Presence Of Dendritic Cells Research Articles

Induced bronchus-associated lymphoid tissue serves as a general priming site for T cells and is maintained by dendritic cells

Mucosal vaccination via the respiratory tract can elicit protective immunity in animal infection models, but the underlying mechanisms are still poorly understood. We show that a single intranasal application of the replication-deficient modified vaccinia virus Ankara, which is widely used as a recombinant vaccination vector, results in prominent induction of bronchus-associated lymphoid tissue (BALT). Although initial peribronchiolar infiltrations, characterized by the presence of dendritic cells (DCs) and few lymphocytes, can be found 4 d after virus application, organized lymphoid structures with segregated B and T cell zones are first observed at day 8. After intratracheal application, in vitro–differentiated, antigen-loaded DCs rapidly migrate into preformed BALT and efficiently activate antigen-specific T cells, as revealed by two-photon microscopy. Furthermore, the lung-specific depletion of DCs in mice that express the diphtheria toxin receptor under the control of the CD11c promoter interferes with BALT maintenance. Collectively, these data identify BALT as tertiary lymphoid structures supporting the efficient priming of T cell responses directed against unrelated airborne antigens while crucially requiring DCs for its sustained presence.

Functionally Mature CD4 and CD8 TCRαβ Cells Are Generated in OP9-DL1 Cultures from Human CD34+ Hematopoietic Cells

Human CD34(+) hematopoietic precursor cells cultured on delta-like ligand 1 expressing OP9 (OP9-DL1) stromal cells differentiate to T lineage cells. The nature of the T cells generated in these cultures has not been studied in detail. Since these cultures do not contain thymic epithelial cells which are the main cell type mediating positive selection in vivo, generation of conventional helper CD4(+) and cytotoxic CD8(+) TCRalphabeta cells is not expected. Phenotypically mature CD27(+)CD1(-) TCRgammadelta as well as TCRalphabeta cells were generated in OP9-DL1 cultures. CD8 and few mature CD4 single-positive TCRalphabeta cells were observed. Mature CD8 single-positive cells consisted of two subpopulations: one expressing mainly CD8alphabeta and one expressing CD8alphaalpha dimers. TCRalphabeta CD8alphaalpha and TCRgammadelta cells both expressed the IL2Rbeta receptor constitutively and proliferated on IL-15, a characteristic of unconventional T cells. CD8alphabeta(+) and CD4(+) TCRalphabeta cells were unresponsive to IL-15, but could be expanded upon TCR stimulation as mature CD8alphabeta(+) and CD4(+) T cells. These T cells had the characteristics of conventional T cells: CD4(+) cells expressed ThPOK, CD40L, and high levels of IL-2 and IL-4; CD8(+) cells expressed Eomes, Runx3, and high levels of granzyme, perforin, and IFN-gamma. Induction of murine or human MHC class I expression on OP9-DL1 cells had no influence on the differentiation of mature CD8(+) cells. Similarly, the presence of dendritic cells was not required for the generation of mature CD4(+) or CD8(+) T cells. These data suggest that positive selection of these cells is induced by interaction between T precursor cells.

In Vivo Confocal Microscopic Evaluation of Corneal Changes in Chronic Stevens-Johnson Syndrome and Toxic Epidermal Necrolysis

To describe corneal changes visible on in vivo confocal microscopy, in patients with debilitating ocular sequelae because of toxic epidermal necrolysis (TEN) or Stevens-Johnson syndrome (SJS). Forty-one eyes of 25 consecutive patients suffering from chronic TEN or SJS were studied using in vivo confocal microscopy. Severe dry eye syndrome with no associated limbal stem cell deficiency (25 eyes, 16 patients, 61%) was the most frequent clinical pattern. Limbal stem cell deficiency was noted in 16 eyes (12 patients, 39%). Three patients had asymmetric disease. Confocal microscopy showed a consistent change in the superficial epithelial cells in both clinical presentations. Patients with dry eye syndrome had frequent pathological nerve damages, and the presence of dendritic cells was prevalent (65%). Inflammatory cells were observed in a large number in 4 of the 12 patients presenting neovascularization of the cornea. The corneas of patients with chronic ocular sequelae linked to SJS and TEN present a number of abnormalities. In vivo confocal microscopy is a potentially useful tool for therapeutic indications and for follow-up of the debilitating chronic ocular problems associated with these diseases.

Intracerebral granulocyte‐macrophage colony‐stimulating factor induces functionally competent dendritic cells in the mouse brain

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a hematopoietic growth factor and a proinflammatory cytokine. While GM-CSF is lacking in normal brain tissue, it is expressed under pathological conditions and correlates with the presence of dendritic cells (DC). However, the role of GM-CSF for the onset of immune responses in the brain is still unclear. To analyze the role of GM-CSF for the induction and functional activity of immune cells in the brain, we performed chronic intracerebroventricular administration of GM-CSF to the brains of adult mice. After GM-CSF administration, intracerebral leukocytes (ICL) were characterized by means of flow cytometry, immunohistochemistry, and an ex vivo functional assay. GM-CSF treatment significantly increased the number of leukocytes expressing high levels of CD45, indicative of peripheral, blood-derived cells. The infiltrating cells were preferentially DC of the myeloid lineage (CD45(high) CD11c+ CD11b+) with an activated phenotype characterized by upregulated expression of MHCII and the costimulatory ligand CD80. Furthermore, DC from GM-CSF treated mice were fully competent to activate naive allogeneic T cells in a mixed leukocyte reaction. In contrast, intracerebroventricular IFN-gamma administration stimulated MHCII expression on cells resembling resident microglia, but did not induce comparable presence of DC. Taken together, intracerebroventricular GM-CSF treatment results in high numbers of DC in the brain. Moreover, these GM-CSF-induced DC display an activated phenotype and exhibit the capacity to act as fully competent DC even without a further inflammatory stimulus.

Requirements for Co-Stimulation in T-Cell Dependent and T-Cell Independent Re-Stimulation of FVIII-Specific Memory B Cells

Memory B cells specific for factor VIII (FVIII) are critical for maintaining FVIII inhibitors in patients with hemophilia A. They are precursors of anti-FVIII antibody-producing plasma cells and are highly efficient antigen-presenting cells for the activation of T cells. The eradication of FVIII-specific memory B cells will be a prerequisite for any successful new approach to induce immune tolerance in patients with FVIII inhibitors. Little is known about the regulation of these cells. Previously we showed that ligands for toll-like receptors (TLR) 7 and 9 are able to re-stimulate FVIII-specific memory B cells in the absence of T-cell help. However, alternative “helper cells” such as dendritic cells are essential for providing help to memory B cells under such conditions. Based on these findings, we asked which co-stimulatory interactions are required for the restimulation of memory B cells in the presence of dendritic cells and ligands for TLR and whether these co-stimulatory interactions are the same as those required for the restimulation of memory B cells in the presence of activated T cells. We used spleen cells from hemophilic mice treated with human FVIII to generate highly purified populations of memory B cells, CD4+ T cells and dendritic cells. The required purity was achieved by a combination of magnetic bead separation and fluorescence-activated cell sorting. The memory B cell compartment was specified by the expression of CD19 together with IgG and the absence of surface IgM and IgD. Memory B cells were cultured in the presence of FVIII to stimulate their differentiation into anti-FVIII antibody-producing plasma cells. Different combinations of CD4+ T cells, ligands for TLR 7 and 9 and dendritic cells were added to the memory-B-cell cultures. Blocking antibodies and competitor proteins were used to specify the co-stimulatory interactions required for the re-stimulation of memory B cells in the presence of either CD4+ T cells or dendritic cells and ligands for TLR 7 and 9. Our results demonstrate that the blockade of B7-1 and B7-2 as well as the blockade of CD40L inhibit the re-stimulation of FVIII-specific memory B cells and their differentiation into anti-FVIII antibody-producing plasma cells in the presence of T-cell help. Similar requirements apply for the re-stimulation of memory B cells in the presence of dendritic cells and ligands for TLR 7 or 9. Dendritic cells in the absence of ligands for TLR are not able to provide help for the re-stimulation of memory B cells, which indicates that dendritic cells need to be activated. Furthermore, ligands for TLR 7 or 9 were not able to re-stimulate memory B cells in the complete absence of dendritic cells. Based on these results we conclude that dendritic cells activated by ligands for TLR 7 or 9 can substitute for activated CD4+ T cells in providing co-stimulatory help for memory-B-cell re-stimulation. CD40-CD40L interactions seem to be the most important co-stimulatory interactions for the re-stimulation of memory B cells, not only in the presence of activated CD4+ T cells but also in the presence of ligands for TLR and dendritic cells.

The surface protein TIGIT suppresses T cell activation by promoting the generation of mature immunoregulatory dendritic cells

Here we have identified a surface protein, TIGIT, containing an immunoglobulin variable domain, a transmembrane domain and an immunoreceptor tyrosine-based inhibitory motif that was expressed on regulatory, memory and activated T cells. Poliovirus receptor, which is expressed on dendritic cells, bound TIGIT with high affinity. A TIGIT-Fc fusion protein inhibited T cell activation in vitro, and this was dependent on the presence of dendritic cells. The binding of poliovirus receptor to TIGIT on human dendritic cells enhanced the production of interleukin 10 and diminished the production of interleukin 12p40. Knockdown of TIGIT with small interfering RNA in human memory T cells did not affect T cell responses. TIGIT-Fc inhibited delayed-type hypersensitivity reactions in wild-type but not interleukin 10-deficient mice. Our data suggest that TIGIT exerts immunosuppressive effects by binding to poliovirus receptor and modulating cytokine production by dendritic cells.

Novel Heterozygous Mutation 572insA in TACI Identified in a Patient with Selective IgA Deficiency.

Selective IgA deficiency (SIgAD) is relatively common disease in patients with primary immunodeficiency. Recently, mutations in TNFRSF13B, encoding transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI), have been identified in 7~15% of patients with common variable immunodeficiency (CVID) and 10% of those with SIgAD. However, the role of TACI mutations in the molecular pathogenesis underlying SIgAD is still controversial and remains unsolved. We identified a novel heterozygous single adenine insertion, 572insA, in exon 4 of TNFRSF13B in a patient with SIgAD. Although the same mutation was detected in her mother and younger brother, their serum immunoglobulin levels were within normal range. This frameshift mutation produces a truncated form of TACI protein lacking majority of the intracellular domain. Peripheral blood B cells from the patient and her siblings with the TACI mutation were able to produce immunoglobulin without inferiority in the presence of dendritic cells, CD40L and IL-10. However, IgD+ or IgM+ B cells from the patient were impaired to produce Cα mature transcripts in response to APRIL, one of the ligands for TACI, reflecting impairment of class-switch recombination required for differentiation into IgA-producing cells. Thus, the T-cell independent B cell activation is impaired in the patient. We overexpressed cDNAs for wild-type (WT), C104R and 572insA mutant TACI in HEK 293 cells, and analyzed the effect on signal transduction through TACI. Immuno-precipitation analysis revealed that either 572insA or C104R mutant associated with WT TACI and formed an oligomeric complex. C104R TACI has been previously shown to exert a dominant-negative effect on ligand-induced TACI signaling, since it forms an oligomeric complex with WT TACI and disrupts ligand-induced receptor signaling. Similarly, since 572insA TACI lacks an intracellular domain that is important for signal transduction, it may exert a dominant-negative effect on ligand-induced TACI signaling. Although the heterozygous 572insA TACI mutation disrupts ligand-induced TACI signaling, not all individuals carrying the mutation display humoral immune deficiency. Thus, an additional genetic modifier and/or an environmental factor may be required for the development of immunological abnormalities due to TACI mutations.

The development of endometriosis is dependent on the presence of dendritic cells in mice

OBJECTIVE: Endometriosis affects 10-15% of women of reproductive age and is associated with pelvic pain and infertility. Recently, we have shown that dendritic cells (DC) supplementation led to the enhancement of endometriosis lesion growth and of intra-lesion angiogenesis. In this study, we investigated whether the development of endometriosis is dependent on the presence of endogenous DC.DESIGN: Prospective study using a surgical model of endometriosis in mice.MATERIALS AND METHODS: Surgical induction of endometriosis was performed in 8-10 week old female mice. Briefly, under anesthesia, a midline laparotomy was performed, uterine horns were removed, and 2-mm biopsy punch samples were sutured to the peritoneal wall. After 8 days, mice were killed, their abdomens were opened through the same incision and lesions were measured in two perpendicular diameters (D1 and D2) and the cross-sectional lesion area calculated using the formula D1xD2xπ/4. Single cell suspensions were analyzed by flow cytometry. In order to ablate DC, we utilized CD11c-DTR/GFP transgenic mice (C57Bl/6 background), in which DC can be ablated in vivo. Administration of diphtheria toxin (DT) into the transgenic mice that express the primate DT receptor (DTR), resulted in depletion of CD11c+DC. Endometriosis was induced in 10 CD11c-DTR/GFP transgenic mice and in 10 C57Bl/6 control mice. The mice were allocated to 4 groups (n=5 mice per group): I: C57Bl/6 treated with vehicle; II: C57Bl/6 treated with DT; III: CD11c-DTR/GFP transgenic mice treated with DT; IV: CD11c-DTR/GFP transgenic mice treated with vehicle. On day 0, endometriosis was induced. On days 2, 5, and 9, DT or vehicle was administered. All mice were sacrificed on day 10, the lesions were measured, and surgically removed from the abdominal cavity.RESULTS: In CD11c-DTR/GFP transgenic mice that were treated with DT (Group III) the endometriotic lesions were significantly smaller than that of all the other groups (ANOVA) (i.e. Group I: 5.25±1.18 mm2; Group II: 4.73±1.11 mm2; Group III: 1.09 ± 0.38 mm2; Group IV: 5.12±1.09 mm2, p<0.001). Single cell suspensions of endometriosis lesions were analyzed by flow cytometry, showing a near complete ablation of DC in Group III while no ablation was observed in the other 3 groups of mice.CONCLUSIONS: Our results indicate that endometriosis is dependent on the infiltrating DC population. Therapies designed to inhibit DC infiltration could target endometriosis in the future. This possibility warrants further studies. OBJECTIVE: Endometriosis affects 10-15% of women of reproductive age and is associated with pelvic pain and infertility. Recently, we have shown that dendritic cells (DC) supplementation led to the enhancement of endometriosis lesion growth and of intra-lesion angiogenesis. In this study, we investigated whether the development of endometriosis is dependent on the presence of endogenous DC. DESIGN: Prospective study using a surgical model of endometriosis in mice. MATERIALS AND METHODS: Surgical induction of endometriosis was performed in 8-10 week old female mice. Briefly, under anesthesia, a midline laparotomy was performed, uterine horns were removed, and 2-mm biopsy punch samples were sutured to the peritoneal wall. After 8 days, mice were killed, their abdomens were opened through the same incision and lesions were measured in two perpendicular diameters (D1 and D2) and the cross-sectional lesion area calculated using the formula D1xD2xπ/4. Single cell suspensions were analyzed by flow cytometry. In order to ablate DC, we utilized CD11c-DTR/GFP transgenic mice (C57Bl/6 background), in which DC can be ablated in vivo. Administration of diphtheria toxin (DT) into the transgenic mice that express the primate DT receptor (DTR), resulted in depletion of CD11c+DC. Endometriosis was induced in 10 CD11c-DTR/GFP transgenic mice and in 10 C57Bl/6 control mice. The mice were allocated to 4 groups (n=5 mice per group): I: C57Bl/6 treated with vehicle; II: C57Bl/6 treated with DT; III: CD11c-DTR/GFP transgenic mice treated with DT; IV: CD11c-DTR/GFP transgenic mice treated with vehicle. On day 0, endometriosis was induced. On days 2, 5, and 9, DT or vehicle was administered. All mice were sacrificed on day 10, the lesions were measured, and surgically removed from the abdominal cavity. RESULTS: In CD11c-DTR/GFP transgenic mice that were treated with DT (Group III) the endometriotic lesions were significantly smaller than that of all the other groups (ANOVA) (i.e. Group I: 5.25±1.18 mm2; Group II: 4.73±1.11 mm2; Group III: 1.09 ± 0.38 mm2; Group IV: 5.12±1.09 mm2, p<0.001). Single cell suspensions of endometriosis lesions were analyzed by flow cytometry, showing a near complete ablation of DC in Group III while no ablation was observed in the other 3 groups of mice. CONCLUSIONS: Our results indicate that endometriosis is dependent on the infiltrating DC population. Therapies designed to inhibit DC infiltration could target endometriosis in the future. This possibility warrants further studies.

Dendritic cells derived from bone marrow cells fail to acquire and present major histocompatibility complex antigens from other dendritic cells

Dendritic cells stimulate primary T-cell responses and a major activation route is via presentation of antigens pre-processed by other dendritic cells. This presentation of pre-processed antigens most likely proceeds through transfer of functional major histocompatibility complex (MHC) antigens through exosomes, 'live nibbling' or apoptotic vesicles. We hypothesized that not all dendritic cell populations may both donate MHC antigen to dendritic cells and present antigens acquired from other dendritic cells. All populations tested, including those derived from bone marrow precursor cells stimulated primary, allogeneic T-cell responses and acted as accessory cells for mitogen stimulation. Populations of responder type, splenic dendritic cells promoted allogeneic responses indirectly but those derived from bone marrow cells blocked rather than promoted T-cell proliferation. To identify mechanisms underlying this difference we studied transfer of I-A antigens between cells. Active, two-way transfer of allogeneic I-A occurred between splenic primary antigen presenting cells including CD8alpha+ lymphoid dendritic cells, CD8alpha- myeloid dendritic cells and B220+ cells; all these cell types donated as well as acquired MHC molecules. By contrast, the bone marrow-derived dendritic cells donated I-A antigens but acquired negligible amounts. Thus, dendritic cells derived directly from bone marrow cells may stimulate primary T-cell responses through transferring functional MHC to other dendritic cells but may not be able to acquire and present antigens from other dendritic cells. The evidence suggests that T-cell activation may be blocked by the presence of dendritic cells that have not matured through lymphoid tissues which are unable to acquire and present antigens pre-processed by other dendritic cells.

CD4+ t cells stimulated in the presence of dendritic cells transduced with human papillomavirus GFP-L1, showed a decreased progression towards s and m phase of the cell cycle

CD4+ t cells stimulated in the presence of dendritic cells transduced with human papillomavirus GFP-L1, showed a decreased progression towards s and m phase of the cell cycle

Early Increases in Superantigen-Specific Foxp3+Regulatory T Cells during Mouse Mammary Tumor Virus Infection

Mouse mammary tumor virus (MMTV) is a milk-borne betaretrovirus that has developed strategies to exploit and subvert the host immune system. Here, we show in a natural model of MMTV infection that the virus causes early and progressive increases in superantigen (SAg)-specific Foxp3(+) regulatory T cells (T(reg)) in Peyer's patches (PP). These increases were shown to be dependent on the presence of dendritic cells. CD4(+) CD25(+) T cells from the PP of infected mice preferentially suppress the proliferative response of T cells to SAg-expressing antigen-presenting cells ex vivo. We investigated the influence of the depletion of CD25(+) cells at different stages of the infection. When CD25(+) cells were depleted before MMTV infection, an increase in the number of PP SAg-cognate Foxp3(-) T cells was found at day 6 of infection. Since the SAg response is associated with viral amplification, the possibility exists that T(reg) cells attenuate the increase in viral load at the beginning of the infection. In contrast, depletion of CD25(+) cells once the initial SAg response has developed caused a lower viral load, suggesting that at later stages T(reg) cells may favor viral persistence. Thus, our results indicated that T(reg) cells play an important and complex role during MMTV infection.

Primary Peripheral T Cells Become Susceptible to 2,3,7,8-Tetrachlorodibenzo-p-Dioxin-Mediated Apoptosis in Vitro upon Activation and in the Presence of Dendritic Cells

Although the toxic effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on T cells in vivo have been well characterized, attempts to reproduce these findings in vitro have not been successful. In the current study, we examined whether activation or the presence of dendritic cells (DCs) would make primary naive T cells from C57BL/6 mice susceptible to TCDD-induced apoptosis in vitro. Although nonactivated primary T cells cultured with 10 to 1000 nM TCDD were relatively resistant to apoptosis, they became sensitive to apoptosis upon activation with concanavalin A (ConA). Moreover, ConA-activated T cells cultured in the presence of DCs showed highest levels of TCDD-induced apoptosis. Likewise, primary T cells from OT.II.2a mice cultured with specific ovalbumin peptide and syngeneic DCs showed higher levels of apoptosis compared with similar nonactivated T cells. T-cell activation led to up-regulation of aryl hydrocarbon receptor (AhR), Fas, and Fas-ligand (FasL) expression. In addition, DC maturation and culture with TCDD caused significant induction of FasL. TCDD-mediated apoptosis in activated peripheral T cells was AhR-dependent. Analysis of why nonactivated T cells are more resistant, whereas activated T cells are sensitive to TCDD-induced apoptosis revealed that TCDD treatment of activated but not nonactivated T cells led to down-regulation of cellular FLICE inhibitory protein (c-FLIP), an inhibitor of apoptosis. Moreover, down-regulation of c-FLIP using small interfering RNA in nonactivated T cells made them sensitive to TCDD-induced apoptosis. The current study demonstrates for the first time that TCDD can induce apoptosis in vitro in peripheral T cells upon activation and in the presence of DCs and that this may be mediated by down-regulation of c-FLIP.

Donor age intensifies the early immune response after transplantation

Increasing donor age is associated with reduced graft function. We wondered if donor age may not only affect intrinsic function but also alter the immune response of the recipient. Kidneys from young and old F-344 rats (3 vs 18 months) were transplanted into bilaterally nephrectomized young Lewis recipients and compared with age-matched controls (follow-up: 6 months). Renal function and structural changes were assessed serially in both native kidneys and allografts. Host alloreactivity, graft-infiltrating cells, and their inflammatory products were determined at intervals to examine the correlation of immune response and donor age. Functional and structural deterioration had advanced significantly in older allografts compared with age-matched native controls, whereas differences between young allografts and native controls of similar age were only minor. Changes in grafts from elderly rats were associated with a more intense host immune response early post-transplant (up to 1 month) reflected by significantly higher numbers of peripheral T and B cells, increased T-cell alloreactivity and modified cytokine patterns associated with elevated frequencies of intragraft dendritic cells, B cells, and CD31+ cells. By 6 months, recipients of young donor grafts produced comparable or more intense alloantigen-specific immune responses. Older donor grafts elicit a stronger immune response in the early period after transplantation.

Plasma and Mucosal Fluid from HIV Type 1-Infected Patients But Not from HIV Type 1-Exposed Uninfected Subjects Prevent HIV Type 1-Exposed DC from Infecting Other Target Cells

Highly exposed persistently seronegative (HEPS) individuals have previously been shown to mount HIV-1-specific humoral and cellular immune responses in the mucosa, despite their uninfected status. It is thus possible that HEPS individuals are protected from HIV-1 infection at the mucosal level. Recent work supports the hypothesis that dendritic cells are involved in the establishment of a mucosal HIV-1 infection as well as the dissemination to other target cells. However, no previous study has investigated if samples collected from HEPS individuals have the capacity to prevent HIV-1 infection in the presence of dendritic cells in vitro. We therefore established an assay that measures HIV-1 neutralization in cocultures of HIV-1-exposed dendritic cells (DC) and PBMC. Plasma and cervicovaginal lavage (CVL) samples from HIV-1-infected patients and HEPS individuals, enrolled in a well-characterized sex worker cohort in Kenya, were evaluated. Most plasma and CVL samples of HIV-1-infected patients neutralized HIV-1 in the DC/PBMC cocultures. Neither plasma nor CVL samples of most HEPS individuals had this capacity. However, they readily neutralized HIV-1 infection of PBMC alone. This may suggest that protection against HIV-1 infection in HEPS individuals occurs prior to interaction between HIV-1-exposed DC and other target cells.

Direct exosome stimulation of peripheral humanT cells detected by ELISPOT

Exosomes from APC are nano-vesicles that can induce antigen-specific T cell responses and are presently explored as therapeutic tools in different clinical settings. Investigations of the capacity of exosomes to stimulate T cells in vitro have mostly been performed on T cell hybridomas, clones or lines. Whether exosomes can stimulate T cells directly or need the presence of dendritic cells (DC) is debated. We could detect exosome-induced antigen-specific CD8(+) T cell responses in peripheral blood from humans. Exosomes from monocyte-derived DC (MDDC) were loaded with a mix of 23 immunogenic peptides from EBV, CMV and influenza virus, and added to autologous peripheral CD8(+) T cells. IFN-gamma-producing cells were detected by enzyme-linked immunospot assay (ELISPOT). MDDC-exosomes induced IFN-gamma production in CD8(+) T cells without addition of DC. The response was exosome dose dependent, and dependent on exosomal MHC class I. Furthermore, we detected an enhanced T cell stimulatory capacity by exosomes from lipopolysaccharide-matured MDDC compared to exosomes from immature MDDC. Exosomes could also induce TNF-alpha production. These results show, for the first time, that exosomes can directly stimulate human peripheral CD8(+) T cells in an antigen-specific manner and that ELISPOT is a suitable method for detecting exosome-induced peripheral T cell responses. This system may provide a useful tool when developing exosomes as therapeutic agents.

Toll-dependent selection of microbial antigens for presentation by dendritic cells

Dendritic cells constitutively sample the tissue microenvironment and phagocytose both microbial and host apoptotic cells. This leads to the induction of immunity against invading pathogens or tolerance to peripheral self antigens, respectively. The outcome of antigen presentation by dendritic cells depends on their activation status, such that Toll-like receptor (TLR)-induced dendritic cell activation makes them immunogenic, whereas steady-state presentation of self antigens leads to tolerance. TLR-inducible expression of co-stimulatory signals is one of the mechanisms of self/non-self discrimination. However, it is unclear whether or how the inducible expression of co-stimulatory signals would distinguish between self antigens and microbial antigens when both are encountered by dendritic cells during infection. Here we describe a new mechanism of antigen selection in dendritic cells for presentation by major histocompatibility complex class II molecules (MHC II) that is based on the origin of the antigen. We show that the efficiency of presenting antigens from phagocytosed cargo is dependent on the presence of TLR ligands within the cargo. Furthermore, we show that the generation of peptide-MHC class II complexes is controlled by TLRs in a strictly phagosome-autonomous manner.

Human embryonic stem cells are immunogenic in allogeneic and xenogeneic settings

Recent studies have suggested that human embryonic stem cells (HESC) are immune-privileged and may thereby circumvent rejection. The expression of immunologically active molecules was studied by DNA microarray analysis and by flow cytometry. HESC were transplanted into immunologically competent mice and traced by fluorescence in-situ hybridization (FISH) and immunohistochemistry. The ability of HESC to directly and indirectly induce immune responses in CD4 + T-cells from naive and transplanted mice was studied. Their ability to induce immune responses of human CD4 + T-cells, when cultured in the presence of dendritic cells (DC) syngeneic to responder T-cells, was also analysed. HESC demonstrated expression of HLA class I and HLA class II genes, but the cell surface expression of HLA class II molecules was low even after incubation with IFNγ. In wild-type mice, HESC could be demonstrated by FISH until 3 days after transplantation and were surrounded by heavy infiltrates of T-cells and macrophages. HESC induced a similar immune response as human fibroblast cells (HFib) on naive and immunized T-cells, both directly and in the presence of syngeneic DC. A similar response was observed in the allogeneic setting. It is concluded that HESC are immunologically inert and do not inhibit immune responses during direct or indirect antigen presentation, and they were acutely rejected in a xenogeneic setting.

Dendritic-NK Cell Interactions in P. gingivalis-specific Responses

Patients with localized aggressive periodontitis have type-1 cytokines in gingival crevicular fluid and high titers of IFN-gamma-dependent IgG2 reactive with P. gingivalis in gingival crevicular fluid and serum. Localized aggressive periodontitis monocytes spontaneously differentiate into dendritic cells that can stimulate IFN-gamma production by NK cells. These relationships prompted the hypothesis that P. gingivalis-dendritic cell-NK cell interactions might promote type-1 cytokine responses. Although P. gingivalis is not a potent inducer of Th1 responses, it stimulated strong IL-12 responses by monocyte-derived dendritic cells in the presence of IFN-gamma, and IFN-gamma was produced by NK cells within 24 hrs in the presence of dendritic cells. Anti-P. gingivalis IgG2 responses were enhanced by dendritic cells, and removal of NK cells reduced IFN-gamma- and P. gingivalis-specific IgG2. Thus, P. gingivalis-dendritic cell-NK cell interactions apparently resulted in reciprocal stimulation and increased type-1 cytokine production by both dendritic cells and NK cells, and increased P. gingivalis-specific IgG2.

Terapias emergentes en artritis reumatoide

The use of biological agents such as etanercept, infliximab, adalimumab and anakinra has been recently approved for the treatment of rheumatoid arthritis. All are effective controlling signs and symptoms and inhibiting disease progression. To overcome the problems generated by their high costs and possible participation in reactivating latent infections, other therapeutic tools are being developed. Gene therapy using expression vectors carrying genes coding for specific proteins, may interfere in key points involved in the pathogenesis of the disease. Intra-articular administration of cDNA coding for soluble TNF receptors, IL-1, or IL-1Ra decreases signs of the disease in animal models. Vectors, expressing inhibitors of signal transduction pathways involving to NF-kB and JAK-STAT-3, are effective in modulating joint inflammation in mice. The use of antigen-pulsed antigen presenting cells or dendritic cells (DC) bound to apoptosis-inducing molecules, specifically eliminates autoreactive T cells. Other novel approach attempts the development of T regulatory-inducing tolerogenic DC-based vaccines that inhibit autoreactive T cells, through the secretion of suppressing cytokines or by other mechanisms to be elucidated. Oral tolerance induction to auto-antigens is also a successful experimental strategy under study. Current research aims to control peripheral tolerance in rheumatoid arthritis patients.

Glucocerebroside treatment ameliorates ConA hepatitis by inhibition of NKT lymphocytes

Concanavalin A (ConA) induces natural killer T (NKT) cell-mediated liver damage. Glucocerebroside (GC) is a naturally occurring glycolipid. Our aims were to determine the effect of GC in a murine model of ConA-induced hepatitis. Mice in groups A and B were treated with GC 2 h before and 2 h following administration of ConA, respectively; group C mice were treated with ConA; group D mice was treated with GC; group E mice did not receive any treatment. Liver damage was evaluated by serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels and liver histology. The immune effect of GC was determined by fluorescence-activated cell sorter analysis of intrahepatic and intrasplenic NKT lymphocytes, measurement of cytokine levels, and Western blot analysis for STAT 1, 4, 6, and NF-kappaB expression. The effect of GC on NKT cell proliferation was assessed in vitro. Serum AST and ALT levels were markedly reduced in GC-treated group A mice compared with nontreated group C animals, and histological damage was markedly attenuated in group A. The beneficial effect of GC was associated with a 20% decrease of intrahepatic NKT lymphocytes, significant lowering of serum IFN-gamma levels, and decreased STAT1 and STAT6 expression. In vitro administration of GC led to a 42% decrease of NKT cell proliferation in the presence of dendritic cells but not in their absence. Intraperitoneally administered radioactive GC was detected in the liver and bowel. Administration of GC led to amelioration of ConA hepatitis associated with an inhibitory effect on NKT lymphocytes. GC holds promise as a new immune-modulatory agent.