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Probiotics modulate mast cell degranulation and reduce stress-induced barrier dysfunction in vitro
Linköping University, Department of Clinical and Experimental Medicine, Division of Clinical Sciences. Linköping University, Faculty of Health Sciences.
Linköping University, Department of Clinical and Experimental Medicine. Linköping University, Faculty of Health Sciences. Gastrointestinal Research Unit, Department of Surgery, University Medical Center, Utrecht, The Netherlands.
Gastrointestinal Research Unit, Department of Surgery, University Medical Center, Utrecht, The Netherlands..
Gastrointestinal Research Group, Department of Physiology & Pharmacology, The Calvin, Phoebe and Joan Snyder Institute of Infection, Inflammation and Immunology, University of Calgary, Calgary, Alberta, Canada.
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2013 (English)Manuscript (preprint) (Other academic)
Abstract [en]

BACKGROUND: Stress has well-established deleterious effects on intestinal barrier function and stressful life events are known to contribute to the development and perpetuation of inflammatory bowel diseases. Mast cells play a pivotal role in pathogenesis of stressinduced barrier dysfunction due to the release of barrier-disruptive content. Conversely, they also have recently been suggested to contribute to barrier protective properties of probiotics, through the release of 15d-PGJ2 and enhanced epithelial PPAR-γ activity. However, mechanisms remain to be elucidated.

AIM: To study if probiotics can modulate mast cell mediator release, resulting in amelioration of stress-induced barrier dysfunction in vitro.

METHODS: Confluent monolayers of the human colon-derived T84 epithelial cell line were co-cultured with rat basophilic leukemia (RBL)-2H3 mast cells and pretreated with probiotics (125x104 CFU/ml, 1hr) before addition of 100nM CRF to activate mast cells. Release of beta hexosaminidase, TNF-α and 15d-PGJ2 from mast cells was determined. Transepithelial resistance (TER), and permeability to microspheres (0.2μm) were measured over a 24h period. To determine dependence of PPAR-γ, monolayers were incubated with the specific PPAR-γ antagonist T0070907 before treatment with probiotics.

RESULTS: CRF-induced activation of mast cells resulted in decreased TERs and increased permeability to microspheres. Both pretreatment with probiotics and filter-sterilized probiotic supernatant resulted in lower levels of mast cell-released beta hexosaminidase and TNF-α, and increased 15d-PGJ2. Furthermore, probiotics ameliorated epithelial barrier dysfunction in monolayers exposed to CRF-activated mast cells. However, when T84 monolayers were exposed to conditioned medium of CRF-activated mast cells or were incubated with T0070907, probiotics showed little or no effect.

CONCLUSIONS: Probiotics modulate mast cell mediator release to a more barrier protective profile, resulting in amelioration of stress-induced epithelial barrier dysfunction, which is putatively mediated by a PPAR-γ dependent pathway.

Place, publisher, year, edition, pages
2013.
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-100768OAI: oai:DiVA.org:liu-100768DiVA: diva2:663515
Available from: 2013-11-12 Created: 2013-11-12 Last updated: 2014-03-25Bibliographically approved
In thesis
1. Role of mast cells and probiotics in the regulation of intestinal barrier function
Open this publication in new window or tab >>Role of mast cells and probiotics in the regulation of intestinal barrier function
2013 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The intestinal mucosa is the largest contact area and one of the most important barriers to the outside environment. It is highly specialized in aiding us digest and absorb nutrients. It is daily exposed to several potentially dangerous substances and microorganisms, which if they were allowed to pass into the body, could give rise to diseases. Throughout the small intestine certain sites specialized in antigen sampling are found. These sites are named Peyer’s patches and are lymphoid follicles. The epithelium covering the Peyer’s patches is called follicle-associated epithelium and is specialized in antigen sampling and uptake. The special epithelium enables presentation of luminal antigen to immune cells in the underlying follicle.

Persistent life stress and stressful life events affect the course of irritable bowel syndrome (IBS) and inflammatory bowel disease (IBD) through largely unknown mechanisms. Regulation of epithelial permeability to antigens is crucial for the balance between inflammation and immune-surveillance, and increased intestinal permeability has been shown in patients with ulcerative colitis and Crohns disease. Vasoactive intestinal polypeptide (VIP) and corticotropin-releasing factor have been implicated as important mediators of stress-induced abnormalities in intestinal mucosal functions in animal models. Both of these mediators have been reported to regulate bowel ion secretion in humans during stress and uptake of horseradish peroxidase in rodents. Probiotics have been shown to ameliorate the deleterious effects of stress on intestinal function, but mechanisms remain to be elucidated.

The aim of this thesis was to elucidate whether mast cells play an important role in intestinal barrier function during stress and inflammation. Moreover, we wanted to determine whether probiotics can ameliorate the mucosal barrier integrity during stress and inflammation.

To study the function of mast cells we conducted in vitro experiments on cell lines and ex vivo experiments in Ussing chambers on mouse, rat and human intestinal tissue. The Ussing chamber technique measures electrophysiological properties of the tissue and also gives the possibility to study transcellular and paracellular passage of markers and bacteria. Immunohistology and confocal microscopy have been used to identify mast cells and receptors of interest.

Our results show that stress affects the follicle-associated epithelium barrier by mechanisms involving VIP and mast cells. These findings were corroborated by the localization of VIP receptors on mucosal mast cells. Furthermore, pretreatment with probiotics was effective in protecting the gut against stress-induced intestinal barrier dysfunction and mucosal inflammation. This protection appeared to involve a mast cell and peroxisome proliferatoractivated receptor-γ dependent mechanism.

Place, publisher, year, edition, pages
Linköping: Linköping University Electronic Press, 2013. 62 p.
Series
Linköping University Medical Dissertations, ISSN 0345-0082 ; 1363
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-100770 (URN)10.3384/diss.diva-100770 (DOI)978-91-7519-630-5 (ISBN)
Public defence
2013-12-12, Berzeliussalen, Campus US, Linköpings universitet, Linköping, 13:15 (Swedish)
Opponent
Supervisors
Available from: 2013-11-12 Created: 2013-11-12 Last updated: 2013-11-12Bibliographically approved

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Carlsson, Anders H.Lutgendorff, FemkeSöderholm, Johan D.

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