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alpha-Galactosylceramide (alpha-GalCer) is the prototype compound for studying the presentation of glycolipids on CD1d molecules to natural killer T (NKT) lymphocytes. A single i.v. dose of glycolipid triggers a cascade of events involving the production of several cytokines over the course of a day, a short-lived activation of NKT and natural killer (NK) cells, and a more prolonged adaptive T cell immune response if certain antigens are given together with alpha-GalCer. We find that a recently described analogue, alpha-C-galactosylceramide (alpha-C-GalCer), more potently induces these innate and adaptive immune responses in mice. alpha-C-GalCer acts as a more effective trigger for IL-12 and IFN-gamma production, although it minimally elicits IL-4 and TNF-alpha release into the serum. Also, alpha-C-GalCer better mobilizes NKT and natural killer cells to resist B16 melanoma. To help understand these effects, we find that alpha-C-GalCer binds more stably to dendritic cells than alpha-GalCer and that dendritic cells loaded with alpha-C-GalCer induce larger and more long lasting NKT cell responses in vivo. When glycolipid is targeted to dendritic cells in spleen together with antigens in dying cells, such as irradiated tumor cells, alpha-C-GalCer is active as an adjuvant for T cell-mediated immunity at lower doses, just 20 ng per mouse, where it is also able to up-regulate the required CD40L costimulatory molecule on NKT cells. Therefore, alpha-C-GalCer represents a glycolipid that binds more stably to dendritic cells and acts as a more effective link between innate and adaptive immunity in vivo.

Dendritic cells (DCs) are located at body surfaces such as the skin, respiratory and genital tracts, and intestine. To further analyze intestinal DCs, we adapted an epidermal sheet separation technique and obtained two intestinal layers, facing the lumen and serosa. Unexpectedly, immunolabeling of the layer toward the serosa revealed a regular, dense, planar network of cells with prominent dendritic morphology within the external muscular layer and with increasing frequency along the length of the intestine. Direct examination of the serosal-disposed layers showed a significant fraction of the DCs to express DEC-205/CD205, CD11c, Langerin/CD207, Fcgamma receptor/CD16/32, CD14, and low levels of activation markers, CD25, CD80, CD86, and CD95. By more sensitive FACS analyses, cells from this layer contained two CD11c(+) populations of CD45(+) CD205(+), CD19(-) leukocytes, MHC II(+) and MHC II(-). When ovalbumin conjugated to an anti-DEC-205 antibody was injected into mice, the conjugate targeted to these DCs, which upon isolation were able to stimulate ovalbumin-specific, CD4(+) and CD8(+) T cell antigen receptor-transgenic T cells. In vivo, these DCs responded to two microbial stimuli, systemic LPS and oral live bacteria, by up-regulating CD80, CD86, DEC-205, and Langerin within 12 h. This network of DCs thus represents a previously unrecognized antigen-presenting cell system in the intestine.

We find that CD11c(+) cells with many markers of dendritic cells (DCs) are a major cell type in the skin lesions of psoriasis. These CD11c(+) cells, which are evident in both epidermis and dermis, are the sites for the expression of two mediators of inflammation, inducible nitric oxide synthase (iNOS) and TNF-alpha in diseased skin. These cells express HLA-DR, CD40, and CD86, lack the Langerin and CD14 markers of Langerhans cells and monocytes, respectively, and to a significant extent express the DC maturation markers DC-LAMP and CD83. Treatment of psoriasis with efalizumab (anti-CD11a, Raptiva) strongly reduces infiltration by these DCs in patients responding to this agent. Disease activity after therapy was more related to DC infiltrates and iNOS mRNA levels than T cell infiltrates, and CD11c(+) cells responded more quickly to therapy than epidermal keratinocytes. Our results suggest that a type of DC, which resembles murine "Tip-DCs" that can accumulate during infection, has proinflammatory effects in psoriasis through nitric oxide and TNF-alpha production, and can be an important target for suppressive therapies.

Induction of antiviral cytotoxic T lymphocytes (CTLs) has been proposed to require cross-presentation of viral antigens derived from infected extralymphatic host cells by antigen-presenting cells (APC). This postulated mechanism of cross-priming is thought to be essential for CTL responses against viruses that do not infect professional APC, e.g., because of absence of the specific virus receptor. Here, we show for the human pathogen poliovirus that naturally nonpermissive murine APC acquire viral RNA in vivo independently of the cellular virus receptor. Uptake of poliovirus or polioviral RNA initiated neosynthesis of viral antigen to an extent sufficient to prime CTLs in vivo, which were detectable 2-3 wk after infection. Our results do not only indicate that experiments studying cross-presentation and cross-priming by using potentially amplifiable or translatable materials need careful examination, but they also question the general biological importance of cross-presentation and cross-priming in antiviral CTL responses.