Langerin expressing cells promote skin immune responses under defined conditions

There are conflicting data in the literature regarding the role of epidermal Langerhans cells (LC) in promoting skin immune responses. On one hand, LC can be extremely potent APCs in vitro, and are thought to be involved in contact hypersensitivity (CHS). On the other hand, it seems counterintuitive that a cell type continually exposed to pathogens at the organism's barrier surfaces should readily trigger potent T cell responses. Indeed, LC depletion in one model led to enhanced contact hypersensitivity, suggesting they play a negative regulatory role. However, apparently similar LC depletion models did not show enhanced CHS, and in one case showed reduced CHS. In this study we found that acute depletion of mouse LC reduced CHS, but the timing of toxin administration was critical: toxin administration 3 days before priming did not impair CHS, whereas toxin administration 1 day before priming did. We also show that LC elimination reduced the T cell response to epicutaneous immunization with OVA protein Ag. However, this reduction was only observed when OVA was applied on the flank skin, and not on the ear. Additionally, peptide immunization was not blocked by depletion, regardless of the site. Finally we show that conditions which eliminate epidermal LC but spare other Langerin(+) DC do not impair the epicutaneous immunization response to OVA. Overall, our results reconcile previous conflicting data in the literature, and suggest that Langerin(+) cells do promote T cell responses to skin Ags, but only under defined conditions.     

Multiple dendritic cell populations activate CD4+ T cells after viral stimulation

Dendritic cells (DC) are a heterogeneous cell population that bridge the innate and adaptive immune systems. CD8alpha DC play a prominent, and sometimes exclusive, role in driving amplification of CD8(+) T cells during a viral infection. Whether this reliance on a single subset of DC also applies for CD4(+) T cell activation is unknown. We used a direct ex vivo antigen presentation assay to probe the capacity of flow cytometrically purified DC populations to drive amplification of CD4(+) and CD8(+) T cells following infection with influenza virus by different routes. This study examined the contributions of non-CD8alpha DC populations in the amplification of CD8(+) and CD4(+) T cells in cutaneous and systemic influenza viral infections. We confirmed that in vivo, effective immune responses for CD8(+) T cells are dominated by presentation of antigen by CD8alpha DC but can involve non-CD8alpha DC. In contrast, CD4(+) T cell responses relied more heavily on the contributions of dermal DC migrating from peripheral lymphoid tissues following cutaneous infection, and CD4 DC in the spleen after systemic infection. CD4(+) T cell priming by DC subsets that is dependent upon the route of administration raises the possibility that vaccination approaches could be tailored to prime helper T cell immunity.     

CD4+ T cell responses elicited by different subsets of human skin migratory dendritic cells

Skin dendritic cells (DC) are professional APC critical for initiation and control of adaptive immunity. In the present work we have analyzed the CD4+ T cell stimulatory function of different subsets of DC that migrate spontaneously from human skin explants, including CD1a+CD14- Langerhans' cells (LC), CD1a-CD14- dermal DC (DDC), and CD1a-CD14+ LC precursors. Skin migratory DC consisted of APC at different stages of maturation-activation that produced IL-10, TGF-beta1, IL-23p19, and IL-12p40, but did not release IL-12p70 even after exposure to DC1-driving stimuli. LC and DDC migrated as mature/activated APC able to stimulate allogeneic naive CD4+ T cells and to induce memory Th1 cells in the absence of IL-12p70. The potent CD4+ T cell stimulatory function of LC and DDC correlated with their high levels of expression of MHC class II, adhesion, and costimulatory molecules. The Th1-biasing function of LC and DDC depended on their ability to produce IL-23. By contrast, CD1a-CD14+ LC precursors migrated as immature-semimature APC and were weak stimulators of allogeneic naive CD4+ T cells. However, and opposite of a potential tolerogenic role of immature DC, the T cell allostimulatory and Th1-biasing function of CD14+ LC precursors increased significantly by augmenting their cell number, prolonging the time of interaction with responding T cells, or addition of recombinant human IL-23 in MLC. The data presented in this study provide insight into the function of the complex network of skin-resident DC that migrate out of the epidermis and dermis after cutaneous immunizations, pathogen infections, or allograft transplantation.     

Characterization of dendritic cells subpopulations in skin and afferent lymph in the swine model

Transcutaneous delivery of vaccines to specific skin dendritic cells (DC) subsets is foreseen as a promising strategy to induce strong and specific types of immune responses such as tolerance, cytotoxicity or humoral immunity. Because of striking histological similarities between human and pig skin, pig is recognized as the most suitable model to study the cutaneous delivery of medicine. Therefore improving the knowledge on swine skin DC subsets would be highly valuable to the skin vaccine field. In this study, we showed that pig skin DC comprise the classical epidermal langerhans cells (LC) and dermal DC (DDC) that could be divided in 3 subsets according to their phenotypes: (1) the CD163(neg)/CD172a(neg), (2) the CD163(high)CD172a(pos) and (3) the CD163(low)CD172a(pos) DDC. These subtypes have the capacity to migrate from skin to lymph node since we detected them in pseudo-afferent lymph. Extensive phenotyping with a set of markers suggested that the CD163(high) DDC resemble the antibody response-inducing human skin DC/macrophages whereas the CD163(neg)CD172(low) DDC share properties with the CD8(+) T cell response-inducing murine skin CD103(pos) DC. This work, by showing similarities between human, mouse and swine skin DC, establishes pig as a model of choice for the development of transcutaneous immunisation strategies targeting DC.     

Inducible ablation of mouse Langerhans cells diminishes but fails to abrogate contact hypersensitivity

Langerhans cells (LC) form a unique subset of dendritic cells (DC) in the epidermis but so far their in vivo functions in skin immunity and tolerance could not be determined, in particular in relation to dermal DC (dDC). Here, we exploit a novel diphtheria toxin (DT) receptor (DTR)/DT-based system to achieve inducible ablation of LC without affecting the skin environment. Within 24 h after intra-peritoneal injection of DT into Langerin-DTR mice LC are completely depleted from the epidermis and only begin to return 4 wk later. LC deletion occurs by apoptosis in the absence of inflammation and, in particular, the dDC compartment is not affected. In LC-depleted mice contact hypersensitivity (CHS) responses are significantly decreased, although ear swelling still occurs indicating that dDC can mediate CHS when necessary. Our results establish Langerin-DTR mice as a unique tool to study LC function in the steady state and to explore their relative importance compared with dDC in orchestrating skin immunity and tolerance.     

Lymphoid organ dendritic cells: beyond the Langerhans cells paradigm

The immune system has developed mechanisms to detect and initiate responses to a continual barrage of immunological challenges. Dendritic cells (DC), a heterogeneous population of leucocytes, play a major role as immunosurveillance agents. To accomplish this function, DC are equipped with highly efficient mechanisms to detect pathogens, to capture, process and present antigens, and to initiate T-cell responses. These mechanisms are developmentally regulated during the DC life cycle in a process termed 'maturation', which was originally defined using Langerhans cells (LC), a DC type of the epidermis. LC exist in the skin in an immature state dedicated to capturing antigens, and in the subcutaneous lymph nodes in a mature state dedicated to presenting those antigens to T cells. The phenotypic changes undergone by LC during maturation, and the correlation of these changes with tissue localization, have been generally considered a paradigm for all DC. However, studies of the multiple DC types found in the lymphoid organs of mice and humans have revealed that most DC subsets do not follow the life cycle typified by LC. In this review we discuss the limitations of the 'LC paradigm' and suggest that this model should be revised to accommodate the heterogeneity of the DC system. We also discuss the implications of the maturational status of the DC subsets contained in the lymphoid organs for their putative roles in the induction of immune responses and the maintenance of peripheral tolerance.     

Burn injury suppresses human dermal dendritic cell and Langerhans cell function

Human skin contains epidermal Langerhans cells (LCs) and dermal dendritic cells (DCs) that are key players in induction of adaptive immunity upon infection. After major burn injury, suppressed adaptive immunity has been observed in patients. Here we demonstrate that burn injury affects adaptive immunity by altering both epidermal LC and dermal DC functions. We developed a human ex vivo burn injury model to study the function of DCs in thermally injured skin. No differences were observed in the capacity of both LCs and dermal DCs to migrate out of burned skin compared to unburned skin. Similarly, expression levels of co-stimulatory molecules were unaltered. Notably, we observed a strong reduction of T cell activation induced by antigen presenting cell (APC) subsets that migrated from burned skin through soluble burn factors. Further analyses demonstrated that both epidermal LCs and dermal DCs have a decreased T cell stimulatory capacity after burn injury. Restoring the T cell stimulatory capacity of DC subsets might improve tissue regeneration in patients with burn wounds.     

Impaired Humoral Immunity and Tolerance in K14-VEGFR-3-Ig Mice That Lack Dermal Lymphatic Drainage

Lymphatic vessels transport interstitial fluid, soluble Ag, and immune cells from peripheral tissues to lymph nodes (LNs), yet the contribution of peripheral lymphatic drainage to adaptive immunity remains poorly understood. We examined immune responses to dermal vaccination and contact hypersensitivity (CHS) challenge in K14-VEGFR-3-Ig mice, which lack dermal lymphatic capillaries and experience markedly depressed transport of solutes and dendritic cells from the skin to draining LNs. In response to dermal immunization, K14-VEGFR-3-Ig mice produced lower Ab titers. In contrast, although delayed, T cell responses were robust after 21 d, including high levels of Ag-specific CD8(+) T cells and production of IFN-γ, IL-4, and IL-10 upon restimulation. T cell-mediated CHS responses were strong in K14-VEGFR-3-Ig mice, but importantly, their ability to induce CHS tolerance in the skin was impaired. In addition, 1-y-old mice displayed multiple signs of autoimmunity. These data suggest that lymphatic drainage plays more important roles in regulating humoral immunity and peripheral tolerance than in effector T cell immunity.     

Th17 cells and activated dendritic cells are increased in vitiligo lesions

Background

Vitiligo is a common skin disorder, characterized by progressive skin de-pigmentation due to the loss of cutaneous melanocytes. The exact cause of melanocyte loss remains unclear, but a large number of observations have pointed to the important role of cellular immunity in vitiligo pathogenesis.

Methodology/Principal Findings

In this study, we characterized T cell and inflammation-related dermal dendritic cell (DC) subsets in pigmented non-lesional, leading edge and depigmented lesional vitiligo skin. By immunohistochemistry staining, we observed enhanced populations of CD11c+ myeloid dermal DCs and CD207+ Langerhans cells in leading edge vitiligo biopsies. DC-LAMP+ and CD1c+ sub-populations of dermal DCs expanded significantly in leading edge and lesional vitiligo skin. We also detected elevated tissue mRNA levels of IL-17A in leading edge skin biopsies of vitiligo patients, as well as IL-17A positive T cells by immunohistochemistry and immunofluorescence. Langerhans cells with activated inflammasomes were also noted in lesional vitiligo skin, along with increased IL-1ß mRNA, which suggest the potential of Langerhans cells to drive Th17 activation in vitiligo.

Conclusions/Significance

These studies provided direct tissue evidence that implicates active Th17 cells in vitiligo skin lesions. We characterized new cellular immune elements, in the active margins of vitiligo lesions (e.g. populations of epidermal and dermal dendritic cells subsets), which could potentially drive the inflammatory responses.     

Calcitonin gene-related peptide biases Langerhans cells toward Th2-type immunity

Langerhans cells (LC) are epidermal dendritic cells capable, in several experimental systems, of Ag-presentation for stimulation of cell-mediated immunity. LC have been considered to play a key role in initiation of cutaneous immune responses. Additionally, administration of donor T cells to bone marrow chimeric mice with persistent host LC, but not mice whose LC have been replaced by donor cells, exhibit marked skin graft-vs-host disease, demonstrating that LC can trigger graft-vs-host disease. However, experiments with transgenic mice in which regulatory elements from human langerin were used to drive expression of diphtheria toxin, resulting in absence of LC, suggest that LC may serve to down-regulate cutaneous immunity. LC are associated with nerves containing the neuropeptide calcitonin gene-related peptide (CGRP), and CGRP inhibits LC Ag-presentation in several models including presentation to a Th1 clone. We now report that CGRP enhances LC function for stimulation of Th2 responses. CGRP exposure enhanced LC Ag presentation to a Th2 clone. Upon presentation of chicken OVA by LC to T cells from DO11.10 chicken OVA TCR transgenic mice, pretreatment with CGRP resulted in increased IL-4 production and decreased IFN-gamma production. CGRP also inhibited stimulated production of the Th1 chemokines CXCL9 and CXCL10 but induced production of the Th2 chemokines CCL17 and CCL22 by a dendritic cell line and by freshly obtained LC. Changes in production of these chemokines correlated with the effect of CGRP on mRNA levels for these factors. Exposure of LC to nerve-derived CGRP in situ may polarize them toward favoring Th2-type immunity.     

Murine spleen contains a diversity of myeloid and dendritic cells distinct in antigen presenting function

The spleen contains multiple subsets of myeloid and dendritic cells (DC). DC are important antigen presenting cells (APC) which induce and control the adaptive immune response. They are cells specialized for antigen capture, processing and presentation to naïve T cells. However, DC are a heterogeneous population and each subset differs subtly in phenotype, function and location. Similarly, myeloid cell subsets can be distinguished which can also play an important role in the regulation of immunity. This review aims to characterize splenic subsets of DC and myeloid cells to better understand their individual roles in the immune response.     

Significant virus replication in Langerhans cells following application of HIV to abraded skin: relevance to occupational transmission of HIV

The cellular events that occur following occupational percutaneous exposure to HIV have not been defined. In this study, we studied relevant host cellular and molecular targets used for acquisition of HIV infection using split-thickness human skin explants. Blockade of CD4 or CCR5 before R5 HIV application to the epithelial surface of skin explants completely blocked subsequent HIV transmission from skin emigrants to allogeneic T cells, whereas preincubation with C-type lectin receptor inhibitors did not. Immunomagnetic bead depletion studies demonstrated that epithelial Langerhans cells (LC) accounted for >95% of HIV dissemination. When skin explants were exposed to HIV variants engineered to express GFP during productive infection, GFP+ T cells were found adjacent to GFP+ LC. In three distinct dendritic cell (DC) subsets identified among skin emigrants (CD1a+langerin+DC-specific intercellular adhesion molecule grabbing non-integrin (SIGN)- LC, CD1a+langerin-DC-SIGN- dermal DC, and CD1a-langerin-DC-SIGN+ dermal macrophages), HIV infection was detected only in LC. These results suggest that productive HIV infection of LC plays a critical role in virus dissemination from epithelium to cells located within subepithelial tissue. Thus, initiation of antiretroviral drugs soon after percutaneous HIV exposure may not prevent infection of LC, which is likely to occur rapidly, but may prevent or limit subsequent LC-mediated infection of T cells.     

Calcitonin gene-related peptide is an important regulator of cutaneous immunity: effect on dendritic cell and T cell functions

Some cutaneous inflammations are induced by percutaneous exposure to foreign Ags, and many chemical mediators regulate this inflammation process. One of these mediators, calcitonin gene-related peptide (CGRP), is a neuropeptide released from nerve endings in the skin. CGRP binds to its receptors composed of receptor activity-modifying protein 1 and calcitonin receptor-like receptor to modulate immune cell function. We show that CGRP regulates skin inflammation under physiological conditions, using contact hypersensitivity (CHS) models of receptor activity-modifying protein 1-deficient mice. CGRP has different functions in CHS responses mediated by Th1 or Th2 cells; it inhibits Th1-type CHS, such as 2,4,6-trinitrochlorobenzene-induced CHS, but promotes Th2-type CHS, such as FITC-induced CHS. CGRP inhibits the migration of Langerin(+) dermal dendritic cells to the lymph nodes in 2,4,6-trinitrochlorobenzene-induced CHS, and upregulates IL-4 production of T cells in the draining lymph nodes in FITC-CHS. These findings suggest that CGRP regulates several types of CHS reactions under physiological conditions and plays an important role in cutaneous immunity.     

Skin langerin+ dendritic cells transport intradermally injected anti-DEC-205 antibodies but are not essential for subsequent cytotoxic CD8+ T cell responses

Incorporation of Ags by dendritic cells (DCs) increases when Ags are targeted to endocytic receptors by mAbs. We have previously demonstrated in the mouse that mAbs against C-type lectins administered intradermally are taken up by epidermal Langerhans cells (LCs), dermal Langerin(neg) DCs, and dermal Langerin(+) DCs in situ. However, the relative contribution of these skin DC subsets to the induction of immune responses after Ag targeting has not been addressed in vivo. We show in this study that murine epidermal LCs and dermal DCs transport intradermally injected mAbs against the lectin receptor DEC-205/CD205 in vivo. Skin DCs targeted in situ with mAbs migrated through lymphatic vessels in steady state and inflammation. In the skin-draining lymph nodes, targeting mAbs were found in resident CD8α(+) DCs and in migrating skin DCs. More than 70% of targeted DCs expressed Langerin, including dermal Langerin(+) DCs and LCs. Numbers of targeted skin DCs in the nodes increased 2-3-fold when skin was topically inflamed by the TLR7 agonist imiquimod. Complete removal of the site where OVA-coupled anti-DEC-205 had been injected decreased endogenous cytotoxic responses against OVA peptide-loaded target cells by 40-50%. Surprisingly, selective ablation of all Langerin(+) skin DCs in Langerin-DTR knock-in mice did not affect such responses independently of the adjuvant chosen. Thus, in cutaneous immunization strategies where Ag is targeted to DCs, Langerin(+) skin DCs play a major role in transport of anti-DEC-205 mAb, although Langerin(neg) dermal DCs and CD8α(+) DCs are sufficient to subsequent CD8(+) T cell responses.     

Dermal fibroblasts induce maturation of dendritic cells

To trigger an effective T cell-mediated immune response in the skin, cutaneous dendritic cells (DC) migrate into locally draining lymph nodes, where they present Ag to naive T cells. Little is known about the interaction of DC with the various cellular microenvironments they encounter during their migration from the skin to lymphoid tissues. In this study, we show that human DC generated from peripheral blood monocytes specifically interact with human dermal fibroblasts via the interaction of beta(2) integrins on DC with Thy-1 (CD90) and ICAM-1 on fibroblasts. This induced the phenotypic maturation of DC reflected by expression of CD83, CD86, CD80, and HLA-DR in a TNF-alpha- and ICAM-1-dependent manner. Moreover, fibroblast-matured DC potently induced T cell activation reflected by CD25 expression and enhanced T cell proliferation. Together these data demonstrate that dermal fibroblasts that DC can encounter during their trafficking from skin to lymph node can act as potent regulators of DC differentiation and function, and thus may actively participate in the regulation and outcome of DC-driven cutaneous immune responses.     

CD4-8- dendritic cells prime CD4+ T regulatory 1 cells to suppress antitumor immunity

It is clear that dendritic cells (DCs) are essential for priming of T cell responses against tumors. However, the distinct roles DC subsets play in regulation of T cell responses in vivo are largely undefined. In this study, we investigated the capacity of OVA-presenting CD4-8-, CD4+8-, or CD4-8+ DCs (OVA-pulsed DC (DC(OVA))) in stimulation of OVA-specific T cell responses. Our data show that each DC subset stimulated proliferation of allogeneic and autologous OVA-specific CD4+ and CD8+ T cells in vitro, but that the CD4-8- DCs did so only weakly. Both CD4+8- and CD4-8+ DC(OVA) induced strong tumor-specific CD4+ Th1 responses and fully protective CD8+ CTL-mediated antitumor immunity, whereas CD4-8- DC(OVA), which were less mature and secreted substantial TGF-beta upon coculture with TCR-transgenic OT II CD4+ T cells, induced the development of IL-10-secreting CD4+ T regulatory 1 (Tr1) cells. Transfer of these Tr1 cells, but not T cells from cocultures of CD4-8- DC(OVA) and IL-10-/- OT II CD4+ T cells, into CD4-8+ DC(OVA)-immunized animals abrogated otherwise inevitable development of antitumor immunity. Taken together, CD4-8- DCs stimulate development of IL-10-secreting CD4+ Tr1 cells that mediated immune suppression, whereas both CD4+8- and CD4-8+ DCs effectively primed animals for protective CD8+ CTL-mediated antitumor immunity.     

Compartmental imbalance and aberrant immune function of blood CD123+ (plasmacytoid) and CD11c+ (myeloid) dendritic cells in atopic dermatitis

Atopic dermatitis (AD) is a pruritic, chronically relapsing skin disease in which Th2 cells play a crucial role in cutaneous and extracutaneous immune reactions. In humans, CD11c+CD123- myeloid dendritic cells (mDC) and CD11c-CD123+ plasmacytoid DC (pDC) orchestrate the decision-making process in innate and acquired immunity. Since the number and function of these blood dendritic cell (DC) subsets reportedly reflect the host immune status, we studied the involvement of the DC subsets in the pathogenesis of AD. Patients with AD had an increased DC number and a low mDC:pDC ratio with pDC outnumbering mDC in the peripheral blood compared with normal subjects and psoriasis patients (a Th1 disease model group). The mDC:pDC ratio was correlated with the total serum IgE level, the ratio of IFN-gamma-producing blood cells:IL-4-producing blood cells, and the disease severity. In vitro allogeneic stimulation of naive CD4+ cells with atopic DC showed that the ability of pDC for Th1 induction was superior or comparable to that of mDC. In skin lesions, pDC infiltration was in close association with blood vessels expressing peripheral neural addressins. Therefore, compartmental imbalance and aberrant immune function of the blood DC subsets may deviate the Th1/Th2 differentiation and thus induce protracted allergic responses in AD.     

Dendritic cells of mucosa and skin: "recruited for vaccination"

Mucosae and skin are exposed to environmental antigens and are natural entry routes for most infectious agents. To maintain immunological tolerance and ensure protective immunity against pathogens, epithelial surfaces are surveyed permanently by antigen-presenting dendritic cells (DCs). Many DC subsets have been described in epithelial tissues, depending on the inflammatory state and the type of epithelium. Identification of the DC subset able to induce cytotoxic CD8+ T cells against antigens delivered via mucosae or skin, is a major issue for the development of efficient anti-infectious and anti-tumoral vaccines. Until recently, it was commonly accepted that Langerhans cells (LC), the prototype of immature DCs residing in skin and certain mucosae, can capture and process antigens and, in response to danger signals, undergo a maturation program allowing their migration to the draining lymph nodes for priming of na?ve T cells. This concept likely needs to be revisited. Recent evidence from animal models revealed that resident epithelial tissue DCs, including LCs, do not play a direct role in T cell priming, but may contribute to maintenance of peripheral tolerance. Alternatively, DCs newly recruited into muco-cutaneous tissues exposed to pro-inflammatory stimuli are responsible for efficient priming and differentiation of CD8+ T cells into cytolytic effectors. These DC originate from blood monocytes and can cross-present protein antigens to CD8+ T cells, which subsequently give rise to specific CTL effectors. Remarkably, components derived from bacteria, virus and chemicals capable to enhance CCL20 production in epithelia, promote CCR6-dependent DC recruitment and behave as adjuvants allowing for cross-primed CD8+ CTL. These advances in the dynamic and function of epithelial tissue DC provide a rationale for the screening of novel CD8+ T cell adjuvants and the design of novel mucosal and skin vaccines.     

Protective immunity to UV radiation-induced skin tumours induced by skin grafts and epidermal cells

There is little evidence that cutaneous dendritic cells (DC), including epidermal Langerhans cells (LC), can induce immunity to UV radiation (UVR)-induced skin tumours. Here, it is shown that cells within skin can induce protective antitumour immunity against a UVR-induced fibrosarcoma. Transplantation of the skin overlying subcutaneous tumours onto na?ve recipients could induce protective antitumour immunity, probably because the grafting stimulated the tumour Ag-loaded DC to migrate to local lymph nodes. This suggests that cutaneous APC can present tumour Ag to induce protective antitumour immunity. Previously, it has been shown that immunization of mice with MHC class II+ epidermal cells (EC) pulsed with tumour extracts could induce delayed-type hypersensitivity against tumour cells. Here, this same immunization protocol could induce protective immunity against a minimum tumorigenic dose of UVR-induced fibrosarcoma cells, but not higher doses. Epidermal cells obtained from semiallogeneic donors and pulsed with tumour extract could also induce protective immunity. However, presentation of BSA Ag from the culture medium was found to contribute to this result using semiallogeneic EC. The results suggest that LC overlying skin tumours may be able to induce protective immunity to UVR-induced tumours if stimulated to migrate from the skin.     

Mycophenolate mofetil impairs the maturation and function of murine dendritic cells

The immunosuppressive drug, mycophenolate mofetil (MMF), has been successfully introduced in allogeneic transplantation medicine and, more recently, in the treatment of autoimmune skin disorders. MMF inhibits lymphocyte proliferation via a blockade of the enzyme inosine 5'-monophosphate dehydrogenase, an enzyme on which lymphocytes solely depend to generate the purines necessary for DNA/RNA synthesis. To investigate the effects of MMF on cutaneous immune responses, a murine model of contact hypersensitivity (CHS) was used, with oxazolone or trinitrochlorobenzene as a contact allergen. Compared with the respective vehicle, i.p. applied MMF significantly inhibited the elicitation and, surprisingly, the induction of CHS responses. This prompted further studies into the effects of MMF on Ag presentation. Bone marrow-derived dendritic cells (DC) were cultured with GM-CSF and IL-4 in the presence of MMF and were tested for their Ag-presenting capacity. Sensitization and elicitation of CHS and delayed-type hypersensitivity responses by s. c. injected haptenated DC were reduced upon preincubation of DC with MMF. CHS responses were not impaired upon resensitization, indicating that MMF does not induce hapten-specific immunotolerance. In addition, MMF decreased the ability of DC to stimulate allogeneic T cells in MLR assays. Accordingly, flow cytometric analyses revealed a dose-dependent reduction of the expression of CD40, CD80, CD86, I-A, and ICAM-1 on DC with a concurrent reduction of IL-12 production. These data suggest that MMF, in addition to affecting T lymphocytes, directly affects APC, resulting in an impairment of immune responses. They furthermore point to a possible role of inosine 5'-monophosphate dehydrogenase in the maturation of DC.