylococcal enterotoxin B (SEB) administered with ovalbumin (OVA) resulted in visceral hypersensitivity upon re-exposure to OVA, indicating the involvement of an aberrant bystander immune response to OVA.
Aim
In the present study, we wanted to further explore the role of mast cells, B and T cells in SEB-induced visceral hypersensitivity, and investigate to what extent S. aureus, S. pyogenes and their superantigens are present in fecal samples of IBS patients.
Methods Wild type (WT) Balb/C, mast cell deficient CPA3-Cre, T and B cell deficient SCID and T cell deficient nude mice received SEB in the presence of OVA for 3 consecutive days. 5 weeks later, mice were re-exposed to OVA by oral gavage every other day. Visceral pain was assessed prior to SEB/OVA administration (baseline) and after 4 OVA challenges by recording of the visceromotor response to colorectal distension using abdominal muscle electromyography. SCID and nude control groups did not receive SEB or OVA. To evaluate the presence of S. aureus and S. pyogenes, fecal samples of 84 well-characterized IBS patients (66 females, 38 years IQR [25-50]) and 64 healthy subjects (35 females, 49 years IQR [32-58]) were collected and DNA was extracted. S. aureus and S. pyogenes (nuc and spy1258 genes, respectively) and genes encoding superantigens (sea, seb, sec, sed, seg, seh, sell, selk, selm, selo, selp, selq) were assessed by qPCR.
Results Balb/C WT mice receiving SEB in the presence of OVA developed visceral hypersensitivity upon OVA re-exposure, however no increase in pain perception was observed in mast cell deficient mice (CPA3-Cre) (Mann Whitney test, p<0.01). SCID and nude mice receiving SEB/OVA did not develop increased visceromotor response upon OVA re-exposure compared to control mice. 19 of the 84 fecal samples of IBS patients (23%) were positive for S. aureus, compared with 6 of the 64 healthy subjects (9%) (Fisher’s exact test, p<0.05). 9 of the 19 positive IBS samples were positive for one or more superantigen (47%) compared to 1 of the 6 of the healthy subject (17%). No samples were positive for S. pyogenes.
Conclusions SEB-induced visceral hypersensitivity in mice re-exposed to OVA is mediated by T and B cells indicating the involvement of an adaptive immune response to OVA, most likely leading to antigen-specific activation of mast cells. Of note, Staphylococcus aureus, including superantigen-producing strains, are more frequently present in fecal samples of IBS patients compared to healthy controls, suggesting that similar to a bacterial infection, superantigens may be involved in installing visceral hypersensitivity in IBS.
