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Histamine receptor 2 is a key influence in immune responses to intestinal histamine-secreting microbes

2014; Elsevier BV; Volume: 134; Issue: 3 Linguagem: Inglês

10.1016/j.jaci.2014.04.034

1097-6825

Ruth Ferstl, Remo Frei, Elisa Schiavi, Patrycja Konieczna, Weronika Barcik, Mario Ziegler, Roger Lauener, Christophe Chassard, Christophe Lacroix, Cezmi A. Akdiş, Liam O’Mahony,

Mast cells and histamine

It is now widely accepted that the microbiota is important for optimal host development and for ongoing immune homeostasis. The composition and metabolic activity of the microbiota has profound effects on mucosal tolerance and pathological responses.1Pfefferle P.I. Prescott S.L. Kopp M. Microbial influence on tolerance and opportunities for intervention with prebiotics/probiotics and bacterial lysates.J Allergy Clin Immunol. 2013; 131: 1453-1463Abstract Full Text Full Text PDF PubMed Scopus (50) Google Scholar, 2Frei R. Lauener R.P. Crameri R. O'Mahony L. Microbiota and dietary interactions: an update to the hygiene hypothesis?.Allergy. 2012; 67: 451-461Crossref PubMed Scopus (103) Google Scholar, 3Konieczna P. Akdis C.A. Quigley E.M. Shanahan F. O'Mahony L. Portrait of an immunoregulatory Bifidobacterium.Gut Microbes. 2012; 3: 261-266Crossref PubMed Scopus (94) Google Scholar One microbial-derived metabolic factor that is receiving increased attention is histamine. Histamine exerts its immunoregulatory effects via the activation of 4 different histamine receptors (named H1R to H4R).4O'Mahony L. Akdis M. Akdis C.A. Regulation of the immune response and inflammation by histamine and histamine receptors.J Allergy Clin Immunol. 2011; 128: 1153-1162Abstract Full Text Full Text PDF PubMed Scopus (234) Google Scholar Activation of histamine receptor 2 (H2R) is associated with potent immunoregulatory effects, and the anti-inflammatory effects of a histamine-secreting L rhamnosus strain were lost in H2R-deficient animals, suggesting that histamine derived from the microbiota could be immunoregulatory.5Frei R. Ferstl R. Konieczna P. Ziegler M. Simon T. Rugeles T.M. et al.Histamine receptor 2 modifies dendritic cell responses to microbial ligands.J Allergy Clin Immunol. 2013; 132: 194-204Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar To extend this observation and determine whether this phenomenon is a common feature of histamine-secreting microbes, we performed in vitro and murine studies with another Lactobacillus that is well recognized for its ability to secrete high levels of histamine, L saerimneri strain 30a (ATCC 33222).6Romano A. Trip H. Campbell-Sills H. Bouchez O. Sherman D. Lolkema J.S. et al.Genome sequence of Lactobacillus saerimneri 30a (formerly Lactobacillus sp. strain 30a), a reference lactic acid bacterium strain producing biogenic amines.Genome Announc. 2013; 1 (e00097-12)Crossref PubMed Scopus (12) Google Scholar In vitro, the strain performed as expected. Coculture of the L saerimneri 30a with THP-1 cells resulted in significantly reduced nuclear factor kappa B (NF-κB) responses to LPS (Fig 1, A). In addition, this inhibitory effect was H2R dependent because famotidine blocked the suppression of NF-κB activation. HPLC analysis of the bacterial culture supernatant revealed high levels of histamine secretion, and we also detected secretion of another decarboxylation product, cadaverine (Fig 1, B). Murine studies were performed in wild-type and H2R-deficient mice to assess the in vivo effect of histamine secretion on mucosal inflammatory responses. Weight was measured daily for each animal, and macroscopically the general health of each animal was recorded daily. Health scores are described in detail in this article's Methods section in the Online Repository at www.jacionline.org. Surprisingly, oral treatment of H2R-deficient and wild-type animals with L saerimneri 30a resulted in significant weight loss and animals showed signs of deteriorating health. Animals were initially hyperactive, which progressed to a hypoactive state and deterioration of coat quality, which was more pronounced in H2R-deficient animals (Fig 2, A and B). A similar trend was observed in wild-type mice administered famotidine, an H2R antagonist. Mice coadministered L saerimneri 30a and famotidine displayed more severe symptoms than did mice administered L saerimneri 30a alone (see Fig E1, A and B, in this article's Online Repository at www.jacionline.org). Quantification of the L saerimneri 30a histidine decarboxylase gene in fecal samples, using specific primers, revealed a 5-6–fold increase over the baseline level only in animals that were gavaged with this microbe, suggesting that L saerimneri 30a transited the murine gut. Control animals did not lose weight, and health scores remained as 0. Peyer patches (PPs) were removed from the mice and stimulated in vitro with phorbol 12-myristate 13-acetate and ionomycin to determine cytokine secretion from mucosal cells. PP secretion of IL-4, IL-6, and IL-17 was significantly increased for H2R-deficient mice after the administration of L saerimneri 30a (Fig 2, C), which was not observed in wild-type animals. Famotidine treatment of wild-type mice also resulted in elevated IL-17 secretion in response to L saerimneri 30a (see Fig E1, C). In contrast, IL-17 and IFN-γ secretion significantly decreased in wild-type animals after the administration of L saerimneri 30a, which was not observed in H2R-deficient mice (Fig 2, C). Intracellular cytokine staining revealed significantly reduced proportions of IL-17+CD4+ cells within the PP of wild-type animals, after exposure to L saerimneri 30a, with a trend toward reduced percentage of IFN-γ positive cells (see Fig E2, A, in this article's Online Repository at www.jacionline.org). However, IL-17+ and IFN-γ+ CD4 lymphocytes did not increase in H2R-deficient animals, suggesting that CD4 cells are not the cellular source for elevated cytokine secretion. No changes in IL-17+ or IFN-γ+ lymphocytes was observed in mesenteric lymph node cells (see Fig E2, B). The percentage of Foxp3+ cells within PPs was similar in wild-type and H2R-deficient animals and levels remained unaltered after L saerimneri 30a gavage (results not shown). The biological consequences of biogenic amine secretion in vivo by the resident microbiota are largely unknown. Histamine can have both proinflammatory and anti-inflammatory effects on immunoregulatory processes, depending on which histamine receptor is activated. With the exception of scombroid poisoning, it is currently unknown whether histamine secretion by the microbiota is altered during, or contributes to, mucosal inflammatory disorders.7Smolinska S. Jutel M. Crameri R. O'Mahony L. Histamine and gut mucosal immune regulation.Allergy. 2014; 69: 273-281Crossref PubMed Scopus (134) Google Scholar Interestingly, L saerimneri 30a secretes approximately 100-fold higher levels of histamine, compared with the levels previously described for L rhamnosus.5Frei R. Ferstl R. Konieczna P. Ziegler M. Simon T. Rugeles T.M. et al.Histamine receptor 2 modifies dendritic cell responses to microbial ligands.J Allergy Clin Immunol. 2013; 132: 194-204Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar This suggests that the amount of histamine secreted by a microbe may be critical in determining its pathological versus protective effects. Maintenance of mucosal homeostasis is heavily dependent on appropriate sampling and processing of microbial ligands by the innate immune system, in particular dendritic cells. In vitro studies have demonstrated that TLR responses to microbial ligands are significantly influenced by histamine signaling through H2R.5Frei R. Ferstl R. Konieczna P. Ziegler M. Simon T. Rugeles T.M. et al.Histamine receptor 2 modifies dendritic cell responses to microbial ligands.J Allergy Clin Immunol. 2013; 132: 194-204Abstract Full Text Full Text PDF PubMed Scopus (94) Google Scholar, 8Thomas C.M. Hong T. van Pijkeren J.P. Hemarajata P. Trinh D.V. Hu W. et al.Histamine derived from probiotic Lactobacillus reuteri suppresses TNF via modulation of PKA and ERK signaling.PLoS One. 2012; 7: e31951Crossref PubMed Scopus (347) Google Scholar In this report, we demonstrate that the H2R is required for IL-6 and IL-17 mucosal responses, suggesting that H2R is a critical immunoregulatory receptor that significantly influences the in vivo immune response to histamine-secreting microbes within the intestine. These observations suggest that histamine from the enteric microbes might exert immunoregulatory effects in vivo; however, it remains to be determined, on a case-by-case basis, whether these effects are protective or pathological. This report also highlights the need to carefully select and screen putative probiotic bacterial strains before human consumption. It is unknown whether the pathological effects seen in mice after the administration of L saerimneri 30a are solely due to histamine secretion because this bacterium also secretes cadaverine, which was previously demonstrated to be toxic in rodents.9Til H.P. Falke H.E. Prinsen M.K. Willems M.I. Acute and subacute toxicity of tyramine, spermidine, spermine, putrescine and cadaverine in rats.Food Chem Toxicol. 1997; 35: 337-348Crossref PubMed Scopus (197) Google Scholar Regardless of the mechanism, this is a cautionary note, which clearly stresses that not all Lactobacilli strains can be considered safe. LPS and histamine were obtained from Sigma (Buchs, Switzerland) and used at concentrations of 500 ng/mL and 1 × 10−5 mol/L, respectively. In addition, the H2R antagonist famotidine was obtained from Sigma. Inhibitors were added 30 minutes before stimulation. L saerimneri 30a was grown in deMan, Rogosa and Sharpe (MRS) medium (Merck, VWR International, Dietikon, Switzerland) containing 0.05% cysteine (Sigma) and 0.2% histidine (Chemie Brunschwig, Basel, Switzerland) under anaerobic conditions. THP-1-XBlueTM cells containing the NF-κB/AP-1 reporter system were obtained from Invivogen (Carlsbad, Calif) and routinely cultured in RPMI containing 10% FCS at 37°C and 5%CO2. THP-1 cells contain an NF-κB–inducible secreted embryonic alkaline phosphatase reporter construct. The levels of NF-κB–induced secreted embryonic alkaline phosphatase in the cell culture supernatant were quantified using an alkaline phosphatase detection medium (Quanti-Blue, Invivogen) and spectrophotometry. Histamine was determined by HPLC analysis (Hitachi LaChrome; Merck, Dietikon, Switzerland). Filter-sterilized culture supernatant from L saerimneri 30a 24-hour cultures was used for analysis. Samples were derivatized as previously described using the Dansyl method and further filtered through a 4-mm Titan HPLC filter with a 0.2-μm nylon membrane (Infochroma AG, Zug, Switzerland) directly in vials, which were immediately sealed and analyzed. The analysis was performed at a flow rate of 1 mL/min at 25°C, with A: 0.1 mol/L C2H3O2NH4/B: C2H3N as gradient eluents (starting at 50%A/50%B, changing during the next 10 minutes to 20%A/80%B, the next minute to 10%A/90%B, and then back to 50%A/50%B within a minute). Total duration of a run was 15 minutes, and the injection volume was 40 μL. UV detection (254 nm) was performed using a diode array detector L-7455 (Hitachi LaChrome), and the results are expressed in milligrams per liter. Wild-type female BALB/c mice were obtained from Charles River Laboratories (Sulzfeld, Germany), and H2R-deficient animals were provided by Prof Takeshi (Watanabe, Osaka, Japan). Wild-type and knockout mice were cohoused and littermates generated at the AO Research Institute (Davos, Switzerland) in individually ventilated cages for the duration of the study. All experimental procedures were carried out in accordance with Swiss law. Viable L saerimneri 30a cells (1 ×109) were administered to each mouse daily by oral gavage. Additional mice received 5 mg/kg famotidine per day, 30 minutes before gavage with L saerimneri 30a. PP single-cell suspensions were generated using GentleMACS tubes according to manufacturer's instructions (Miltenyi Biotec, Bergisch Gladbach, Germany). PP cells were stimulated with phorbol 12-myristate 13-acetate/ionomycin for 24 hours, and cytokine secretion was quantified using the Bioplex Multiplex Suspension Array System. Weight and animal health were monitored daily. Animal model macroscopic health scores included feces condition (1, wet; 2, diarrhea; 3, bloody diarrhea or rectal prolapse), activity (1, isolated, abnormal position; 2, huddled, hypoactive, or hyperactive; 3, unconscious), coordination of movement (1, slightly uncoordinated; 2, very uncoordinated; 3, paralysis), and fur quality (1, reduced grooming; 2, disheveled; 3, hair loss). Observers were blinded as to the identity of the mice (ie, wild-type vs H2R-deficient) and to the mouse treatment limb (ie, control gavage vs bacterial gavage and famotidine-treated vs non–famotidine-treated). Antimouse CD3 (Biolegend, Lucerna-Chem, Luzerna, Switzerland), CD25, Foxp3, IL-17A, and IFN-γ antibodies (eBioscience San Diego, Calif) were used to characterize lymphocyte phenotypes. Cells for intracellular cytokine staining were prestimulated for 4 hours with phorbol 12-myristate 13-acetate (50 ng/mL, Sigma) and ionomycin (500 ng/mL, Sigma) in the presence of Brefeldin A (1 μg/mL, eBioscience). Flow cytometric analysis was performed using a 10-color Galios flow cytometer (Beckman Coulter, Brea, Calif). Kaluza (Beckman Coulter) was used for data analysis. Graphing and statistical analysis of normally distributed data was performed using Prism 5 (Graph Pad Software, San Diego, Calif). Data are expressed as mean ± SEM and are analyzed for significance using the Student t test.Fig E2CD4+ lymphocyte cytokine response to L saerimneri 30a. Intracellular staining of IL-17+ and IFN-γ+ CD4 lymphocytes was performed for PP (A) and mesenteric lymph nodes (B). The percentage of IL-17+ lymphocytes within the PP decreased after the administration of L saerimneri to both wild-type and H2R-deficient animals, while no significant differences were observed in the mesenteric lymph nodes. A nonstatistically significant trend toward reduced IFN-γ+ lymphocytes after the administration of L saerimneri 30a was observed only in PP cells isolated from wild-type animals (n = 6 per group, *P < .05).View Large Image Figure ViewerDownload Hi-res image Download (PPT)