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Allergen specific tolerance induction efficiently ameliorates subsequent allergen induced inflammatory responses. The underlying regulatory mechanisms have been attributed mainly to interleukin (IL)-10 produced by diverse hematopoietic cells, while targets of IL-10 in allergen specific tolerance induction have not yet been well defined. Here, we investigate potential cellular targets of IL-10 in allergen specific tolerance induction using mice with a cell type specific inactivation of the IL-10 receptor gene. Allergic airway inflammation was effectively prevented by tolerance induction in mice with IL-10 receptor (IL-10R) deficiency in T or B cells. Similarly, IL-10R on monocytes/macrophages and/or neutrophils was not required for tolerance induction. In contrast, tolerance induction was impaired in mice that lack IL-10R on dendritic cells: those mice developed an allergic response characterized by a pronounced neutrophilic lung infiltration, which was not ameliorated by tolerogenic treatment. In conclusion, our results show that allergen specific tolerance can be effectively induced without a direct impact of IL-10 on cells of the adaptive immune system, and highlight dendritic cells, but not macrophages nor neutrophils, as the main target of IL-10 during tolerance induction., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Regulatory mechanisms initiated by allergen-specific immunotherapy are mainly attributed to T cell derived IL-10. However, it has not been shown that T cell derived IL-10 is required for successful tolerance induction (TI). Here, we analyze cellular sources and the functional relevance of cell type specific IL-10 during TI in a murine model of allergic airway inflammation. While TI was effective in IL-10 competent mice, neutralizing IL-10 prior to tolerogenic treatment completely abrogated the beneficial effects. Cellular sources of IL-10 during TI were identified by using transcriptional reporter mice as T cells, B cells, and to a lesser extent DCs. Interestingly, TI was still effective in mice with T cell, B cell, B and T cell, or DC-specific IL-10 deficiency. In contrast, TI was not possible in mice lacking IL-10 in all hematopoetic cells, while it was effective in bone marrow (BM) chimera that lacked IL-10 only in nonhematopoetic cells. Taken together, allergen-specific tolerance depends on IL-10 from hematopoetic sources. The beneficial effects of allergen-specific immunotherapy cannot solely be attributed to IL-10 from T cells, B cells, or even DCs, suggesting a high degree of cellular redundancy in IL-10-mediated tolerance., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Dendritic cells (DC) presenting tumor antigens are crucial to induce potent T cell-mediated anti-tumor immune responses. Therefore DC-based cancer vaccines have been established for therapy, however clinical outcomes are often poor and need improvement. Using a mouse model of B16 melanoma, we found that the route of preventive DC vaccination critically determined tumor control. While repeated DC vaccination did not show an impact of the route of DC application on the prevention of tumor growth, a single DC vaccination revealed that both the imprinting of skin homing receptors and an enhanced proliferation state of effector T cells was seen only upon intracutaneous but not intravenous or intraperitoneal immunization. Tumor growth was prevented only by intracutaneous DC vaccination. Our results indicate that under suboptimal conditions the route of DC vaccination crucially determines the efficiency of tumor defense. DC-based strategies for immunotherapy of cancer should take into account the immunization route in order to optimize tissue targeting of tumor antigen specific T cells.

Background: Allergic contact dermatitis is one of the most common occupational diseases. A main protective mechanism in those who do not develop allergic contact dermatitis is tolerance induction by repeated exposure to low doses of contact allergen, which is termed low zone tolerance (LZT). The mechanisms that determine the tolerance induction in subjects with LZT are still elusive., Objective: We performed analysis of the role of CD4(+)CD25(+) forkhead box protein 3 (FOXP3)-positive regulatory T (Treg) cells and dendritic cells (DCs) in mice with LZT., Methods: Mechanisms of tolerance induction were analyzed in a murine model of LZT by using FOXP3 and IL-10 reporter mice, as well as mice that allow the selective depletion of Treg cells or DCs., Results: Depletion of CD4(+)CD25(+)FOXP3(+) Treg cells during tolerance induction completely abolishes the development of LZT, resulting in a pronounced contact hypersensitivity response. Adoptive transfer experiments, depletion studies, and use of cell type-specific deficient mice revealed that IL-10 production is critical for the suppressor function of Treg cells in mice with LZT and that tolerogenic CD8(+)CD11c(+) DCs located in the skin-draining lymph nodes are essential for LZT. In the absence of Treg cells, DCs did not develop tolerogenic functions, indicating that activated IL-10(+) Treg cells might imprint the tolerogenic DC phenotype. Cell communication analysis revealed that the education of tolerogenic DCs might involve a direct interaction with Treg cells mediated by gap junctions. Subsequently, induction of tolerogenic CD11c(+) DCs leads to the generation of hapten-specific CD8(+) Treg cells, which protect against contact hypersensitivity., Conclusions: Our data demonstrate critical interactions between CD4(+)CD25(+)FOXP3(+) Treg cells and tolerogenic CD8(+)CD11c(+) DCs during the induction of LZT., (Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.)

Diesel exhaust particles (DEP) were described as potent adjuvant in the induction and maintenance of allergic diseases, suggesting that they might play a role in the increase of allergic diseases in the industrialized countries. However, the cellular basis by which these particles enhance allergic immune responses is still a matter of debate. Thus, we exposed immature murine bone marrow-derived dendritic cells (BMDC) to different particles or particle-associated organic compounds in the absence or presence of the maturation stimuli lipopolysaccharide (LPS) and analyzed the cellular maturation, viability, and cytokine production. Furthermore, we monitored the functionality of particle-exposed BMDC to suppress B cell isotype switching to immunoglobulin (Ig) E. Only highly polluted DEP (standard reference material 1650a [SRM1650a]) but not particle-associated organic compounds or less polluted DEP from modern diesel engines were able to modulate the dendritic cell phenotype. SRM1650a particles significantly suppressed LPS-induced IL-12p70 production in murine BMDC, whereas cell-surface marker expression was not altered. Furthermore, SRM1650a-exposed immature BMDC lost the ability to suppress IgE isotype switch in B cells. This study revealed that highly polluted DEP not only interfere with dendritic cell maturation but also additionally with dendritic cell function, thus suggesting a role in T(h)2 immune deviation.

In a humid milieu such as mucosal surfaces, pollen grains do not only release allergens but also proinflammatory and immunomodulatory lipids, termed pollen-associated lipid mediators. Among these, the E(1)-phytoprostanes (PPE(1)) were identified to modulate dendritic cell (DC) function: PPE(1) inhibit the DC's capacity to produce IL-12 and enhance DC mediated T(H)2 polarization of naive T cells. The mechanism(s) by which PPE(1) act on DC remained elusive. We thus analyzed candidate signaling elements and their role in PPE(1)-mediated regulation of DC function. Aqueous birch pollen extracts induced a marked cAMP response in DC that could be blocked partially by EP2 and EP4 antagonists. In contrast, PPE(1) hardly induced cAMP and the inhibitory effect on IL-12 production was mostly independent of EP2 and EP4. Instead, PPE(1) inhibited the LPS-induced production of IL-12 p70 by a mechanism involving the nuclear receptor PPAR-gamma. Finally, PPE(1) efficiently blocked NF-kappaB signaling in DCs by inhibiting IkappaB-alpha degradation, translocation of p65 to the nucleus, and binding to its target DNA elements. We conclude that pollen-derived PPE(1) modulate DC function via PPAR-gamma dependent pathways that lead to inhibition of NFkappaB activation and result in reduced DC IL-12 production and consecutive T(H)2 polarization.

The immune response of atopic individuals against allergens is characterized by increased levels of Th2 cytokines and chemokines. However, the way in which the cytokine/chemokine profile is matched to the type of invading allergen, and why these profiles sometimes derail and lead to disease, is not well understood. We recently demonstrated that pollen modulates dendritic cell (DC) function in a way that results in an enhanced capacity to initiate Th2 responses in vitro. Here, we examined the effects of aqueous birch pollen extracts (Bet.-APE) on chemokine receptor expression and chemokine production by human monocyte-derived DCs. Bet.-APE strongly induced expression and function of CXCR4 and reduced CCR1 and CCR5 expression on immature DCs. In addition, DC treatment with Bet.-APE significantly reduced LPS-induced production of CXCL10/IP-10, CCL5/RANTES; induced CCL22/macrophage-derived chemokine; and did not significantly change release of CCL17/thymus and activation-regulated chemokine. At a functional level, Bet.-APE increased the capacity of LPS-stimulated DCs to attract Th2 cells, whereas the capacity to recruit Th1 cells was reduced. Bet.-APE significantly and dose-dependently enhanced intracellular cAMP, suggesting that water-soluble factors from pollen grains bind a G(alphas)-protein-coupled receptor. E(1)-Phytoprostanes were identified to be one player in the Th2-polarizing potential of aqueous pollen extracts. In summary, our results demonstrate that pollen itself releases regulatory mediators which generate a Th2-promoting micromilieu with preferential recruitment of Th2 cells to the site of pollen exposure.

Pollen grains induce allergies in susceptible individuals by release of allergens upon contact with mucosal membranes of the upper respiratory tract. We recently demonstrated that pollen not only function as allergen carriers but also as rich sources of bioactive lipids that attract cells involved in allergic inflammation such as neutrophils and eosinophils. Here we demonstrate that soluble factors from birch (Betula alba L.) pollen activate human dendritic cells (DCs) as documented by phenotypical and functional maturation and altered cytokine production. Betula alba L. aqueous pollen extracts (Bet.-APE) selectively inhibited interleukin (IL)-12 p70 production of lipopolysaccharide (LPS)- or CD40L-activated DC, whereas IL-6, IL-10, and TNFalpha remained unchanged. Presence of Bet.-APE during DC activation resulted in DC with increased T helper type 2 (Th2) cell and reduced Th1 cell polarizing capacity. Chemical analysis of Bet.-APE revealed the presence of phytoprostanes (dinor isoprostanes) with prostaglandin E(1)-, F(1)-, A(1)-, or B(1)-ring systems of which only E(1)-phytoprostanes dose dependently inhibited the LPS-induced IL-12 p70 release and augmented the Th2 cell polarizing capacity of DC. These results suggest that pollen-derived E(1)-phytoprostanes not only resemble endogenous prostaglandin E(2) structurally but also functionally in that they act as regulators that modulate human DC function in a fashion that favors Th2 cell polarization.

The question “What makes an allergen an allergen?” has puzzled generations of researchers, and we still do not have a conclusive answer. Despite increasing knowledge about the molecular and functional characteristics of allergens that have been identified, we still do not fully understand why some proteins are clinically relevant allergens and most are not. Different approaches have been taken to identify the structural and functional features of allergens, aiming at developing methods to predict allergenicity and thus to identify allergens. However, none of these methods has allowed a reliable discrimination between allergenic and nonallergenic compounds on its own. This review sums up diverse determinants that contribute to the phenomenon of allergenicity and outlines that in addition to the structure and function of the allergen, factors derived from allergen carriers, the environment, and the susceptible individual are of importance. [Copyright &y& Elsevier]

Pollen allergy is characterized by a TH2-biased immune response to pollen-derived allergens. However, pollen-exposed epithelia do not encounter pure allergen but rather a plethora of protein and non-protein substances. We demonstrated that pollen liberate lipids with chemical and functional similarities to leukotriens and prostaglandins - the pollen associated lipid mediators (PALMs). To date, two main groups of PALMs have been characterized: The immunostimulatory PALMs activating innate immune cells such as neutrophils and eosinophils, and the immunomodulatory E1-phytoprostanes blocking IL-12 production of dendritic cells, resulting in the preferential induction of TH2 responses. This article reviews our work in the field of PALMs and their effects on cells of the innate and adoptive immune system. From recent results a general picture starts to emerge in which PALMs (and possibly other pollen-associated substances) may - independently from protein allergens - propagate an overall TH2 favoring micromilieu in pollen exposed tissue of predisposed individuals. [ABSTRACT FROM AUTHOR]

Background: We recently demonstrated that pollen not only function as allergen carriers but also as rich sources of bioactive lipids, such as phytoprostanes, that modulate human dendritic cell (DC) function in a way that results in an enhanced TH2 polarization in vitro. Objective: Here we analyzed the immunomodulatory capacities of Betula alba (white birch) aqueous pollen extracts (Bet-APEs) and pollen-associated phytoprostanes in the murine system in vitro and in vivo. Methods: DC function was analyzed in vitro by using BALB/c bone marrow–derived DCs. T-cell responses were analyzed with DO11.10 peptide 323-339 from chicken ovalbumin (OVA)–specific CD4 T cells as responder cells. For in vivo studies, OVA-specific CD4 T cells were adoptively transferred into BALB/c mice. Twenty-four hours later, mice were challenged by means of intranasal application of OVA in the absence or presence of Bet-APEs or phytoprostanes. Polarization of T-cell responses in vivo was analyzed in draining lymph node cells. Results: In vitro Bet-APEs and E1–phytoprostanes dose-dependently inhibited LPS-induced IL-12p70 of DCs. In addition, Bet-APEs induced a TH2 polarization in vitro. Similarly, intranasal instillation of Bet-APEs in vivo, together with the antigen, lead to increased IL-4, IL-5, and IL-13 secretion and decreased IFN-γ secretion from antigen-specific T cells in the draining lymph nodes. In contrast, intranasal E1- and F1–phytoprostanes downregulated both TH1 and TH2 cytokine production in vivo. Conclusion: Pollen release water-soluble factors that display TH2-polarizing capacities in vivo independently of E1- and F1–phytoprostanes. Clinical implications: Identification of the underlying mechanisms might open new approaches for pharmacologic intervention. [Copyright &y& Elsevier]

Background: Unmethylated CpG sequences in bacterial DNA act as adjuvants selectively inducing Th1 predominant immune responses during genetic vaccination or when used in conjunction with protein Ag. The precise mechanism of this adjuvant effect is unknown. Because dendritic cells (DC) are thought to be crucially involved in T cell priming and Th1/Th2 education during vaccination via skin, we characterized the effects of bacterial DNA and CpG–containing oligodeoxynucleotides (CpG ODN) on cutaneous DC. Methods and Results: Stimulation with CpG ODN 1826 (6 μg/ml) induced activation of immature Langerhans cell (LC)–like DC as determined by an increased expression of MHC class II and costimulatory molecules, loss of E–cadherin–mediated adhesion and increased ability to stimulate allogeneic T cells. Composition–matched control ODN 1911 lacking CpG sequences at equal concentrations was without effect. In comparison to LPS and ODN 1911, CpG ODN 1826 selectively stimulated DC to release large amounts of IL–12 (p40) and little IL–6 or TNF–α within 18 h and detectable levels of IL–12 p70 within 72 h. Stimulation with Escherichia coli DNA, but not calf thymus DNA, similarly induced DC maturation and IL–12 p40 production. Injection of CpG ODN into murine dermis induced enhanced expression of MHC class II and CD86 by LC in the overlying epidermis and intracytoplasmic IL–12 p40 accumulation in a subpopulation of activated LC. Conclusion: Bacterial DNA and CpG ODN stimulate DC in vitro and in vivo and may preferentially elicit Th1–predominant immune responses because they can activate and mobilize DC, inducing them to produce IL–12. [ABSTRACT FROM AUTHOR]