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Upon inhalation, house dust mite (HDM) allergens are deposited at the nasal and oral mucosa, where IgA is produced abundantly. IgA subclasses have been linked to protection against respiratory allergy previously. It is currently not known whether and how the human IgA subclasses IgA1 and IgA2 contribute to the clinical status of house dust mite-allergic patients. Saliva and serum samples were collected, and HDM-specific, IgE, IgG4, IgA1 and IgA2 levels were determined. HDM-specific levels of IgA in serum were similar to levels measured in nonallergic controls, but HDM-specific levels of IgA2 in saliva were decreased in allergic subjects. HDM-allergic patients who suffered from rhinitis and eczema showed a significant decrease in IgA2-levels compared to patients who suffered from rhinitis only. Taken together, our findings indicate that HDM-specific IgA2, but not IgA1, levels in serum and saliva are reduced in HDM-allergic patients suffering from eczema.

Cow's milk proteins cause allergic symptoms in 2-3% of all infants. In these individuals, the tolerogenic state of the intestinal immune system is broken, which can lead to sensitization against antigens and eventually to allergic responses. Although a true treatment for food allergy is not available, symptoms can be avoided by providing the infants with hydrolyzed proteins. Hydrolyzed proteins are proteins that are enzymatically degraded. They lack typical allergenic IgE-binding epitopes but are also thought to play a pertinent role in other mechanisms inducing hypoallergenic effects. This review discusses the mechanisms and evidence for immunomodulating properties of cow's milk hydrolysates. Hydrolysates are found to strengthen the epithelial barrier, modulate T-cell differentiation, and decrease inflammation. Some studies suggest a role for hydrolysates in manipulating pathogen recognition receptors signaling as underlying mechanism. Peptides from hydrolysates have been shown to bind to TLR2 and TLR4 and influence cytokine production in epithelial cells and macrophages. Current insight suggests that hydrolysates may actively participate in modulating the immune responses in subjects with cow's milk allergy and those at risk to develop cow's milk allergy. However, more research is required to design effective and reproducible means to develop targeting strategies to modulate the immune response.

Oral cholera vaccination is used to induce immune responses in the intestines to protect against cholera infection. However, oral vaccination may also affect immune responses in other mucosal tissues. To study this, tissue-specific homing potential and kinetics of B-cell responses were characterized after oral cholera vaccination. Healthy adult volunteers received two doses of Dukoral® and blood, saliva, nasal wash, and fecal samples were collected over time to detect vaccine-specific antibodies. Additionally, homing potential of lymphocytes to small intestine, colon, airways, skin, and periphery was measured by expression of Integrin β1 and β7, CCR9, CCR10, CCR7, and CLA. After vaccination, antibody responses to cholera toxin B (CTB) and Dukoral® were detected in serum and nasal wash. CTB-specific memory B cells in peripheral blood and tissue homing profiles of memory B cells peaked at day 18. IgA+ memory B cells expressed markers that enable homing to the airways and colon, while IgA− memory B cells primarily expressed small-intestine-homing markers. These data show that oral cholera vaccination has a differential effect on immune responses in various mucosal sites, including the respiratory tract.

It is well established that both the production of IgE by B lymphocytes and the maturation and recruitment of eosinophils in late-phase reactions are dependent on the activation of allergen-specific type-2 T-helper cells. What is less well known is the fact that efficient activation of allergen-specific T cells upon low-dose exposure to allergens is critically dependent on IgE-mediated or -facilitated allergen presentation. In fact, changes in the level of IgE-mediated allergen presentation may account for many of the immunological effects described for specific immunotherapy or anti-IgE treatment. This review aims to summarize the current knowledge, and will discuss the clinical relevance of blocking IgG antibodies induced by specific immunotherapy and anti-IgE monoclonal antibodies that both interfere with IgE-mediated allergen presentation.

Background The role of allergen-specific CD4+ T lymphocytes in the pathophysiology of atopic disease is well established. Previous studies on allergen-specific T-cell responses have focused on the recognition of single major allergens to identify T-cell epitopes. Objective However, it is not clear whether immune responses to allergen extracts are exclusively targeted at major allergens or whether additional proteins are recognized. Methods Here we describe the Phleum pratense-specific immunoglobulin E (IgE) and T-cell responses of six allergic rhinitis patients. Reactivity was measured to size-separated fractions of a P. pratense extract as well as to the purified major allergens Phl p 1, Phl p 2/3 and Phl p 5. Results The specificity of the patients' serum IgE, measured in a fluid phase assay, was restricted to one or two of the major allergens. Even though the majority of the patients had IgE antibodies reactive with a single major allergen, one patient reacted with both Phl p 5 and with Phl p 2/3. Analysis of the T-cell repertoire with P. pratense-specific T-cell lines (TCLs) and CD4+ T-cell clones (TCCs) revealed that at least six different proteins were recognized, including the three major allergens, most notably Phl p 5. Simultaneous production of IL-5 and interferon (IFN) -γ was detected in supernatants of the TCLs stimulated with P. pratense extract and the major allergens. Conclusion These results indicate that allergic rhinitis patients have a large pool of circulating allergen-specific CD4+ T cells that recognize many different proteins in an allergenic extract, whereas only a small number of these proteins are recognized by serum IgE.

The CC-chemokine receptor 5 (CCR5) has recently been described as a surface marker of human T cells producing type 1 (Th1) cytokines. Here we confirm that CCR5 is expressed on human Th1 but not on Th2 T-cell clones. Using intracellular cytokine staining, we show that alloantigen specific CD4+ T-cell lines derived from a CCR5-deficient individual (delta32 allele homozygote) contain high numbers of both interferon gamma (IFN-gamma) and interleukin (IL)-2 producing cells, low numbers of IL-10 producing cells and no IL4 or IL-5 producing cells when stimulated with phorbol ester and ionomycin in vitro. These results were similar to those obtained from alloantigen specific CD4+ T-cell lines derived from CCR5 expressing individuals. An enzyme-linked immunoabsorbent assay (ELISA) confirmed that the Th1 cytokine-positive cells from the CCR5-deficient individual were able to produce equal amounts of cytokines when compared to T-cell lines from CCR5-expressing individuals, These results demonstrate that CCR5-negative T cells display the same capacity of Th1 T-cell differentiation as T cells derived from CCR5-expressing individuals. Thus, CCR5 expression is not essential for differentiation of human Th1 T cells.

Background The involvement of CD4+ T cells in the pathophysiology of atopic disease is well established. Objective To gain further insight into the activation requirements for allergen-specific T cells, we characterized epitope specificity, HLA restriction and T-cell receptor (TCR) usage for T cells specific to Phl p 5, the group 5 major allergen of the grass Phleum pratense. Methods To identify the T-cell epitopes of Phl p 5, three Phl p 5-specific T-cell lines (TCLs) and 15 T-cell clones (TCCs) generated from the peripheral blood of three grass-allergic patients were tested with recombinant truncated Phl p 5a fragments and synthetic Phl p 5b peptides representing these two different recombinant Phl p 5 isoallergens. Additional activation experiments with HLA-subtyped antigen-presenting cells and flow cytometry analysis with TCR V-specific mAb were performed to further characterize the activation requirements for Phl p 5-specific TCCs. Results At least nine distinct T-cell specificities were identified and the T-cell epitopes recognized differed considerably among the three patients. Most of the epitopes found were isoform-specific, whereas three epitopes were shared between Phl p 5a and 5b. Several human leucocyte antigen (HLA) class II molecules were involved in the recognition of Phl p 5. Different HLA restriction specificities were even found among TCCs specific to the same epitope region. All TCCs were TCR-alpha/beta positive, and an overrepresentation of TCR Vbeta 3.1+ clones among TCCs specific to Phl p 5 appear to exists as 31% (4/13) of the TCCs expressed TCR Vbeta 3.1 (compared with 5% TCR Vbeta 3.1+ T cells in human peripheral blood) with no correlation with epitope specificity or HLA restriction. Conclusion The T-cell reactivity of the three grass-allergic patients investigated shows that isoallergen-specific T-cell epitopes are found throughout the peptide backbone of Phl p 5a and Phl p 5b, and dominant T-cell epitopes of Phl p 5 were not identified. This indicates that a mixture of at least full-length rPhl p 5a and rPhl p 5b may be required to target the total Phl p 5-specific T-cell response of atopic patients.

Background: Pollinosis, caused by grasses of the Poaceae family, is a problem worldwide. The relative importance of grass groups 1 and 5 major allergens is well established. However, not much is known about the recognition of these allergens by T cells and whether this T-cell reactivity correlates with skin reactivity and serum IgE levels. Objectives: The aim of this study was to characterize the cross-reactive, grass allergen–specific T-cell responses from patients allergic to grass and nonatopic individuals and to investigate whether these responses correlate with grass-specific IgE and skin reactivity. Methods: Skin prick test wheal areas and grass-specific serum IgE levels were determined in all patients ( n = 21) and nonallergic control donors ( n = 20). Peripheral blood mononuclear cells were isolated and stimulated with grass allergen extracts (Phleum pratense, Poa pratensis, Lolium perenne) and immunoaffinity-purified group 5 allergens, and the production of type 1 and type 2 cytokines was determined in the patient group. Results: Donors allergic to grass showed increased T-cell–proliferative responses to grass allergens compared with nonatopic control subjects. We find it interesting that the magnitude of the patients' T-cell responses could not be correlated with the individual skin prick test areas and specific serum IgE levels, and several patients with allergies to grass had group 5–specific T-cell responses in the absence of group 5–specific IgE. The absence of a correlation between T-cell proliferation and IgE levels or skin prick test results may in part be explained by the finding that patients predominantly produced IL-5 in response to Phl p 5, the major allergen, and predominantly IFN-γ in response to Phl p extract. In general, all donors responded equally well to all three grasses. Additional experiments with Phl p 5–specific T-cell lines indicated that the equal proliferation of peripheral blood mononuclear cells to all three species is the direct result of cross-reactivity. Conclusions: Grass allergen–specific T-cell responses are highly cross-reactive, and patients with allergies exhibit higher responses than nonallergic donors, suggesting that T cells are involved in the allergic reaction to grass group 5 allergens. However, group 5–specific T-cell responses are also found in donors without group 5–specific IgE, and the patients' grass-specific T-cell responses and cytokine production do not correlate to skin reactivity or to concentrations of grass-specific IgE. (J Allergy Clin Immunol 1998;101:241-9.)

Summary Background Allergen-specific T lymphocytes play an important role in the pathophysiology of atopic disease. Detailed studies of their epitope-specificity and crossreactivity are required for the development of novel approaches for specific immunotherapy. Objectives The aim of the study was to characterize the fine specificity of Bet v 1-specific T cells from allergic donors. Methods Polyclonal T-cell lines (TCL) and T-cell clones (TCC), specific for Bet v 1, the major birch (Betula verrucosa) pollen allergen, were isolated from the peripheral blood of three birch-allergic patients. Their epitope-specificity was studied using overlapping synthetic peptides, and crossreactivity with other tree pollen allergens of the Fagales order was evaluated. In addition, the Bet v 1-specific TCC were studied for their phenotype and cytokine production. Results All isolated Bet v 1-specific TCC (19/21 CD4+, 2/21 CD8+) reacted with affinity purified Bet v 1, but showed different reactivities with recombinant Bet v 1 (rBet v 1), and with group 1 allergens from other Fagales species. Epitope mapping of rBet v 1-reactive TCC with synthetic peptides of Bet v 1 showed the presence of four T-cell epitopes. Polyclonal T-cell lines reacted with 13 different peptides, and displayed even broader crossreactivity with group 1 pollen allergens from other Fagales members. Conclusion This study demonstrates that apart from T-cell epitopes of rBet v 1, many other crossreactive or Bet v 1 isoallergen-specific epitopes exist. This indicates that isoallergenic variation plays an important role in the induction of Bet v 1-specific and crossreactive T-cell responsiveness to allergens.

Non-inflammatory dendritic cell (DC) subsets play an essential role in preventing massive inflammation in mucosal tissues. We investigated whether mucosa-related factors, namely retinoic acid (RA) and transforming growth factor-β (TGF-β1), can induce such DC types. DCs were differentiated from monocytes in the absence or presence TGF-β1 and RA. The phenotype as well as responsiveness to bacterial ligands was studied in detail. Compared to monocyte-derived DCs (moDCs), the expression of co-stimulatory molecule CD86 and DC maturation marker CD83 were strongly reduced by RA and TGF-β1. In addition, both RA- and TGF-β1-induced DCs showed strongly decreased responsiveness to stimulation with the bacterial ligands lipopolysaccharide and peptidoglycan, and produced significantly lower levels of the pro-inflammatory cytokines IL-12 and TNF-α compared to moDCs, whilst IL-10 production was not significantly reduced. DCs differentiated under the influence of RA uniquely expressed markers related to intestinal homing (CD103 and integrin β7). In addition, CCR7, which mediates homing to lymph nodes, was expressed by DCs differentiated in the presence of RA, and also to a lesser extent by the other DC types. Furthermore, whereas moDCs and TGF-β1-derived moDCs expressed high levels of CD32, RA-derived DCs lacked CD32 expression but expressed high levels of CD64, suggesting that RA-DCs may primarily respond to soluble proteins and moDCs, and TGF-β DCs to immune complexes. The data presented here support the hypothesis that the mucosal factors TGF-β1 and RA, which can also be provided through dietary intake of dairy products, result in functionally and phenotypically distinct DC types with non-inflammatory properties.

Basophils account for only 0.1-1% of all peripheral blood leukocytes. They were considered to be a redundant cell type for a long time. However, several findings show a non-redundant role for basophils in type 2 T-helper cell (Th2) immune responses in helminth infections, allergy and autoimmunity. Both immunoglobulin-E-dependent and -independent pathways have been described to contribute to basophil activation. In addition, several recent studies reported that basophils can function as antigen-presenting cells and are important in the initiation of Th2 immune responses. However, there are also conflicting studies that do not corroborate the importance of basophils in Th2 immune responses. This review discusses the role of basophils in Th2 immune responses in view of these recent findings. Copyright (c) 2012 S. Karger AG, Basel

Background: As a result of the very efficient capture of allergens by IgE that focuses to CD23 on B cells or FcΕRI on dendritic cells, allergen-specific T cells can be activated after exposure to very low levels of allergens. This IgE-mediated allergen presentation is 100- to 1,000-fold more efficient than fluid phase endocytosis. The aim of the present study was to determine whether humanized anti-IgE mAb Hu-901 can prevent the activation of allergen-specific T cells by inhibiting IgE-mediated allergen presentation. Methods: A house dust mite major allergen Der p 1-specific T cell line was generated from an allergic asthma patient, and a model was set up to show IgE-facilitated allergen presentation via CD23 on EBV-transformed B cells. In addition, experiments were performed by FACS analysis, detecting the presence of IgE-allergen complexes bound to EBV-B cells by polyclonal FITC-labeled anti-IgE antisera. Results: The anti-IgE mAb Hu-901 inhibited proliferation of allergen-specific T cells at low allergen concentrations. Inhibition was dose-dependent. This effect could be explained by Hu-901 inhibition of binding of allergen-IgE complexes to CD23 expressed on EBV-transformed B lymphocytes. Conclusions: These data clearly indicate that anti-IgE antibodies for the treatment of allergy exert their effect not only by inhibiting mast cell/basophil degranulation, but also by preventing T cell activation, which possibly explains the effect of anti-IgE treatment on late-phase reactions noted in clinical studies.

BACKGROUND The central role of specific IgE in cow's milk allergy (CMA) is well documented. However, less is known about the function of other immunoglobulin isotypes in allergy and tolerance to cow's milk proteins (CMPs). OBJECTIVE To determine differences in the antibody responses that are associated with allergy and tolerance to cow's milk in allergic, atopic and non-atopic individuals of different age groups. METHODS Nineteen infants (

ummary Background The clinical efficacy of specific allergy vaccination (SAV), previously called specific immunotherapy is well documented. The working mechanism of this treatment is not completely known at present. Allergen-specific CD4+ T lymphocytes are activated at extremely low allergen concentrations in vivo possibly as a result of serum IgE-facilitated allergen presentation (S-FAP). Previously, we have shown that this process can be inhibited by long-term birch SAV sera. Methods In the present study, we have analysed sera from birch-allergic patients in a randomized double-blind, placebo-controlled clinical trial for their ability to mediate S-FAP. Birch-specific IgE levels were not changed after SAV. Bet v 1-specific IgE levels, however, were significantly decreased (P

Background: At present, recombinant allergens are considered for the diagnosis and treatment of atopic allergies. To evaluate the theoretical impact of isoallergen variation on the selection of isoallergens for specific allergy vaccination, we characterized the T–cell response of allergic patients to the Phleum pratense major allergen, Phl p 5, and five of its recombinant isoallergens. Methods: Phl p 5–specific T cell lines (TCLs) were isolated from the peripheral blood of 3 allergic rhinitis patients, and their reactivity patterns were studied in detail. Results: The TCLs were highly crossreactive with related grasses. The crossreactivity with Poa pratensis was more extensive than with Lolium perenne, directly reflecting the sequence identity between Phl p 5, Poa p 5, and Lol p 5. The TCLs produced IFN–γ and IL–4 simultaneously, resembling a Th0–like cytokine production profile. Interestingly, when T cell clones were tested with natural Phl p 5 and five rPhl p 5 isoallergens, a differential recognition pattern was found. One of the TCLs exclusively reacted with Phl p 5b, another reacted with all isoforms tested, and the third reacted strongly with native purified Phl p 5, but only weakly with all five recombinant isoallergens. Conclusion: These results indicate that Phl p 5–specific T cells are highly heterogeneous, and that they differentially recognize isoallergens. This indicates that when recombinant Phl p 5 is considered for allergy vaccination, a mixture of isoallergens representing the different isoallergen groups may clinically prove to be more effective than single isoallergens.