Expression of langerin/CD207 reveals dendritic cell heterogeneity between inbred mouse strains.

Langerin/CD207 is expressed by a subset of dendritic cells (DC), the epithelial Langerhans cells. However, langerin is also detected among lymphoid tissue DC. Here, we describe striking differences in langerin-expressing cells between inbred mouse strains. While langerin+ cells are observed in comparable numbers and with comparable phenotypes in the epidermis, two distinct DC subsets bear langerin in peripheral, skin-draining lymph nodes of BALB/c mice (CD11c(high) CD8alpha(high) and CD11c(low) CD8alpha(low)), whereas only the latter subset is present in C57BL/6 mice. The CD11c(high) subset is detected in mesenteric lymph nodes and spleen of BALB/c mice, but is virtually absent from C57BL/6 mice. Similar differences are observed in other mouse strains. CD11c(low) langerin+ cells represent skin-derived Langerhans cells, as demonstrated by their high expression of DEC-205/CD205, maturation markers, and recruitment to skin-draining lymph nodes upon imiquimod-induced inflammation. It will be of interest to determine the role of lymphoid tissue-resident compared to skin-derived langerin+ DC.

Dendritic cells rapidly recruited into epithelial tissues via CCR6/CCL20 are responsible for CD8+ T cell crosspriming in vivo.

The nature of dendritic cell(s) (DC[s]) that conditions efficient in vivo priming of CD8+ CTL after immunization via epithelial tissues remains largely unknown. Here, we show that myeloid DCs rapidly recruited by adjuvants into the buccal mucosa or skin are essential for CD8+ T cell crosspriming. Recruitment of circulating DC precursors, including Gr1+ monocytes, precedes the sequential accumulation of CD11c+ MHC class II+ DCs in dermis and epithelium via a CCR6/CCL20-dependent mechanism. Remarkably, a defect in CCR6, local neutralization of CCL20, or depletion of monocytes prevents in vivo priming of CD8+ CTL against an innocuous protein antigen administered with adjuvant. In addition, transfer of CCR6-sufficient Gr1+ monocytes restores CD8+ T cell priming in CCR6( degrees / degrees ) mice via a direct Ag presentation mechanism. Thus, newly recruited DCs likely derived from circulating monocytes are responsible for efficient crosspriming of CD8+ CTL after mucosal or skin immunization.

Mouse lymphoid tissue contains distinct subsets of langerin/CD207 dendritic cells, only one of which represents epidermal-derived Langerhans cells.

Langerin/CD207 is a C-type lectin associated with formation of Birbeck granules (BG) in Langerhans cells (LC). Here, we describe a monoclonal antibody (mAb 205C1) recognizing the extracellular domain of mouse langerin. Cell-surface langerin was detected in all epidermal LC, which presented a uniform phenotype. Two subpopulations of langerin+ cells were identified in peripheral lymph nodes (LN). One population (subset 1) was CD11c(low/+)/CD8alpha(-/low)/CD11b+/CD40+/CD86+. The other population (subset 2) was CD11c(high)/CD8alpha+/CD11b(low), and lacked CD40 and CD86. Only subset 1 was fluorescein 5-isothiocyanate (FITC+) following painting onto epidermis, and the appearance of such FITC+ cells in draining LN was inhibited by pertussis toxin. Mesenteric LN, spleen, and thymus contained only a single population of langerin+ DC, corresponding to peripheral LN subset 2. Unexpectedly, BG were absent from spleen CD8alpha+ DC despite expression of langerin, and these organelles were not induced by mAb 205C1. Collectively, we demonstrate that two langerin+ DC populations (subsets 1 and 2) co-exist in mouse lymphoid tissue. Subset 1 unequivocally identifies epidermal LC-derived DC. The distribution of subset 2 indicates a non-LC origin of these langerin+ cells. These findings should facilitate our understanding of the role played by langerin in lymphoid organ DC subsets.

Disruption of the langerin/CD207 gene abolishes Birbeck granules without a marked loss of Langerhans cell function.

Langerin is a C-type lectin expressed by a subset of dendritic leukocytes, the Langerhans cells (LC). Langerin is a cell surface receptor that induces the formation of an LC-specific organelle, the Birbeck granule (BG). We generated a langerin(-/-) mouse on a C57BL/6 background which did not display any macroscopic aberrant development. In the absence of langerin, LC were detected in normal numbers in the epidermis but the cells lacked BG. LC of langerin(-/-) mice did not present other phenotypic alterations compared to wild-type littermates. Functionally, the langerin(-/-) LC were able to capture antigen, to migrate towards skin draining lymph nodes, and to undergo phenotypic maturation. In addition, langerin(-/-) mice were not impaired in their capacity to process native OVA protein for I-A(b)-restricted presentation to CD4(+) T lymphocytes or for H-2K(b)-restricted cross-presentation to CD8(+) T lymphocytes. langerin(-/-) mice inoculated with mannosylated or skin-tropic microorganisms did not display an altered pathogen susceptibility. Finally, chemical mutagenesis resulted in a similar rate of skin tumor development in langerin(-/-) and wild-type mice. Overall, our data indicate that langerin and BG are dispensable for a number of LC functions. The langerin(-/-) C57BL/6 mouse should be a valuable model for further functional exploration of langerin and the role of BG.

In vivo manipulation of dendritic cell migration and activation to elicit antitumour immunity.

Two approaches have been pursued to elicit antitumour immunity: (i) induce recruitment of immature dendritic cells or their precursors at a site of antigen delivery, and (ii) induce activation of tumour-infiltrating dendritic cells (DCs). The recruitment of selected DC subtype conditions the class of the immune response. Each immature DC population displays a unique spectrum of chemokine responsiveness. For examples, Langerhans cells (LCs) migrate selectively in response to CCL20/MIP-3alpha (through CCR6), blood CD11c+ DC to MCP chemokines (through CCR2). All these chemokines are inducible in response to inflammatory stimuli. CCL20/MIP-3alpha in particular is only detected within inflamed epithelium, at the site of antigen entry, which is infiltrated by immature DCs. Furthermore, to reach the site of injury, sequential responsiveness might operate, blood DC precursors are recruited by a set of chemokines (MIP, MCP) while within the tissue other chemokines will direct their navigation (CCL20/MIP-3alpha). Of interest, when injected in vivo together with antigen, MCP-4/CCL13, but not CCL20/MIP-3alpha, recruits blood monocytes or blood DC precursors that promptly differentiate into typical DCs and that improve antitumour immune responses. After antigen uptake, DCs acquire, upon maturation, responsiveness to CCR7 ligands (CCL21/SLC/6Ckine, CCL19/ELC/MIP-3beta) due to receptor up-regulation. In particular, in the periphery, CCL21/SLC/6Ckine expressed by lymphatic vessels may direct into the lymph stream, antigen-loaded maturing DCs leaving the site of infection; while within lymph-node, CCL21/SLC/6Ckine plays a critical role in the entry of naïve T cells from the blood through HEV. In regard to its central role, we decided to investigate whether the expression of CCL21/SLC/6Ckine in tumour may lead to antitumour immune responses. C26 colon carcinoma tumour cell line transduced with CCL21/SLC/6Ckine showed reduced tumorigenicity when injected in vivo into immunocompetent mice. The protection was CD8 dependent and associated with an important intratumoral infiltration of DCs. Most tumour infiltrating DCs (TIDCs) had an immature phenotype, were able to present TAA in the context of MHC class I, but were refractory to stimulation with the combination of LPS, IFNgamma and anti-CD40 antibody. TIDC paralysis could be reverted, however, by in vitro or in vivo stimulation with the combination of a CpG immunostimulatory sequence and an anti-interleukin 10 receptor (IL10R) antibody. CpG or anti-IL10R alone were inactive in TIDC, while CpG triggered activation in normal DC. In particular, CpG plus anti-IL10R enhanced the TAA-specific immune response and triggered de novo IL-12 production. Subsequently, CpG plus anti-IL10R treatment showed robust antitumour therapeutic activity exceeding by far that of CpG alone, and elicited antitumour immune memory.

Reversal of tumor-induced dendritic cell paralysis by CpG immunostimulatory oligonucleotide and anti-interleukin 10 receptor antibody.

Progressing tumors in man and mouse are often infiltrated by dendritic cells (DCs). Deficient antitumor immunity could be related to a lack of tumor-associated antigen (TAA) presentation by tumor-infiltrating DCs (TIDCs) or to a functional defect of TIDCs. Here we investigated the phenotype and function of TIDCs in transplantable and transgenic mouse tumor models. Although TIDCs could encompass various known DC subsets, most had an immature phenotype. We observed that TIDCs were able to present TAA in the context of major histocompatibility complex class I but that they were refractory to stimulation with the combination of lipopolysaccharide, interferon gamma, and anti-CD40 antibody. We could revert TIDC paralysis, however, by in vitro or in vivo stimulation with the combination of a CpG immunostimulatory sequence and an anti-interleukin 10 receptor (IL-10R) antibody. CpG or anti-IL-10R alone were inactive in TIDCs, whereas CpG triggered activation in normal DCs. In particular, CpG plus anti-IL-10R enhanced the TAA-specific immune response and triggered de novo IL-12 production. Subsequently, CpG plus anti-IL-10R treatment showed robust antitumor therapeutic activity exceeding by far that of CpG alone, and elicited antitumor immune memory.

Sequential involvement of CCR2 and CCR6 ligands for immature dendritic cell recruitment: possible role at inflamed epithelial surfaces.

To reach the site of antigen deposition at epithelial surfaces, dendritic cells (DC) have to traverse the endothelial barrier, progress through the tissue (i.e., dermis) and cross the dermo-epithelial junction (basal membrane). In the present study, we demonstrate that (1) circulating blood DC and monocytes express high levels of CCR2 and primarily respond to monocyte chemotactic protein (MCP) and not to macrophage inflammatory protein (MIP)-3alpha/CCL20; (2) while the CD34(+) hematopoietic progenitor cells (HPC)-derived CD1a(+) precursors committed to Langerhans cell differentiation primarily respond to MIP-3alpha/CCL20, the HPC-derived CD14(+) precursors respond to both MCP and MIP-3alpha/CCL20; (3) in concordance with the sequential expression of CCR2 and CCR6, the HPC-derived CD14(+) precursors initially acquire the ability to migrate in response to MCP-4/CCL13 and subsequently in response to MIP-3alpha/CCL20; and (4) in vivo, in inflamed epithelium, MCP-4/CCL13 and MIP-3alpha/CCL20 form complementary gradients, with MCP-4/CCL13 expressed in basal epithelial cells at the contact of blood vessels, while MIP-3alpha/CCL20 expression is restricted to epithelial cells bordering the external milieu. These observations suggest that the recruitment of DC to the site of infection is controlled by the sequential action of different chemokines: (i) CCR2(+) circulating DC or DC precursors are mobilized into the tissue via the expression of MCP by cells lining blood vessels, and (ii) these cells traffic from the tissue to the site of pathogen invasion via the production of MIP-3alpha/CL20 by epithelial cells and the up-regulation of CCR6 in response to the tissue environment.

Identification of mouse langerin/CD207 in Langerhans cells and some dendritic cells of lymphoid tissues.

Human (h)Langerin/CD207 is a C-type lectin of Langerhans cells (LC) that induces the formation of Birbeck granules (BG). In this study, we have cloned a cDNA-encoding mouse (m)Langerin. The predicted protein is 66% homologous to hLangerin with conservation of its particular features. The organization of human and mouse Langerin genes are similar, consisting of six exons, three of which encode the carbohydrate recognition domain. The mLangerin gene maps to chromosome 6D, syntenic to the human gene on chromosome 2p13. mLangerin protein, detected by a mAb as a 48-kDa species, is abundant in epidermal LC in situ and is down-regulated upon culture. A subset of cells also expresses mLangerin in bone marrow cultures supplemented with TGF-beta. Notably, dendritic cells in thymic medulla are mLangerin-positive. By contrast, only scattered cells express mLangerin in lymph nodes and spleen. mLangerin mRNA is also detected in some nonlymphoid tissues (e.g., lung, liver, and heart). Similarly to hLangerin, a network of BG form upon transfection of mLangerin cDNA into fibroblasts. Interestingly, substitution of a conserved residue (Phe(244) to Leu) within the carbohydrate recognition domain transforms the BG in transfectant cells into structures resembling cored tubules, previously described in mouse LC. Our findings should facilitate further characterization of mouse LC, and provide insight into a plasticity of dendritic cell organelles which may have important functional consequences.

Regulation of dendritic cell recruitment by chemokines.

Dendritic cells (DC) are a heterogeneous family of cells that function as sentinels of the immune system. This article summarizes observations suggesting that inflammatory chemokines secreted at the site of pathogen invasion determine the DC subset recruited and influence the class of the immune response initiated. Langerhans cells are selectively recruited by MIP-3alpha/CCL20. In contrast, CCR7 ligands have a key role in the accumulation of antigen-loaded mature DC in T cell-rich areas of the draining lymph node. Improved understanding of the regulation of DC trafficking might offer new opportunities for therapeutic interventions to control immune responses.