Oral Tolerance promotes reduction of inflammatory lesions in mice infected with Leishmania amazonensis.
-
1
Universidade do Estado do Rio de Janeiro (UERJ), Departamento de Genética (DGen), Brazil
-
2
Universidade Federal Fluminense (UFF), Departamento de Imunobiologia (GIM), Brazil
-
3
Universidade Federal do Rio de Janeiro (UFRJ), Instituto de Microbiologia Prof. Paulo de Góes, Departamento de Imunologia., Brazil
-
4
Universidade do Estado do Rio de Janeiro (UERJ), Departamento de Microbiologia, Imunologia e Parasitologia., Brazil
Introduction:
The Oral Tolerance is a physiological mechanism of systemic hyporeactivity to an immunogen previously ingested and bystander suppression is an inhibited response to a second immunogen when it is presented along with the immunogen for which it was established oral tolerance. We use for bystander suppression study, two strains of mice genetically selected for extreme phenotypes of susceptibility (TS) and resistance (TR) to oral tolerance (Silva, AC et al. 1998). TR strain presents good inflammatory responses and a non-tolerogenic profile while TS strain presents non-inflammatory but high-tolerogenic profile, with high percentages of regulatory T cells (Treg cells, CD4+CD25+Foxp3+). Furthermore, TS strain is able to produce high levels of inhibitory citokynes such as IL-10 (Silva, MF et al. 2010). Previous studies in mice strains with extreme phenotypes to susceptibility (TS strain) and resistance (TR strain) to Oral Tolerance showed that when infected with Leishmania amazonensis, TR strain develops an exacerbate lesion while a regulatory activity of the TS strain depresses the inflammation and avoid acute lethal response (Tavares, D et al. 2006).
Material and Methods:
TR and TS strains of mice from the F25 generation, obtained by two-way genetic selection according to susceptibility (TS) or resistance (TR) to ovalbumin oral tolerance (Silva, AC et al. 1998). Bystander suppression was done by modification of Miller et al. (1991). Female mice were gavaged with 2 mg OVA (Sigma Chemical Co., St Louis, MO) in PBS during a two-consecutive-day protocol (total dose of 4 mg). Seven days afterwards, OVA-gavaged mice were challenged with 50 µg of soluble OVA in PBS 8 h after subcutaneously infected with 10 million viable L. amazonensis stationary promastigotes in a volume of 25 µl in the left hind footpad. Evolution of the lesion was monitored by weekly measuring of footpad thickness with a Mitutoyo caliper (MTI Co USA) and expressed as the difference in thickness between the infected and the uninfected counterlateral footpad. The Committee for the Care and Use of Laboratory Animals of the Universidade do Estado do Rio de Janeiro, approved the protocols of the experiments described in this work (CEUA/013/2012/UERJ).
Treg cells from bystander draining lymph nodes were evaluated by flow cytometry according Silva, MF et al. (2010). Briefly, 106 cells were fixed and permeabilized using the mouse regulatory T-cell staining kit (eBioscience), following the manufacturer’s instructions. Data were acquired on a FACSCalibur (BD Biosciences, San Jose, CA) and analyzed using CELLQUEST (BD Biosciences) software.
Results:
The bystander suppression reduced the inflammatory lesion in TR strain similarly to the swelling of infected TS strain (Figure 1A). Non infected (normal group) TS strain presents a higher proportion of Treg cells than TR strain (Figure 2B). In both strains, infected mice present a higher proportion of Treg cells than non infected mice (Figure 2B). Mice previously tolerized and immunized (bystander suppression group) showed an increase of CD4+ CD25+ FoxP3+ cells (Figure 1B).
Discussion:
The suppression of immune reactivity is currently attributed to the regulatory T cells, population dedicated to maintaining peripheral tolerance to self-antigens or prevent harmful immunopathological responses, by inhibiting the release of cytokines such as IL-10 and TGF-β or by mechanisms contact-dependent.
The TS strain presents intrinsically a higher proportion of Treg cells that the TR strain (Figure 2B), and initially, the difference of the inflammatory profiles of these strains could be attributed to a greater or lesser amount of Treg cells.
Bystander suppression by oral tolerance has been described as a regulatory T cell mechanism producing IL-10 and TGF-b that reduces inflammation (Miller A et al. 1991). It has been reported that T CD4+ cells induced by oral antigen, use IL-4 and IL-10 to “educate” Dendritic Cells (DCs), which in turn induce naive T cells to produce the same cytokines as those produced by the orally tolerized memory T cells (Alpan et al, 2004). These CD4+ T cells do not need to recognize the same antigen recognized by naive T cell to which the “educated DCs” send the message. Thus, “educated DCs” by orally tolerized memory T cells induce naive T cells that recognize Leishmania antigens to produce anti-inflammatory cytokines. The bystander suppression induced an increase of IL-10 and decrease of IFN-g in draining lymph nodes, when compared with naïve or immunized mice (Unpublished data).
These results suggest the involvement of Treg cells in the Bystander Suppression mechanism and demonstrate their importance in the reduction of inflammatory lesions in animals infected with Leishmania amazonensis.
Figure 1: Inflammatory lesion and percentage of regulatory T cells (CD4+CD25+Foxp3+) in draining lymph nodes of infected mice, treated or not to bystander oral tolerance protocol. Six mice per group, and the figures are representative of two experiments.
Acknowledgements
We are grateful to Drs Cid Couto and Marcia Giesta for care of the animal colonies (Laboratório de Imunobiologia). This work was supported by This work was supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) - Grant 485269/2007-4 - and Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ) - Grant E-26/101.964/2009 and E-26/112.433/2012.
References
Alpan O, Bachelder E, Isil E, Arnheiter H, Matzinger P. 'Educated' dendritic cells act as messengers from memory to naive T helper cells. Nat Immunol (2004) 5(6):615-22.
Silva AC, Souza KW, Machado RC, Silva MFS, Sant'Anna OA. Genetics of immunological tolerance: I. Bidirectional selective breeding of mice for oral tolerance. Res. Immunol (1998) 149: 151-161.
Silva MF, Kamphorst AO, Hayashi EA, Bellio M, Carvalho CR, Faria AM, Sabino KC, Coelho MG, Nobrega A, Tavares D, Silva AC. Innate profiles of cytokines implicated on oral tolerance correlate with low- or high-suppression of humoral response. Immunology (2010) 130(3):447-57.
Miller A., Lider O, Weiner H. Antigen-driven bystander suppression after oral administration of antigens. J. Exp. Med. (1991)174, 791-798.
Tavares D, Ribeiro RC, Silva AC. Inflammatory lesion and parasite load are inversely associated in Leishmania amazonensis infected mice genetically selected according to oral tolerance susceptibility. Microbes Infect (2006) 8(4):957-64.
Keywords:
bystander suppression,
oral tolerance,
Leishmania amazonensis,
Treg cells,
Dendritic Cells,
inflamation
Conference:
15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013.
Presentation Type:
Abstract
Topic:
Host-pathogen interactions
Citation:
Tavares
D,
Santos
JO,
Xavier
AL,
Bellio
M,
Da Silva
A and
Andrade
AF
(2013). Oral Tolerance promotes reduction of inflammatory lesions in mice infected with Leishmania amazonensis..
Front. Immunol.
Conference Abstract:
15th International Congress of Immunology (ICI).
doi: 10.3389/conf.fimmu.2013.02.01087
Copyright:
The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers.
They are made available through the Frontiers publishing platform as a service to conference organizers and presenters.
The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated.
Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed.
For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions.
Received:
29 Jun 2013;
Published Online:
22 Aug 2013.
*
Correspondence:
Prof. Daniel Tavares, Universidade do Estado do Rio de Janeiro (UERJ), Departamento de Genética (DGen), Rio de Janeiro, Rio de Janeiro, 20550-013, Brazil, tavares.uerj@gmail.com