Robust Anti-viral Immunity Requires Multiple Distinct T Cell-Dendritic Cell Interactions.
ABSTRACT: Host defense against viruses and intracellular parasites depends on effector CD8(+) T cells, whose optimal clonal expansion, differentiation, and memory properties require signals from CD4(+) T cells. Here, we addressed the role of dendritic cell (DC) subsets in initial activation of the two T cell types and their co-operation. Surprisingly, initial priming of CD4(+) and CD8(+) T cells was spatially segregated within the lymph node and occurred on different DCs with temporally distinct patterns of antigen presentation via MHCI versus MHCII molecules. DCs that co-present antigen via both MHC molecules were detected at a later stage; these XCR1(+) DCs are the critical platform involved in CD4(+) T cell augmentation of CD8(+) T cell responses. These findings delineate the complex choreography of cellular interactions underlying effective cell-mediated anti-viral responses, with implications for basic DC subset biology, as well as for translational application to the development of vaccines that evoke optimal T cell immunity.
Project description:Adaptive cellular immunity is initiated by antigen-specific interactions between T lymphocytes and dendritic cells (DCs). Plasmacytoid DCs (pDCs) support antiviral immunity by linking innate and adaptive immune responses. Here we examined pDC spatiotemporal dynamics during viral infection to uncover when, where, and how they exert their functions. We found that pDCs accumulated at sites of CD8+ T cell antigen-driven activation in a CCR5-dependent fashion. Furthermore, activated CD8+ T cells orchestrated the local recruitment of lymph node-resident XCR1 chemokine receptor-expressing DCs via secretion of the XCL1 chemokine. Functionally, this CD8+ T cell-mediated reorganization of the local DC network allowed for the interaction and cooperation of pDCs and XCR1+ DCs, thereby optimizing XCR1+ DC maturation and cross-presentation. These data support a model in which CD8+ T cells upon activation create their own optimal priming microenvironment by recruiting additional DC subsets to the site of initial antigen recognition.
Project description:Evidence suggests that distinct splenic dendritic cell (DC) subsets activate either CD4+ or CD8+ T cells in vivo. This bias has been partially ascribed to differential antigen presentation; however, all DC subsets can activate both T cell lineages in vitro. Therefore, we tested whether the organization of DC and T cell subsets in the spleen dictated this preference. We discovered that CD4+ and CD8+ T cells segregated within splenic T cell zones prior to immunization. After intravenous immunization, the two major conventional DC populations, distinguished by 33D1 and XCR1 staining, migrated into separate regions of the T cell zone: 33D1+ DCs migrated into the CD4+ T cell area, whereas XCR1+ DCs migrated into the CD8+ T cell area. Thus, the post-immunization location of each DC subset correlated with the T cell lineage it preferentially primes. Preventing this co-localization selectively impaired either CD4+ or CD8+ T cell immunity to blood-borne antigens.
Project description:Dendritic cells (DCs) are antigen-presenting cells specialized for activating T cells to elicit effector T-cell functions. Cross-presenting DCs are a DC subset capable of presenting antigens to CD8(+) T cells and play critical roles in cytotoxic T-cell-mediated immune responses to microorganisms and cancer. Although their importance is known, the spatiotemporal dynamics of cross-presenting DCs in vivo are incompletely understood. Here, we study the T-cell zone in skin-draining lymph nodes (SDLNs) and find it is compartmentalized into regions for CD8(+) T-cell activation by cross-presenting DCs that express the chemokine (C motif) receptor 1 gene, Xcr1 and for CD4(+) T-cell activation by CD11b(+) DCs. Xcr1-expressing DCs in the SDLNs are composed of two different populations: migratory (CD103(hi)) DCs, which immigrate from the skin, and resident (CD8?(hi)) DCs, which develop in the nodes. To characterize the dynamic interactions of these distinct DC populations with CD8(+) T cells during their activation in vivo, we developed a photoconvertible reporter mouse strain, which permits us to distinctively visualize the migratory and resident subsets of Xcr1-expressing DCs. After leaving the skin, migratory DCs infiltrated to the deep T-cell zone of the SDLNs over 3 d, which corresponded to their half-life in the SDLNs. Intravital two-photon imaging showed that after soluble antigen immunization, the newly arriving migratory DCs more efficiently form sustained conjugates with antigen-specific CD8(+) T cells than other Xcr1-expressing DCs in the SDLNs. These results offer in vivo evidence for differential contributions of migratory and resident cross-presenting DCs to CD8(+) T-cell activation.
Project description:Graft-versus-host disease (GVHD) is a major cause of morbidity and mortality in allogeneic hematopoietic stem cell transplantation (alloSCT). By static microscopy, cutaneous GVHD lesions contain a mix of T cells and myeloid cells. We used 2-photon intravital microscopy to investigate the dynamics of CD4+ and CD8+ T cells and donor dendritic cells (DCs) in cutaneous GVHD lesions in an MHC-matched, multiple minor histocompatibility antigen-mismatched (miHA) model. The majority of CD4 and CD8 cells were stationary, and few cells entered and stopped or were stopped and left the imaged volumes. CD8 cells made TCR:MHCI-dependent interactions with CD11c+ cells, as measured by the durations that CD8 cells contacted MHCI+ vs MHCI- DCs. The acute deletion of Langerin+CD103+ DCs, which were relatively rare, did not affect CD8 cell motility and DC contact times, indicating that Langerin-CD103- DCs provide stop signals to CD8 cells. CD4 cells, in contrast, had similar contact durations with MHCII+ and MHCII- DCs. However, CD4 motility rapidly increased after the infusion of an MHCII-blocking antibody, indicating that TCR signaling actively suppressed CD4 movements. Many CD4 cells still were stationary after anti-MHCII antibody infusion, suggesting CD4 cell heterogeneity within the lesion. These data support a model of local GVHD maintenance within target tissues.
Project description:Cross-presenting Xcr1+CD8? DCs are attractive APCs to target for therapeutic cancer vaccines, as they are able to take up and process antigen from dying tumor cells for their MHCI-restricted presentation to CD8 T cells. To this aim, we developed fusion proteins made of the Xcr1 ligand Xcl1 fused to an OVA synthetic long peptide (SLP) and IgG1 Fc fragment. We demonstrated the specific binding and uptake of the Xcl1 fusion proteins by Xcr1+ DCs. Most importantly, their potent adjuvant effect on the H-2Kb/OVA specific T cell response was associated with a sustained tumor control even against the poorly immunogenic B16-OVA melanoma tumor. The increased tumor protection correlated with higher tumor infiltration of antigen-specific CD8+ T cells, increased IFN? production and degranulation potential. Altogether, these results demonstrate that therapeutic cancer vaccines may be greatly improved by the combination of SLP antigen and Xcl1 fusion proteins.
Project description:Human BDCA3+ dendritic cells (DCs) were suggested to be homologous to mouse CD8alpha+ 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 CD8alpha+ 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 CD8alpha+, 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 CD8alpha+ DCs in higher vertebrates and promotes their ability to activate early CD8+ T cell defenses against an intracellular pathogenic bacteria.
Project description:Naive CD8(+) T cell priming during tumor development or many primary infections requires cross-presentation by XCR1(+) dendritic cells (DCs). Memory CD8(+) T lymphocytes (mCTLs) harbor a lower activation threshold as compared with naive cells. However, whether their recall responses depend on XCR1(+) DCs is unknown. By using a new mouse model allowing fluorescent tracking and conditional depletion of XCR1(+) DCs, we demonstrate a differential requirement of these cells for mCTL recall during secondary infections by different pathogens. XCR1(+) DCs were instrumental to promote this function upon secondary challenges with Listeria monocytogenes, vesicular stomatitis virus, or Vaccinia virus, but dispensable in the case of mouse cytomegalovirus. We deciphered how XCR1(+) DCs promote mCTL recall upon secondary infections with Listeria. By visualizing for the first time the in vivo choreography of XCR1(+) DCs, NK cells and mCTLs during secondary immune responses, and by neutralizing in vivo candidate molecules, we demonstrate that, very early after infection, mCTLs are activated, and attracted in a CXCR3-dependent manner, by NK cell-boosted, IL-12-, and CXCL9-producing XCR1(+) DCs. Hence, depending on the infectious agent, strong recall of mCTLs during secondary challenges can require cytokine- and chemokine-dependent cross-talk with XCR1(+) DCs and NK cells.
Project description:During an immune response, antigen-bearing dendritic cells (DCs) migrate to the local draining lymph node and present antigen to CD4(+) helper T cells. Antigen-activated CD4(+) T cells then up-regulate TNF superfamily members including CD40 ligand and lymphotoxin (LT)??. Although it is well-accepted that CD40 stimulation on DCs is required for DC licensing and cross-priming of CD8(+) T-cell responses, it is likely that other signals are integrated into a comprehensive DC activation program. Here we show that a cognate interaction between LT?? on CD4(+) helper T cells and LT? receptor on DCs results in unique signals that are necessary for optimal CD8(+) T-cell expansion via a type I IFN-dependent mechanism. In contrast, CD40 signaling appears to be more critical for CD8(+) T-cell IFN? production. Therefore, different TNF family members provide integrative signals that shape the licensing potential of antigen-presenting DCs.
Project description:The chemokine (C motif) receptor 1 (XCR1) and its ligandXCL1 have been intensively studied in the mouse and human immune systems. Here, we determined the molecular characteristics of cattle XCR1 and XCL1 and their distribution among peripheral blood cells. Cattle XCR1 mRNA expression was mainly restricted to CD26+CADM1+CD205+MHCII+CD11b- cells in blood that were otherwise lineage marker negative (lin-); these represented a subset of classic dendritic cells (DCs), not plasmacytoid DCs. Some of these DCs expressed CD11a, CD44, CD80 and CD86, but they did not express CD4, CD8, CD163 or CD172a. Cattle XCL1 was expressed in quiescent NK cells and in activated CD8+ T cells. Cattle XCR1+ DCs migrated chemotactically in response to mouse, but not to human, XCL1. The distribution characters of cattle XCR1 and XCL1 suggested a vital role in regulation of acquired immune responses and indicated a potential for a DC targeted veterinary vaccine in cattle using XCL1 fused antigens.
Project description:DC-SIGN is an antigen uptake receptor expressed on dendritic cells (DCs) with specificity for glycans present on a broad variety of pathogens and is capable of directing its cargo to MHC-I and MHC-II pathways for the induction of CD8+ and CD4+ T cell responses, respectively. Therefore, DC-SIGN is a very promising target for the delivery of antigen for anti-cancer vaccination. Although the endocytic route leading to MHC-II presentation is characterized to a large extent, the mechanisms controlling DC-SIGN targeted cross-presentation of exogenous peptides on MHC-I, are not completely resolved yet. In this paper, we used imaging flow cytometry and antigen-specific CD8+ T cells to investigate the intracellular fate of DC-SIGN and its cargo in human DCs. Our data demonstrates that immature DCs and toll-like receptor 4 (TLR4) stimulated DCs had similar internalization capacity and were both able to cross-present antigen targeted via DC-SIGN. Interestingly, simultaneous triggering of TLR4 and DC-SIGN on DCs resulted in the translocation of cargo to the cytosol, leading to proteasome-dependent processing and increased CD8+ T cell activation. Understanding the dynamics of DC-SIGN-mediated uptake and processing is essential for the design of optimal DC-SIGN-targeting vaccination strategies aimed at enhancing CD8+ T cell responses.