Abstract
(Deutsch)
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Harnessing interferon-lambda as a mucosal adjuvant of the respiratory tract in pigsBackground: Interferon-? (IFN-?) is an antiviral cytokine, which is secreted by a variety of cell types in response to viral infection. IFN-? acts on a heterodimeric cell surface receptor consisting of the ubiquitously expressed IL-10 receptor ß-chain and a second IFN-?-specific component designated IFNLR1, which is primarily expressed by epithelial cells. IFN-? induces an antiviral response in epithelial tissues and protects against infection with viruses that preferentially replicate in the mucosal tissues of the respiratory or gastrointestinal tract. The role of IFN-? in adaptive immunity is less well understood, but recent work with wild-type and IFNLR1-/- mice suggests that IFN-? enhances mucosal immunity by boosting the release of thymic stromal lymphopoietin (TSLP), which in turn acts on migratory dendritic cells (DCs). Nasal immunisation of mice with influenza subunit vaccines along with either recombinant murine IFN-?2 or murine TSLP resulted in enhanced IgA and IgG1 antibody production, whereas subcutaneous or intraperitoneal routes of immunization did not cause such effects. Mucosal immunization improved survival of mice following lethal challenge infection with influenza A virus and also reduced influenza A virus transmission to contact animals, suggesting that the adjuvant activity of IFN-? might be employed to improve the efficacy of mucosal vaccines in general. Goal of the study: Type III IFNs are found in all mammalian species and play important role in the innate immune response of epithelial tissues. However, it is not known whether the ability of IFN-? to stimulate adaptive mucosal immunity, which has recently been observed in the mouse, would also be seen with other mammalian species including humans. This project aims at analyzing adjuvant activity of porcine IFN-? with respect to stimulating mucosal adaptive immunity to mucosal vaccination of pigs with influenza virus antigen. Specific aims: (1) Produce recombinant, biologically active porcine IFN-? and TSLP; (2) Analyze the induction of IFN-? of porcine respiratory epithelial cells grown in air-liquid interface (ALI) cultures by single-cycle vesicular stomatitis virus (VSV) lacking host shut-off activity (VSVMq?G) or encoding porcine IFN-? (VSV?G-IFN-?); (3) Analyze mucosal IgA and IgG responses following mucosal vs. intramuscular immunisation of pigs with: (a) Inactivated influenza vaccine supplemented with either pIFN-?, TSLP, VSVMq?G or VSV?G-IFN-?, (b) Single-cycle VSV vector encoding either porcine influenza HA antigen (VSV?G-HA), HA antigen and IFN-? (VSV?G-HA, IFN-?), IFN-inducing vector (VSVMq?G-HA), (c) Live attenuated NS1-deficient porcine influenza viruses; (4) Analyze replication of homologous or heterologous porcine influenza A viruses in the respiratory tract of vaccinated pigs; (5) Investigate effects of vaccination on virus transmission to sentinel animals.Methods: Generation and purification of recombinant porcine IFN-? and TSLP; generation of recombinant VSV vectors; analysis of antigen expression (Western blot, flow cytometry); experimental vaccination and infection of pigs (BSL-2); RT-qPCR; detection of mucosal IgA and IgG antibodies (ELISA); virus titration; virus neutralisation test; ALI cultures.Expected results: This project will show whether recombinant porcine IFN-? and TSLP will enhance porcine adaptive mucosal immunity in response to vaccination with influenza antigen. In addition, this project will provide proof of principle for IFN-? adjuvant activity if the cytokine is induced or directly expressed by viral vectors that are delivered via the mucosal route. Finally, we will provide experimental evidence for the power of IFN-? adjuvant in terms of mucosal immunity-mediated inhibition of influenza virus transmission in a relevant host. Importance: Mucosal adaptive immunity may efficiently prevent infection of respiratory and gastrointestinal epithelial tissues, the major entry sites for the majority of pathogens. However, induction of mucosal adaptive immunity by vaccination is normally low and short-lived. Therefore, the development of adjuvants that are able to strongly trigger mucosal adaptive immune responses is of high relevance in veterinary as well as human medicine.
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