Negative Regulation of NKG2D Expression by IL-4 in Memory CD8 T Cells

International audienceIL-4 is one of the main cytokines produced during Th2-inducing pathologies. This cytokine has been shown to affect a number of immune processes such as Th differentiation and innate immune responses. However, the impact of IL-4 on CD8 T cell responses remains unclear. In this study, we analyzed the effects of IL-4 on global gene expression profiles of Ag-induced memory CD8 T cells in the mouse. Gene ontology analysis of this signature revealed that IL-4 regulated most importantly genes associated with immune responses. Moreover, this IL-4 signature overlapped with the set of genes preferentially expressed by memory CD8 T cells over naive CD8 T cells. In particular, IL-4 downregulated in vitro and in vivo in a STAT6-dependent manner the memory-specific expression of NKG2D, thereby increasing the activation threshold of memory CD8 T cells. Furthermore, IL-4 impaired activation of memory cells as well as their differentiation into effector cells. This phenomenon could have an important clinical relevance as patients affected by Th2 pathologies such as parasitic infections or atopic dermatitis often suffer from viral-induced complications possibly linked to inefficient CD8 T cell responses

Identification of Nascent Memory CD8 T Cells and Modeling of Their Ontogeny

International audiencePrimary immune responses generate short-term effectors and long-term protective memory cells. The delineation of the genealogy linking naive, effector and memory cells has been complicated by the lack of phenotypes discriminating effector from memory differentiation stages. Using transcriptomics and phenotypic analyses, we identify a novel marker combination that allows us to track nascent memory cells within the effector phase. We then use a formal approach based on mathematical models describing the dynamics of population-size evolutions to test potential progeny links and demonstrate that most cells follow a linear naive-early effector-late effector-memory pathway. Moreover, our mathematical modelallows long-term prediction of memory cell numbers from a few early experimental measurements. Our work thus provides a phenotypic means to identify effector and memory cells, as well as a mathematical framework to investigate the ontology of their generation and to predict the outcome of immunization regimens in terms of memory cell numbers generated

Antigen specific activation of cytotoxic CD8+ T cells by Staphylococcus aureus infected dendritic cells

International audienceStaphylococcus aureus ( S. aureus ) is a pathogen associated with a wide variety of diseases, from minor to life-threatening infections. Antibiotic-resistant strains have emerged, leading to increasing concern about the control of S. aureus infections. The development of vaccines may be one way to overcome these resistant strains. However, S. aureus ability to internalize into cells – and thus to form a reservoir escaping humoral immunity – is a challenge for vaccine development. A role of T cells in the elimination of persistent S. aureus has been established in mice but it remains to be established if CD8 + T cells could display a cytotoxic activity against S. aureus infected cells. We examined in vitro the ability of CD8 + T cells to recognize and kill dendritic cells infected with S. aureus. We first evidenced that both primary mouse dendritic cells and DC2.4 cell line can be infected with S. aureus . We then generated a strain of S. aureus expressing a model CD8 epitope and transgenic F5 CD8 + T cells recognizing this model epitope were used as reporter T cells. In response to S. aureus -infected dendritic cells, F5 CD8 + T cells produced IFN-γ in an antigen-specific manner and displayed an increased ability to kill infected cells. Altogether, these results demonstrate that cells infected by S. aureus display bacteria-derived epitopes at their surface that are recognized by CD8 + T cells. This paves the way for the development of CD8 + T cell-based therapies against S. aureus

Poly-functional and long-lasting anticancer immune response elicited by a safe attenuated Pseudomonas aeruginosa vector for antigens delivery

International audienceLive-attenuated bacterial vectors for antigens delivery have aroused growing interest in the field of cancer immunotherapy. Their potency to stimulate innate immunity and to promote intracellular antigen delivery into antigen-presenting cells could be exploited to elicit a strong and specific cellular immune response against tumor cells. We previously described genetically-modified and attenuated Pseudomonas aeruginosa vectors able to deliver in vivo protein antigens into antigen-presenting cells, through Type 3 secretion system of the bacteria. Using this approach, we managed to protect immunized mice against aggressive B16 melanoma development in both a prophylactic and therapeutic setting. In this study, we further investigated the antigen-specific CD8(+) T cell response, in terms of phenotypic and functional aspects, obtained after immunizations with a killed but metabolically active P. aeruginosa attenuated vector. We demonstrated that P. aeruginosa vaccine induces a highly functional pool of antigen-specific CD8+ T cell able to infiltrate the tumor. Furthermore, multiple immunizations allowed the development of a long-lasting immune response, represented by a pool of predominantly effector memory cells which protected mice against late tumor challenge. Overall, killed but metabolically active P. aeruginosa vector is a safe and promising approach for active and specific antitumor immunotherapy

ASC controls IFN-γ levels in an IL-18-dependent manner in caspase-1-deficient mice infected with Francisella novicida.

Remerciements ECOFECTInternational audienceThe inflammasome is a signaling platform that is central to the innate immune responses to bacterial infections. Francisella tularensis is a bacterium replicating within the host cytosol. During F. tularensis subspecies novicida infection, AIM2, an inflammasome receptor sensing cytosolic DNA, activates caspase-1 in an ASC-dependent manner, leading to both pyroptosis and release of the proinflammatory cytokines IL-1β and IL-18. Activation of this canonical inflammasome pathway is key to limit F. novicida infection. In this study, by comparing the immune responses of AIM2 knockout (KO), ASC(KO), and Casp1(KO) mice in response to F. novicida infection, we observed that IFN-γ levels in the serum of Casp1(KO) mice were much higher than the levels observed in AIM2(KO) and ASC(KO) mice. This difference in IFN-γ production was due to a large production of IFN-γ by NK cells in Casp1(KO) mice that was not observed in ASC(KO) mice. The deficit in IFN-γ production observed in ASC(KO) mice was not due to a reduced Dock2 expression or to an intrinsic defect of ASC(KO) NK cells. We demonstrate that in infected Casp1(KO) mice, IFN-γ production is due to an ASC-dependent caspase-1-independent pathway generating IL-18. Furthermore, we present in vitro data suggesting that the recently described AIM2/ASC/caspase-8 noncanonical pathway is responsible for the caspase-1-independent IL-18 releasing activity. To our knowledge, this study is the first in vivo evidence of an alternative pathway able to generate in a caspase-1-independent pathway bioactive IL-18 to boost the production of IFN-γ, a cytokine critical for the host antibacterial response

Generation of a C57BL/6J mouse strain expressing the CD45.1 epitope to improve hematopoietic stem cell engraftment and adoptive cell transfer experiments

Adoptive cell transfer between genetically identical hosts relies on the use of a congenic marker to distinguish the donor cells from the host cells. CD45, a glycoprotein expressed by all hematopoietic cells, is one of the main congenic markers used because its two isoforms, CD45.1 and CD45.2, can be discriminated by flow cytometry. As a consequence, C57BL/6J (B6; CD45.2) and B6.SJL-Ptprca Pepcb/BoyJ (B6.SJL; CD45.1) mice are widely used in adoptive cell transfer experiments, under the presumption that they differ only at the CD45 (Ptprc) locus. However, recent studies have identified genetic variations between these congenic strains and have notably highlighted a differential expression of cathepsin E (CTSE). The B6.SJL mouse presents a number of functional differences in hematopoietic stem cell engraftment potential and immune cell numbers compared with the B6 mouse. In this study, we showed that B6 and B6.SJL mice also differ in their CD8+ T cell compartment and CD8+ T cell responses to viral infection. We identified Ctse as the most differentially expressed gene between CD8+ T cells of B6 and B6.SJL and demonstrated that the differences reported between these two mouse strains are not due to CTSE. Finally, using CRISPR–Cas9 genome editing, we generated a CD45.1-expressing B6 mouse by inserting one nucleotide mutation (A904G) leading to an amino acid change (K302E) in the Ptprc gene of the B6 mouse. We showed that this new B6-Ptprc^{em(K302E)Jmar/}J mouse resolves the experimental biases reported between the B6 and B6.SJL mouse lines and should thus represent the new gold standard for adoptive cell transfer experiments in B6

Generation of a C57BL/6J mouse strain expressing the CD45.1 epitope to improve hematopoietic stem cell engraftment and adoptive cell transfer experiments

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T inflammatory memory CD8 T cells participate to antiviral response and generate secondary memory cells with an advantage in XCL1 production

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