Immune responses after immunization with plasmid DNA encoding Bet v 1, the major allergen of birch pollen (original) (raw)
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Journal of allergy and …, 2003
Background: In mouse, IgG1 has been reported to make up 2 functionally distinct phenotypes that also differ in their induction requirements. One of these phenotypes lacks anaphylactic activity. Objective: We hypothesized that nonanaphylactic IgG1 could modulate allergic reactions and investigated whether such antibodies are induced by DNA immunization. Methods: Mice were immunized with allergen-encoding plasmid DNA or with recombinant allergens and alum. Sera were analyzed for IgG subclasses by ELISA for anaphylactic IgE by rat basophil degranulation, and after heat inactivation of IgE for anaphylactic IgG by passive cutaneous anaphylaxis assay. IFN-γ and IL-5 from in-vitro restimulated spleen cells were quantitated by ELISA. Results: After protein immunization, mice produced IgG1 and IgE, whereas DNA immunization elicited IgG1 and IgG2a but no IgE. However, all sera were positive for non-IgE-mediated passive cutaneous anaphylaxis. In the presence of anaphylactic IgG1, the additional occurrence of nonanaphylactic IgG1 cannot be strictly ruled out. To circumvent this problem, we immunized IL-4 receptor-deficient mice against Bet v 1a, because anaphylactic but not nonanaphylactic IgG1 has been reported to depend on IL-4. These animals produced only low amounts of IgG1, but sera were again positive for non-IgE-mediated anaphylactic activity. Conclusions: Our results revealed no evidence for the production of nonanaphylactic IgG1. Furthermore, our data indicate that the development of non-IgE-mediated anaphylaxis does not require IL-4 receptor signaling. (J Allergy Clin Immunol 2003;112:579-84.)
Proceedings of the National Academy of Sciences of the United States of America, 1996
We compared the antigen-specific antibody isotypes and lymphokine secretion by CD4+ T cells in BALB/c mice immunized intradermally with either Escherichia coli 13-galactosidase (13-gal) or plasmid DNA (pDNA) encoding ,8-gal in a cytomegalovirus-based expression vector (pCMV-LacZ). pCMV-LacZ induced mainly IgG2a, whereas 13-gal in saline or alum induced IgGI and IgE 1-gal-specific antibodies. In addition, splenic CD4+ T helper (Th) cells isolated from pDNA-immunized mice secreted interferon-y but not interleukin (IL)-4 and IL-5, whereas Th cells from 13-gal-injected mice secreted IL-4 and IL-5 but not interferon-y after in vitro stimulation with antigen. Together these data demonstrate that pDNA immunization induced a T helper type 1 (Thl) response, whereas protein immunization induced a T helper type 2 (Th2) response to the same antigen. Interestingly, priming of mice with pCMV-LacZ prevented IgE antibody formation to a subsequent i.p. 13-gal in alum injection. This effect was antigen-specific, because priming with pCMV-LacZ did not inhibit IgE anti-ovalbumin antibody formation. Most importantly, intradermal immunization with pCMV-LacZ (but not pCMV-OVA) of 13-gal in alum-primed mice caused a 66-75% reduction of the IgE anti-,B-gal titer in 6 weeks. Also, pCMV-LacZ induced specific IgG2a antibody titers and interferon-y secretion by Th cells in the 13-gal in alum-primed mice. The data demonstrate that gene immunization induces a Th, response that dominates over an ongoing proteininduced Th2 response in an antigen-specific manner. This suggests that immunization with pDNA encoding for allergens may provide a novel type of immunotherapy for allergic diseases.
International Immunopharmacology, 2007
Immunotherapy using T-cell epitope peptides or CpG DNA conjugated with allergenic protein is useful, although the mechanisms of these therapies differ. However, the combination of CpG DNA and peptide, but not protein, had not been documented. Therefore, we investigated CpG DNA conjugated with peptide to obtain positive synergistic effects. In the first experiment, mice were vaccinated with a conjugate of CpG DNA and Cry j 2 T-cell epitope peptide p246-259 (CpG-peptide); a mixture of CpG DNA and peptide (CpG+peptide); peptide alone, or PBS alone, and immunized with Cry j 2. In the second experiment, mice were immunized with Cry j 2 and injected with CpG-peptide, CpG+peptide, peptide only, or PBS only. In both experiments, Cry j 2-specific IgE, IL-4, and IL-5 were significantly lower in mice given CpG-peptide, versus those given CpG+peptide, peptide alone, or PBS alone. However, IgG2a, IgG2b and IFN-γ did not increase in mice injected with CpG-peptide. In the third experiment, CpG-peptide significantly attenuated nasal symptoms (sneezing and nasal rubbing) compared to CpG+peptide, peptide alone, or PBS alone. Mice were also injected with a conjugate of CpG DNA and Cry j 2 protein (CpG-Cry j 2) or CpG-peptide to compare prime responses. Mice vaccinated with CpG-Cry j 2 generated Cry j 2-specific IgG1, whereas those vaccinated with CpG-peptide did not produce IgG1. This study demonstrated, for the first time, that immunotherapy with CpG DNA conjugated with a T-cell peptide is useful in preventing and treating allergic conditions.
The Journal of …, 1996
We dissected the T cell activation potency and the immunoglobulin (Ig) E-binding properties (allergenicity) of nine isoforms of Bet v 1 (Bet v 1a-Bet v 11), the major birch pollen allergen. Immunoblot experiments showed that Bet v 1 isoforms differ in their ability to bind IgE from birch pollen-allergic patients. All patients tested displayed similar IgE-binding patterns toward each particular isoform. Based on these experiments, we grouped Bet v 1 isoforms in three classes: molecules with high IgE-binding activity (isoforms a, e, and j), intermediate IgE-binding (isoforms b, c, and 0, and low/no IgE-binding activity (isoforms d, g, and 1). Bet v la, a recombinant isoform selected from a cDNA expression library using IgE immunoscreening, exhibited the highest IgE-binding activity. Isoforms a, b, d, e, and 1 were chosen as representatives from the three classes for experimentation. The potency of each isoallergen to activate T lymphocytes from birch pollen-allergic patients was assayed using peripheral blood mononuclear cells, allergen-specific T cell lines, and peptide-mapped allergen-specific T cell clones. Among the patients, some displayed a broad range of T cell-recognition patterns for Bet v 1 isoforms whereas others seemed to be restricted to particular isoforms. In spite of this variability, the highest scores for T cell proliferative responses were observed with isoform d (low IgE binder), followed by b, 1, e, and a. In vivo (skin prick) tests showed that the potency of isoforms d and 1 to induce typical urticarial type I reactions in Bet v 1-allergic individuals was significantly lower than for isoforms a, b, and e. Taken together, our results indicate that hypoallergenic Bet v 1 isoforms are potent activators of allergen-specific T lymphocytes, and Bet v 1 isoforms with high in vitro IgE-binding activity and in vivo allergenicity can display low T cell antigenicity. Based on these findings, we propose a novel approach for immunotherapy of type I allergies: a treatment with high doses of hypoallergenic isoforms or recombinant variants ofatopic allergens. We proceed on the assumption that this measure would modulate the quality of the T helper cell response to allergens in vivo. The therapy form would additionally implicate a reduced risk ofanaphylactic side effects. J. Exp.