A trematode parasite derived growth factor binds and exerts influences on host immune functions via host cytokine receptor complexes

The trematode Fasciola hepatica is responsible for chronic zoonotic infection globally. Despite causing a potent T-helper 2 response, it is believed that potent immunomodulation is responsible for rendering this host reactive non-protective host response thereby allow- ing the parasite to remain long-lived. We have previously identified a growth factor, FhTLM, belonging to the TGF superfamily can have developmental effects on the parasite. Herein we demonstrate that FhTLM can exert influence over host immune functions in a host receptor specific fashion. FhTLM can bind to receptor members of the Transforming Growth Factor (TGF) superfamily, with a greater affinity for TGF-β RII. Upon ligation FhTLM initiates the Smad2/3 pathway resulting in phenotypic changes in both fibroblasts and macrophages. The formation of fibroblast CFUs is reduced when cells are cultured with FhTLM, as a result of TGF-β RI kinase activity. In parallel the wound closure response of fibroblasts is also delayed in the presence of FhTLM. When stimulated with FhTLM blood monocyte derived macrophages adopt an alternative or regulatory phenotype. They express high levels interleukin (IL)-10 and arginase-1 while displaying low levels of IL-12 and nitric oxide. Moreover they also undergo significant upregulation of the inhibitory recep- tor PD-L1 and the mannose receptor. Use of RNAi demonstrates that this effect is depen- dent on TGF-β RII and mRNA knock-down leads to a loss of IL-10 and PD-L1. Finally, we demonstrate that FhTLM aids newly excysted juveniles (NEJs) in their evasion of antibody- dependent cell cytotoxicity (ADCC) by reducing the NO response of macrophages—again dependent on TGF-β RI kinase. FhTLM displays restricted expression to the F. hepatica gut resident NEJ stages. The altered fibroblast responses would suggest a role for damp- ened tissue repair responses in facilitating parasite migration. Furthermore, the adoption of a regulatory macrophage phenotype would allow for a reduced effector response targetingjuvenile parasites which we demonstrate extends to an abrogation of the ADCC response. Thus suggesting that FhTLM is a stage specific evasion molecule that utilises host cytokine receptors. These findings are the first to clearly demonstrate the interaction of a helminth cytokine with a host receptor complex resulting in immune modifications that facilitate the non-protective chronic immune response which is characteristic of F. hepatica infection

A mouse model to study immunity against pseudorabies virus infection: significance of CD4+ and CD8+ cells in protective immunity

In this study we firstly established a vaccination/challenge model to study pseudorabies virus infection in mice. The mouse model was used to investigate the significance of CD4 and CD8 cells and of IFN production in protective immunity. Functional depletion of CD4 and CD8 and IFN was obtained in vivo by intraperitoneal injection of alginate-encapsulated anti-CD4, -CD8 or -IFN producing hybridoma's before and at the moment of vaccination. The observed protective immunity was correlated with underlying immunologic responses such as PRV-specific DTH reactivity, lymphoproliferation and cytotoxicity. The significance of CD4 and CD8 cells and of IFN production was also investigated for these immunological responses by the same in vivo depletion technique. The results demonstrated that protective vaccination of mice, that could be induced by immunization with 107 plaque forming units of the avirulent PRV mutant NIA3 TK−, was characterized by a typical anti-viral Th1 type immune response. A clear PRV-specific, CD4-dependent DTH reactivity and a classical CD8-dependent, MHC-restricted cytotoxicity was induced after protective immunization and the humoral immune response had a bias towards PRV-specific IgG2a formation. In vivo treatment with anti-CD8 and anti-IFN demonstrated that the cytotoxic response and humoral IgG2a response, respectively, were strongly reduced, whereas protection against lethal challenge was unaffected. On the other hand anti-CD4 treatment reduced the induced protection so that 30 f the mice died after lethal challenge. The results of our study demonstrated that CD4 , DTH like effector cells are a crucial effector mechanism for protective immunity against PRV

Carbohydrate-based particles: a new adjuvant for allergen-specific immunotherapy

The occurrence of systemic anaphylactic side-effects in the course of allergen-specific immunotherapy has been strongly reduced by the adsorption of allergens to aluminium hydroxide, the most frequently used adjuvant in humans. Using the major timothy grass pollen allergen, Phl p 5b, in its recombinant form for immunization of mice, we demonstrate that carbohydrate-based particles (CBP) exhibit several potential advantages over aluminium-hydroxide as adjuvant for immunotherapy. Similar to alum-bound rPhl p 5b, CBP-bound rPhl p 5b induced a stronger antibody and cytokine response than unbound rPhl p 5b after subcutaneous injection in mice. The antibodies induced by CBP-bound rPhl p 5b, exhibited potentially beneficial activities as they cross-reacted with group 5 allergens from five other grass species and inhibited the binding of grass pollen allergic patients IgE to Phl p 5b. Alum-bound rPhl p 5b induced a preferential allergen-specific Th2-response characterized by high immunoglobulin G1 (IgG1) antibody levels and elevated interleukin (IL)-4 and IL-5 production in cultured splenocytes. By contrast, CBP-bound rPhl p 5b, but not rPhl p 5b alone or coadministered with CBP, induced a mixed allergen-specific T helper 1 (Th1)/Th2 immune response characterized by the additional production of allergen-specific IgG2a/b antibody responses and elevated interferon-γ production. Conjugation of rPhl p 5b to CBP yielded a stable vaccine formulation with preserved immunogenic features of the allergen and, in contrast to alum, induced no granulomatous tissue reactions. Based on these results, CBP is suggested as a potentially useful adjuvant for specific immunotherapy of IgE-mediated allergies