Transgenic mice expressing the T cell antigen receptor specific for an immunodominant epitope of a major allergen of house dust mite develop an asthmatic phenotype on exposure of the airways to allergen.

BACKGROUND: Current studies on mechanisms underlying allergen-induced pulmonary inflammation and asthma are hampered by the lack of appropriate physiological in vivo models that reflect the natural route of allergen exposure and sensitization. OBJECTIVE: To generate and phenotype a transgenic mouse strain expressing the T cell receptor (TCR) specific for an immunodominant domain of the major inhalant allergen Dermatophagoides pteronyssinus species of house dust mite (Der p 1), for the development of an in vivo model of allergic asthma. METHODS: Der p 1 transgenic mice were generated using TCR-alphabeta derived from a CD4+ T cell hybridoma reactive with Der p 1 residues p 110-131. The frequency and functional activity of peripheral T cells were determined and parameters of airway inflammation assessed following allergen challenge of the airways with Der p 1. RESULTS: CD4+ T cells are functionally active, exhibiting dose-dependent proliferation and IL-4 production on primary stimulation with Der p 1 or Der p 1, p 110-131 in vitro, independent of in vivo antigen priming. On sensitization of the airways with allergen, in the absence of systemic priming or the application of adjuvants, the TCR transgenic mice develop airway inflammation characterized by a marked lymphocytic and eosinophilic infiltrate with goblet cell hyperplasia and enhanced mucin production. CONCLUSION: The Der p 1 TCR transgenic mice provide a model for investigating the pathophysiological mechanisms of pulmonary inflammation following sensitization by exposure of the airways to allergen and for investigating the mode of action and efficacy of novel immunotherapeutics.

Deficient cytokine response of human allergen-specific T lymphocytes from humanized SCID mice and reconstitution by professional antigen-presenting cells.

BACKGROUND: Hu-PBL-SCID mice generated by the transfer of PBMCs from atopic individuals may provide a physiologic in vivo model for investigating human responses to allergens and potential approaches toward immunotherapy. OBJECTIVE: This study was undertaken to investigate the functional activity and cytokine profile of human allergen-reactive T lymphocytes isolated from hu-PBL-SCID mice. METHODS: PBMCs from allergic individuals were coinjected with allergen into SCID mice. Human lymphocyte migration and phenotype were established by reverse transcription-PCR and immunohistochemistry, IgE levels in sera were determined, and the frequency of allergen-reactive cytokine-producing T lymphocytes was established. RESULTS: After immunization with allergen, specific IgE levels in hu-PBL-SCID sera were comparable with levels in donor sera. Although the majority of lymphocytes remained in the peritoneum, significant numbers of T lymphocytes were located in the spleen, where human IL-4, IL-5, and IFN-gamma messenger RNA expression was detected after stimulation with PHA and phorbol myristate acetate. Failure to induce cytokine production by human T lymphocytes isolated from the peritoneum and spleen of hu-PBL-SCID mice by allergen was reversed by stimulating with allergen in the presence of exogenously added IL-2 and antigen-presenting cells (APC), particularly CD14(+) monocytes. Under these conditions, allergen-reactive T cells expressed a T(H)2-like phenotype. CONCLUSIONS: These data suggest that, after initial activation and induction of antibody production, human T lymphocytes enter a state of unresponsiveness, arising from a loss of human professional APC, in hu-PBL-SCID mice. The use of hu-PBL-SCID mouse models in studies on therapeutic approaches for allergy may benefit from the additional transfer of human professional APC.

Short-term preseasonal birch pollen allergoid immunotherapy influences symptoms, specific nasal provocation and cytokine levels in nasal secretions, but not peripheral T-cell responses, in patients with allergic rhinitis.

BACKGROUND: [corrected] Birch pollen allergic rhinitis can be sufficiently treated with specific subcutaneous allergoid immunotherapy (IT). However, little is known about the clinical and immunological effects of short-term therapy protocols. OBJECTIVE: To investigate the clinical efficacy of a birch pollen allergoid IT using seven preseasonal injections and to evaluate immunological parameters that might explain clinical findings. METHODS: Thirty-seven patients were included into the study and randomized to either a symptomatic treatment or allergoid IT plus symptomatic treatment. Patients were examined during the pre-IT season, at two extraseasonal visits both before and after IT and during the post-IT season. At each visit, nasal secretion samples were taken and analysed for levels of IL-4, IL-5 and IFNgamma. In addition, short-term birch-specific T-cell lines (TCLs) were cultured from peripheral blood mononuclear cells of 10 patients of the IT group, both before and after IT, and the ratios of lymphocyte subpopulations were determined. Cytokine production by TCLs (IL-4, IL-5, IFNgamma, IL-10) and proliferation of TCLs in response to stimulation with birch pollen allergen were measured. RESULTS: It was possible to evaluate 27 patients in accordance with the study protocol. Clinical symptoms and medication intake were reduced as a result of the IT as were nasal secretion levels of IL-5 (P = 0.007). IFNgamma was increased in nasal secretions (P = 0.01), while IL-4 was not measurable in most samples. No effect was found on proliferation of birch pollen-reactive TCLs, cytokine production by TCLs and the frequency and ratio of CD4+ and CD8bright or CD45RA+ and CD45RO+ cells in peripheral blood (all P > 0.05). Conclusion Preseasonal IT with a birch pollen allergoid is clinically effective in allergic rhinitis and influences cytokine production in the nose, but does not modulate the measured responses of peripheral blood T cells.

Inhibition of murine IgE and immediate cutaneous hypersensitivity responses to ovalbumin by the immunomodulatory agent leflunomide.

Leflunomide has been identified as an immunoregulatory and anti-inflammatory compound. Allergic disease is characterized by elevated serum IgE levels, production of allergen-specific IgE and the release of inflammatory mediators from mast cells and granulocytes. Here we demonstrate, using an in vivo murine model, the ability of leflunomide to down-regulate levels of total and allergen-specific serum IgE production. Mice receiving leflunomide (45 mg/kg) orally at the time of primary immunization with ovalbumin adsorbed to aluminium hydroxide adjuvant, showed a reduction in total serum IgE levels of 95%, 41% and 32% following primary, secondary and tertiary immunizations, respectively (P < 0.05). When leflunomide was administered both at the time of primary and subsequent immunizations, reductions in total and specific serum IgE levels of > 80% and > 38%, respectively, were observed (P < 0.05). Administration of leflunomide to mice which had already developed an IgE response resulted in reductions in total and specific serum IgE levels of > 80% and > 45%, respectively (P < 0.05). Following leflunomide treatment, animals failed to develop immediate cutaneous hypersensitivity responses when challenged intradermally with allergen. Down-regulation of immunoglobulin production was not restricted to IgE, since levels of allergen-specific IgG1 and IgG2a in serum were also reduced. The finding of significant reductions in total and allergen-specific IgM suggests that the mechanism of action does not involve selective inhibition of immunoglobulin class switching. A loss in production of the T helper cell-derived B cell differentiation factor IL-5 may account for the reduction in immunoglobulin levels. In adoptive transfer experiments leflunomide did not induce tolerance in allergen-reactive Th2 populations, contrary to animal disease models of transplantation and autoimmunity, where leflunomide was shown to induce tolerance in the effector T cell population.

Responses of human birch pollen allergen-reactive T cells to chemically modified allergens (allergoids).

BACKGROUND: Allergoids are widely used in specific immunotherapy for the treatment of IgE-mediated allergic diseases. OBJECTIVE: The aim of this study was to analyse whether a modification of birch pollen allergens with formaldehyde affects the availability of T-cell epitopes. METHODS: Efficient modification of the allergens was verified by determining IgE and IgG binding activity using ELISA inhibition tests. T-cell responses to birch pollen allergoids were analysed in polyclonal systems, using peripheral blood mononuclear cells (PBMC) of five birch pollen-allergic individuals, as well as birch pollen extract-reactive T-cell lines (TCL), established from the peripheral blood of 14 birch pollen-allergic donors. To determine whether the modification of natural (n)Bet v 1 with formaldehyde or maleic anhydride results in epitope-specific changes in T-cell reactivities, 22 Bet v 1-specific T-cell clones (TCC), established from nine additional birch pollen-allergic individuals, were tested for their reactivity with these products. RESULTS: The majority of PBMC and TCL showed a reduced response to the birch pollen extract allergoid. Bet v 1-specific TCC could be divided into allergoid-reactive and -non-reactive TCC. No simple correlation between possible modification sites of formaldehyde in the respective T-cell epitopes and the stimulatory potential of the allergoid was observed. Mechanisms of suppression or of anergy induction were excluded as an explanation for the non-reactivity of representative TCC. All TCC could be stimulated by maleylated and unmodified nBet v 1 to a similar extent. CONCLUSION: These results demonstrate differences in the availability of T-cell epitopes between allergoids and unmodified allergens, which are most likely due to structural changes within the allergen molecule.

Inhibition of human T-cell responses to house dust mite allergens by a T-cell receptor peptide.

Recent analysis of the usage of T-cell receptor (TcR) beta chain variable region (V beta) gene elements by house dust mite (HDM)-reactive T cells from an atopic donor suggested that TcR-V beta 3 gene products may form a major component of the human T-cell repertoire reactive to this common allergen. In this study a peptide analog of the TcR-V beta 3 complementarity determining region 2 (CDR2) is shown to inhibit the polyclonal human T-cell response to HDM; this effect is specific because inhibition is dependent on the presence of V beta 3 + T cells. This experimental approach has been used to determine whether the pattern seen in T-cell clones derived from one atopic donor reflects TcR-V beta usage in the polyclonal response to allergen in the general population. Inhibition of more than 50% of the polyclonal response to allergen by V beta 3-CDR2 peptide was observed in 16 of 21 donors tested, suggesting that TcR-V beta 3 gene usage may form a major component of the human HDM repertoire and as such offer a suitable target for T cell-directed specific immunotherapy in HDM-allergic individuals. Depletion of CD8+ T cells abolishes peptide-mediated inhibition of CD4+ T-cell proliferation to HDM, suggesting that induction of a CD8+ regulatory T-cell subset by the CDR2 peptide may modulate HDM-specific allergic T-cell responses.