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Query: UMLS:C0033036 (APC)
 10,214 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effect of radiation on three discrete Ag-presentation functions in resting B cells was examined: 1) Ag uptake and processing, 2) expression of processed Ag in the context of functional class II molecules, and 3) provision of necessary co-stimulatory, or "second," signals. Analysis of radiation's effect on B cell presentation of intact vs fragmented Ag or its effect on presentation by Ag-pulsed B cells indicated that damage to Ag uptake and processing could not account for the bulk of the radiation-induced Ag-presentation defect. Experiments with phosphatidylinositol hydrolysis as an indirect measure of TCR occupancy suggested that irradiation caused a fairly rapid (within 1 to 2 h) decrease in the ability of the B cell APC to display a stimulatory combination of Ag and class II molecule. Ag dose-response analyses demonstrated that when presenting a fragment of the Ag pigeon cytochrome c to a T cell clone, 3000 rad-treated B cell APC were able to stimulate approximately 50% as much phosphatidylinositol turnover as unirradiated B cells. It was also found that, in contrast to their inability to initiate T cell proliferation, and similarly to chemically cross-linked splenocytes, heavily irradiated resting B cells plus Ag induced a state of Ag hyporesponsiveness in T cell clones. This effect on T cells had the same Ag- and MHC-specificity as did receptor occupancy required for proliferation, indicating that heavily irradiated resting B cells bear functional class II molecules. Co-culture of T cells with allogeneic B cells and syngeneic heavily irradiated B cells or chemically cross-linked splenic APC plus Ag resulted in T cell proliferation and interfered with the induction of the hyporesponsive state. This co-stimulatory function was radiosensitive in resting allogeneic B cells. Together, these data support the hypothesis that the major functional consequences of radiation to resting B cell APC are a reduction in the effective display of Ag plus class II molecules and, probably what is more important, a loss in the ability to provide APC-derived co-stimulatory signals.
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PMID:Effect of gamma radiation on resting B lymphocytes. II. Functional characterization of the antigen-presentation defect. 284 2

The experiments described here were designed to investigate the role of intracellular acidic organelles in the processing of foreign antigens. Simultaneous addition of antigen and the inactive precursor of an acid protease, cathepsin L, greatly diminished the stimulation of a T-cell hybridoma specific for pigeon cytochrome c. The effect occurred during antigen processing as the presentation of a peptide fragment was unaffected by precursor-cathepsin L. The effect required uptake of the enzyme via mannose-6-PO4 receptors and was specific for antigenic determinants cleavable by the enzyme. The results confirm, without any pharmacologic manipulation, that foreign antigen moves into an acidic environment during some phase of its processing. We also explored the role in antigen processing of one acidic compartment, the early endosome, by examining CHO cells expressing a temperature-sensitive defect in endosomal acidification. These cells were transfected with MHC class II genes to convert them into APC for CD4+T cells. When these cells were incubated at the nonpermissive temperature, their ability to process antigen was impaired. There was no negative effect on presentation of antigenic peptides nor on the uptake and degradation of the antigen. Wildtype CHO cells similarly transfected and tested did not show any aberrancy in antigen processing. The inhibition of processing by the mutant cells was only partial, even under stringent conditions, and the residual processing was eliminated by chloroquine. Our findings provide the first evidence for a contribution by acidified early endosomes to the pathway of antigen processing and suggest that other acidified compartments are also involved.
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PMID:The role of intracellular acidification in antigen processing. 307 87

Here we review our current results studying B cells as APC and the mechanisms by which processed antigen is transported to and held on the cell surface for recognition by the specific T cell along with the MHC class II molecules. These studies were carried out using the globular protein cytochrome c as antigen for which the T-cell antigenic determinant was localized to a C-terminal 10-amino acid peptide fragment. For certain analyses, native cytochrome c or antigenic peptide fragments were covalently coupled to antibodies directed toward B-cell surface structures, allowing the targeting of antigen to the APC surface. Our findings indicate that all B cells function as APC and that the APC function is not differentially regulated in defined B-cell subpopulations. Using cytochrome c-antibody conjugates, it was shown that the surface Ig plays two significant roles in augmenting the B-cell APC function following antigen binding: signalling for enhanced APC function and concentrating antigen for subsequent internalization and processing. Both IgM and IgD appear to function identically in facilitating antigen processing in both immune and nonimmune B-cell populations. Furthermore, the surface Ig does not appear to be specially differentiated to function in concentrating antigen, as antigen artificially bound to other B-cell surface structures including MHC class I and class II molecules is also effectively presented. Lastly, evidence is presented that a previously described B-cell activating factor activity is strongly associated with the membranes of activated but not unactivated helper T cells, providing a mechanism by which the T-cell helper function can be focused on the specific antigen-presenting B cell. Concerning the mechanism by which processed antigen is presented at the B-cell surface, evidence is presented suggesting a role of peptide-binding chaperone proteins which may function to transport peptide to the APC surface and facilitate its association with the appropriate Ia. One candidate protein, PBP72/74, is described which binds peptides but not native antigens, is a member of the hsp70 family and appears to play a role in antigen presentation by the ability of antisera raised against it to block APC functions. Peptide-antibody conjugates were used to explore the spacial restrictions on MHC-restricted peptide presentation and it was shown that peptides covalently coupled to antibodies specific for Ig, class I or class II molecules are effective antigens in vitro even in the absence of processing.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Antigen-presenting function of B lymphocytes. 307 88

Efficient initiation of a CD4 T cell response requires both activation through the TCR and costimulation provided by molecules on APC with counterreceptors on the T cell. We investigated the relative contribution of the ICAM-1:LFA-1 and B7:CD28/CTLA-4 costimulatory pathways in naive T cell activation, using either anti-CD28 Ab or fibroblast cell lines transfected with I-Ek, which express either no costimulatory molecules, ICAM-1 alone, B7-1 alone, or ICAM-1 and B7-1 together. Peptide Ag or immobilized anti-CD3 was used to provide the TCR signal. CD4 T cells from mice transgenic for the V beta 3/V alpha 11 TCR, which recognize a peptide of pigeon cytochrome c complexed to I-Ek, were used as a source of naive T cells. Naive T cells stimulated with Ag or anti-CD3 responded well to high numbers of APC expressing either ICAM-1 alone or B7-1 alone. However, APC expressing both ICAM-1 and B7-1 were much better stimulators of proliferation and IL-2 secretion at low cell numbers, and were far superior inducers of IL-2 at higher numbers, indicating a synergy between the two pathways. Stimulation provided by ICAM-1 could not be solely attributed to adhesive strengthening of other pathways, since costimulation was seen when immobilized anti-CD3 was used and when ICAM-1 only APC were added, indicating that ICAM-1 was in fact acting as a classic costimulatory molecule. Both the magnitude of the response and the amount of costimulation required for response were dependent on the intensity of TCR interaction. These results suggest that an efficient naive T cell response requires both a strong TCR signal and more than one costimulatory signal that will synergize with the TCR signal. This offers an explanation as to why APC such as dendritic cells and activated B cells, which express high levels of multiple costimulatory/adhesion molecules, are the only APC that elicit naive T cell responses.
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PMID:Costimulatory requirements of naive CD4+ T cells. ICAM-1 or B7-1 can costimulate naive CD4 T cell activation but both are required for optimum response. 754 26

The Ag, pigeon cytochrome c, was coupled to human ferric transferrin by a heteroligation technique to target Ag into the endosomal transport pathway via transferrin receptors. The ability of various types of APC that do or do not express transferrin receptors to process exogenous Ag in their endosomes was investigated by the stimulation of Ag-specific CD4+ T cells with the transferrin-Ag conjugate in a serum-free assay. When two B lymphoma cells were the source of APC, the conjugate was significantly more potent than native Ag in activating the T cells, agreeing with our previous finding using a third B lymphoma cell. The conjugate and Ag were similarly presented by splenic B cells that lack transferrin receptors to the T cells. However, both a macrophage hybridoma and a MHC class II-L cell transfectant hardly elicited a T cell response to the conjugate, although a response to native Ag was readily observed. These findings could not be attributed to an absence of transferrin receptors or receptor-mediated internalization of the conjugate, nor to differential expression of MHC class II molecules or li chain by the APC. The poor presentation of the conjugate by the L cell transfectants was associated with diminished catabolism of the conjugate, however, the macrophage hybridoma rapidly degraded the conjugate, similar to the B lymphoma cell. Peritoneal macrophages, which lack transferrin receptors, and the macrophage hybridoma induced a response to the conjugate only at concentrations that allowed internalization by fluid phase pinocytosis. The lower potency of the conjugate compared with native Ag with non-B-presenting cells suggest that these cell types process the conjugate by a different mechanism than used by B cells. Differences in the mechanism of Ag processing used by APC of distinct cell lineages may possibly influence immune responsiveness.
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PMID:Differences among various lineages of antigen-presenting cells in processing exogenous antigen internalized through transferrin receptors. 790 98

Secondary responses to Ag in vivo are characterized by more rapid kinetics and greatly enhanced magnitude compared with primary responses. For CD4+ T cells, this is in part due to a greater frequency of Ag-specific memory cells, and may also reflect differences in responsiveness of memory vs naive cells to stimulation. To compare activation requirements and the role of accessory cells, naive and memory cells were stimulated with immobilized anti-CD3 in the presence or absence of APC. With anti-CD3 alone, naive cells proliferated slightly but produced no detectable IL-2, whereas memory cells proliferated well with significant IL-2 production. Increasing numbers of T-depleted APC greatly enhanced responses of naive cells to levels equivalent to those of memory cells, whereas for memory cells only IL-2 production increased slightly. The response of naive cells was equivalent in magnitude and kinetics to that of memory cells when low density APC, enriched in dendritic cells and depleted of resting B cells, were used with anti-CD3. To directly compare naive and memory responses in an Ag-specific model, we examined CD4+ cells specific for a peptide of pigeon cytochrome c fragment isolated from TCR-alpha beta transgenic mice. Naive cells were compared with 4-day activated blasts (effectors) and memory cells generated by adoptive transfer of effectors to adult thymectomized bone marrow reconstituted mice, in which the cells return to a resting state but still respond to recall Ag. Naive cells responded to Ag on dendritic cells and activated B cells but not on resting B cells or macrophages. In contrast, both memory cells and effectors were stimulated by all APCs, including resting B cells and macrophage to a limited extent. The ability of memory cells to respond to all APC types was confirmed using Ag-specific cells generated by in vivo priming with keyhole limpet hemocyanin. These results suggest that memory cells are considerably less dependent on accessory cell costimulation than naive cells, but that naive cells can respond equivalently in both magnitude and kinetics if Ag is presented on costimulatory APCs such as dendritic cells. In addition, these studies suggest that the enhanced secondary T cell response is due to a combination of the increased frequency of Ag-specific cells and their ability to react to Ag presented on a wider range of APC types, rather than an inherent capacity of memory T cells to respond better and faster.
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PMID:Naive versus memory CD4 T cell response to antigen. Memory cells are less dependent on accessory cell costimulation and can respond to many antigen-presenting cell types including resting B cells. 790 1

The role of endosomes in exogenous Ag processing was investigated by targeting Ag into the endosomal transport pathway via transferrin receptors. The Ag, pigeon cytochrome c and chicken OVA, were coupled to human ferric transferrin by a heteroligation technique. The conjugates were significantly more efficient than native Ag in stimulating Ag-specific CD4+ T cells, when the APC expressed transferrin receptors. The addition of ferric transferrin eliminated the enhanced response. Paraformaldehyde-fixed APC did not present the conjugates, indicating that the conjugates still required processing to activate T cells. An augmented level of T cell activation was not observed when the APC lacked transferrin receptors or when the conjugate contained the apoenzyme form of transferrin, which does not bind the receptor. The conjugate followed an intracellular pathway similar to that for transferrin, remaining in low density vesicles. Degraded conjugate appeared rapidly in culture supernatants, within 5 min, and peaked by 20 min; under these conditions a T cell response to the conjugate was elicited that was consistent with an early processing compartment. Our results suggest that antigenic peptide fragments can be generated in the early endosomes, without delivery of these Ag to the lysosomes. Thus, various Ag may have differential processing requirements, dictated by their molecular nature, that determine the site of Ag processing.
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PMID:Exogenous antigens internalized through transferrin receptors activate CD4+ T cells. 809 28

The hypothesis that reduction of disulfide bonds and exposure to low pH may be sufficient to allow intact proteins to bind class II MHC was addressed in this study. Functional assays were used to determine minimal conditions sufficient to bypass the requirement for Ag processing. Fixed APC, pulsed with HEL at pH 5 in the presence of a reducing agent, were observed to stimulate I-Ed-restricted T cell hybridomas. Activity required both low pH and reducing agent. Fixed APC also stimulated cytochrome c-specific T cells after exposure to Ag at pH 5 but not pH 7. No reducing agent was required. Peptide was considerably more potent in these assays than intact cytochrome c. By contrast, highly purified cytochrome c was equal in potency with a peptide in competition binding assays using purified I-Ek. The protein did not bind I-Ad or I-Ak. This suggested that cytochrome c can efficiently bind I-Ek at low pH without the requirement for proteolytic cleavage. The capacity of HEL to bind purified I-Ed was strictly dependent on the presence of reducing agent. The reducing agent alone had no effect. A small panel of proteins was screened for class II binding at low pH in competition assays. Horse myoglobin and human hemoglobin were particularly potent in their capacity to inhibit binding of biotin-labeled peptide to purified class II. Unlabeled peptide inhibited binding of labeled peptide to I-Ad at pH 5 and pH 7. By contrast, intact horse myoglobin was active only at pH 5. This result suggested that low pH may enhance the binding of horse myoglobin to I-Ad through effects on the structure of both proteins. Our observations support the hypothesis that low pH and disulfide reduction can be sufficient to allow partially unfolded or structurally destabilized proteins to bind class II MHC.
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PMID:Acidification and disulfide reduction can be sufficient to allow intact proteins to bind class II MHC. 838 84

The Ag, pigeon cytochrome c, was delivered to the early endosomes in a form coupled to human ferric transferrin that entered APC through transferrin receptors. The processing of the transferrin-Ag conjugate by B lymphoma cells was compared with that of unconjugated native Ag that entered APC by a nonreceptor-mediated mechanism. Within 5 min after internalization, catabolized conjugate was detected in isolated early endosomes and did not accumulate in these organelles. Analysis of the rapid catabolism of the conjugate demonstrated that the Ag, not the transferrin, portion of the molecule was degraded by the APC, suggesting that similar proteases may mediate the processing of the conjugate and native Ag. The processing mechanisms of these molecules shared similarities. Treatment of APC with chloroquine or paraformaldehyde interfered with the stimulation of Ag-specific CD4+ T cells by both transferrin-Ag conjugate and native Ag. However, the T cell responses to the conjugate and native Ag were different in two important respects. First, T cell activation by the conjugate began at an earlier time point and occurred at a faster rate than T cell stimulation by the same concentration of native Ag during a 3-h time course. Second, the T cell response to the conjugate, but not to native Ag, was diminished by treating APC with cycloheximide, a reversible protein synthesis inhibitor. This partial inhibition of the conjugate response by cycloheximide could not be attributed to significant effects on transferrin receptor expression, or on internalization, recycling, or degradation of the conjugate. The differential cycloheximide-sensitivity of the T cell responses indicates that the processing pathways of the two molecules are different. Our findings suggest that the early endosomes may function as an Ag-processing compartment, and that more than one pathway may lead to productive processing in B lymphoma cells.
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PMID:Antigen presentation by B lymphoma cells. Requirements for processing of exogenous antigen internalized through transferrin receptors. 840 20

Residues 46 and 54 on pigeon cytochrome c 43-58 (p43-58) analogues function as agretopes (sites bound to MHC molecules). Phenylalanine (F) and alanine (A) at positions 46 and 54 on p43-58 respectively bind to I-Ab. Aspartic acid (D) and alanine at positions 46 and 54 respectively bind to I-Ak. To determine the allele specific binding sites (desetope (s)) on class II molecules that are correspondent to the agretopes of peptide antigen (Ag), we analyzed directly binding capacity of p43-58 analogues with glutamic acid (E) at the epitopic position 50 (50E) to L cell transfectants expressing recombinant I-A molecules between b and k types. An Ak binding peptide, 46D50E54A, bound to transfectant possessing amino acid sequence of k type on N-terminal half of alpha-helix of A alpha-chain irrespective of the b type sequence on the other part, whereas an Ab binding peptide, 46F50E54A, did not bind to these transfects. Thus, agretopic residue 46 of 46D50E54A peptides appeared to bind to N-terminal half of alpha-helix of A alpha-chain. To define critical residues for the allele specific peptide binding, we then analyzed peptide binding capacity of Ak mutants substituted one of four polymorphic residues between Ak and Ab molecules. An Ak mutant, Ak alpha(56A), where arginine (R) at position 56 of the Ak alpha-chains was substituted with alanine located at the same position 56 of the Ab alpha-chains hardly bound 46D50E54A. By contrast, the Ak alpha(56A) bound 46F50E54A. Furthermore, Ak restricted T cell hybridomas responded to 46F50E54A but not to 46D50E54A in the presence of the Ak alpha(56A) APC. Thus, an amino acid on the position 56 of A alpha-chain determines critically specificity of the allele specific peptide binding (desetope).
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PMID:[Analysis of the allele specific Ag-binding site on murine class II MHC]. 864 75


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