Rapid Sequestration of Leishmania mexicana by Neutrophils Contributes to the Development of Chronic Lesion.

The protozoan Leishmania mexicana parasite causes chronic non-healing cutaneous lesions in humans and mice with poor parasite control. The mechanisms preventing the development of a protective immune response against this parasite are unclear. Here we provide data demonstrating that parasite sequestration by neutrophils is responsible for disease progression in mice. Within hours of infection L. mexicana induced the local recruitment of neutrophils, which ingested parasites and formed extracellular traps without markedly impairing parasite survival. We further showed that the L. mexicana-induced recruitment of neutrophils impaired the early recruitment of dendritic cells at the site of infection as observed by intravital 2-photon microscopy and flow cytometry analysis. Indeed, infection of neutropenic Genista mice and of mice depleted of neutrophils at the onset of infection demonstrated a prominent role for neutrophils in this process. Furthermore, an increase in monocyte-derived dendritic cells was also observed in draining lymph nodes of neutropenic mice, correlating with subsequent increased frequency of IFNγ-secreting T helper cells, and better parasite control leading ultimately to complete healing of the lesion. Altogether, these findings show that L. mexicana exploits neutrophils to block the induction of a protective immune response and impairs the control of lesion development. Our data thus demonstrate an unanticipated negative role for these innate immune cells in host defense, suggesting that in certain forms of cutaneous leishmaniasis, regulating neutrophil recruitment could be a strategy to promote lesion healing.

Polyethylenimine-based polyplex delivery of self-replicating RNA vaccines

Self-amplifying replicon RNA (RepRNA) are large molecules (12-14kb); their self-replication amplifies mRNA template numbers, affording several rounds of antigen production, effectively increasing vaccine antigen payloads. Their sensitivity to RNase-sensitivity and inefficient uptake by dendritic cells (DCs) - absolute requirements for vaccine design - were tackled by condensing RepRNA into synthetic, nanoparticulate, polyethylenimine (PEI)-polyplex delivery vehicles. Polyplex-delivery formulations for small RNA molecules cannot be transferred to RepRNA due to its greater size and complexity; the N:P charge ratio and impact of RepRNA folding would influence polyplex condensation, post-delivery decompaction and the cytosolic release essential for RepRNA translation. Polyplex-formulations proved successful for delivery of RepRNA encoding influenza virus hemagglutinin and nucleocapsid to DCs. Cytosolic translocation was facilitated, leading to RepRNA translation. This efficacy was confirmed in vivo, inducing both humoral and cellular immune responses. Accordingly, this paper describes the first PEI-polyplexes providing efficient delivery of the complex and large, self-amplifying RepRNA vaccines.

Is The Allergen Really Needed in Allergy Immunotherapy?

Immunotherapy for type I allergies is well established and is regarded to be the most efficient treatment option besides allergen avoidance. As of today, different forms of allergen preparations are used in this regard, as well as different routes of application. Virus-like particles (VLPs) represent a potent vaccine platform with proven immunogenicity and clinical efficacy. The addition of toll-like receptor ligands and/or depot-forming adjuvants further enhances activation of innate as well as adaptive immune responses. CpG motifs represent intensively investigated and potent direct stimulators of plasmacytoid dendritic cells and B cells, while T cell responses are enhanced indirectly through increased antigen presentation and cytokine release. This article will focus on the function of VLPs loaded with DNA rich in nonmethylated CG motifs (CpGs) and the clinical experience gained in the treatment of allergic rhinitis, demonstrating clinical efficacy also if administered without allergens. Several published studies have demonstrated a beneficial impact on allergic symptoms by treatment with CpG-loaded VLPs. Subcutaneous injection of VLPs loaded with CpGs was tested with or without the adjuvant alum in the presence or absence of an allergen. The results encourage further investigation of VLPs and CpG motifs in immunotherapy, either as a stand-alone product or as adjuvants for allergen-specific immunotherapy.

Immune Responses Against Classical Swine Fever Virus: Between Ignorance and Lunacy.

Classical swine fever virus infection of pigs causes disease courses from life-threatening to asymptomatic, depending on the virulence of the virus strain and the immunocompetence of the host. The virus targets immune cells, which are central in orchestrating innate and adaptive immune responses such as macrophages and conventional and plasmacytoid dendritic cells. Here, we review current knowledge and concepts aiming to explain the immunopathogenesis of the disease at both the host and the cellular level. We propose that the interferon type I system and in particular the interaction of the virus with plasmacytoid dendritic cells and macrophages is crucial to understand elements governing the induction of protective rather than pathogenic immune responses. The review also concludes that despite the knowledge available many aspects of classical swine fever immunopathogenesis are still puzzling.

A frequent hypofunctional IRAK2 variant is associated with reduced spontaneous hepatitis C virus clearance.

UNLABELLED Patients carrying very rare loss-of-function mutations in interleukin-1 receptor-associated kinase 4 (IRAK4), a critical signaling mediator in Toll-like receptor signaling, are severely immunodeficient, highlighting the paramount role of IRAK kinases in innate immunity. We discovered a comparatively frequent coding variant of the enigmatic human IRAK2, L392V (rs3844283), which is found homozygously in ∼15% of Caucasians, to be associated with a reduced ability to induce interferon-alpha in primary human plasmacytoid dendritic cells in response to hepatitis C virus (HCV). Cytokine production in response to purified Toll-like receptor agonists was also impaired. Additionally, rs3844283 was epidemiologically associated with a chronic course of HCV infection in two independent HCV cohorts and emerged as an independent predictor of chronic HCV disease. Mechanistically, IRAK2 L392V showed intact binding to, but impaired ubiquitination of, tumor necrosis factor receptor-associated factor 6, a vital step in signal transduction. CONCLUSION Our study highlights IRAK2 and its genetic variants as critical factors and potentially novel biomarkers for human antiviral innate immunity.

The Gas6-Axl Interaction Mediates Endothelial Uptake of Platelet Microparticles.

Upon activation, platelets release plasma-membrane derived microparticles (PMPs) exposing phosphatidylserine (PS) on their surface. The function and clearance mechanism of these MPs are incompletely understood. As they are pro-coagulant and potentially pro-inflammatory, rapid clearance from the circulation is essential for prevention of thrombotic diseases. The tyrosine kinase receptors Tyro3, Axl and Mer (TAMs) and their ligands protein S and Gas6 are involved in the uptake of PS-exposing apoptotic cells in macrophages and dendritic cells. Both TAMs and their ligands are expressed in the vasculature, the functional significance of which is poorly understood. In this study we investigated how vascular TAMs and their ligands may mediate endothelial uptake of PMPs. PMPs, generated from purified human platelets, were isolated by ultracentrifugation and labeled with biotin or PKH67. The uptake of labeled MPs in the presence of protein S and Gas6 in human aortic endothelial cells (HAEC) and human umbilical vein endothelial cells (HUVEC) was monitored by flow cytometry, western blotting and confocal/electron microscopy. We found that both endothelial cell types can phagocytose PMPs, and using TAM-blocking antibodies or siRNA knock-down of individual TAMs we show that the uptake is mediated by endothelial Axl and Gas6. As circulating PMPs-levels were not altered in Gas6-/- mice compared to Gas6+/+ mice, we hypothesize that the Gas6-mediated uptake is not a means to clear the bulk of circulating PMPs but may serve to phagocytose PMPs locally generated at sites of platelet activation and as a way to affect endothelial responses.