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Lactobacilli Regulate Staphylococcus aureus 161:2-Induced Pro-Inflammatory T-Cell Responses In Vitro
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
Stockholm University, Faculty of Science, Department of Molecular Biosciences, The Wenner-Gren Institute.
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2013 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 10Article in journal (Refereed) Published
Abstract [en]

There seems to be a correlation between early gut microbiota composition and postnatal immune development. Alteration in the microbial composition early in life has been associated with immune mediated diseases, such as autoimmunity and allergy. We have previously observed associations between the presence of lactobacilli and Staphylococcus (S.) aureus in the early-life gut microbiota, cytokine responses and allergy development in children. Consistent with the objective to understand how bacteria modulate the cytokine response of intestinal epithelial cell (IEC) lines and immune cells, we exposed IEC lines (HT29, SW480) to UV-killed bacteria and/or culture supernatants (-sn) from seven Lactobacillus strains and three S. aureus strains, while peripheral blood mononuclear cells (PBMC) and cord blood mononuclear cells (CBMC) from healthy donors were stimulated by bacteria-sn or with bacteria conditioned IEC-sn. Although the overall IEC response to bacterial exposure was characterized by limited sets of cytokine and chemokine production, S. aureus 161: 2-sn induced an inflammatory response in the IEC, characterized by CXCL1/GROa and CXCL8/IL-8 production, partly in a MyD88-dependent manner. UV-killed bacteria did not induce a response in the IEC line, and a combination of both UV-killed bacteria and the bacteria-sn had no additive effect to that of the supernatant alone. In PBMC, most of the Lactobacillus-sn and S. aureus-sn strains were able to induce a wide array of cytokines, but only S. aureus-sn induced the T-cell associated cytokines IL-2, IL-17 and IFN-gamma, independently of IEC-produced factors, and induced up regulation of CTLA-4 expression and IL-10 production by T-regulatory cells. Notably, S. aureus-sn-induced T-cell production of IFN-gamma and IL-17 was down regulated by the simultaneous presence of any of the different Lactobacillus strains, while the IEC CXCL8/IL-8 response was unaltered. Thus these studies present a possible role for lactobacilli in induction of immune cell regulation, although the mechanisms need to be further elucidated.

Place, publisher, year, edition, pages
2013. Vol. 8, no 10
National Category
Biological Sciences Mathematics
Research subject
Immunology
Identifiers
URN: urn:nbn:se:su:diva-96642DOI: 10.1371/journal.pone.0077893ISI: 000326029300123OAI: oai:DiVA.org:su-96642DiVA: diva2:668042
Note

AuthorCount:12;

Available from: 2013-11-28 Created: 2013-11-25 Last updated: 2017-12-06Bibliographically approved
In thesis
1. THE INFLUENCE OF LACTOBACILLI AND STAPHYLOCOCCUS AUREUS ON IMMUNE RESPONSIVENESS IN VITRO
Open this publication in new window or tab >>THE INFLUENCE OF LACTOBACILLI AND STAPHYLOCOCCUS AUREUS ON IMMUNE RESPONSIVENESS IN VITRO
2013 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

Alteration of gut microbiota has been associated with development of immune mediated diseases, such as allergy. In part, this could be due to the influence of microbes in shaping the immune response. In paper I, we investigated the association of early-life gut colonization with bacteria, and numbers of IL-4, IL-10 and IFN-γ producing cells at two years of age in response to PBMC stimulation with phytohemagglutinin (PHA) in vitro. Early Staphylococcus (S) aureus colonization was directly proportional to increased numbers of IL-4 and IL-10 secreting cells, while early co-colonization with lactobacilli and S. aureus associated with a decrease in IL-4, IL-10 and IFN-γ secreting cells compared to S. aureus alone. This was also confirmed in in vitro stimulations of PBMC with Lactobacillus and/or S. aureus strains, where S. aureus-induced IFN-γ production by Th cells was down regulated by co-stimulation with Lactobacillus. In paper II, we investigated the effects of UV-killed and/or culture supernatant (sn) of Lactobacillus strains and S. aureus strains on IEC and immune cell responses. IEC exposed to S. aureus-sn produced CXCL-1/GRO-α and CXCL-8/IL-8, while UV-killed bacteria had no effect. Further, PBMC from healthy donors exposed to Lactobacillus-sn and S. aureus-sn were able to produce a plethora of cytokines, but only S. aureus induced the T-cell associated cytokines: IL-2, IL-17, IFN-γ and TNF-α; which were down regulated by co-stimulation with any of the different Lactobacillus strains. Intracellular staining verified S. aureus-induced IFN-γ and IL-17 production by Th cells, and increased CTLA-4 expression and IL-10 production by T reg cells.

In conclusion, we show that colonization with gut microbiota at early age modulates the cytokine response in infancy. In addition, bacterial species influence cytokine response in a species-specific manner and we demonstrate that lactobacilli modulate S. aureus-induced immune response away from an inflammatory phenotype.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, 2013. 44 p.
Keyword
Cytokine, T cells, immune response, HT-29, Intestinal epithelial cells, Lactobacillus, Staphylococcus aureus, bacteria and infants
National Category
Immunology in the medical area
Research subject
Immunology
Identifiers
urn:nbn:se:su:diva-94326 (URN)
Presentation
2013-10-25, F564, Arrheniuslaboratorierna, Stockholm, 10:00 (English)
Supervisors
Available from: 2013-10-06 Created: 2013-10-03 Last updated: 2013-11-28Bibliographically approved
2. Lactobacilli- and Staphylococcus aureus mediated modulation of immune responses in vitro
Open this publication in new window or tab >>Lactobacilli- and Staphylococcus aureus mediated modulation of immune responses in vitro
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The human gut harbors a vast number of microbes. These microbes are not passive bystanders. They are important in modulating the immune system. We have previously shown that early colonization with lactobacilli and Staphylococcus (S.) aureus differentially associates with allergy development and/or immune profile at early ages. Here we focus on understanding how these microbes modulate the response of intestinal epithelial cells and immune cells in vitro. In paper I, we investigated the impact of UV-killed and/or cell free supernatant (CFS) of different Lactobacillus (L.) species and S. aureus strains on cytokine production from intestinal epithelial cells (IEC) and immune cells. Enterotoxin-expressng S. aureus 161:2-CFS triggered CXCL-1/GRO-α and CXCL-8/IL-8 production by IEC. S. aureus-induced CXCL-8/IL-8 production was hampered by MyD88 gene silencing of IEC, indicating the importance of TLR signaling. Further, lactobacilli-CFS and S. aureus-CFS were able to induce the production of a number of cytokines by peripheral blood mononuclear cells (PBMC) from healthy donors, but only S. aureus triggered T-cell associated cytokines: IL-2, IL-17, IFN-γ and TNF-α; which were dampened by the co-treatment with S. aureus and any of the different Lactobacillus strains. Flow cytometry of the stimulated PBMC further verified IFN-γ and IL-17 production by T cells upon treatment with S. aureus-CFS, which also induced CTLA-4 expression and IL-10 production by Treg cells. In paper II, we investigated the influence of CFS of L. reuteri and S. aureus on the differentiation of monocyte to DC and subsequently how the generated DC influence T cell response. DC generated in the presence of L. reuteri exhibited an increase in expression of surface markers (HL-DR, CD86, CD83, CCR7) and cytokine production (IL-6, IL-10 and IL-23), but had a decreased phagocytic capacity compared with conventional Mo-DC, showing a more mature phenotype. However, upon LPS stimulation, DC generated in the presence of L. reuteri-CFS displayed a more regulatory phenotype, with a reduced cytokine response both at mRNA and protein levels. On the contrary, DC generated in the presence of S. aureus-CFS resembled the control Mo-DC both at mRNA and protein expression, but SA-DC was more efficient in inducing cytokine production in autologous T cells. In paper III, we studied the influence of L. reuteri-CFS on the retinoic acid (RA)-driven mucosal-like DCs’ phenotype and function to modulate T regulatory cells (Treg) in vitro. DC generated in the presence of RA showed a mucosal-like regulatory-DC phenotype with its CD103 expression, high IL10 production and decreased expression of genes associated with inflammation (NFκB1, RELB and TNF). Further, treatment with L. reuteri-CFS enhanced the regulatory phenotype of RA-DC by increasing the production of several chemokines, such as CXCL1, CXCL5, CCL3, CCL15 and CCL20, which are involved in gut homeostasis, while dampening the expression of most chemokine receptor genes. L. reuteri-CFS also increased CCR7 expression on RA-DC.  RA-DC co-cultured with T cell increased IL10 and FOXP3 expression in Treg. However L. reuteri-CFS pre-conditioning of the RA-DC did not improve the Treg phenotype. In conclusion, bacteria-CFS can have an impact on the response of IEC, differentiation and function of DC and, subsequently the T cell response, when taken together in the context of gut; these can have an impact on the health and disease of the host.

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2016. 70 p.
Keyword
lactobacilli, staphylococcus aureus, dendritic cells, retinoic acid, epithelial cells, T cells
National Category
Immunology
Research subject
Immunology
Identifiers
urn:nbn:se:su:diva-127399 (URN)978-91-7649-365-6 (ISBN)
Public defence
2016-04-29, Vivi Täckholmsalen (Q-salen) NPQ-huset, Svante Arrhenius väg 20, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 2: Submitted.

Available from: 2016-04-06 Created: 2016-03-03 Last updated: 2017-02-17Bibliographically approved

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Haileselassie, YenenehJohansson, Maria A.Björkander, SophiaPetursdottir, Dagbjort H.Petersson, MikaelPersson, Jan-OlovFernández, CarmenHolmlund, UlrikaSverremark-Ekström, Eva
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