Our results indicate that BP-SIT may induce Bet v 1-specific IgG4 antibodies that cross-react with related food allergens and inhibit IgE binding by epitope competition.

Accumulating evidence shows that IL-17 is critically involved in diverse autoimmune diseases. However, its effect on the induction and progression of the humoral immune response is not fully understood. Using a preclinical model of IL-17-mediated dry eye disease (DED), we demonstrate that upon encountering both the B cell receptor and a secondary T cell signal, IL-17 can enhance B cell proliferation and germinal center formation in DED mice, suggesting that a stable antigen-dependent T-B cell interaction is required. In addition, IL-17 also promotes the differentiation of B cells into isotype-switched B cells and plasma cells. Furthermore, we show that Th17 cells are more effective than Th1 cells to provide B cell help. Reduced B cell response correlates with significant reduction in clinical disease after in vivo IL-17A neutralization. In conclusion, our findings demonstrate a new role of IL-17 in promoting autoimmunity in part through directly enhancing B cell proliferation, differentiation and plasma cell generation.

Background Corneal neovascularization increases the risk of T cell mediated allograft rejection. Here, we investigate whether T cells promote angiogenesis in transplantation. Methods Conventional effector T cells were collected from draining lymph nodes (DLNs) of allogeneic or syngeneic corneal transplanted BALB/c mice. T cells were either cocultured with vascular endothelial cells (VECs) to assess VEC proliferation or used in a Mixed Lymphocyte Reaction assay. mRNA expression of vascular endothelial growth factor (VEGF)-A, -C, and VEGF receptor 2 (VEGF-R2) in VECs was assessed by real-time PCR. VEGF-A protein expression was determined by ELISA. Flow cytometry was used to analyze VEGF-R2 expression in corneal CD31+ cells, and VEGF-A and IFNγ expression in corneal CD4+ T cells. Results Allogeneic T cells from high-risk (HR) grafted mice induced more VEC proliferation than those from syngeneic transplant recipients (p=0.03). VEGF-A mRNA and protein expression were higher in T cells from DLNs (p=0.03 and p=0.04, respectively) and cornea (protein; p=0.04) of HR compared to low-risk (LR) grafted hosts. VEGF-A, VEGF-C and VEGF-R2 mRNA expression were increased in VECs when cocultured with T cells from HR transplants compared to LR transplants and naïve mice. In addition, IFNγ blockade in T cell/VEC coculture increased VEC proliferation and VEGF-A protein expression, whereas blocking VEGF-A significantly reduced VEC proliferation (p=0.04). Conclusions Allogeneic T cells from corneal transplant hosts promote VEC proliferation, probably via VEGF-A signaling, while IFNγ shows an antiangiogenic effect. Our data suggest that T cells are critical mediators of angiogenesis in transplantation.

Summary Background Allergen‐specific immunotherapy (AIT) with birch pollen generates Bet v 1‐specific immunoglobulin (Ig)G4 which blocks IgE‐mediated hypersensitivity mechanisms. Whether IgG4 specific for Bet v 1a competes with IgE for identical epitopes or whether novel epitope specificities of IgG4 antibodies are developed is under debate. Objective We sought to analyze the epitope specificities of IgE and IgG4 antibodies from sera of patients who received AIT. Methods 15 sera of patients (13/15 received AIT) with Bet v 1a‐specific IgE and IgG4 were analyzed. The structural arrangements of recombinant (r)Bet v 1a and rBet v 1a_11x, modified in five potential epitopes, were analyzed by circular dichroism and nuclear magnetic resonance spectroscopy. IgE binding to Bet v 1 was assessed by ELISA and mediator release assays. Competitive binding of monoclonal antibodies specific for Bet v 1a and serum IgE/IgG4 to rBet v 1a and serum antibody binding to a non‐allergenic Bet v 1‐type model protein presenting an individual epitope for IgE was analyzed in ELISA and western blot. Results rBet v 1a_11x had a Bet v 1a – similar secondary and tertiary structure. Monomeric dispersion of rBet v 1a_11x was concentration and buffer‐dependent. Up to 1500‐fold increase in the EC50 for IgE‐mediated mediator release induced by rBet v 1a_11x was determined. The reduction of IgE and IgG4 binding to rBet v 1a_11x was comparable in 67% (10/15) of sera. Bet v 1a‐specific monoclonal antibodies inhibited binding of serum IgE and IgG4 to 66.1% and 64.9%, respectively. Serum IgE and IgG4 bound specifically to an individual epitope presented by our model protein in 33% (5/15) of sera. Conclusion and Clinical Relevance Patients receiving AIT develop Bet v 1a‐specific IgG4 which competes with IgE for partly identical or largely overlapping epitopes. The similarities of epitopes for IgE and IgG4 might stimulate the development of epitope‐specific diagnostics and therapeutics.

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Birch pollen (BP)-allergic patients sensitized to the major BP allergen, Bet v 1, often show IgE-cross-reactivity with Mal d 1 in apple and Cor a 1 in hazelnut. We recently found that specific immunotherapy (SIT) with BP induces high titers of Bet v 1-specific IgG4. In parallel, many BP-SIT-treated patients developed food-reactive IgG4. However, around 25% of patients showed no detectable food-specific IgG4 response. The diversity of epitope recognition of IgG4 antibodies (abs) in BP-SIT-treated patients developing or lacking food-reactive IgG4 were compared. Sera containing Bet v 1-, Mal d 1- and Cor a 1-specific IgE and Bet v 1-specific IgG4 with and without food-reactive IgG4 abs were included. Blocking capacity of the sera was analysed in facilitated antigen-binding and basophil activation assays. Potential epitopes recognized by allergen-specific IgE and IgG4 abs were screened by competitive immunoscreening with a random phage-displayed 12-mer peptide library. Patients showing food reactive-IgG4 with blocking capacity recognized a higher number of putative epitopes distributed over the surface of allergens. In contrast, a restricted number of putative epitopes was recognized by sera from patients who had failed to develop food-reactive IgG4. The induction of polyclonal IgG4 responses during BP-SIT may be a prerequisite for the development of abs that cross-react with Bet v 1-related food allergens and consequently, block food-reactive IgE abs.