Elsevier

Vaccine

Volume 31, Issue 46, 4 November 2013, Pages 5405-5412
Vaccine

Neonatal colonization of germ-free mice with Bifidobacterium longum prevents allergic sensitization to major birch pollen allergen Bet v 1

https://doi.org/10.1016/j.vaccine.2013.09.014Get rights and content

Highlights

  • B. longum was tested for perinatal intervention against the allergic sensitization.

  • Neonatal mother-to-offspring mono-colonization model was used.

  • B. longum suppressed Bet v 1-specific immune responses on humoral and cellular level.

  • B. longum induced increased levels of regulatory cytokines IL-10 and TGF-β.

  • B. longum induced production of IL-10 in TLR2-, MyD88-, and MAPK-dependent manner.

Abstract

The main goal in reversing the allergy epidemic is the development of effective prophylactic strategies. We investigated the prophylactic effect of neonatal mother-to-offspring mono-colonization with Bifidobacterium longum ssp. longum CCM 7952 on subsequent allergic sensitization. Adult male and female germ-free (GF) mice were mono-colonized with B. longum, mated and their offspring, as well as age-matched GF controls, were sensitized with the major birch pollen allergen Bet v 1. Furthermore, signaling pathways involved in the recognition of B. longum were investigated in vitro. Neonatal mono-colonization of GF mice with B. longum suppressed Bet v 1-specific IgE-dependent β-hexosaminidase release as well as levels of total IgE and allergen-specific IgG2a in serum compared to sensitized GF controls. Accordingly, Bet v 1-induced production of both Th1- and Th2-associated cytokines in spleen cell cultures was significantly reduced in these mice. The general suppression of Bet v 1-specific immune responses in B. longum-colonized mice was associated with increased levels of regulatory cytokines IL-10 and TGF-β in serum. In vitro, B. longum induced low maturation status of bone marrow-derived dendritic cells and production of IL-10 in TLR2-, MyD88-, and MAPK-dependent manner. Our data demonstrate that neonatal mono-colonization with B. longum reduces allergic sensitization, likely by activation of regulatory responses via TLR2, MyD88, and MAPK signaling pathways. Thus, B. longum might be a promising candidate for perinatal intervention strategies against the onset of allergic diseases in humans.

Introduction

There has been a remarkable increase in allergic diseases over the past few decades, especially in Western countries [1]. Accumulating evidence suggests that environmental changes, rather than genetic factors, are driving the epidemic character of allergy, and events acting within a narrow window of opportunity, either prenatally or early in life, might have major physiological effects [2], [3].

The germ-free status of fetus changes rapidly after birth and the composition of colonizing microbiota can be influenced by multiple factors such as the mode of delivery, dietary changes, high hygiene or over-use of antibiotics [4]. The exposure to microbial stimuli is crucial for the development, maturation and function of the immune system and association between intestinal dysbiosis and allergic disease has been proposed [5]. Several prospective studies have shown lower neonatal colonization by bifidobacteria and lactobacilli species accompanied by higher counts of Clostridium difficile in neonates who developed allergy later in life [6], [7], [8].

Due to the ability of bifidobacteria and lactobacilli to interact with the host immune system and to modulate host immune responses, they have been used with some success in prevention or treatment of allergic disease in infants at risk [9]. Several randomized clinical trials have shown that combined prenatal/postnatal probiotic interventions reduced the cumulative incidence of eczema while less beneficial effects were found in trials using exclusively postnatal treatment approaches (recently reviewed in [10]). These data highlight the importance of the correct timing of probiotic interventions, for which the prenatal period seems to be a significant component.

Dendritic cells (DC) are pivotal in early bacterial recognition through engagement of TLR and C-type lectins, which leads to induction of distinct innate responses that shape the type of T helper cell responses [11], [12]. In this respect, Konieczna et al. recently showed that preconditioning of DC with probiotic Bifidobacterium infantis led to induction of Foxp3 positive regulatory T (Treg) cells with enhanced IL-10 production [13]. Due to the fact that allergic diseases have been associated with a deficiency in Treg cells numbers and/or function [14], specific probiotic strains inducing regulatory immune responses might be beneficial in the prevention of allergic disorders. Indeed, we have previously shown that perinatal administration of Lactobacillus paracasei to pregnant/lactating mice protected against the development of airway inflammation in offspring by activating regulatory pathways [15]. Similarly, neonatal application of Lactobacillus rhamnosus GG or Bifidobacterium lactis Bb-12 suppressed allergic sensitization and airway inflammation in mice by induction of Treg cells associated with increased levels of TGF-β [16].

Germ-free (GF) animals represent a powerful model to study the interaction of a single bacterial strain or a defined mixture of microbial strains with the host immune system [17]. Taking the advantage of this model we have previously shown that neonatal mono-colonization of GF mice with the recombinant bacterial strain Lactobacillus plantarum producing major birch pollen allergen Bet v 1 reduced the development of allergic responses upon systemic sensitization with the same allergen [18].

Bifidobacterium longum subspecies longum CCM 7952 (B. longum) is a commensal bacterial strain originally isolated from the feces of a healthy breast fed infant. This strain has been shown to induce regulatory responses in vitro and to suppress the inflammatory responses in a mouse model of experimental colitis (Srutkova, unpublished results). In the present study we investigated whether neonatal mother-to-offspring mono-colonization of GF mice with B. longum protects the offspring against allergic sensitization to allergen Bet v 1 in a model of type I allergy.

Section snippets

Animals

Germ-free BALB/c mice were kept under sterile conditions and were supplied with water and sterile pellet diet ST1 (Bergman, Czech Republic) ad libitum. Fecal samples were weekly controlled for microbial contamination as previously described [19]. Wild-type (WT) and TLR2−/−, TLR4−/−, and MyD88−/− mice on a C57BL/6 background, obtained from Prof. M. Müller (Vienna, Austria), were kept under SPF conditions. All animal experiments were approved by the local ethics committee.

Bacterial strain

Probiotic strain B.

Neonatal mono-colonization with B. longum prevents systemic sensitization to Bet v 1

Colonization of mice with B. longum remained stable throughout the experiments and reached the level of approximately 5 × 109 CFU/g of feces (data not shown). Sensitization with Bet v 1 induced high levels of allergen-specific antibodies in serum of GF mice. Neonatal mono-colonization with B. longum prevented the development of allergic sensitization as the production of Bet v 1-specific Th2-related IgE and IgG1 as well as Th1-related IgG2a antibodies in serum were reduced in these mice compared

Discussion

In the current study, we demonstrate that neonatal mother-to-offspring mono-colonization with B. longum CCM 7952 significantly reduced the development of allergen-specific immune responses in a gnotobiotic mouse model of type I allergy, which was associated with induction of regulatory milieu. Furthermore we show that TLR2, MyD88 and MAPK signaling pathways are crucial for the recognition of B. longum and induction of IL-10.

Clinical and experimental studies differ greatly regarding the outcome

Acknowledgements

We would like to thank Prof. M. Müller, University of Veterinary Medicine, Vienna, Austria, for providing TLR2-, TLR4- and MyD88-deficient mice. Excellent technical assistance of J. Jarkovska, A. Smolova, I. Grimova, B. Drabonova and O. Ul-Haq is gratefully acknowledged. Supported by grants 303/09/0449 of the Czech Science Foundation, CZ.3.22/2.1.00/09.01574, WTZ CZ16 of the OEAD, NR12-0101-10/2011 of the Republic of Poland, SFB F46 of the Austrian Science Fund (FWF), 7AMB12AT020 of the

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