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All investigated strains of S. The responsible protease, Ide Ssuis , is a highly protective antigen against S. The animals were immunized with either recombinant Ide Ssuis or placebo plac, adjuvant only intramuscularly with a booster immunization 2 weeks later Figure 6 A. Two weeks after the boost, PBMCs and serum samples were obtained from both groups.

Stimulation of PBMCs with ctr-ag also induced T helper cell reactivity in some of the animals but was not restricted to the immunized group. A successful vaccination requires activation of T helper cells capable of costimulating B cells for generating antibodies that protect against the disease of interest. Figure 6. A Piglets were primed with rIde Ssuis followed by booster immunization 2 weeks later. Blood samples for the investigation of antigen-specific Th cells were taken 2 week post-booster immunization. The frequency of antigen-specific Th cells was calculated as the difference of specified cells from antigen-restimulated and medium-cultivated PBMC, respectively.

However, with the current tools available at the moment, we could not directly prove SLAII-TCR interaction as demonstrated in mouse and human studies 27 , Regarding the percentage of C. In the second step, we amplified antigen-responsive cells by in vitro expansion to optimize and study time- and dose-dependent responses in more detail during autologous, antigen-specific restimulation.

CFSE-dilution analysis of the expansion culture allowed for a subgroup analysis to characterize cells that subsequently expressed CD upon restimulation.

Fundamental Immunology

Using expansion and restimulation, our results indicate that both superantigen-activated and fungus-induced CD expression follow a similar kinetic, starting between 2 and 4 h after antigenic stimulation. This is in line with kinetic studies with mouse 30 , 55 and human PBMCs Notably, in contrast to those studies we found no indication that CD expression peaked around 6 h after restimulation in expanded swine CD4 T cells.

But, 6 h after MoDC restimulation, we already found a substantial population of antigen-responsive cells that was sufficient for downstream phenotyping of fungus-reactive T cells and therefore avoided longer restimulation times.

It should be noted that restimulation times and antigen dosing have to be carefully adapted according to any pathogen, infection stage, or sample handling e. For priming MoDC with C. For a basic understanding and also for modulation or intervention of the pathogen-specific T cell response, characterizing the responding T cell repertoire is crucial. The detection of cytokines enables identification of functional subsets of reactive cells. Their data highlight the rareness of antigen-activated cytokine producers detectable in peripheral blood, never exceeding 0.

However, defining antigen specificity based on preselected cytokines may potentially underestimate the complete frequency and complexity of antigen-specific T cell responses. Our data on the T cell response of outbred pigs to C. However, when we amplified antigen-responsive T cells by expansion and analyzed the T cell response during antigen-primed MoDC restimulation, we were able to increase the resolution between background levels and antigen-induced cytokine producers.

One very important benefit of using a human-relevant large animal model is the possibility to easily dissect organ tissue to study immune cells that are directly in contact with a given pathogen. We therefore selected a parasitic infection that not only has a significant, economic impact in pig production but also is zoonotic and highly prevalent in human populations in tropical and subtropical areas with poor hygiene management—the roundworm Ascaris sp.

A major hallmark of Ascaris sp. A massive larval infiltration of the lung causes tissue destruction, pulmonary eosinophilia, and infiltration of inflammatory cells that can lead to acute pneumonia By analyzing the pathogen-specific T cell responses during experimental A. And third, comparing whole worm lysates with worm-derived excretory—secretory products as antigenic source, the latter exhibited a higher frequency of antigen-responding cells identified in affected lungs after trickle infection.

These findings could be explained by recalling that proteins released by live parasites are directly targeting host barrier and host immune cells contain highly immunogenic antigens and are known to potently interfere with every aspect of host immunity For the first time, we could identify antigen-reactive T helper cells by immunization with a recombinant antigen of S.

In previous S. But until now, data about T cell response in S.

Progress in Immunology Vol. VIII

Though not significant, we observed a putative T cell reactivity to this protein. Noteworthy, conserved T-cell epitopes especially in the C-terminal fibronectin-binding domain of SfbI were identified, recognized by three different mouse strains Therefore, we suggest a possible cross-reactivity with other fibronectin-binding proteins 65 expressed by bacteria on the mucosal surfaces of those piglets that show reactivity toward SfbI.

Thus, application of CD expression following antigen-specific stimulation as a marker for protein-specific T helper cells will provide new perspectives to investigate vaccination-responsive T cell immunity in pig. All animal studies were performed according to the principles outlined in the European Convention for the Protection of Vertebrate Animals used for Experimental and other Scientific Purposes.

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FE designed experiments, carried out experiments, evaluated data, and wrote the manuscript. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The authors further thank Prof. Alexander Scheffold and Dr. Petra Bacher for their valuable methodical advice.

Karoline Rieckmann and Anna Seydel are acknowledged for their work with the piglets in the rIde Ssuis vaccination experiment.

Original Research ARTICLE

Lunney JK. Advances in swine biomedical model genomics. Int J Biol Sci — Nutritional programming of gastrointestinal tract development. Is the pig a good model for man? Nutr Res Rev — The pig: a model for human infectious diseases. Trends Microbiol —7. The evolutionary basis for differences between the immune systems of man, mouse, pig and ruminants. Vet Immunol Immunopathol —9. Anat Embryol Berl — Dev Comp Immunol — The porcine innate immune system: an update. Mol Immunol — Influenza A virus infection in pigs attracts multifunctional and cross-reactive T cells to the lung.

J Virol — Detection of Foxp3 protein expression in porcine T lymphocytes. Vet Immunol Immunopathol — Natural and inducible Tregs in swine: Helios expression and functional properties. A novel lineage transcription factor based analysis reveals differences in T helper cell subpopulation development in infected and intrauterine growth restricted IUGR piglets.

Porcine CD identification, expression and functional aspects in lymphocyte subsets in swine. CD27 expression discriminates porcine T helper cells with functionally distinct properties. Vet Res Opportunities for bead-based multiplex assays in veterinary diagnostic laboratories. J Vet Diagn Invest — Structural and functional annotation of the porcine immunome. BMC Genomics T-cell quality in memory and protection: implications for vaccine design. Nat Rev Immunol — T cell fate at the single-cell level. Annu Rev Immunol — Antigen-specific T cells and cytokines detection as useful tool for understanding immunity against zoonotic infections.

Clin Dev Immunol Bacher P, Scheffold A.

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Flow-cytometric analysis of rare antigen-specific T cells: flow-cytometric analysis of rare antigen-specific T cells. Cytometry A 83A— J Exp Med — Antigen-reactive T cell enrichment for direct, high-resolution analysis of the human naive and memory Th cell repertoire. J Immunol — J Immunol Methods — New technologies for monitoring human antigen-specific T cells and regulatory T cells by flow-cytometry.

Curr Opin Pharmacol — T cell responses specific for subregions of allogeneic MHC molecules. Peptide-independent recognition by alloreactive cytotoxic T lymphocytes CTL. Crystal structure of a T cell receptor bound to an allogeneic MHC molecule. Nat Immunol ; 1: - Self peptide requirement for class II major histocompatibility complex allorecognition.

Alloreactive T cells discriminate among a diverse set of endogenous peptides. On the nature of peptides involved in T cell alloreactivity. Alloreactive cytotoxic T lymphocytes generated in the presence of viral-derived peptides show exquisite peptide and MHC specificity. J Immunol ; 1 - Alloreactive cytotoxic T lymphocytes focus on specific major histocompatibility complex-bound peptides.

The role of peptides in T cell alloreactivity is determined by self-major histocompatibility complex molecules. J Exp Med ; 41 - Direct recognition of foreign MHC determinants by naive T cells mobilizes specific V b families without skewing of the complementarity-determining region 3 length distribution. Shapes of MHC restriction. Immunity ; 1 - 7.

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Responses against single histocompatibility antigens. Analysis of cloned helper T cells. Tekolf WA, Shaw S. Primary in vitro generation of cytotoxic cells specific for human minor histocompatibility antigens between HLA-identical siblings. Induction of minor alloantigen-specific T cell subsets in vivo: recognition of processed antigen by helper but not by cytotoxic T cell precursors.