Silvia Corinti

Regulatory Activity of Autocrine IL-10 on Dendritic Cell Functions

IL-10 is a critical cytokine that blocks the maturation of dendritic cells (DCs), but the relevance of autocrine IL-10 on DC functions has not been investigated. In this study, we found that immature monocyte-derived DCs released low but sizeable amounts of IL-10. After stimulation with bacteria, LPS, lipoteichoic acid, or soluble CD40 ligand, DCs secreted high levels of IL-10. Addition of an anti-IL-10-neutralizing Ab to immature DCs as well as to soluble CD40 ligand- or LPS-maturing DCs led to enhanced expression of surface CD83, CD80, CD86, and MHC molecules and markedly augmented release of TNF-α and IL-12, but diminished IL-10 mRNA expression. Moreover, DCs treated with anti-IL-10 Ab showed an increased capacity to activate allogeneic T cells and primed naive T cells to a more prominent Th1 polarization. DC maturation and IL-10 neutralization were associated with enhanced accumulation of the IL-10 receptor binding chain (IL-10R1) mRNA and intracellular IL-10R1 protein. In contrast, surface IL-10R1 and IL-10 binding activity diminished in mature DCs. These results indicate that autocrine IL-10 prevents spontaneous maturation of DCs in vitro, limits LPS- and CD40-mediated maturation, and increases IL-10 production by DCs. Moreover, IL-10R expression appears to be regulated by both transcriptional and posttranscriptional mechanisms. Endogenous IL-10 and IL-10R can be relevant targets for the manipulation of DC functions.

Oral therapeutic administration of a probiotic mixture suppresses established Th2 responses and systemic anaphylaxis in a murine model of food allergy.

To cite this article: Schiavi E, Barletta B, Butteroni C, Corinti S, Boirivant M, Di Felice G. Oral therapeutic administration of a probiotic mixture suppresses established Th2 responses and systemic anaphylaxis in a murine model of food allergy. Allergy 2011; 66: 499–508.

Ligand activation of nerve growth factor receptor TrkA protects monocytes from apoptosis

Nerve growth factor (NGF) receptors are expressed in different cell types outside the nervous system, and increasing evidence indicates that NGF can act as a regulatory molecule during inflammatory and immune responses. In this study, we show that triggering of the high‐affinity NGF receptor TrkA with agonists protects monocytes from apoptosis induced by gliotoxin or UVB radiation. TrkA stimulation up‐regulates the expression of the anti‐apoptotic Bcl‐2 family members, Bcl‐2, Bcl‐XL, and Bfl‐1. On the other hand, TrkA stimulation does not change the expression of MHC, CD80, CD86, CD40, and CD54 molecules, nor the antigen‐presenting function of monocytes. In addition, during in vitro monocyte to dendritic cell differentiation TrkA expression is progressively lost, suggesting that NGF selectively affects monocyte but not dendritic cell survival.

Interferon-γ promotes exaggerated cytokine production in keratinocytes cultured from patients with atopic dermatitis

Abstract Recent studies suggest that skin keratinocytes from patients with atopic dermatitis (AD) and nonatopic subjects differ in their intrinsic ability to respond to proinflammatory stimuli. In this study keratinocyte cultures established from the normal-looking skin of six adult patients with AD and six healthy, nonatopic control subjects were compared in their response to interferon (IFN)-γ, a potent proinflammatory lymphokine whose expression is increased in chronic AD lesions. Basal expression of IFN-γ receptor as well as IFN-γ–induced membrane expression of HLA-DR and intercellular adhesion molecule (ICAM)-1 were evaluated by flow cytometry. Keratinocyte release of IL-1α, IL-1 receptor antagonist (IL-1ra), granulocyte-macrophage colony stimulating factor (GM-CSF), and tumor necrosis factor (TNF)-α were measured by ELISA on culture supernatants after treatment with IFN-γ or medium alone. Expression of membrane IFN-γ receptor was similar in keratinocytes cultured from nonatopic subjects and subjects with AD. IFN-γ (10 to 500 U/ml) induced comparable levels of membrane HLA-DR and ICAM-1 in both groups of keratinocytes. In contrast, spontaneous release of IL-1α, IL-1ra, GM-CSF, and TNF-α was increased in the supernatants of unstimulated keratinocytes from patients with AD compared with keratinocytes from control subjects, with IL-1ra and GM-CSF reaching statistically significant difference. Moreover, IFN-γ–induced release of all the cytokines tested was much higher for keratinocytes from patients with AD, but the IL-1ra/IL-1α ratio for the two groups of keratinocytes was not substantially different, either basally or after IFN-γ stimulation. The results indicate that keratinocytes from patients with AD are hyperresponsive to IFN-γ in terms of cytokine release.

Evaluation of allergenicity of genetically modified soybean protein extract in a murine model of oral allergen-specific sensitization

Background With the development of genetically modified crop plants there has been a growing interest in the approaches available to assess the potential allergenicity of novel gene products. For additional assessment of the potential allergenicity of expressed proteins, informative data can be generated using animal models. Soybean is one of the major source of protein in human and animal nutrition, and has also been well characterized as a major allergenic source. Advances in biotechnology have resulted in an increasing number of genetically engineered foods, and among these soybean is one of the most widespread.

The recombinant major allergen of Parietaria judaica and its hypoallergenic variant: in vivo evaluation in a murine model of allergic sensitization.

Background  Par j 1 represents the major allergenic component of Parietaria judaica pollen. Its three‐dimensional structure is stabilized by four disulphide bridges. A family of three‐dimensional mutants of the recombinant Par j 1 (rPar j 1) allergen, showing reduced allergenicity and retained T cell recognition has been recently developed by site‐directed mutagenesis.

Oral sensitization with shrimp tropomyosin induces in mice allergen-specific IgE, T cell response and systemic anaphylactic reactions

Appropriate murine models of shrimp tropomyosin (ST) allergy would be useful in investigating the mechanisms underlying food allergy in human subjects, as well as for the pre-clinical evaluation of efficacy and safety of novel therapeutic approaches. These models should mimic immune and clinical features of human disease, including anaphylactic response. We sensitized C3H/HeJ mice by the oral route with purified ST using cholera toxin (CT) as adjuvant. ST-specific IgE, IgG1, IgG2a and IgA responses were evaluated by ELISA. Spleen cell proliferation and cytokine production by allergen-specific activation were assessed. Jejunum and colon fragments were collected to evaluate the local expression of cytokine genes by PCR. Local and systemic anaphylactic reactions induced by oral ST challenge were scored according to symptoms observed. Faecal samples were collected to assess local IgA production and histamine levels. Oral sensitization with ST plus CT induced in mice significant levels of serum IgE and IgG1 and faecal IgA. ST-specific cell proliferation and IL-4, IL-13 and IFN-gamma cytokine production were induced in the spleen. After oral challenge, 100% of mice had anaphylactic symptoms while no symptoms were observed in challenged naive mice. Faecal histamine content after ST challenge appeared significantly increased in sensitized mice when compared with that observed in pre-immune mice. Jejunum mRNA expression of T(h)2 cytokines was up-regulated by ST sensitization. These results support the importance of the oral way of sensitization and of the in-depth characterization of the anaphylactic response for the development of a suitable in vivo model of food allergy.

A Hybrid Expressing Genetically Engineered Major Allergens of the Parietaria Pollen as a Tool for Specific Allergy Vaccination

Background: Allergy is an immunological disorder affecting about 25% of the population living in the industrialized countries. Specific immunotherapy is the only treatment with a long-lasting relief of allergic symptoms and able to reduce the risk of developing new allergic sensitizations and inhibiting the development of clinical asthma in children treated for allergic rhinitis. Methods: By means of DNA recombinant technology, we were able to design a head to tail dimer expressing disulphide bond variants of the major allergen of the Parietaria pollen. IgE binding activity was studied by Western blot, ELISA inhibition assays and the skin prick test. T cell recognition was studied by peripheral blood mononuclear cell proliferation. The immunogenicity of the hybrid was studied in a mouse model of sensitization. Results: In vitro and in vivo analysis showed that the disruption of specific cysteine residues in both allergens caused a strong reduction in IgE binding activity of the PjEDcys hybrid. In addition,we were able to show that a reduction in the IgE epitope content profoundly reduced the anaphylactic activity of the hybrid (from 100 to 1,000 times less than wild-type allergens) without interfering with the T cell recognition. Sera from BALB/c mice immunized with the hybrid were able to bind the natural Parietaria allergens and to inhibit the binding of human IgE to wild-type Par j 1 and Par j 2 allergens up to 90%. Conclusion: Our results demonstrate that hybrid-expressing disulphide bond variants of the major allergens of the Parietaria pollen displayed reduced allergenicity and maintained T cell reactivity for induction of protective antibodies.

Effects of Live and Inactivated VSL#3 Probiotic Preparations in the Modulation of in vitro and in vivo Allergen-Induced Th2 Responses

Background: The immunological mechanisms responsible for the immunomodulatory and anti-allergic effects of probiotic bacteria are still poorly defined. The combined effects of mixtures of different species of probiotic bacteria have been explored only in part. The present study describes the immunomodulatory activity of the VSL#3 probiotic preparation in in vitro and in vivo systems. Methods: The activation and cytokine production by in vitro probiotic-stimulated bone-marrow dendritic cells (BM-DCs) and spleen cells isolated from naïve or Par j 1-sensitized mice were investigated. Mice were intranasally administered a sonicate preparation of VSL#3 before immunization with rPar j 1. Serum antibody levels and cytokine expression in the lung were determined. Results: Both live and sonicated VSL#3 preparations induced maturation and cytokine production by BM-DCs. Cytokine production by spleen cells from naïve or Par j 1-sensitized mice was modulated by the probiotic preparations towards a Treg/Th0 profile, characterized by increased IL-10 and IFN-γ production. In vivo prophylactic treatment with VSL#3 induced a significant reduction of serum specific IgG1. At lung level, VSL#3 pre-treatment remarkably reduced IL-13 and IL-4 mRNA expression and increased IL-10 expression. Conclusions: The VSL#3 preparations have not only the capacity to bias primary immune responses towards a Treg/Th0-type profile, but also to modify in the same way the functional characteristics of established in vitro Th2 responses. In vivo studies on a mouse model of Par j 1 sensitization indicate that the prophylactic intranasal treatment with probiotic bacteria is able to modulate the development of Th2-biased responses.

Regulatory Role of Nitric Oxide on Monocyte-Derived Dendritic Cell Functions

Nitric oxide (NO) has an established role in the defense against bacterial infections and exerts multiple modulatory activities on both inflammatory and immune responses. However, the relevance of NO on dendritic cell (DC) functions has been poorly investigated. In this study, we found that addition of the NO donor S-nitrosoglutathione (GSNO) to monocyte-derived DCs matured by lipopolysaccharide (LPS) or soluble CD40 ligand led to a decreased capacity to activate naive allogeneic T cells but a more prominent Th1 polarization, with increased interferon-gamma (IFN-gamma) secretion and reduced interleukin-5 (IL-5) release. The presence of GSNO during maturation of DCs caused a reduced expression of surface CD86, whereas CD80, CD83, and MHC molecule expression was not affected. Moreover, GSNO induced a dose-dependent decrease of IL-10 and enhancement of tumor necrosis factor-alpha (TNF-alpha) release from mature DCs. In parallel, a marked reduced production of IL-12 p40 subunit but no significant perturbation of the bioactive IL-12 p70 production was observed. Finally, GSNO significantly reduced the release of IP-10/CXCL10 and RANTES/CCL5 but not IL-8/CXCL8 by mature DCs. Although GSNO can strengthen the capacity of mature DCs to induce type 1 polarization of T lymphocytes, our data suggest that it elicits distinct anti-inflammatory functions, eventually reducing T lymphocyte proliferation and recruitment.