A common pathway for dendritic cell and early B cell development

B cells and dendritic cells (DCs) each develop from poorly described progenitor cells in the bone marrow (BM). Although a subset of DCs has been proposed to arise from lymphoid progenitors, a common developmental pathway for B cells and BM-derived DCs has not been clearly identified. To address this possibility, we performed a comprehensive analysis of DC differentiative potential among lymphoid and B lymphoid progenitor populations in adult mouse BM. We found that both the common lymphoid progenitors (CLPs), shown here and elsewhere to give rise exclusively to lymphocytes, and a down-stream early B-lineage precursor population devoid of T and NK cell precursor potential each give rise to DCs when exposed to the appropriate cytokines. This result contrasts with more mature B-lineage precursors, all of which failed to give rise to detectable numbers of DCs. Significantly, both CLP and early B-lineage-derived DCs acquired several surface markers associated with functional DCs, and CLP-derived DCs readily induced proliferation of allogeneic CD4(+) T cells. Surprisingly, however, DC differentiation from both lymphoid-restricted progenitors was accompanied by up-regulation of CD11b expression, a cell surface molecule normally restricted to myeloid lineage cells including putative myeloid DCs. Together, these data demonstrate that loss of DC developmental potential is the final step in B-lineage commitment and thus reveals a previously unrecognized link between early B cell and DC ontogeny.     

Antitumor effect of intratumoral administration of dendritic cell combination with vincristine chemotherapy in a murine fibrosarcoma model

A new antitumor therapeutic strategy utilizing the combined effect of chemotherapy and DC (dendritic cell)-based immunotherapy was designed, and the effect of intratumoral injections of unpulsed, immature DCs was evaluated after in vivo pretreatment of vincristine on tumor growth in a murine fibrosarcoma tumor model. Vincristine exerted a much more potent apoptosis/necrosis-inducing effect on MCA-102 tumor cells than on DCs both in vitro and in vivo. Moreover, CD11c, CD40, CD80 and CD86 molecules on DCs were not downregulated after treatment with vincristine either in vitro or in vivo. The growth of tumor significantly regressed in the group which received the combined vincristine chemotherapy with intratumoral administration of DCs in contrast to the untreated group, the group treated with DCs alone, and the group treated with vincristine alone. In particular, an upregulated expression of CD40, CD80 and CD86 molecules on DCs was found in the combination treatment group. Furthermore, the number of CD4+ and CD8+ T cells and the staining intensity of their CD4 and CD8 surface molecules also increased after the combination treatment. Therefore, our results indicate the feasibility of this combination therapy with vincristine chemotherapy and DC-based immunotherapy as an efficient antitumor strategy for the treatment of fibrosarcoma.     

Human dendritic cells very efficiently present a heterologous antigen expressed on the surface of recombinant gram-positive bacteria to CD4+ T lymphocytes.

Recombinant Streptococcus gordonii expressing on the surface the C-fragment of tetanus toxin was tested as an Ag delivery system for human monocyte-derived dendritic cells (DCs). DCs incubated with recombinant S. gordonii were much more efficient than DCs pulsed with soluble C-fragment of tetanus toxin at stimulating specific CD4+ T cells as determined by cell proliferation and IFN-gamma release. Compared with DCs treated with soluble Ag, DCs fed with recombinant bacteria required 102- to 103-fold less Ag and were at least 102 times more effective on a per-cell basis for activating specific T cells. S. gordonii was internalized in DCs by conventional phagocytosis, and cytochalasin D inhibited presentation of bacteria-associated Ag, but not of soluble Ag, suggesting that phagocytosis was required for proper delivery of recombinant Ag. Bacteria were also very potent inducers of DC maturation, although they enhanced the capacity of DCs to activate specific CD4+ T cells at concentrations that did not stimulate DC maturation. In particular, S. gordonii dose-dependently up-regulated expression of membrane molecules (MHC I and II, CD80, CD86, CD54, CD40, CD83) and reduced both phagocytic and endocytic activities. Furthermore, bacteria promoted in a dose-dependent manner DC release of cytokines (IL-6, TNF-alpha, IL-1beta, IL-12, TGF-beta, and IL-10) and of the chemokines IL-8, RANTES, IFN-gamma-inducible protein-10, and monokine induced by IFN-gamma. Thus, recombinant Gram-positive bacteria appear a powerful tool for vaccine design due to their extremely high capacity to deliver Ags into DCs, as well as induce DC maturation and secretion of T cell chemoattractans.     

Induction of CD83+CD14+ nondendritic antigen-presenting cells by exposure of monocytes to IFN-alpha

IFN-alpha is a well-known agent for treatment of viral and malignant diseases. It has several modes of actions, including direct influence on the immune system. We investigated IFN-alpha effects on PBMC in terms of dendritic cell (DC) differentiation, as PBMC are exposed to high IFN-alpha levels during treatment of infections and cancers. We show that in vitro IFN-alpha exposure induced rapid and strong up-regulation of the DC-maturation markers CD80, CD86, and CD83 in bulk PBMC. Consistently, IFN-alpha induced up-regulation of these molecules on purified monocytes within 24 h. Up-regulation of CD80 and CD83 expression was IFN-alpha concentration-dependent. In contrast to GM-CSF + IL-4-generated DCs, most of the IFN-alpha-challenged CD83(+) cells coexpressed the monocyte marker CD14. Despite a typical mature DC immunophenotype, IFN-alpha-treated monocytes conserved phagocytic activity and never acquired a dendritic morphology. In mixed lymphocyte reactions IFN-alpha-treated monocytes were less potent than GM-CSF + IL-4-generated DCs but significantly more potent than untreated monocytes to induce T cell proliferation in bulk PBMC. However, only GM-CSF + IL-4-generated DCs were able to induce a significant proliferation of naive CD4(+) T cells. Notably, autologous memory CD4(+) T cells proliferated when exposed to tetanus toxoid-pulsed IFN-alpha-treated monocytes. At variance with untreated or GM-CSF + IL-4-exposed monocytes, those challenged with IFN-alpha showed long-lasting STAT-1 phosphorylation. Remarkably, CD83(+)CD14(+) cells were present in varicella skin lesions in close contact with IFN-alpha-producing cells. The present findings suggest that IFN-alpha alone promptly generates nondendritic APCs able to stimulate memory immune responses. This may represent an additional mode of action of IFN-alpha in vivo.     

Dendritic cell-based vaccines suppress metastatic liver tumor via activation of local innate and acquired immunity

Background  Dendritic cell (DC)-based vaccines have been applied clinically in the setting of cancer, but tumor-associated antigens (TAAs) have not yet been enough identified in various cancers. In this study, we investigated whether preventive vaccination with unpulsed DCs or peptide-pulsed DCs could offer anti-tumor effects against MC38 or BL6 liver tumors. Methods  Mice were subcutaneously (s.c.) immunized with unpulsed DCs or the recently defined TAA EphA2 derived peptide-pulsed dendritic cells (Eph-DCs) to treat EphA2-positive MC38 and EphA2-negative BL6 liver tumors. Liver mononuclear cells (LMNCs) from treated mice were subjected to ⁵¹Cr release assays against YAC-1 target cells. In some experiments, mice were injected with anti-CD8, anti-CD4 or anti-asialo GM1 antibody to deplete each lymphocyte subsets. Results  Immunization with unpulsed DCs displayed comparable efficacy against both MC38 and BL6 liver tumors when compared with Eph-DCs. Both DC-based vaccines significantly augmented the cytotoxicity of LMNCs against YAC-1 cells. In vivo antibody depletion studies revealed that NK cells, as well as, CD4+ and CD8+ T cells play critical roles in the anti-tumor efficacy associated with either DC-based modality. Tumor-specific cytotoxic T lymphocyte (CTL) activity was generally higher if mice had received Eph-DCs versus unpulsed DCs. Importantly, the mice that had been protected from MC38 liver tumor by either unpulsed DCs or Eph-DCs became resistant to s.c. MC38 rechallenge, but not to BL6 rechallenge. Conclusions  These results demonstrate that unpulsed DC vaccines might serve as an effective therapy for treating metastatic liver tumor, for which TAA has not yet been identified. Shinjiro Yamaguchi and Tomohide Tatsumi contributed equally to this work.     

Inhibition of myeloid dendritic cell accessory cell function and induction of T cell anergy by alcohol correlates with decreased IL-12 production

Alcohol consumption inhibits accessory cell function and Ag-specific T cell responses. Myeloid dendritic cells (DCs) coordinate innate immune responses and T cell activation. In this report, we found that in vivo moderate alcohol intake (0.8 g/kg of body weight) in normal volunteers inhibited DC allostimulatory capacity. Furthermore, in vitro alcohol treatment during DC differentiation significantly reduced allostimulatory activity in a MLR using naive CD4(+) T cells, and inhibited tetanus toxoid Ag presentation by DCs. Alcohol-treated DCs showed reduced IL-12, increased IL-10 production, and a decrease in expression of the costimulatory molecules CD80 and CD86. Addition of exogenous IL-12 and IL-2, but not neutralization of IL-10, during MLR ameliorated the reduced allostimulatory capacity of alcohol-treated DCs. Naive CD4(+) T cells primed with alcohol-treated DCs showed decreased IFN-gamma production that was restored by exogenous IL-12, indicating inhibition of Th1 responses. Furthermore, CD4(+) T cells primed with alcohol-treated DCs were hyporesponsive to subsequent stimulation with the same donor-derived normal DCs, suggesting the ability of alcohol-treated DCs to induce T cell anergy. LPS-induced maturation of alcohol-treated immature DCs partially restored the reduced allostimulatory activity, whereas alcohol given only during DC maturation failed to inhibit DC functions, suggesting that alcohol primarily impairs DC differentiation rather than maturation. NFkappaB activation, a marker of DC maturation was not affected by alcohol. Taken together, alcohol both in vitro and in vivo can impair generation of Th1 immune responses via inhibition of DC differentiation and accessory cell function through mechanisms that involve decreased IL-12 induction.     

Monocyte-derived dendritic cells induce a house dust mite-specific Th2 allergic inflammation in the lung of humanized SCID mice: involvement of CCR7

In rodents, airway dendritic cells (DCs) capture inhaled Ag, undergo maturation, and migrate to the draining mediastinal lymph nodes (MLN) to initiate the Ag-specific T cell response. However, the role of human DCs in the pathogenesis of the Th2 cell-mediated disease asthma remains to be clarified. Here, by using SCID mice engrafted with T cells from either house dust mite (HDM)-allergic patients or healthy donors, we show that DCs pulsed with Der p 1, one of the major allergens of HDM, and injected intratracheally into naive animals migrated into the MLN. In the MLN, Der p 1-pulsed DCs from allergic patients induced the proliferation of IL-4-producing CD4(+) T cells, whereas those from healthy donors induced IFN-gamma-secreting cells. In reconstituted human PBMC-reconstituted SCID mice primed with pulsed DCs from allergic patients, repeated exposure to aerosols of HDM induced 1) a strong pulmonary inflammatory reaction rich in T cells and eosinophils, 2) an increase in IL-4 and IL-5 production in the lung lavage fluid, and 3) increased IgE production compared with that in mice primed with unpulsed DCs. All these effects were reduced following in vivo neutralization of the CCR7 ligand secondary lymphoid tissue chemokine. These data in human PBMC-reconstituted SCID mice show that monocyte-derived DCs might play a key role in the pathogenesis of the pulmonary allergic response by inducing Th2 effector function following migration to the MLN.     

Abnormal priming of CD4(+) T cells by dendritic cells expressing hepatitis C virus core and E1 proteins

Patients infected with hepatitis C virus (HCV) have an impaired response against HCV antigens while keeping immune competence for other antigens. We hypothesized that expression of HCV proteins in infected dendritic cells (DC) might impair their antigen-presenting function, leading to a defective anti-HCV T-cell immunity. To test this hypothesis, DC from normal donors were transduced with an adenovirus coding for HCV core and E1 proteins and these cells (DC-CE1) were used to stimulate T lymphocytes. DC-CE1 were poor stimulators of allogeneic reactions and of autologous primary and secondary proliferative responses. Autologous T cells stimulated with DC-CE1 exhibited a pattern of incomplete activation characterized by enhanced CD25 expression but reduced interleukin 2 production. The same pattern of incomplete lymphocyte activation was observed in CD4(+) T cells responding to HCV core in patients with chronic HCV infection. However, CD4(+) response to HCV core was normal in patients who cleared HCV after alpha interferon therapy. Moreover, a normal CD4(+) response to tetanus toxoid was found in both chronic HCV carriers and patients who had eliminated the infection. Our results suggest that expression of HCV structural antigens in infected DC disturbs their antigen-presenting function, leading to incomplete activation of anti-HCV-specific T cells and chronicity of infection. However, presentation of unrelated antigens by noninfected DC would allow normal T-cell immunity to other pathogens.     

DC-SIGN and CD150 have distinct roles in transmission of measles virus from dendritic cells to T-lymphocytes

Measles virus (MV) is among the most infectious viruses that affect humans and is transmitted via the respiratory route. In macaques, MV primarily infects lymphocytes and dendritic cells (DCs). Little is known about the initial target cell for MV infection. Since DCs bridge the peripheral mucosal tissues with lymphoid tissues, we hypothesize that DCs are the initial target cells that capture MV in the respiratory tract and transport the virus to the lymphoid tissues where MV is transmitted to lymphocytes. Recently, we have demonstrated that the C-type lectin DC-SIGN interacts with MV and enhances infection of DCs in cis. Using immunofluorescence microscopy, we demonstrate that DC-SIGN+ DCs are abundantly present just below the epithelia of the respiratory tract. DC-SIGN+ DCs efficiently present MV-derived antigens to CD4+ T-lymphocytes after antigen uptake via either CD150 or DC-SIGN in vitro. However, DC-SIGN+ DCs also mediate transmission of MV to CD4+ and CD8+ T-lymphocytes. We distinguished two different transmission routes that were either dependent or independent on direct DC infection. DC-SIGN and CD150 are both involved in direct DC infection and subsequent transmission of de novo synthesized virus. However, DC-SIGN, but not CD150, mediates trans-infection of MV to T-lymphocytes independent of DC infection. Together these data suggest a prominent role for DCs during the initiation, dissemination, and clearance of MV infection.     

Mature dendritic cells derived from human monocytes within 48 hours: a novel strategy for dendritic cell differentiation from blood precursors

It is widely believed that generation of mature dendritic cells (DCs) with full T cell stimulatory capacity from human monocytes in vitro requires 5-7 days of differentiation with GM-CSF and IL-4, followed by 2-3 days of activation. Here, we report a new strategy for differentiation and maturation of monocyte-derived DCs within only 48 h of in vitro culture. Monocytes acquire immature DC characteristics by day 2 of culture with GM-CSF and IL-4; they down-regulate CD14, increase dextran uptake, and respond to the inflammatory chemokine macrophage inflammatory protein-1alpha. To accelerate DC development and maturation, monocytes were incubated for 24 h with GM-CSF and IL-4, followed by activation with proinflammatory mediators for another 24 h (FastDC). FastDC expressed mature DC surface markers as well as chemokine receptor 7 and secreted IL-12 (p70) upon CD40 ligation in the presence of IFN-gamma. The increase in intracellular calcium in response to 6Ckine showed that chemokine receptor 7 expression was functional. When FastDC were compared with mature monocyte-derived DCs generated by a standard 7-day protocol, they were equally potent in inducing Ag-specific T cell proliferation and IFN-gamma production as well as in priming autologous naive T cells using tetanus toxoid as a model Ag. These findings indicate that FastDC are as effective as monocyte-derived DCs in stimulating primary, Ag-specific, Th 1-type immune responses. Generation of FastDC not only reduces labor, cost, and time required for in vitro DC development, but may also represent a model more closely resembling DC differentiation from monocytes in vivo.     

CD40 ligand and CTLA-4 are reciprocally regulated in the Th1 cell proliferative response sustained by CD8(+) dendritic cells

Subsets of murine dendritic cells (DCs) from the spleen differ in their ability to induce proliferative responses in both primary and secondary CD4(+) T cells. Recent evidence indicates that lymphoid-related CD8(+) DCs fail to provide appropriate signals to freshly isolated secondary CD4(+) T cells to sustain their proliferation in vitro. In the present study, we examined peptide-pulsed CD8(-) and CD8(+) DCs for ability to stimulate Th1 and Th2 cell clones with the same Ag specificity. Defective ability to induce proliferation was selectively shown by CD8(+) DCs presenting Ag to the Th1 clone. The deficiency in CD8(+) DCs was overcome by CD40 triggering before peptide pulsing. When exposed to CD8(+) DCs in the absence of CD40 activation, the Th1 clone expressed low levels of CD40 ligand and high levels of surface CTLA-4. Neutralization of CTLA-4 during the DC/T cell coculture resulted in increased CD40 ligand expression and proliferation of T cells. Remarkably, the activation of CD40 on DCs under conditions that would increase Th1 cell proliferation, also resulted in down-regulation of surface CTLA-4. These results confirm differential effects of CD8(+) and CD8(-) DCs in the stimulation of Ag-primed Th cells. In addition, they suggest that reciprocal regulation of CD40 ligand and CTLA-4 expression occurs in Th1 cells exposed to CD8(+) DCs.     

CD8alpha+ and CD11b+ dendritic cell-restricted MHC class II controls Th1 CD4+ T cell immunity

The activation, proliferation, differentiation, and trafficking of CD4 T cells is central to the development of type I immune responses. MHC class II (MHCII)-bearing dendritic cells (DCs) initiate CD4(+) T cell priming, but the relative contributions of other MHCII(+) APCs to the complete Th1 immune response is less clear. To address this question, we examined Th1 immunity in a mouse model in which I-A(beta)(b) expression was targeted specifically to the DCs of I-A(beta)b-/- mice. MHCII expression is reconstituted in CD11b(+) and CD8alpha(+) DCs, but other DC subtypes, macrophages, B cells, and parenchymal cells lack of expression of the I-A(beta)(b) chain. Presentation of both peptide and protein Ags by these DC subsets is sufficient for Th1 differentiation of Ag-specific CD4(+) T cells in vivo. Thus, Ag-specific CD4(+) T cells are primed to produce Th1 cytokines IL-2 and IFN-gamma. Additionally, proliferation, migration out of lymphoid organs, and the number of effector CD4(+) T cells are appropriately regulated. However, class II-negative B cells cannot receive help and Ag-specific IgG is not produced, confirming the critical MHCII requirement at this stage. These findings indicate that DCs are not only key initiators of the primary response, but provide all of the necessary cognate interactions to control CD4(+) T cell fate during the primary immune response.     

Flt3-ligand, IL-4, GM-CSF, and adherence-mediated isolation of murine lung dendritic cells: assessment of isolation technique on phenotype and function

Lung dendritic cells (DCs) are difficult to study due to their limited quantities and the complexities required for isolation. Although many procedures have been used to overcome this challenge, the effects of isolation techniques on lung DCs have not been reported. The current study shows that freshly isolated DCs (CD11c+) have limited ability to induce proliferation in allogeneic T cells, and are immature as indicated by low cell surface expression of costimulatory molecules compared with liver or splenic DCs. DCs isolated after overnight culture or from mice treated with Flt3L are phenotypically mature and potent stimulators of allogeneic T cells. DCs could not be propagated from lung mononuclear cells in response to IL-4 and GM-CSF. Contrary to data reported for nonpulmonary DCs, expression of CCR6 was decreased on mature lung DCs, and only a subset of mature DCs expressed higher levels of CCR7. Absence of CD8alpha expression indicates that freshly isolated DCs are myeloid-type, whereas mature DCs induced by overnight culture are both "lymphoid" (CD8alpha+) and "myeloid" (CD8alpha-). DCs from mice genetically deficient in CD8alpha expression were strong simulators of allogeneic T cells which was consistent with data showing that CD8alpha- DCs from CD8alpha-sufficient mice are better APCs compared with CD8alpha+ DCs from the same mice. These data show that freshly isolated lung DCs are phenotypically and functionally distinct, and that the isolation technique alters the biology of these cells. Therefore, lung DC phenotype and function must be interpreted relative to the technique used for isolation.     

Unique functions of CD11b+, CD8 alpha+, and double-negative Peyer's patch dendritic cells

We have recently demonstrated the presence of three populations of dendritic cells (DC) in the murine Peyer's patch. CD11b(+)/CD8alpha(-) (myeloid) DCs are localized in the subepithelial dome, CD11b(-)/CD8alpha(+) (lymphoid) DCs in the interfollicular regions, and CD11b(-)/CD8alpha(-) (double-negative; DN) DCs at both sites. We now describe the presence of a novel population of intraepithelial DN DCs within the follicle-associated epithelium and demonstrate a predominance of DN DCs only in mucosal lymphoid tissues. Furthermore, we demonstrate that all DC subpopulations maintain their surface phenotype upon maturation in vitro, and secrete a distinct pattern of cytokines upon exposure to T cell and microbial stimuli. Only myeloid DCs from the PP produce high levels of IL-10 upon stimulation with soluble CD40 ligand(-) trimer, or Staphylococcus aureus and IFN-gamma. In contrast, lymphoid and DN, but not myeloid DCs, produce IL-12p70 following microbial stimulation, whereas no DC subset produces IL-12p70 in response to CD40 ligand trimer. Finally, we show that myeloid DCs from the PP are particularly capable of priming naive T cells to secrete high levels of IL-4 and IL-10, when compared with those from nonmucosal sites, while lymphoid and DN DCs from all tissues prime for IFN-gamma production. These findings thus suggest that DC subsets within mucosal tissues have unique immune inductive capacities.     

Dendritic cell-induced activation of adaptive and innate antitumor immunity

While studying Ag-pulsed syngeneic dendritic cell (DC) immunization, we discovered that surprisingly, unpulsed DCs induced protection against tumor lung metastases resulting from i.v. injection of a syngeneic BALB/c colon carcinoma CT26 or a syngeneic C57BL/6 lung carcinoma LL/2. Splenocytes or immature splenic DCs did not protect. The protection was mediated by NK cells, in that it was abrogated by treatment with anti-asialo-GM1 but not anti-CD8, and was induced by CD1(-/-) DCs unable to stimulate NKT cells, but did not occur in beige mice lacking NK cells. Protection correlated with increased NK activity, and increased infiltration of NK but not CD8(+) cells in lungs of tumor-bearing mice. Protection depended on the presence of costimulatory molecules CD80, CD86, and CD40 on the DCs, but surprisingly did not require DCs that could make IL-12 or IL-15. Unexpectedly, protection sensitive to anti-asialo-GM1 and increased NK activity were still present 14 mo after DC injection. As NK cells lack memory, we found by depletion that CD4(+) not CD8(+) T cells were required for induction of the NK antitumor response. The role of DCs and CD4(+) T cells provides a novel mechanism for NK cell induction and innate immunity against cancer that may have potential in preventing clinical metastases.     

In vitro anti-tumor immune response induced by dendritic cells transfected with EBV-LMP2 recombinant adenovirus

Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) is a high-incidence tumor in southern China. Latent membrane proteins 2 (LMP2) is a subdominant antigen of EBV. The present study was to develop a dendritic cells (DCs)-based cancer vaccine (rAd-LMP2-DC) and to study its biological characteristics and its immune functions. Our results showed that LMP2 gene transfer did not alter the typical morphology of mature DC, and the representative phenotypes of mature DC (CD80, CD83, and CD86) were highly expressed in rAd-LMP2-DCs. The expression of LMP2 in rAd-LPM2-DCs was about 84.54%, which suggested efficient gene transfer. Transfected DCs markedly increased antigen-specific T-cell proliferation. The specific cytotoxicity against NPC cell was significantly higher than that in controls (p < 0.05), and enhanced with increased stimulations by transfected DCs. In addition, phenotypic analysis demonstrated that the LMP2-specific CTLs consisted of both CD4(+) and CD8(+) T cells. These results showed that development of DC-based vaccine by transfection with malignancy-associated virus antigens could elicit potent CTL response and provide a potential strategy of immunotherapy for EBV-associated NPC.     

Chronic helminth infection promotes immune regulation in vivo through dominance of CD11cloCD103- dendritic cells

Gastrointestinal helminth infections are extremely prevalent in many human populations and are associated with downmodulated immune responsiveness. In the experimental model system of Heligmosomoides polygyrus, a chronic infection establishes in mice, accompanied by a modulated Th2 response and increased regulatory T cell (Treg) activity. To determine if dendritic cell (DC) populations in the lymph nodes draining the intestine are responsible for the regulatory effects of chronic infection, we first identified a population of CD11c(lo) nonplasmacytoid DCs that expand after chronic H. polygyrus infection. The CD11c(lo) DCs are underrepresented in magnetic bead-sorted preparations and spared from deletion in CD11c-diptheria toxin receptor mice. After infection, CD11c(lo) DCs did not express CD8, CD103, PDCA, or Siglec-H and were poorly responsive to TLR stimuli. In DC/T cell cocultures, CD11c(lo) DCs from naive and H. polygyrus-infected mice could process and present protein Ag, but induced lower levels of Ag-specific CD4(+) T cell proliferation and effector cytokine production, and generated higher percentages of Foxp3(+) T cells in the presence of TGF-β. Treg generation was also dependent on retinoic acid receptor signaling. In vivo, depletion of CD11c(hi) DCs further favored the dominance of the CD11c(lo) DC phenotype. After CD11c(hi) DC depletion, effector responses were inhibited dramatically, but the expansion in Treg numbers after H. polygyrus infection was barely compromised, showing a significantly higher regulatory/effector CD4(+) T cell ratio compared with that of CD11c(hi) DC-intact animals. Thus, the proregulatory environment of chronic intestinal helminth infection is associated with the in vivo predominance of a newly defined phenotype of CD11c(lo) tolerogenic DCs.     

Measles virus induces abnormal differentiation of CD40 ligand-activated human dendritic cells

Measles virus (MV) infection induces a profound immunosuppression responsible for a high rate of mortality in malnourished children. MV can encounter human dendritic cells (DCs) in the respiratory mucosa or in the secondary lymphoid organs. The purpose of this study was to investigate the consequences of DC infection by MV, particularly concerning their maturation and their ability to generate CD8+ T cell proliferation. We first show that MV-infected Langerhans cells or monocyte-derived DCs undergo a maturation process similarly to the one induced by TNF-alpha or LPS, respectively. CD40 ligand (CD40L) expressed on activated T cells is shown to induce terminal differentiation of DCs into mature effector DCs. In contrast, the CD40L-dependent maturation of DCs is inhibited by MV infection, as demonstrated by CD25, CD69, CD71, CD40, CD80, CD86, and CD83 expression down-regulation. Moreover, the CD40L-induced cytokine pattern in DCs is modified by MV infection with inhibition of IL-12 and IL-1alpha/beta and induction of IL-10 mRNAs synthesis. Using peripheral blood lymphocytes from CD40L-deficient patients, we demonstrate that MV infection of DCs prevents the CD40L-dependent CD8+ T cell proliferation. In such DC-PBL cocultures, inhibition of CD80 and CD86 expression on DCs was shown to require both MV replication and CD40 triggering. Finally, for the first time, MV was shown to inhibit tyrosine-phosphorylation level induced by CD40 activation in DCs. Our data demonstrate that MV replication modifies CD40 signaling in DCs, thus leading to impaired maturation. This phenomenon could play a pivotal role in MV-induced immunosuppression.     

Distinct maturation of, but not migration between, human monocyte-derived dendritic cells upon ingestion of apoptotic cells of early or late phases

Cell death via apoptosis is a normal physiological process. Rapid, but silent, removal of apoptotic cells (ACs) plays an essential role in maintaining homeostasis in the immune system. Defective clearance of ACs allows ACs to accumulate and undergo late phase apoptosis, also known as secondary necrosis, which may generate danger signals, leading to inflammation or autoimmunity. In this study we investigate the outcome of dendritic cells (DCs), which are potent APCs, on the interaction with ACs of early or late phase. Immature DCs internalized ACs of both early and late phases with similar efficiency. However, DCs that had taken up ACs of early phase acquired a non-fully mature DC phenotype, expressing low MHC class II complex, costimulatory molecule CD40, and mature DC-restricted marker CD83, and had a low capacity to stimulate allogeneic CD4+ T cell proliferation, whereas DCs that had taken up ACs of late phase acquired a mature DC phenotype with enhanced T cell stimulatory capacity. Ingestion of either early or late ACs induced minimal production of IL-12 and modulated CC chemokine and CCR expression in DCs. In particular, there was down-regulation of CCR5 and up-regulation of CCR7, resulting in switches in responsiveness from inflammatory to lymphoid chemokines. We conclude from these data that after taking up ACs of either early or late phases, DCs acquire the capability of homing to draining lymph nodes, and the distinct maturation between DCs taking up early or late ACs may contribute to DC function in the induction of T cell tolerance or Ag-specific T cell response, respectively.     

In situ replication of immediate dendritic cell (DC) precursors contributes to conventional DC homeostasis in lymphoid tissue

The developmental biology of dendritic cells (DC) under physiological conditions remains unclear. In this study, we show that mouse CD11c(+) MHC class II(-)lineage(-) cells are immediate precursors of conventional DC and are widely distributed in both bone marrow and lymphoid tissues. These precursors have a high clonal efficiency, and when cocultured on a supportive stromal monolayer or adoptively transferred in vivo, generate a population CD11c(+)MHC class II(+) DC that retain limited proliferation capacity. During steady state conditions, a small proportion of immediate DC precursors (DCp) and DCs are dividing actively in bone marrow and spleen. Cytokines that initiate and support proliferation of immediate DCp were defined. Collectively, our findings provide evidence of a distinct development pathway for conventional DC in both bone marrow and lymphoid tissues and highlight the importance of in situ replication of immediate DCp and DC in maintaining conventional DC populations.