Antigen processing for presentation to CD4+ T cells.

The immune system surveys the organism for the presence of foreign or abnormal structures. An important role in the immune response is assumed by T lymphocytes that recognize foreign antigen while tolerating self-proteins. T lymphocytes can recognize only peptide fragments that are presented to them by molecules of the major histocompatibility complex (MHC). Antigen processing for presentation to T cells involves distinct cellular compartments where peptides and MHC molecules interact. Whereas class I MHC molecules (recognized by CD8+ cytotoxic T cells) acquire peptides in an early biosynthetic compartment, class II molecules (recognized by CD4+ helper T cells) acquire peptides most efficiently in an endocytic compartment. It has emerged recently that the class II processing compartment can be fed not only from the outside with exogenous antigen but also from endogenous sources, including membrane-associated and cytosolic proteins. The potential sources of proteins that can trigger a helper T cell response during viral infections and that can induce self-tolerance are thus much wider than previously anticipated.     

Adherent Ia+ murine cell lines with characteristics of dendritic cells. II. Characteristics of I region-restricted antigen presentation

The ability of an adherent Ia+, interleukin 1+ (IL-1) tumor cell line (P388AD) to present turkey gamma-globulin (TGG) to primed T lymphocytes was demonstrated and compared with normal antigen-presenting cells (APC) found in mouse spleen. P388AD tumor cells presented TGG to long-term cultures of TGG-reactive T cells (LTTC) and to lymph node-derived T cells which were enriched on nylon wool columns and subsequently depleted of endogenous antigen-presenting cells with anti-Ia antisera and complement. MHC-restricted antigen presentation by P388AD was observed when long-term cultures of TGG-reactive T cells were used as the responding T-cell population. Furthermore, antisera directed against I-region determinants expressed on the P388AD tumor cells inhibited TGG-specific T-cell proliferation in a dose-related fashion, suggesting a functional role for the tumor cell-associated Ia molecules. The kinetics of antigen presentation to LTTC by P388AD were similar to the kinetics observed for splenic APC, although the magnitude of the proliferative response to LTTC to TGG was generally lower when antigen (Ag) was presented by the tumor cells compared to splenic antigen-presenting cells (APC). However, the magnitude of T-cell proliferation of immune lymph node (LN) T cells was comparable when Ag was presented on tumor cells or splenic APC. Several experiments suggested that Ag uptake and/or processing may be less effective in P388AD tumor cells as compared to normal splenic APC. A nonadherent Ia+, IL-1- tumor cell line (P388NA), which was isolated from the same parental tumor as P388AD, was also tested for the ability to present Ag to primed T lymphocytes and Ag-reactive LTTC. In contrast, to P388AD, the nonadherent tumor cell failed to present TGG under identical culture conditions even though Ia molecules were expressed on the tumor cells and Ag uptake had occurred. However, the defect in Ag presentation by P388NA could be corrected if an exogenous source of purified interleukin 1 was supplied to the cultures. A unique opportunity thus exists with both the P388AD and P388NA tumor cell lines to decipher some of the molecular interactions leading to T-cell proliferation during antigen presentation.     

Antigen presentation in the rat: role of a nonadherent, nonphagocytic, W3/13, OX-6 positive T cell in the presentation of antigen to primed T lymphocytes

The nature of accessory cells in rat lymph nodes which can present antigen to primed T cells was investigated. Removal of adherent, phagocytic cells from antigen-primed lymph node cells by passage over glass-bead and nylon wool columns followed by treatment with carbonyl iron did not abrogate the antigen-specific proliferative response to keyhole limpet hemocyanin (KLH) or to the synthetic polypeptide L-glutamic acid-L-alanine-L-tyrosine (GAT). This T cell-enriched population was free of contaminating macrophages as determined by latex bead ingestion and morphological criteria during a 4-day culture period. Treatment of the T cell preparation with rabbit anti-rat IgG and complement or rosetting with IgG-coated sheep erythrocytes to remove any remaining B cells or macrophages did not significantly affect the proliferative response to antigen. Analysis of the T cell preparation by panning techniques with monoclonal antibodies to T cell surface markers suggested that both the responding T cell and the antigen-presenting cell were positive for the rat T cell marker, W3/13. The KLH-primed LN T cell-enriched fraction contained two distinct cell populations that were separable on the basis of their reactivity to OX-6 antibody. Two populations, an OX-6+ and an OX-6-, interacted synergistically in a KLH-dependent in vitro proliferative response. The cells within the T cell-enriched fraction that were positive for the OX-6 marker functioned primarily as the APCs, while the OX-6- cell fraction contained cells that proliferated to antigen when OX-6+ cells from either the T cell fraction or the adherent fraction were present. The implications of these findings are discussed.     

CD8+ T cells can be primed in vitro to produce IL-4.

IL-4 production by T lymphocytes from naive mice in response to stimulation by plate-bound anti-CD3 is concentrated among CD4+ T cells. In vitro stimulation of lymph node T cells with anti-CD3 plus IL-2 and IL-4 strikingly increases the frequency of cells that produce IL-4 in response to subsequent stimulation with anti-CD3 plus IL-2. Separation of these primed cell populations into CD4+ and CD8+ T cell by cell sorting reveals that the frequency of IL-4-producing cells in both population is similar. Verification that CD8+ T cells produce IL-4 is provided by the capacity of anti-IL-4 mAb to inhibit the response of the indicator cell line to the growth factor produced by the primed cells and by detection of IL-4 by an IL-4-specific ELISA. The in vitro "priming" of CD8+ T cells to produce IL-4 is not dependent on the presence of CD4+ T cells because highly purified CD8+ T cells can be stimulated to develop into cells capable of producing IL-4 by culture with plate-bound anti-CD3 plus IL-2 and IL-4.     

Murine thyroid follicular epithelial cells can be induced to express class II (Ia) gene products but fail to present antigen in vitro

To determine whether thyroid follicular epithelial cells (TFEC) might be involved in the induction of autoimmune thyroiditis, they were tested for their potential to express Ia antigens, and for their ability to present antigen in vitro. Results showed that Ia antigens, absent on normal TFEC, could be readily induced with interferon gamma, as detected by immunofluorescence. Maximal expression of Ia antigens in over 50% of TFEC was observed after 4 days of culture in the presence of IFN-gamma, and was quantitatively comparable to spleen cells by cytofluorometric analysis. Moreover, primary TFEC in culture secreted thyroglobulin (tg) and interleukin 1. However, TFEC consistently failed to stimulate various populations of T cells. These included lymph node cells sensitized to tg, a T-cell clone specific for azo-benzene-arsonate tyrosine (ABA), and a hybridoma specific for beef insulin. Likewise, Ia-positive TFEC did not stimulate T-cell hybridomas restricted to the class II alloantigen I-Ab, while stimulating a hybridoma specific for the class I alloantigen Kb. T-cell unresponsiveness could not be explained by inhibitory activity of TFEC, released either into the culture supernatant or exerted by cell contact. The data indicate that Ia-positive TFEC failed to serve as class II-restricted antigen-presenting cells (APC) in vitro and thus argue against a primary role for these cells in the inductive phase of thyroiditis.     

Antigen presentation by Toxoplasma gondii-infected cells to CD4+ proliferative T cells and CD8+ cytotoxic cells

Cytotoxic cells specific for Toxoplasma gondii-infected cells were detected in the peripheral blood leukocytes from a patient with acute toxoplasmosis. The cytotoxicity was mediated by CD5+, CD4-, CD8+ cells. The cytotoxic T cells lysed Toxoplasma-infected target cells with HLA class I restriction. Two types of T cell clones were established from peripheral blood leukocytes of a patient with chronic toxoplasmosis; one was a CD5+, CD4-, CD8+ cytotoxic cell specific for Toxoplasma-infected cells, and the other was a CD5+, CD4+, CD8- proliferative cell that responded to Toxoplasma antigen. Toxoplasma-infected cell-specific cytotoxic cloned T cells recognize the infected target cells in the context of the HLA class I molecules, and the CD8 molecule was involved in the cytotoxicity. Toxoplasma antigen-specific proliferative cloned T cells were stimulated by Toxoplasma antigen-pulsed or Toxoplasma-infected cells in conjunction with HLA-DR molecule on the target cells. Thus, antigen presentation by Toxoplasma-infected cells for activation of both cytotoxic and proliferative T cells has been demonstrated.     

Antigen presentation to human T lymphocytes. II. Requirements for Mac-120+ macrophages and responsiveness to interleukin 2

To investigate mechanisms by which antigen, macrophages, and interleukin 2 (IL2) participate in the induction of secondary T-cell proliferative responses, trinitrophenyl (TNP) was presented in three distinct modes: (i) TNP-modified peripheral blood mononuclear cells (TNP-PBMC), (ii) TNP-PBMC cell sonicates, and (iii) TNP-ovalbumin (TNP-OVA). Stimulators were depleted of Mac-120+ macrophages using Mac-120 monoclonal antibody plus complement. TNP-Mac-120 macrophages stimulated primed T cells nearly as well as TNP-unfractionated macrophages (which were about 40% Mac-120+). In contrast, although greater than 70% DR+, Mac-120- macrophages plus either TNP-OVA or TNP-PBMC sonicate elicited minimal responses compared to unfractionated macrophages plus antigen. After 21-28 days of in vitro priming, macrophage-depleted T cells were not stimulated to proliferate by either IL2 alone or sonicates alone. IL2 plus TNP-PBMC sonicates, however, stimulated significant proliferation. Furthermore, this response was considerably greater than that to IL2 plus either TNP-T cell sonicates or TNP-mouse spleen sonicates. Thus, the Mac-120+ macrophage population may have an important antigen-presenting and/or accessory function in the stimulation of primed T cells by soluble or particulate antigen, although it is unnecessary for responses to intact TNP-Ia+ PBMC. In addition, the data suggest that Ia+ sonicates alone may suffice for induction of IL2 responsiveness, but not for endogenous IL2 production and subsequent proliferation by primed T cells.     

Antigen presentation by hapten-specific B lymphocytes. IV. Comparative ability of B cells to present specific antigen and anti-immunoglobulin antibody

The ability of trinitrophenyl (TNP)-binding murine B lymphocytes to present native rabbit IgG (RGG), TNP-modified RGG, and rabbit anti-mouse Ig (RAMG) to an Ia-restricted, RGG-specific helper/inducer T cell clone was compared. By three independent assays (lymphokine secretion, T cell proliferation, and B cell differentiation), TNP-RGG was presented at 10(2)- to 10(3)-fold lower concentrations than RGG, and RAMG at 10(2)- to 10(3)-fold lower concentrations than TNP-RGG. The available data suggest that the efficiency of antigen presentation is dependent primarily on the avidity of binding of a ligand to B cell surface Ig and/or the extent of subsequent endocytosis (modulation). Despite the observed quantitative differences between anti-Ig (RAMG) and specific antigen (TNP-RGG), these results demonstrate that qualitatively both are essentially similar in their ability to mediate specific T-B interactions. Thus, anti-Ig antibodies are valid models for analyzing cognate interactions between antigen-specific B and helper T lymphocytes.     

Induction of peripheral T cell tolerance by antigen-presenting B cells. II. Chronic antigen presentation overrules antigen-presenting B cell activation

Ag presentation in the absence of danger signals and Ag persistence are the inductive processes of peripheral T cell tolerization proposed so far. Nevertheless, it has never been definitively shown that chronic Ag presentation per se can induce T cell tolerance independent of the state of activation of APCs. In the present work, we investigated whether chronic Ag presentation by either resting or activated B cells can induce tolerance of peripheral Ag-specific T cells. We show that CD4(+) T cells that re-encounter the Ag for a prolonged period, presented either by resting or activated Ag-presenting B cells, become nonfunctional and lose any autoimmune reactivity. Thus, when the main APCs are B cells, the major mechanism responsible for peripheral T cell tolerization is persistent Ag exposure, independent of the B cell activation state.     

Arsonate-specific murine T cell clones. V. Antigen presentation by L cells transfected with normal and mutant class II genes

Class II-restricted murine T cell clones specific for the immunogenic determinant L-tyrosine-p-azobenzenearsonate failed to proliferate to Ag presented by L cell lines transfected with and expressing the appropriate class II genes, but are activated to kill the APC in an Ag-dependent, MHC-restricted manner. Inhibition of APC proliferation was used as an assay to determine the relative contributions of polymorphic sites on the class II alpha- and beta-chains to MHC-restricted activation of I-A beta k-restricted cloned T cells. Transfectants expressing A beta k in conjunction with the alpha chain of k, u, or d were equally effective APCs, whereas transfectants expressing A beta u were completely ineffective, implicating the beta-chain as more critical for the presentation of L-tyrosine-p-azobenzenearsonate. Site-directed mutagenesis of polymorphic positions in the beta chain revealed a remarkable stringency for the k haplotype, in contrast to the relaxed alpha-chain requirement. These results, in conjunction with others, indicate that the relative contribution of polymorphic sites on class II alpha- and beta-chains to T cell Ag recognition can differ markedly, and, furthermore, may vary as a function of the Ag.     

LILRA2 activation inhibits dendritic cell differentiation and antigen presentation to T cells

The differentiation of monocytes into dendritic cells (DC) is a key mechanism by which the innate immune system instructs the adaptive T cell response. In this study, we investigated whether leukocyte Ig-like receptor A2 (LILRA2) regulates DC differentiation by using leprosy as a model. LILRA2 protein expression was increased in the lesions of the progressive, lepromatous form vs the self-limited, tuberculoid form of leprosy. Double immunolabeling revealed LILRA2 expression on CD14+, CD68+ monocytes/macrophages. Activation of LILRA2 on peripheral blood monocytes impaired GM-CSF induced differentiation into immature DC, as evidenced by reduced expression of DC markers (MHC class II, CD1b, CD40, and CD206), but not macrophage markers (CD209 and CD14). Furthermore, LILRA2 activation abrogated Ag presentation to both CD1b- and MHC class II-restricted, Mycobacterium leprae-reactive T cells derived from leprosy patients, while cytokine profiles of LILRA2-activated monocytes demonstrated an increase in TNF-alpha, IL-6, IL-8, IL-12, and IL-10, but little effect on TGF-beta. Therefore, LILRA2 activation, by altering GM-CSF-induced monocyte differentiation into immature DC, provides a mechanism for down-regulating the ability of the innate immune system to activate the adaptive T cell response while promoting an inflammatory response.     

Antigen presentation by Langerhans cells in vivo: donor-derived Ia+ Langerhans cells are required for induction of delayed-type hypersensitivity but not for cytotoxic T lymphocyte responses to alloantigens

T cell activation in response to allogeneic stimulation and hapten-specific delayed-contact hypersensitivity responses in vivo can be initiated by Ia-bearing epidermal Langerhans cells (LC). By using a murine heterotopic corneal allograft model, we have investigated the requirement for allogeneic LC as antigen-presenting cells (APC) in the in vivo induction of delayed-type hypersensitivity (DTH) and cytolytic T lymphocyte (CTL) responses to alloantigens in fully allogeneic and H-2 I region-disparate strain combinations. LC-deficient, avascular central corneal allografts from BALB/c donors failed to induce DTH responsiveness when grafted to a subdermal bed on C57BL/6 recipients (p greater than 0.05), yet antigen-specific primary CTL reactivity developed within 7 days after grafting. LC-containing corneal-limbus allografts or central corneal allografts containing a latex bead-induced infiltrate of LC resulted in intense DTH as well as CTL responsiveness when grafted in this same strain combination. Similarly, LC-containing but not LC-deficient corneal allografts from A.TL donors induced DTH responsiveness in I region-disparate A.TH hosts despite the fact that these grafts survived for prolonged duration (less than 28 days). By contrast, CTL induction in I region-disparate hosts was independent of the presence of allogeneic LC. Corneal epithelial cells of grafts removed from I region-disparate hosts 7 days posttransplantation were shown by immunohistology to express the Iak antigens of donor origin. The possibility that bone marrow-derived allogeneic LC were a sufficient requirement for DTH induction was confirmed in experiments performed with CB6F1----B6 bone marrow chimeras used as corneal allograft donors. Corneal-limbus grafts obtained from mice 90 days after chimerization were shown by immunohistology to contain Iad-bearing CB6F1 LC as a sole source of class II alloantigens. When grafted to C57BL/6 recipients, LC-containing chimeric corneas induced DTH responsiveness that was similar in magnitude to that observed in C57BL/6 mice grafted with chimeric skin, yet no DTH response to LC-deficient chimeric central corneal grafts was observed. Moreover, in all cases, the chimeric corneal and skin allografts survived for prolonged duration (greater than 28 days). These results demonstrate that donor-derived LC act as APC in the induction of DTH responsiveness to allogeneic tissue; however, there was no apparent requirement for allogeneic LC in the induction of CTL responses to class I or class II MHC alloantigens.(ABSTRACT TRUNCATED AT 400 WORDS)     

Accessory cell function of human endothelial cells. I. A subpopulation of Ia positive cells is required for antigen presentation

The expression of class I and class II HLA antigens on preparations of human endothelial cells, isolated from umbilical cord veins, was investigated by immunofluorescence. While virtually all endothelial cells expressed class I antigens, less than 1% were positive for class II antigens, as detected with a panel of 10 different monoclonal antibodies. Antigen specific T cell lines proliferated in response to mumps antigen in the presence of endothelial cells or blood monocytes from HLA-DR matched donors. However, these T cell lines failed to respond in the absence of accessory cells or when accessory cells from HLA-D-region mismatched cord donors were used. The ability of both monocytes and endothelial cells to present antigen was abolished by treatment of the cells with monoclonal antibodies specific for either class I or class II HLA antigens plus complement. Similar treatment with monoclonal antibodies specific for monocytes greatly reduced antigen presentation by endothelial cells. These results indicate that preparations of endothelial cells contain a subpopulation of Ia positive cells, distinct from monocytes, which are required for antigen presentation.     

Astrocytes as antigen-presenting cells. I. Induction of Ia antigen expression on astrocytes by T cells via immune interferon and its effect on antigen presentation

Ia antigens seem to control immune responses on at least two levels. First, they influence the antigen recognition repertoire of the T cells. Second, their variable expression on certain antigen-presenting cells is a powerful regulatory mechanism for the local immune reaction. This is particularly important in the central nervous system (CNS) in which no Ia antigens are normally expressed. Recent experiments in this context have shown that astrocytes are able to express Ia antigens during interaction with T cells, and that they function as antigen-presenting cells. The Ia-inducing activity is produced by activated T cells, and can be replaced by immune interferon (IFN-gamma). In this study we report on the functional and kinetic relationship between Ia antigen expression on astrocytes and the immune-specific activation of T cells by astrocytes. Normal resting astrocytes were found to be negative for Ia antigens by immunofluorescence and by biochemical criteria. Moreover, they are only able to stimulate T cells after they have been induced to express Ia antigens by a signal from the T cells, which is probably mediated by IFN-gamma. In conclusion, the immune-specific interaction between astrocytes and T lymphocytes is a sensitively controlled system that might be pivotal to the development of immune responses in the brain. Malfunction of the system could be an important factor in the pathogenesis of aberrant immune reactions in the CNS, e.g., in multiple sclerosis.     

Antigen presentation by liver cells controls intrahepatic T cell trapping, whereas bone marrow-derived cells preferentially promote intrahepatic T cell apoptosis.

Systemic activation and proliferation of CD8(+) T cells result in T cell accumulation in the liver, associated with T cell apoptosis and liver injury. However, the role of Ag and APC in such accumulation is not clear. Bone marrow chimeras were constructed to allow Ag presentation in all tissues or alternatively to restrict presentation to either bone marrow-derived or non-bone marrow-derived cells. OVA-specific CD8(+) T cells were introduced by adoptive transfer and then activated using peptide, which resulted in clonal expansion followed by deletion. Ag presentation by liver non-bone marrow-derived cells was responsible for most of the accumulation of activated CD8(+) T cells. In contrast, Ag presentation by bone marrow-derived cells resulted in less accumulation of T cells in the liver, but a higher frequency of apoptotic cells within the intrahepatic T cell population. In unmodified TCR-transgenic mice, Ag-induced T cell deletion and intrahepatic accumulation of CD8(+) T cells result in hepatocyte damage, with the release of aminotransaminases. Our experiments show that such liver injury may occur in the absence of Ag presentation by the hepatocytes themselves, arguing for an indirect mechanism of liver damage.     

Two roles for Ia in antigen-specific T cell activation. II. Toward a Velcro model of antigen recognition

It has been previously reported that Ia Ag on APC seems to be involved in Ag-specific T cell activation in at least two different ways: one is to associate with foreign Ag to form a neoantigenic determinant (the Ag-specific Ia function), and the second is to interact with T cells in a non-Ag-specific manner. Both Ia functions are required for T cell activation. In the present study we examined whether the T cell structures responsible for the non-Ag-specific Ia interaction were separable from the Ag-specific alpha/beta TCR. Purified protein derivative of tuberculin (PPD)-specific murine hybridoma T cells and polyclonal lymph node T cells were stimulated for IL-2 production by APC pulsed with PPD, glutaraldehyde fixed, and anti-Ia antibody treated, to provide the antigenic PPD/Ia determinant, in the presence of glutaraldehyde-fixed non-Ag-pulsed APC, to provide the non-Ag-specific Ia interactions. However, in several different approaches the T cell structures or activation signals responsible for the Ag-specific recognition and non-Ag-specific Ia interactions seemed to be associated with each other in this experimental system. First, the Ag-specific and non-Ag-specific Ia interactions with T cells were both required simultaneously to initiate T cell activation, and it was not possible to activate T cells by providing either Ia signal subsequent to the other. Second, the T cell structures responsible for the non-Ag-specific Ia interactions appeared to be clonally distributed in PPD-specific lymph node T cells. Third, another T cell hybridoma specific for bovine insulin also showed dual Ia interactions, but the specificity of the non-Ag-specific Ia function was different than that for the PPD-specific T cell response. Fourth, all subclones of PPD-specific T hybridomas that had lost Ag-specific responsiveness also lost functional non-Ag-specific Ia interactions. Taken together, these observations suggest that a single species of TCR may mediate both the Ag-specific and non-Ag-specific Ia interactions. In addition, the non-Ag-specific Ia interaction with T cells augmented the Ag-specific Ia interaction for T cell activation, indicating that both types of interactions may be involved in some T cell responses. Based on these observations, a Velcromodel depicting the synergy between the two Ia functions is proposed in which a matrix of interactions consisting of higher affinity Ag binding and lower affinity Ia-TCR associations provides cooperative sets of signals necessary for cellular activation.     

HLA-DQ-restricted CD4+ T cells specific to streptococcal antigen present in low but not in high responders.

We have found that the low immune response to streptococcal cell wall Ag (SCW) was inherited as a dominant trait and was linked to HLA, as deduced from family analysis. In the present report, HLA class II alleles of healthy donors were determined by serology and DNA typing to identify the HLA alleles controlling low or high immune responses to SCW. HLA-DR2-DQA1*0102-DQB1*0602(DQw6)-Dw2 haplotype or HLA-DR2-DQA1*0103-DQB1*0601(DQw6)-DW12 haplotype was increased in frequency in the low responders and the frequency of HLA-DR4-DRw53-DQA1*0301-DQB1*0401(DQw4)-Dw15 haplotype or HLA-DR9-DRw53-DQA1*0301-DQB1*0303(DQw3)-Dw23 haplotype was increased in the high responders to SCW. Homozygotes of either DQA1*0102 or DQA1*0103 exhibited a low responsiveness to SCW and those of DQA1*0301 were high responders. The heterozygotes of DQA1*0102 or 0103 and DQA1*0301 showed a low response to SCW, thereby confirming that the HLA-linked gene controls the low response to SCW, as a dominant trait. Using mouse L cell transfectants expressing a single class II molecule as the APC, we found that DQw6(DQA1*0103 DQB1*0601) from the low responder haplotype (DR2-DQA1*0103-DQB1*0601(DQw6)-Dw12) activated SCW-specific T cell lines whereas DQw4(DQA1*0301 DQB1*0401) from the high responder haplotype (DR4-DRw53-DQA1*0301-DQB1*0401(DQw4)-Dw15) did not activate T cell lines specific to SCW. However, DR4 and DR2 presented SCW to CD4+ T cells in both the high and low responders to SCW, hence the DR molecule even from the low responder haplotype functions as an restriction molecule in the low responders. Putative mechanisms linked to the association between the existence of DQ-restricted CD4+ T cells specific to SCW, and low responsiveness to SCW are discussed.     

A subset of mouse splenic macrophages can constitutively present alloantigen directly to CD8+ T cells.

About a third of mouse splenic macrophage (M phi) progenitors give rise to cloned progeny that constitutively induce the selective proliferation of naive allogeneic CD8+ T cells in a CD4+ helper cell-independent manner--a response that is inhibited by mAb to the MHC class I molecules present on the M phi. Colony-mixing experiments indicated that the failure of most M phi clones to present allo-Ag was not due to their suppression of the ability of CD8+ cells to respond, nor did the nonpresenting clones interfere with the activity of the allo-Ag presenting M phi. The allo-Ag presenting phenotypes were found to be a stable characteristic in a panel of cell lines derived from individual clones of M phi. Analysis of the cell lines revealed that the differential expression of allo-APC activity could not be attributed to the levels of MHC class I molecules; rather, the cell lines and the primary M phi clones differ in their expression of a cell-associated costimulator molecule that likely functions to induce the expression of the IL-2R on and the secretion of IL-2 from the T cells.     

Salmonella typhimurium coordinately regulates FliC location and reduces dendritic cell activation and antigen presentation to CD4+ T cells

During infection, Salmonella transitions from an extracellular-phase (STEX, growth outside host cells) to an intracellular-phase (STIN, growth inside host cells): changes in gene expression mediate survival in the phagosome and modifies LPS and outer membrane protein expression, including altered production of FliC, an Ag recognized by immune CD4+ T cells. Previously, we demonstrated that systemic STIN bacteria repress FliC below the activation threshold of FliC-specific T cells. In this study, we tested the hypothesis that changes in FliC compartmentalization and bacterial responses triggered during the transition from STEX to STIN combine to reduce the ability of APCs to present FliC to CD4+ T cells. Approximately 50% of the Salmonella-specific CD4+ T cells from Salmonella-immune mice were FliC specific and produced IFN-gamma, demonstrating the potent immunogenicity of FliC. FliC expressed by STEX bacteria was efficiently presented by splenic APCs to FliC-specific CD4+ T cells in vitro. However, STIN bacteria, except when lysed, expressed FliC within a protected intracellular compartment and evaded stimulation of FliC-specific T cells. The combination of STIN-mediated responses that reduced FliC bioavailability were overcome by dendritic cells (DCs), which presented intracellular FliC within heat-killed bacteria; however, this ability was abrogated by live bacterial infection. Furthermore, STIN bacteria, unlike STEX, limited DC activation as measured by increased MHC class II, CD86, TNF-alpha, and IL-12 expression. These data indicate that STIN bacteria restrict FliC bioavailability by Ag compartmentalization, and together with STIN bacterial responses, limit DC maturation and cytokine production. Together, these mechanisms may restrain DC-mediated activation of FliC-specific CD4+ T cells.