Protective Antiviral Immunity Conferred by a Nonintegrative Lentiviral Vector-Based Vaccine

Lentiviral vectors are under intense scrutiny as unique candidate viral vector vaccines against tumor and aggressive pathogens because of their ability to initiate potent and durable specific immune responses. Strategies that alleviate safety concerns will facilitate the clinical developments involving lentiviral vectors. In this respect, the development of integration deficient lentiviral vectors circumvents the safety concerns relative to insertional mutagenesis and might pave the way for clinical applications in which gene transfer is targeted to non-dividing cells. We thus evaluated the potential use of nonintegrative lentiviral vectors as vaccination tools since the main targeted cell in vaccination procedures is the non-dividing dendritic cell (DC). In this study, we demonstrated that a single administration of nonintegrative vectors encoding a secreted form of the envelope of a virulent strain of West Nile Virus (WNV) induces a robust B cell response. Remarkably, nonintegrative lentiviral vectors fully protected mice from a challenge with a lethal dose of WNV and a single immunization was sufficient to induce early and long-lasting protective immunity. Thus, nonintegrative lentiviral vectors might represent a safe and efficacious vaccination platform for the development of prophylactic vaccines against infectious agents

The Immune Cellular Effectors of Terrestrial Isopod Armadillidium vulgare: Meeting with Their Invaders, Wolbachia

Most of crustacean immune responses are well described for the aquatic forms whereas almost nothing is known for the isopods that evolved a terrestrial lifestyle. The latter are also infected at a high prevalence with Wolbachia, an endosymbiotic bacterium which affects the host immune system, possibly to improve its transmission. In contrast with insect models, the isopod Armadillidium vulgare is known to harbor Wolbachia inside the haemocytes.In A. vulgare we characterized three haemocyte types (TEM, flow cytometry): the hyaline and semi-granular haemocytes were phagocytes, while semi-granular and granular haemocytes performed encapsulation. They were produced in the haematopoietic organs, from central stem cells, maturing as they moved toward the edge (TEM). In infected individuals, live Wolbachia (FISH) colonized 38% of the haemocytes but with low, variable densities (6.45±0.46 Wolbachia on average). So far they were not found in hyaline haemocytes (TEM). The haematopoietic organs contained 7.6±0.7×10(3)Wolbachia, both in stem cells and differentiating cells (FISH). While infected and uninfected one-year-old individuals had the same haemocyte density, in infected animals the proportion of granular haemocytes in particular decreased by one third (flow cytometry, Pearson's test = 12 822.98, df = 2, p<0.001).The characteristics of the isopod immune system fell within the range of those known from aquatic crustaceans. The colonization of the haemocytes by Wolbachia seemed to stand from the haematopoietic organs, which may act as a reservoir to discharge Wolbachia in the haemolymph, a known route for horizontal transfer. Wolbachia infection did not affect the haemocyte density, but the quantity of granular haemocytes decreased by one third. This may account for the reduced prophenoloxidase activity observed previously in these animals

Shaping of Monocyte-Derived Dendritic Cell Development and Function by Environmental Factors in Rheumatoid Arthritis

Dendritic cells (DC) are heterogeneous cell populations essential for both inducing immunity and maintaining immune tolerance. Chronic inflammatory contexts, such as found in rheumatoid arthritis (RA), severely affect the distribution and the function of DC, contributing to defective tolerance and fueling inflammation. In RA, the synovial fluid of patients is enriched by a subset of DC that derive from monocytes (Mo-DC), which promote deleterious Th17 responses. The characterization of environmental factors in the joint that impact on the development and the fate of human Mo-DC is therefore of great importance in RA. When monocytes leave the blood and infiltrate inflamed synovial tissues, the process of differentiation into Mo-DC can be influenced by interactions with soluble factors such as cytokines, local acidosis and dysregulated synoviocytes. Other molecular factors, such as the citrullination process, can also enhance osteoclast differentiation from Mo-DC, favoring bone damages in RA. Conversely, biotherapies used to control inflammation in RA, modulate also the process of monocyte differentiation into DC. The identification of the environmental mediators that control the differentiation of Mo-DC, as well as the underlying molecular signaling pathways, could constitute a major breakthrough for the development of new therapies in RA

An unexpected response to rituximab in a patient with rheumatoid arthritis

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Extensive Phenotype of Human Inflammatory Monocyte-Derived Dendritic Cells

Inflammatory monocyte-derived dendritic cells (Mo-DCs) have been described in several chronic inflammatory disorders, such as rheumatoid arthritis (RA), and are suspected to play a detrimental role by fueling inflammation and skewing adaptive immune responses. However, the characterization of their phenotype is still limited, as well as the comprehension of the factors that govern their differentiation. Here, we show that inflammatory Mo-DCs generated in vitro expressed a large and atypical panel of C-type lectin receptors, including isoforms of CD209 and CD206, CD303 and CD207, as well as intracellular proteins at their surfaces such as the lysosomal protein CD208. Combination of these markers allowed us to identify cells in the synovial fluid of RA patients with a close phenotype of inflammatory Mo-DCs generated in vitro. Finally, we found in coculture experiments that RA synoviocytes critically affected the phenotypic differentiation of monocytes into Mo-DCs, suggesting that the crosstalk between infiltrating monocytes and local mesenchymal cells is decisive for Mo-DCs generation

MEK/ERK-dependent uPAR expression is required for motility via phosphorylation of P70S6K in human hepatocarcinoma cells.

International audienceMotility and invasiveness events require specific intracellular signaling cascade activations. In cancer liver cells, one of these mechanisms could involve the MAPK MEK/ERK cascade activation which has been shown over expressed and activated in hepatocellular carcinoma. To study whether the MEK/ERK cascade is involved in the motility of HCC, we examined the effect of MEK inhibitor and ERK2 silencing using monolayer wound-healing assays and fluoroblock invasion systems. Evidence was provided that the MAPK cascade is a key transduction pathway which controls HCC cells motility and invasiveness. We could disconnect proliferation to motility using mitomycin C and we established that RNAi-mediated inhibition of ERK2 led to strongly reduced cell motility. To improve our understanding, we analysed the regulation and the role of urokinase receptor (uPAR) in this process. We provided evidence that uPAR was under a MEK/ERK dependent mechanism and blocking uPAR activity using specific antagonist or inhibiting its expression by RNA interference which resulted in complete inhibition of motility. Moreover, we found in MAPK inhibited cultures and in uPAR silencing cells that p70S6K phosphorylation on residue Thr-389 was significantly reduced, whereas Ser-421/Thr-424 phosphorylation did not change. We highlighted that the FRAP/mTOR pathway did not affect motility and Thr-389 phosphorylation. Furthermore, we demonstrated that p70S6K inhibition by RNA interference completely inhibited hepatocarcinoma cell motility. Therefore, targeting uPAR and/or MEK/ERK/S6K by RNA interference could be a major therapeutic strategy for the future treatment of invasive hepatocarcinoma cells

1-Methyl-tryptophan can interfere with TLR signaling in dendritic cells independently of IDO activity.

International audienceThe compound 1-methyl-tryptophan (1-MT) is a competitive inhibitor of IDO that can break tolerance and induce fetus, graft, and tumor rejection. Because of its broad effect on immune-related mechanisms, the direct action of 1-MT on human monocyte-derived dendritic cells (DC) was analyzed. It is shown here that the effect of 1-MT on DC is dependent on the maturation pathway. Although 1-MT had no effect on DC stimulated by the TLR3 ligand poly(I:C), it strongly enhanced the Th1 profile of DC stimulated with TLR2/1 or TLR2/6 ligands. Drastic changes in the function of DC stimulated by the TLR4 ligand LPS were induced by 1-MT. These cells could still activate allogeneic and syngeneic T cells but stimulation yielded T cells secreting IL-5 and IL-13 rather than IFN-gamma. This action of 1-MT correlated with an increased phosphorylation of p38 and ERK MAPKs and sustained activation of the transcription factor c-Fos. Inhibiting p38 and ERK phosphorylation with synthetic inhibitors blocked the effect of 1-MT on LPS-stimulated DC. Thus, 1-MT can modulate DC function depending on the maturation signal and independently of its action on IDO. This is consistent with previous observations and will help further understanding the mechanisms of DC polarization