Regulation of T helper cell differentiation in vivo by soluble and membrane proteins provided by antigen-presenting cells

The aim of this study was to test whether the nature of the antigen-presenting cell (APC) can influence the Th1/Th2 balance in vivo. Our data show that dendritic cells (DC), pulsed extracorporeally with antigen, induced the development of cells secreting IL-2, IFN-gamma and IL-4 upon antigen rechallenge in vitro. Priming with peritoneal macrophages sensitized cells that produced IL-4 but not IFN-gamma. To identify the factors involved in T helper development, mice were primed with APC with or without treatment with neutralizing antibodies to costimulatory molecules or cytokines. Our results indicate that priming with DC or macrophages is strictly dependent on the CD28-CTLA4/B7 interaction. Of note, CD86 provides the initial signal to induce naive T cells to become IL-4 producers, whereas CD80 is a more neutral differentiation signal. IL-12, released by the DC, appears as a potent and obligatory inducer of differentiation for IFN-gamma-producing cells. IL-6, although produced by both APC populations, is necessary to direct activation of the Th2-type response by macrophages but not by DC.     

Angiotensin II-mediated expression of p27Kip1 and induction of cellular hypertrophy in renal tubular cells depend on the generation of oxygen radicals

Presentation of MHC class I antigens by professional antigen-presenting cells (APC) is an important pathway in priming cytotoxic T lymphocyte responses in vivo. This study sought to identify the nature of the professional APC responsible for indirect class I presentation by examining a special feature of professional APC, namely their ability to process exogenous forms of antigen for class I presentation. Incubation of highly purified bone marrow-derived precursor cells with chicken ovalbumin (OVA) led to the efficient presentation of the major class I-restricted OVA determinant by mature dendritic cells (DC), but not by macrophages (Mphi) derived from the precursor population. DC as well as macrophages were, however, able to mediate class II presentation of OVA, suggesting that macrophages were deficient in class I processing but not in capturing exogenous OVA. The majority of mature DC, i.e. over 80 %, generated from the precursor cells pulsed with OVA, presented the class I OVA epitope. Upon maturation, class I presentation of OVA by DC was greatly reduced, suggesting that class I processing of exogenous antigen is modulated during DC maturation in a manner similar to class II antigen processing. This study shows that bone marrow-derived DC/ME progenitors capture exogenous antigen for class I presentation, and that cells of the DC lineage can be functionally distinguished from cells of the macrophage lineage based on their ability to process exogenous antigen for class I presentation.     

Nonviral transfection of distinct types of human dendritic cells: high-efficiency gene transfer by electroporation into hematopoietic progenitor- but not monocyte-derived dendritic cells

Human dendritic cells (DC) are highly professional antigen presenting cells for the priming of naive cytotoxic T cells. Gene transfer in DC would be a useful strategy to load DC with relevant de novo synthesized antigens for immunotherapeutical purposes. As a first step towards a DC-based gene therapy, we examined the efficiency of nonviral transfection in different types of cultured human dendritic cells with a humanized red-shifted green fluorescent protein reporter gene. Plasmid DNA transfection by electroporation or lipofection was used to transfect CD34+ progenitor cell-derived DC (PC-DC) and Langerhans' cells (PC-LC), as well as monocyte-derived DC (Mo-DC). While lipofection was unsuccessful in all types of DC, we obtained high-efficiency gene transfer by electroporation in PC-LC (16%) and PC-DC (12%). In contrast, electroporation was strikingly less efficient in Mo-DC (< or = 2%). The potent allostimulatory capacity of DC was still retained in electroporated PC-DC and PC-LC. In conclusion, electroporation of antigen expressing plasmid DNA is an efficient tool for nonviral gene transfer in PC-DC and PC-LC, but not in Mo-DC and could be useful for the development of DC-based tumor immunotherapy.     

CD4+ T cell responses in mice lacking MHC class II molecules specifically on B cells

The role of B lymphocytes in initiating and maintaining a CD4+ T cell response has been examined using a variety of strategies, but remains controversial because of weaknesses inherent to each of the approaches. Here, we address this issue by measuring CD4+ T cell priming both in mutant mice devoid of B cells and in chimeric animals lacking major histocompatibility complex class II molecules specifically on B cells. We find that peptide and some protein antigens do not require B cells expressing class II molecules, nor B cells themselves, to efficiently prime. This could be demonstrated by the usual lymph node proliferation assay, a rather indirect in vitro measure of priming, and by a direct ex vivo assay of population expansion and activation marker expression. Interestingly, one protein antigen, conalbumin, could not prime in the absence of B cells, but could in the presence of B cells devoid of class II molecules. This finding constrains the possible mechanisms whereby B lymphocytes contribute to the initiation of a CD4+ T cell response, arguing against the importance of surface immunoglobulin-mediated antigen presentation by B cells.     

Presentation of exogenous protein antigens on major histocompatibility complex class I molecules by dendritic cells: pathway of presentation and regulation by cytokines

Several recent studies have shown that dendritic cells (DC) pulsed with soluble proteins can present peptide epitopes derived from these exogenous antigens on major histocompatability complex (MHC) class I molecules and induce an antigen-specific cytotoxic T lymphocyte (CTL) response. We provide evidence here that DC use macropinocytosis to capture soluble antigens that are then presented on MHC class I molecules. The presentation of an epitope derived from soluble ovalbumin was transporter associated with antigen presentation (TAP)-dependent, brefeldin A-sensitive, blocked by inhibitors of proteasomes, and resistant to chloroquine. These data suggest that exogenous antigens access the cytosol of DC and are proccessed for presentation via the same pathway described for conventional MHC class I-restricted cytosolic antigens. Proinflammatory mediators such as tumor necrosis factor-alpha (TNF-alpha) and lipopolysaccharide (LPS) reduced the efficiency of ovalbumin presentation via this pathway. This reduced presentation was not due to impaired expression of class I molecules because these substances upregulated the cell surface expression of Kb-molecules comparable to levels induced by interferon-gamma (IFN-gamma) treatment. The addition of IFN-gamma increased ovalbumin presentation even in the presence of TNF-alpha or LPS. These results show that DC might be involved in the cross-priming phenomenon. This could offer the immune system an additional pathway for effective priming of cytotoxic T cells and provide the possibility to activate both CD4 and CD8 T-cell responses.     

Uptake of Leishmania major amastigotes results in activation and interleukin 12 release from murine skin-derived dendritic cells: implications for the initiation of anti-Leishmania immunity

Epidermal Langerhans cells (LC) are immature dendritic cells (DC) located in close proximity to the site of inoculation of infectious Leishmania major metacyclic promastigotes by sand flies. Using LC-like DC expanded from C57BL/6 fetal skin, we characterized interactions involving several developmental stages of Leishmania and DC. We confirmed that L. major amastigotes, but not promastigotes, efficiently entered LC-like DC. Parasite internalization was associated with activation manifested by upregulation of major histocompatibility complex (MHC) class I and II surface antigens, increased expression of costimulatory molecules (CD40, CD54, CD80, and CD86), and interleukin (IL)-12 p40 release within 18 h. L. major-induced IL-12 p70 release by DC required interferon gamma and prolonged (72 h) incubation. In contrast, infection of inflammatory macrophages (Mphi) with amastigotes or promastigotes did not lead to significant changes in surface antigen expression or cytokine production. These results suggest that skin Mphi and DC are infected sequentially in cutaneous leishmaniasis and that they play distinct roles in the inflammatory and immune response initiated by L. major. Mphi capture organisms near the site of inoculation early in the course of infection after establishment of cellular immunity, and kill amastigotes but probably do not actively participate in T cell priming. In contrast, skin DC are induced to express increased amounts of MHC antigens and costimulatory molecules and to release cytokines (including IL-12 p70) by exposure to L. major amastigotes that ultimately accumulate in lesional tissue, and thus very likely initiate protective T helper cell type 1 immunity.     

CD40 ligand inhibits Fas/CD95-mediated apoptosis of human blood-derived dendritic cells

Dendritic cells (DC) are considered to be the most potent antigen-presenting cells (APC) in the immune system. In this study, we analyzed the regulation of apoptosis of human peripheral blood-derived DC. DC were generated from adherent peripheral blood mononuclear cells that had been cultured for 7 days with granulocyte-macrophage colony-stimulating factor and interleukin-4. These cells displayed phenotypic properties of DC, including dendritic processes, expression of CD1a and lack of expression of CD14, and were very potent at presenting soluble antigens to T cells. Blood-derived DC were demonstrated to express the Fas/CD95 antigen and an agonist antibody to CD95 strongly induced apoptotic cell death in these cells. Soluble trimeric CD40 ligand potently inhibited both CD95-mediated and spontaneous apoptosis in DC. The data suggest that interactions between members of the tumor necrosis factor family of ligands expressed by T cells with their receptors on DC play an important role in the regulation of apoptosis in DC during antigen presentation and may, therefore, regulate the duration of T cell expansion and cytokine production.     

Regression of tumors in mice vaccinated with professional antigen-presenting cells pulsed with tumor extracts

Vaccination with tumor extracts circumvents the need to identify specific tumor rejection antigens and extends the use of active immunotherapy to the vast majority of cancers, in which specific tumor antigens have not yet been identified. In this study we examined the efficacy of tumor vaccines comprised of unfractionated tumor material presented by professional antigen-presenting cells (APC): dendritic cells (DC) or macrophages (M phi). To enhance the relevance of these studies for human patients we used 2 poorly immunogenic murine tumor models and evaluated the effectiveness of the vaccination protocols in tumor-bearing animals. APC (in particular DC) pulsed with unfractionated extracts from these "poorly immunogenic" tumors were highly effective in eliciting tumor-specific cytotoxic T lymphocytes. A measurable CTL response could be detected after even a single immunization with tumor extract-pulsed DC. DC or M phi pulsed with tumor extract were also effective vaccines in tumor-bearing animals. In the murine bladder tumor (MBT-2) model a modest extension of survival and 40% cure rate was seen in the animal groups immunized with DC or M phi pulsed with MBT-2 tumor extract. DC or M phi pulsed with B16/F10.9 tumor extract were also remarkably effective in the B16 melanoma lung metastasis model, as shown by the observation that treatment with APC caused a significant reduction in lung metastases. Cumulatively, the CTL and immunotherapy data from the two murine tumor systems suggest that APC (in particular DC) pulsed with unfractionated cell extracts as a source of tumor antigen may be equally or more effective than genetically modified tumor vaccines.     

Antigen-pulsed CD8alpha+ dendritic cells generate an immune response after subcutaneous injection without homing to the draining lymph node

Two subsets of murine splenic dendritic cells, derived from distinct precursors, can be distinguished by surface expression of CD8alpha homodimers. The functions of the two subsets remain controversial, although it has been suggested that the lymphoid-derived (CD8alpha+) subset induces tolerance, whereas the myeloid-derived (CD8alpha-) subset has been shown to prime naive T cells and to generate memory responses. To study their capacity to prime or tolerize naive CD4(+) T cells in vivo, purified CD8alpha+ or CD8alpha- dendritic cells were injected subcutaneously into normal mice. In contrast to CD8alpha- dendritic cells, the CD8alpha+ fraction failed to traffic to the draining lymph node and did not generate responses to intravenous peptide. However, after in vitro pulsing with peptide, strong in vivo T cell responses to purified CD8alpha+ dendritic cells could be detected. Such responses may have been initiated via transfer of peptide-major histocompatibility complex complexes to migratory host CD8alpha- dendritic cells after injection. These data suggest that correlation of T helper cell type 1 (Th1) and Th2 priming with injection of CD8alpha+ and CD8alpha- dendritic cells, respectively, may not result from direct T cell activation by lymphoid versus myeloid dendritic cells, but rather from indirect modification of the response to immunogenic CD8alpha- dendritic cells by CD8alpha+ dendritic cells.     

Dual role of dendritic cells in the induction and down-regulation of antigen-specific cutaneous inflammation

We have previously reported that contact sensitivity (CS) to dinitrofluorobenzene (DNFB) in C57BL/6 mice was mediated by MHC class I-restricted CD8+ T cells and down-regulated by MHC class II-restricted CD4+ T cells. In this study, we analyzed the contribution of dendritic cells (DC) in the induction of these two T cell subsets endowed with opposite functions. Hapten-pulsed skin- and bone marrow-derived DC, obtained from either normal C57BL/6 mice or from MHC class II (I+ II-) and MHC class I (I- II+)-deficient mice, were tested for their ability to prime normal mice for CS to dinitrofluorobenzene. Expression of MHC class I molecules by transferred DC was mandatory both for the induction of CS and for the generation of hapten-specific CD8+ T cells in lymphoid organs. I+ II- DC were as potent as I+ II+ DC in priming for CS, demonstrating that activation of effector CD8+ T cells can occur independently of CD4+ T cell help. I- II+ DC could not immunize for CS, although they could sensitize for a delayed-type hypersensitivity reaction to protein Ags. Moreover, I- II+ DC injected simultaneously with cutaneous sensitization down-regulated the inflammatory response, suggesting that hapten presentation by MHC class II molecules could prime regulatory CD4+ T cells. These results indicate that DC can present haptenated peptides by both MHC class I and class II molecules and activate Ag-specific CD8+ effector and CD4+ regulatory T cell subsets, concurrently and independently.     

Induction of antigen-specific antitumor immunity with adenovirus-transduced dendritic cells

Transduction of dendritic cells (DC) can result in presentation of tumor-associated antigens and induction of immunity against undefined epitopes. The present studies demonstrate adenovirus (Ad)-mediated transduction of the beta-galactosidase gene in mouse DC. Similar transductions have been obtained with the gene encoding the DF3/MUC1 tumor-associated antigen. We show that the Ad-transduced DC are functional in primary allogeneic mixed lymphocyte reactions. Mice immunized with Ad-transduced DC develop cytotoxic T lymphocytes that are specific for the beta-galactosidase or DF3/MUC1 antigens. The results also demonstrate that Ad MUC1-transduced DC induce a specific response which inhibits the growth of DF3/MUC1-positive tumors. These findings support the usefulness of Ad-transduced DC for in vivo immunization against tumor-associated antigens.     

Decreased antigen presentation by dendritic cells in patients with breast cancer

We evaluated T-cell responses to mitogens and to defined antigens in breast cancer patients. Significant defects in responses to tetanus toxoid and influenza virus were observed in patients with advanced-stage breast cancer. To define whether these defects were associated with a defect in antigen presentation [dendritic cells (DCs)] or effector function (T cells), these cells were studied separately. Purified DCs from 32 patients with breast cancer demonstrated a significantly decreased ability to stimulate control allogeneic T cells, but stimulation of patient T cells with either control allogeneic DCs or immobilized anti-CD3 antibody resulted in normal T-cell responses, even in patients with stage IV tumors. These data suggest that reduced DC function could be one of the major causes of the observed defect in cellular immunity in patients with advanced breast cancer. We then tested whether stem cells from these patients could give rise to functional DCs after in vitro growth with granulocyte/macrophage colony-stimulating factor and interleukin 4. Normal levels of control allogeneic and tetanus toxoid-dependent T-cell proliferation were observed when DCs obtained from precursors were used as stimulators. Those cells also induced substantially higher levels of influenza virus-specific CTL responses than mature DCs from the peripheral blood of these patients, although responses did not quite reach control values. Thus, defective T-cell function in patients with advanced breast cancer can be overcome by stimulation with DCs generated from precursors, suggesting that these cells may better serve as autologous antigen carriers for cancer immunotherapy than mature peripheral blood DCs.     

Antigen-unspecific B cells and lymphoid dendritic cells both show extensive surface expression of processed antigen-major histocompatibility complex class II complexes after soluble protein exposure in vivo or in vitro

Intravenous (i.v.) injection of high amounts of soluble proteins often results in the induction of antigen-specific tolerance or deviation to helper rather than inflammatory T cell immunity. It has been proposed that this outcome may be due to antigen presentation to T cells by a large cohort of poorly costimulatory or IL-12-deficient resting B cells lacking specific immunoglobulin receptors for the protein. However, previous studies using T cell activation in vitro to assess antigen display have failed to support this idea, showing evidence of specific peptide-major histocompatibility complex (MHC) class II ligand only on purified dendritic cells (DC) or antigen-specific B cells isolated from protein injected mice. Here we reexamine this question using a recently derived monoclonal antibody specific for the T cell receptor (TCR) ligand formed by the association of the 46-61 determinant of hen egg lysozyme (HEL) and the mouse MHC class II molecule I-Ak. In striking contrast to conclusions drawn from indirect T cell activation studies, this direct method of TCR ligand analysis shows that i.v. administration of HEL protein results in nearly all B cells in lymphoid tissues having substantial levels of HEL 46-61-Ak complexes on their surface. DC readily isolated from spleen also display this TCR ligand on their surface. Although the absolute number of displayed ligands is greater on such DC, the relative specific ligand expression compared to total MHC class II levels is similar or greater on B cells. These results demonstrate that in the absence of activating stimuli, both lymphoid DC and antigen-unspecific B cells present to a similar extent class II-associated peptides derived from soluble proteins in extracellular fluid. The numerical advantage of the TCR ligand-bearing B cells may permit them to interact first or more often with naive antigen-specific T cells, contributing to the induction of high-dose T cell tolerance or immune deviation.     

Maturation of Peyer's patch dendritic cells in vitro upon stimulation via cytokines or CD40 triggering

In mouse Peyer's patches (PP), dendritic cells (DC) are localized in T cell areas as NLDC145+ CD11c+ cells, and in the dome and corona region of the follicle as NLDC145- CD11c+ cells, respectively, suggesting the presence of two different DC populations with distinct roles in antigen uptake, processing, and presentation. However, it is not clear how this relates to DC maturation. In this report, we demonstrate that freshly-isolated CD11c+ DC have the properties of immature DC since they endocytose soluble antigens, phagocytose particulate material such as latex beads, synthetize major histocompatibility complex (MHC) class II and invariant chain, but, at the same time, display low stimulatory activity for resting T cells, as shown in mixed-lymphocyte reaction and oxidative mitogenesis assays. When cultured for 24 h in the presence of the cytokines granulocyte-macrophage colony-stimulating factor and tumor necrosis factor or anti-CD40, the cells undergo dramatic phenotypic and functional changes characteristic of DC maturation. After 24 h stimulation in vitro, CD11c+ cells lose the ability to take up proteins such as ovalbumin, and in parallel with this decline, the biosynthesis of MHC class II and invariant chain is dramatically down-regulated or eliminated. On the other hand cells treated in vitro exhibit on the cell surface higher levels of MHC class II, of co-stimulatory molecules (CD80, CD86), of adhesion molecules (CD44, intercellular adhesion molecule-1), and acquire expression of the interdigitating DC surface marker NLDC145. Concomitantly, the ability to stimulate naive T cells drastically increased after in vitro treatment with both stimuli. Taken together, our results indicate that the majority of DC in the PP are immature in terms of their antigen-uptake capacity. These sentinel antigen presenting cells are strategically positioned at the dome region of PP, where antigens are transcytosed via the M cells from the gut lumen. A second population of mature interdigitating NLDC145+ CD11c+ DC stimulates naive unprimed T cells in interfollicular areas by up-regulation of surface ligands and accessory signals.     

Tumor-peptide-pulsed dendritic cells isolated from spleen or cultured in vitro from bone marrow precursors can provide protection against tumor challenge

Dendritic cells (DC) purified from murine spleen or generated in vitro from bone marrow precursors were compared for their respective abilities to stimulate T cell responses and provide tumor protection in vivo. In vitro incubation with synthetic tumor peptide conferred on both DC populations the ability to induce proliferation of tumor-peptide-specific T cells in vitro. Spleen DC were reproducibly about twofold more effective than bone-marrow-derived DC in this assay. Both DC populations could also induce cytotoxic activity in vivo. In vitro cytoxicity assays showed that, while cytotoxic activity induced by immunization with spleen DC was clearly peptide-specific, a high non-specific cytotoxic activity was consistently observed after immunization with bone-marrow-derived DC, whether peptide-pulsed or not. Regardless of such high non-specific activity in vitro, only tumor-peptide-pulsed DC could provide protection against subsequent inoculation of tumor cells. DC not pulsed with tumor peptide were ineffective. We conclude that DC isolated from spleen or generated in vitro from bone marrow precursors are suitable reagents for use in tumor vaccination studies.     

Concentrations and spatial variations of cyclodienes and other organochlorines in herring and perch from the Baltic Sea

Naive T lymphocytes specific for a given primary antigen occur in low frequencies and require the relevant antigen to be presented by specialist antigen presenting cells (APC), i.e., dendritic cells (DC). For these reasons, the in vitro induction of primary T lymphocyte responses remains a significant technical challenge. We have attempted to improve current strategies for generating in vitro responses by optimising (i) isolation and concomitant activation of DC from peripheral blood, (ii) uptake, processing and presentation of antigen by DC and (iii) antigen driven T lymphocyte proliferation. We established that RPMI 1640 media supplemented with 10% autologous serum resulted in the best yield of CMRF-44+, CD14-, CD19- DC after enrichment over a Nycodenz gradient. Optimal presentation of whole protein and peptide antigen was achieved by addition after the purification of the APC, i.e., at the start of the T lymphocyte proliferation assay. RPMI 1640 supplemented with 10% autologous serum or plasma supported the best antigen driven specific T lymphocyte responses. Using these optimised conditions, we compared the efficacy of PBMC and purified blood DC for priming T lymphocyte responses to the chronic myeloid leukaemia (CML) specific bcr-abl (b3a2) peptide. Peptide specific T lymphocyte responses were generated with both purified DC and whole PBMC, suggesting that T lymphocyte precursor frequency was the limiting factor in these experiments. These results will aid in the generation of human T lymphocyte lines to primary antigens, for in vitro and therapeutic applications.     

Peripheral blood dendritic cells reinduce proliferation in in vitro aged T cell populations

Dendritic cells (DC) are professional antigen presenting cells which are essential for the initiation of an immune response. Recently we demonstrated that DC, which had been propagated from the peripheral blood of healthy elderly people, were morphologically and functionally intact. It was the aim of the present study to analyze how DC from young and old healthy individuals could affect T cell responsiveness to antigen in an in vitro senescence model. Tetanus toxoid (TT)-specific T cell lines were derived from 3 young (< 30 years) and 3 old (> 65 years) individuals and were kept in long term culture. T cell proliferation in response to stimulation with antigen presented by either autologous peripheral blood mononuclear cells (PBMC) or DC was assessed at three different time points, once soon after the initiation of the cultures and twice after 20 to 30 population doublings at a stage when growth, was slow and programmed cell death imminent. Antigen presentation by DC enhanced T cell proliferation at each time point and reinduced proliferation in in vitro aged T cell populations which had stopped dividing. Terminal apoptosis was thus prevented. DC from old individuals were as effective as cells from young donors. Our results demonstrate that DC stimulate the clonal expansion and postpone the clonal elimination of antigen-specific T cell populations. As a consequence they may increase immunoreactivity, prolong immunological memory and be of particular importance for the maintenance of the T cell repertoire in old age.     

Distinct populations of dendritic cells are present in the subepithelial dome and T cell regions of the murine Peyer's patch

Despite the fact that the Peyer's patch (PP) is the primary site for antigen uptake in the intestine, the cellular basis of antigen handling after transport into the PP is poorly understood. We performed immunohistology of murine PPs using the dendritic cell (DC)-reactive monoclonal antibodies N418, NLDC-145, M342, and 2A1, as well as antibodies to other T cell, B cell, and macrophage markers. N418+, 2A1+, NLDC-145-, M342- cells form a dense layer of cells in the subepithelial dome (SED), just beneath the follicle epithelium, and are scattered throughout the follicle, sparing the germinal center. In contrast, N418+, 2A1+, NLDC-145+, and M342+ DCs are present in the interfollicular T cell regions (IFR). CD3+ and CD4+, but no CD8+ T cells were present in the SED and the follicle, including the germinal center, while CD3+, CD4+, and CD8+ T cells were present in the IFR. B cells and macrophages were poorly represented in the SED as no B220+ cells, only few Mac-1lo cells, and no F4/80+ cells were present at this site. In contrast, Mac-1hi cells were found in the IFR and lamina propria of intestinal villi, while F4/80+ cells were found only in the latter. In further phenotypic studies, we analyzed surface molecules of PP and spleen DCs by flow cytometry and found that these cells had similar fluorescence profiles when stained with N418, NLDC-145, and 33D1 DC-reactive antibodies, and antibodies to the costimulatory molecules B7-1 (1G10) and B7-2 (GL1). In contrast, PP DCs expressed 5-10-fold higher levels of major histocompatibility complex class II antigens (IEk) than spleen DCs. Finally, in functional studies, we demonstrated that both PP and spleen DCs process soluble protein antigens during overnight culture and induce similar levels of proliferation in CD3+ T cells, and CD4+/Mel 14hi T cells from T cell receptor transgenic mice. The in vivo relevance of such presentation was shown by the fact that PP DCs isolated from Balb/c mice after being fed ovalbumin stimulated proliferation in ovalbumin T cell receptor T cells. Taken together, our data suggest that DCs in the SED of the PP are uniquely positioned for the processing of antigens passed into the PP from the overlying M cell, and that PP DCs are effective at processing and presenting oral antigens to naive T cells.     

Selective elimination of the delayed-type hypersensitivity response by extracorporeal thoracic duct filtration

Calves were sensitized to tuberculin and histoplasmin. The delayed-type hypersensitivity skin response to these antigens was produced and the diameter of induration measured. Repeated skin tests prior to filtration demonstrated that the amount of induration produced by these skin tests was closely reproducible. Histoplasmin or tuberculin-coated columns were then introduced into a closed circuit extracorporeal thoracic duct circulation. A significant (P is less than 0.01) reduction or ablation of the delayed-type hypersensitivity response was obtained to the antigen used to coat the column. In contrast, no significant reduction occurred in the skin test response to the other antigen. Repeated skin tests to both antigens after the cessation of filtration showed a gradual rise toward prefiltration levels in the skin test to the filtered antigen. The results of these experiments indicate that a selective population of T lymphocytes can be removed from an in vivo system. The removal of these cells can selectively reduce a delayed-type hypersensitivity skin test response to a particular antigen. These results are consistent with the hypothesis that a type of in vivo selective immunosuppression can be produced by antigen-coated columns when they are placed in an extracorporeal thoracic duct lymph circulation system.     

Th1 cells induce and Th2 inhibit antigen-dependent IL-12 secretion by dendritic cells

Dendritic cells are the most relevant antigen-presenting cells (APC) for presentation of antigens administered in adjuvant to CD4+ T cells. Upon interaction with antigen-specific T cells, dendritic cells (DC) expressing appropriate peptide-MHC class II complexes secrete IL-12, a cytokine that drives Th1 cell development. To analyze the T cell-mediated regulation of IL-12 secretion by DC, we have examined their capacity to secrete IL-12 in response to stimulation by antigen-specific Th1 and Th2 DO11.10 TCR-transgenic cells. These cells do not differ either in TCR clonotype or CD40 ligand (CD40L) expression. Interaction with antigen-specific Th1, but not Th2 cells, induces IL-12 p40 and p75 secretion by DC. The induction of IL-12 production by Th1 cells does not depend on their IFN-gamma secretion, but requires direct cell-cell contact mediated by peptide/MHC class II-TCR and CD40-CD40L interactions. Th2 cells not only fail to induce IL-12 secretion, but they inhibit its induction by Th1 cells. Unlike stimulation by Th1, inhibition of IL-12 production by Th2 cells is mediated by soluble molecules, as demonstrated by transwell cultures. Among Th2-derived cytokines, IL-10, but not IL-4 inhibit Th1-driven IL-12 secretion. IL-10 produced by Th2 cells appears to be solely responsible for the inhibition of Th1 -induced IL-12 secretion, but it does not account for the failure of Th2 cells to induce IL-12 production by DC. Collectively, these results demonstrate that Th1 cells up-regulate IL-12 production by DC via IFN-gamma-independent cognate interaction, whereas this is inhibited by Th2-derived IL-10. The inhibition of Th1 -induced IL-12 production by Th2 cells with the same antigen specificity represents a novel mechanism driving the polarization of CD4+ T cell responses.