Considering that interleukin 10 (IL10), transforming growth factor beta1 (TGFB1), and interferon gamma (IFNG) are involved in the susceptibility of BeWo trophoblast cells to Toxoplasma gondii infection, the aim of the present study was to investigate the effector mechanisms triggered by these cytokines in the control of T. gondii in BeWo cells. For this purpose, infected/uninfected BeWo cells were treated with IL10, TGFB1 (50 ng/ml), and IFNG (20 or 100 ng/ml) in order to verify the phosphorylation of signal transducers and activators of transcription 1 (STAT1), STAT3, and Smad2, parasite intracellular proliferation, as well as the Th1/Th2/IL17A cytokine production. The treatment of BeWo cells with IL10 and TGFB1 favored T. gondii proliferation, and these findings were associated with STAT3 and Smad2 phosphorylation, respectively (P < 0.05). Also, these cytokine treatments were able to down-modulate TNF alpha (TNFA) and IL6 production (P < 0.05). Low concentration of IFNG was unable to control T. gondii infection but was able to trigger STAT1 phosphorylation and up-regulate IL6 and IL17A production; whereas a high concentration of IFNG was unable to activate STAT1 but down-modulated IL6 and TNFA and increased T. gondii proliferation (P < 0.05). IL10, TGFB1, and IFNG regulate a differential T. gondii proliferation in BeWo cells because they distinctly trigger intracellular signaling pathways and cytokine production, especially IL6 and TNFA. Our data open new windows to understand the mechanisms triggered by IL10, TGFB1, and IFNG at the maternal-fetal interface in the presence of T. gondii, contributing to recognizing the importance of these effector mechanisms involved in the vertical transmission of this parasite.

Because macrophage migration inhibitory factor (MIF) is a key cytokine in pregnancy and has a role in inflammatory response and pathogen defense, the objective of the present study was to investigate the effects of MIF in first-and third-trimester human placental explants infected with Toxoplasma gondii. Explants were treated with recombinant MIF, IL-12, interferon-␥, transforming growth factor-␤1, or IL-10, followed by infection with T. gondii RH strain tachyzoites. Supernatants of cultured explants were assessed for MIF production. Explants were processed for morphologic analysis, immunohistochemistry, and real-time PCR analysis. Comparison of infected and stimulated explants versus noninfected control explants demonstrated a significant increase in MIF release in first-trimester but not third-trimester explants. Tissue parasitism was higher in third-than in first-trimester explants. Moreover, T. gondii DNA content was lower in first-trimester explants treated with MIF compared with untreated explants. However, in third-trimester explants, MIF stimulus decreased T. gondii DNA content only at the highest concentration of the cytokine. In addition, high expression of MIF receptor was observed in first-trimester placental explants, whereas MIF receptor expression was low in third-trimester explants. In conclusion, MIF was up-regulated and demonstrated to be important for control of T. gondii infection in first-trimester explants, whereas lack of MIF up-regulation in third-trimester placentas may be involved in higher susceptibility to infection at this gestational age.

Galectin-3 (Gal 3) is a glycan-binding protein that can be secreted by activated macrophages and mast cells at inflammation sites and plays an important role in inflammatory diseases caused by Bacteria and their products, such as lipopolysaccharides (LPS). Although it is well established that Gal 3 can interact with LPS, the pathophysiological importance of LPS/Gal 3 interactions is not fully understood. Data presented herein demonstrate for the first time that the interaction of Gal 3, either via its carbohydrate binding C-terminal domain or via its N-terminal part, with LPS from different bacterial strains, enhances the LPS-mediated neutrophil activation in vitro. Gal 3 allowed low LPS concentrations (1 µg/mL without serum, 1 ng/mL with serum) to upregulate CD11b expression and reactive oxygen species (ROS) generation on human neutrophils in vitro and drastically enhanced the binding efficiency of LPS to the neutrophil surface. These effects required LPS preincubation with Gal 3, before neutrophil stimulation and involved specific Gal 3/LPS interaction. A C-terminal Gal-3 fragment, which retains the lectin domain but lacks the N-terminal part, was still able to bind both to Escherichia coli LPS and to neutrophils, but had lost the ability to enhance neutrophil response to LPS. This result emphasizes the importance of an N-terminus-mediated Gal 3 oligomerization induced by its interaction with LPS. Finally we demonstrated that Balb/C mice were more susceptible to LPS-mediated shock when LPS was pretreated with Gal 3. Altogether, these results suggest that multimeric interactions between Gal 3 oligomers and LPS potentiate its pro-inflammatory effects on neutrophils.

There is recent evidence that galectin-3 participates in immunity to infections, mostly by tuning cytokine production. We studied the balance of Th1/Th2 responses to P. brasiliensis experimental infection in the absence of galectin-3. The intermediate resistance to the fungal infection presented by C57BL/6 mice, associated with the development of a mixed type of immunity, was replaced with susceptibility to infection and a Th2-polarized immune response, in galectin-3-deficient (gal3−/−) mice. Such a response was associated with defective inflammatory and delayed type hypersensitivity (DTH) reactions, high IL-4 and GATA-3 expression and low nitric oxide production in the organs of infected animals. Gal3−/− macrophages exhibited higher TLR2 transcript levels and IL-10 production compared to wild-type macrophages after stimulation with P. brasiliensis antigens. We hypothesize that, during an in vivo P. brasiliensis infection, galectin-3 exerts its tuning role on immunity by interfering with the generation of regulatory macrophages, thus hindering the consequent Th2-polarized type of response.

Galectin-3 is a b-galactoside-binding lectin implicated in the fine-tuning of innate immunity. Rhodococcus equi, a facultative intracellular bacterium of macrophages, causes severe granulomatous bronchopneumonia in young horses and immunocompromised humans. The aim of this study is to investigate the role of galectin-3 in the innate resistance mechanism against R. equi infection. The bacterial challenge of galectin-3-deficient mice (gal3 À/À ) and their wild-type counterpart (gal3 1/1 ) revealed that the LD 50 for the gal3 À/À mice was about seven times higher than that for the gal3 1/1 mice. When challenged with a sublethal dose, gal3 À/À mice showed lower bacteria counts and higher production of IL-12 and IFN-c production, besides exhibiting a delayed although increased inflammatory reaction. Gal3 À/À macrophages exhibited a decreased frequency of bacterial replication and survival, and higher transcript levels of IL-1b, IL-6, IL-10, TLR2 and MyD88. R. equi-infected gal3 1/1 macrophages showed decreased expression of TLR2, whereas R. equi-infected gal3 À/À macrophages showed enhanced expression of this receptor. Furthermore, galectin-3 deficiency in macrophages may be responsible for the higher IL-1b serum levels detected in infected gal3 À/À mice. Therefore galectin-3 may exert a regulatory role in innate immunity by diminishing IL-1b production and thus affecting resistance to R. equi infection.Key words: Bacterial infections . Galectin-3 . IL-1b . Innate immunity . Toll-like receptor IntroductionActivation of resident macrophages is one of the earliest events in the cellular host response to microbial invasion, and macrophage-derived cytokines play a key role in the initiation and amplification of the inflammatory process as well as in the regulation of the immune response. On the basis of its capacity to recognize carbohydrates and its abundant expression in activated macrophages [1,2], galectin-3 has been considered an important factor in the interaction of host cells with microorganisms [3]. Extracellular galectin-3 is able to activate cells [4][5][6][7][8][9] cell-cell and cell-extracellular matrix interactions [10][11][12], and induce phagocyte migration [13]. However, galectin-3 also functions inside the cells and can contribute to macrophage functions that are essential in the cellular response during the infectious process, such as cell survival [14] and phagocytosis [15].As a result of its ability to recognize glycans containing b-galactoside, galectin-3 binds to glycoconjugates synthesized by several pathogens such as Mycobacterium tuberculosis [16], Leishmania major [17], Trypanosoma cruzi [18], Schistosoma mansoni [19] and Candida albicans [20]. Recently, galectin-3 and TLR2 have been found to be associated in C. albicans-infected differentiated macrophages, an association that has been considered essential for TLR2-dependent cytokine production in response to the fungal infection [21]. Therefore, galectin-3 has been considered as a novel pattern recognition receptor, acting either alone or in ...

GalectinKeywords: Galectin-3 r IL-10 r Leishmania major r T regulatory (Treg) cells r Notch signaling IntroductionGalectins are a family of glycan-binding proteins composed of 15 members that are conserved throughout animal evolution Correspondence: Prof. Gabriel A. Rabinovich e-mail: gabyrabi@gmail.com and share sequence similarities in their carbohydrate-recognition domain [1][2][3]. Galectin-3, a widely distributed member of the family, plays pleiotropic roles in innate and adaptive immunity by regulating cytokine production, phagocytosis, chemotaxis, * These authors contributed equally to this work.C 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu Eur. J. Immunol. 2013. 43: 1806-1817 Immunity to infection 1807 signaling, and survival [4][5][6][7]. Through these mechanisms, galectin-3 has been proposed to control host immunity against several infectious agents [1,[6][7][8]]. Yet, despite considerable evidence on the role of galectin-3 in the control of immune responses, its contribution to T regulatory (T REG ) cell function during microbial attack has not yet been explored. T REG cells, either inducible or naturally occurring, suppress effector T (T EFF )-cell responses through different mechanisms including cell-cell contact and secretion of immunosuppressive cytokines such as IL-10, . Interestingly, galectin-1 and -10 have been proposed to mediate the immunosuppressive activity of Foxp3 + T REG cells [10, 11] Results Leishmania major infected Lgals3−/− mice show increased frequency of CD4To investigate the role of galectin-3 within the T REG cell compartment, we first compared the outcome of L. major infection in Lgals3 −/− and WT mice on BALB/c background. Mice were inoculated with 1 × 10 7 metacyclic promastigotes into one hind footpad, and the development of the lesions was monitored weekly. Lgals3 −/− mice developed more pronounced footpad swelling starting from 35 days postinfection and exhibited an increased parasite burden (at day 35) compared with WT mice (Fig. 1A). To examine the possible mechanisms underlying the increased susceptibility to L. major infection, we examined the impact of galectin-3 deficiency in different immune cell types. We found no significant differences in the frequency of F4 Fig. 2A and B). In addition, real-time RT-PCR analysis showed increased Foxp3 mRNA expression in footpad tissue from Lgals3 −/− -infected animals as compared with their WT counterpart (Fig. 2C). Of note, galectin-3 protein was detected at high levels in footpad tissue from WT mice ( Fig. 2A; infection sites, but also limits the immunosuppressive function of these cells during the course of parasitic protozoa infection. mRNA was substantially upregulated after stimulation with anti-CD3 and anti-CD28 antibodies in both T EFF and T REG WT cells (Fig. 5C). Galectin-3 controls IL-10 production by T REG cells even in the absence of infectionTo further dissect the role of galectin-3 within the T REG -cell compartment during infection, we isolated T EFF and T REG cells from draining LNs...

Extracellular galectin-3 participates in the control of B2 lymphocyte migration and adhesion and of their differentiation into plasma cells. Here, we analyzed the role of galectin-3 in B1-cell physiology and the balance between B1a and B1b lymphocytes in the peritoneal cavity. In galectin-3(-/-) mice, the total number of B1a lymphocytes was lower, while B1b lymphocyte number was higher as compared to wild-type mice. The differentiation of B1a cells into plasma cells was associated with their abnormal adhesion and location on the mesentery. The B220 and CD43, constitutively expressed by B1 lymphocytes, were respectively up- and downregulated in galectin-3(-/-) mice. Mononuclear cells were strongly adhered to the mesenteric membranes of both CD43(-/-) and galectin-3(-/-) mice, but in contrast to CD43(-/-) mice, the accumulation of B1 cells in peritoneal membranes in galectin-3(-/-) mice was accompanied by their functional differentiation into plasma cells. We have shown that in the absence of galectin-3, B1-cell differentiation into plasma cells is favored and the dynamic equilibrium of B1-cell populations in the peritoneum is maintained through a compensatory increase in B1b lymphocytes.

This study provides new insights on the mechanisms of allergen-free immunotherapy by showing that both DNA-HSP65 and CpG/CFP downregulated house dust mite-induced allergic airway inflammation via distinct pathways that involve not only induction of mycobacterial-specific adaptive responses but also signaling via MyD88 and Fas molecules.