The fungal pathogen Candida albicans has a multilayered cell wall composed of an outer layer of proteins glycosylated with N-or O-linked mannosyl residues and an inner skeletal layer of β-glucans and chitin. We demonstrate that cytokine production by human mononuclear cells or murine macrophages was markedly reduced when stimulated by C. albicans mutants defective in mannosylation. Recognition of mannosyl residues was mediated by mannose receptor binding to N-linked mannosyl residues and by TLR4 binding to O-linked mannosyl residues. Residual cytokine production was mediated by recognition of β-glucan by the dectin-1/ TLR2 receptor complex. C. albicans mutants with a cell wall defective in mannosyl residues were less virulent in experimental disseminated candidiasis and elicited reduced cytokine production in vivo. We concluded that recognition of C. albicans by monocytes/macrophages is mediated by 3 recognition systems of differing importance, each of which senses specific layers of the C. albicans cell wall.

Dendritic cells (DC) that express the type II C-type lectin DC-SIGN (CD209) are located in the submucosa of tissues, where they mediate HIV-1 entry. Interestingly, the pathogen Candida albicans, the major cause of hospital-acquired fungal infections, penetrates at similar submucosal sites. Here we demonstrate that DC-SIGN is able to bind C. albicans both in DC-SIGN-transfected cell lines and in human monocyte-derived DC. The binding was shown to be time-as well as concentration-dependent, and live as well as heat-inactivated C. albicans were bound to the same extent. Moreover, in immature DC, DC-SIGN was able to internalize C. albicans in specific DC-SIGN-enriched vesicles, distinct from those containing the mannose receptor, the other known C. albicans receptor expressed by DC. Together, these results demonstrate that DC-SIGN is an exquisite pathogen-uptake receptor that captures not only viruses but also fungi.

Current dendritic cell (DC)-based vaccines are based on ex vivo-generated autologous DCs loaded with antigen prior to readministration into patients. A more direct and less laborious strategy is to target antigens to DCs in vivo via specific surface receptors. Therefore, we developed a humanized antibody, hD1V1G2/G4 (hD1), directed against the C-type lectin DC-specific intercellular adhesion molecule 3-grabbing nonintegrin (DC-SIGN) to explore its capacity to serve as a target receptor for vaccination purposes. hD1 was cross-linked to a model antigen, keyhole limpet hemocyanin (KLH). We observed that the chimeric antibody-protein complex (hD1-KLH) bound specifically to DC-SIGN and was rapidly internalized and translocated to the lysosomal compartment. To determine the targeting efficiency of hD1-KLH, monocyte-derived DCs and peripheral blood lymphocytes (PBLs) were obtained from patients who had previously been vaccinated with KLHpulsed DCs. Autologous DCs pulsed with hD1-KLH induced proliferation of patient PBLs at a 100-fold lower concentration than KLH-pulsed DCs. In addition, hD1- KLH-targeted IntroductionDendritic cells (DCs) are professional antigen-presenting cells (APCs) that play a key role in regulating antigen-specific immunity. DCs capture antigens, process them into peptides, and present these to T cells. 1 The interaction between DC and T-cell controls the type and magnitude of the resulting immune response. Recently, preclinical and clinical studies have exploited DCs in an attempt to improve vaccine efficacy. 2 Most of these studies involve ex vivo antigen loading of autologous monocyte-derived DCs that are readministrated to the patient, a laborious and costly procedure. A more direct strategy involves targeting of antigens specifically to antigen uptake receptors on the DC in vivo. Potential candidate receptors highly expressed by DCs include Fc receptors [3][4][5] and members of the C-type lectin family. 6,7 Whereas Fc receptors are expressed by many different cell types, the expression of some members of the C-type lectin family are more DC restricted. 8 C-type lectins bind sugar residues in a calcium-dependent manner via a highly conserved carbohydrate recognition domain. C-type lectin receptors expressed by DCs are implicated in immunoregulatory processes, such as antigen capture, DC trafficking, and DC-T-cell interactions. 8 Based on the location of the amino (N) terminus, 2 types of membrane-bound C-type lectins can be distinguished on DCs. Type I C-type lectins have their N terminus located outside, while type II C-type lectins have their N terminus located inside the cell. Several studies have been conducted on antigen targeting to C-type lectin receptors for vaccination purposes, mainly focusing on the type I C-type lectins mannose receptor (MR) 9 and DEC-205. 6,10,11 Vaccines based on natural MR ligands have been shown to effectively induce humoral and cellular responses. 9 However, these ligands lack specificity for the MR, and may target multiple lectins with overlapping binding sp...

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