Investigating Evolutionary Conservation of Dendritic Cell Subset Identity and Functions

International audienceDendritic cells (DCs) were initially defined as mononuclear phagocytes with a dendritic morphology and an exquisite efficiency for naïve T-cell activation. DC encompass several subsets initially identified by their expression of specific cell surface molecules and later shown to excel in distinct functions and to develop under the instruction of different transcription factors or cytokines. Very few cell surface molecules are expressed in a specific manner on any immune cell type. Hence, to identify cell types, the sole use of a small number of cell surface markers in classical flow cytometry can be deceiving. Moreover, the markers currently used to define mononuclear phagocyte subsets vary depending on the tissue and animal species studied and even between laboratories. This has led to confusion in the definition of DC subset identity and in their attribution of specific functions. There is a strong need to identify a rigorous and consensus way to define mononuclear phagocyte subsets, with precise guidelines potentially applicable throughout tissues and species. We will discuss the advantages, drawbacks, and com-plementarities of different methodologies: cell surface phenotyping, ontogeny, functional characterization, and molecular profiling. We will advocate that gene expression profiling is a very rigorous, largely unbiased and accessible method to define the identity of mononuclear phagocyte subsets, which strengthens and refines surface phenotyping. It is uniquely powerful to yield new, experimentally testable, hypotheses on the ontogeny or functions of mononuclear phagocyte subsets, their molecular regulation, and their evolutionary conservation. We propose defining cell populations based on a combination of cell surface phenotyping, expression analysis of hallmark genes, and robust functional assays, in order to reach a consensus and integrate faster the huge but scattered knowledge accumulated by different laboratories on different cell types, organs, and species

A new subunit vaccine based on nucleoprotein nanoparticles confers partial clinical and virological protection in calves against bovine respiratory syncytial virus

Human and bovine respiratory syncytial viruses (HRSV and BRSV) are two closely related, worldwide prevalent viruses that are the leading cause of severe airway disease in children and calves, respectively. Efficacy of commercial bovine vaccines needs improvement and no human vaccine is licensed yet. We reported that nasal vaccination with the HRSV nucleoprotein produced as recombinant ringshaped nanoparticles (NSRS) protects mice against a viral challenge with HRSV. The aim of this work was to evaluate this new vaccine that uses a conserved viral antigen, in calves, natural hosts for BRSV. Calves, free of colostral or natural anti-BRSV antibodies, were vaccinated with NSRS either intramuscularly, or both intramuscularly and intranasally using MontanideTM ISA71 and IMS4132 as adjuvants and challenged with BRSV. All vaccinated calves developed anti-N antibodies in blood and nasal secretions and N-specific cellular immunity in local lymph nodes. Clinical monitoring post-challenge demonstrated moderate respiratory pathology with local lung tissue consolidations for the non vaccinated calves that were significantly reduced in the vaccinated calves. Vaccinated calves had lower viral loads than the nonvaccinated control calves. Thus NSRS vaccination in calves provided cross-protective immunity against BRSV infection without adverse inflammatory reaction

Leukocytes migrating in lymph from the oro-nasal mucosae : interest for vaccination and immune tolerance

We developed a novel sheep model of lymphatic catheterisation to collect, in real time, cells circulating in the afferent lymph draining mainly the mucosae in the head. With this model, we were able to collect migrating leukocytes, either in their baseline condition, or after the administration of a vaccine antigen, from the oro-nasal mucosa to the draining lymph nodes where the immune response takes place. We showed that particulate antigens, such as bacteria, are taken from the tissues to the lymph nodes mainly by two types of cells, the monocytes and the granulocytes, and only occasionally by dendritic cells (DC). This finding challenges the general view, common among immunologists, whereby dendritic cells capture the antigen in the tissues before taking it to the lymph node. In addition, our study identified the permanent migration of a subset of DC - expressing the CD26 molecule – which carries cell-derived apoptotic bodies of self. This DC subset could be responsible for self-tolerance, a mechanism involved in control of transplant rejection, autoimmune diseases and vaccination.Un modèle original de cathétérisme lymphatique chez l'ovin, développé dans notre laboratoire, permet d'accéder en temps réel aux cellules en migration dans la lymphe drainant principalement les muqueuses de la tête de l'animal. Ainsi, à l'état basal ou après administration d'un antigène vaccinal, il est possible de collecter des cellules leucocytaires migrant depuis les muqueuses oro-nasales vers les ganglions, sites décisionnels de la réponse immune. Grâce à ce modèle, nous avons montré que des antigènes particulaires, notamment des bactéries, étaient transportés dans la lymphe essentiellement par deux types de cellules, les monocytes et les granulocytes et plus marginalement par les cellules dendritiques (DC). Ces résultats modifient la vision linéaire communément admise selon laquelle les DC capturent, transportent et présentent les antigènes aux lymphocytes naïfs; ils suggèrent que d'autres populations phagocytaires, impliquées dans le transport antigénique, pourraient moduler la réponse immune. Par ailleurs, nous avons mis en évidence la migration permanente d'une sous-population de DC, identifiée par la molécule CD26, qui transporte des débris de cellules du soi. Ce phénomène pourrait être impliqué dans la tolérance périphérique, mécanisme d'importance dans le rejet des greffes, les maladies auto-immunes et la vaccination

Study of type I interferon producing cells in sheep lymph draining oro-nasal mucosae or skin tissues

Interferons (IFN) are natural antiviralmolecules produced by cells as an early response to a viral aggression. To study the early production of IFN at primary sites of viral infections (i.e. mucosal or skin tissues), we used a novel lymphatic cannulation model in sheep, providing real-time access to lymph and lymph cells draining a mucosal or skin area. Following oronasal or cutaneous injection of an IFNinducing oligonucleotide, we were able to detect IFN early on in the lymph draining the injection site, as well as IFN-producing lymph cells migrating from the injection site to the draining lymph node. These rare cells (<1%) share the same features as plasmacytoid dendritic cells (low density, Bneg CD11cneg CD45RBpos, expressing TLR7 and 9, and IRF7 transcripts). They play an important role in the activation of adaptive immunity, and could provide interesting targets for innovative vaccine strategies.Les interférons (IFN) constituent une famille de molécules naturelles antivirales dont la fabrication par les cellules est déclenchée dès les premières phases d'une agression virale. Afin d'étudier cette production précoce au plus près des sites primaires d'infection virale (muqueuses ou peau), nous avons utilisé un modèle original de cathétérisme lymphatique chez le mouton, qui permet d'accéder en temps réel à la lymphe et aux cellules lymphatiques drainant un territoire muqueux ou cutané. Après injection oro-nasale ou cutanée d'un oligonucléotide inducteur d'IFN, nous détectons la présence précoce d'IFN dans la lymphe drainant le site d'injection, ainsi que la présence de cellules lymphatiques productrices d'IFN, en migration du site d'induction vers le ganglion drainant. Ces cellules rares (<1%) ont les caractéristiques de cellules dendritiques plasmacytoïdes (faible densité, Bneg CD11cneg CD45RBpos, exprimant les transcrits des TLR 7 et 9 et de l'IRF7). Ces cellules, déterminantes dans l'activation de l'immunité adaptative, pourraient être ciblées avantageusement par des stratégies vaccinales novatrices

Lymphatic Transport and Lymph Node Location of Microspheres Subcutaneously Injected in the Vicinity of Tumors in a Rabbit Model of Breast Cancer

International audienc

Issues and special features of animal health research

In the rapidly changing context of research on animal health, INRA launched a collective discussion on the challenges facing the field, its distinguishing features, and synergies with biomedical research. As has been declared forcibly by the heads of WHO, FAO and OIE, the challenges facing animal health, beyond diseases transmissible to humans, are critically important and involve food security, agriculture economics, and the ensemble of economic activities associated with agriculture. There are in addition issues related to public health (zoonoses, xenobiotics, antimicrobial resistance), the environment, and animal welfare

The XC chemokine receptor 1 is a conserved selective marker of mammalian cells homologous to mouse CD8α+ dendritic cells

Human BDCA3+ dendritic cells (DCs) were suggested to be homologous to mouse CD8α+ DCs. We demonstrate that human BDCA3+ DCs are more efficient than their BDCA1+ counterparts or plasmacytoid DCs (pDCs) in cross-presenting antigen and activating CD8+ T cells, which is similar to mouse CD8α+ DCs as compared with CD11b+ DCs or pDCs, although with more moderate differences between human DC subsets. Yet, no specific marker was known to be shared between homologous DC subsets across species. We found that XC chemokine receptor 1 (XCR1) is specifically expressed and active in mouse CD8α+, human BDCA3+, and sheep CD26+ DCs and is conserved across species. The mRNA encoding the XCR1 ligand chemokine (C motif) ligand 1 (XCL1) is selectively expressed in natural killer (NK) and CD8+ T lymphocytes at steady-state and is enhanced upon activation. Moreover, the Xcl1 mRNA is selectively expressed at high levels in central memory compared with naive CD8+ T lymphocytes. Finally, XCR1−/− mice have decreased early CD8+ T cell responses to Listeria monocytogenes infection, which is associated with higher bacterial loads early in infection. Therefore, XCR1 constitutes the first conserved specific marker for cell subsets homologous to mouse CD8α+ DCs in higher vertebrates and promotes their ability to activate early CD8+ T cell defenses against an intracellular pathogenic bacteria

Porcine Reproductive and Respiratory Syndrome Virus Type 1.3 Lena Triggers Conventional Dendritic Cells 1 Activation and T Helper 1 Immune Response Without Infecting Dendritic Cells

Porcine Reproductive and Respiratory Syndrome virus (PRRSV) is an arterivirus responsible for highly contagious infection and huge economic losses in pig industry. Two species, PRRSV-1 and PRRSV-2 are distinguished, PRRSV-1 being more prevalent in Europe. PRRSV-1 can further be divided in subtypes. PRRSV-1.3 such as Lena are more pathogenic than PRRSV-1.1 such as Lelystad or Flanders13. PRRSV-1.3 viruses trigger a higher Th1 response than PRRSV-1.1, although the role of the cellular immune response in PRRSV clearance remains ill defined. The pathogenicity as well as the T cell response inductions may be differentially impacted according to the capacity of the virus strain to infect and/or activate DCs. However, the interactions of PRRSV with in vivo-differentiated-DC subtypes such as conventional DC1 (cDC1), cDC2, and monocyte-derived DCs (moDC) have not been thoroughly investigated. Here, DC subpopulations from Lena in vivo infected pigs were analyzed for viral genome detection. This experiment demonstrates that cDC1, cDC2, and moDC are not infected in vivo by Lena. Analysis of DC cytokines production revealed that cDC1 are clearly activated in vivo by Lena. In vitro comparison of 3 Europeans strains revealed no infection of the cDC1 and cDC2 and no or little infection of moDC with Lena, whereas the two PRRSV-1.1 strains infect none of the 3 DC subtypes. In vitro investigation of T helper polarization and cytokines production demonstrate that Lena induces a higher Th1 polarization and IFNγ secretion than FL13 and LV. Altogether, this work suggests an activation of cDC1 by Lena associated with a Th1 immune response polarization