Monocyte function in mixed lymphocyte reactions

Subpopulations of human peripheral blood lymphocytes were isolated by sequential separation techniques. The stimulating and responding capacity of these cells together with the T-cell population remaining after the removal of other populations was studied in one-way allogeneic mixed lymphocyte culture. Incorporation of [³H]thymidine was used as a measure of response. Monocytes, present in the stimulating or responding cell population, were necessary for lymphocyte response. T cells stimulated responding T-cell populations containing monocytes but not B cells. Stimulation by T cells could be inhibited with DRW antisera. Response was also inhibited by sera detecting DRw antigens on the monocytes of the responding cell population. It is concluded that monocytes play an important functional role in mixed lymphocyte reactions. In addition, it appears that the combination of anti-DRw sera and monocytes influences mixed lymphocyte reactions by an active process in that inhibition of response cannot be explained entirely by blocking DRw determinants.     

Alteration of the antigenicity of human lymphocytes by plant lectins and short-term tissue culture.

Human lymphocytes studied after being placed in culture for 1–6 wk progressively lost stimulating ability, i.e., lymphocyte defined antigens, when tested in one way mixed lymphocyte culture (MLC) but retained several other identifiable membrane components as well as the capacity to respond to mitogenic stimuli. Lymphocytes placed in culture with motogenic doses of PHA and Con-A after 1 and 2 wk strongly stimulated autologous responding fresh lymphocytes, but the MLC response of allogeneic fresh lymphocytes to stimulating lectin treated cells was even lower than the response to stimulating allogeneic cultured lymphocytes. The HL-A antigens on lectin treated cells or on lymphocytes through 6 wk in culture were clearly identifiable. Assays for T cell rosettes and B cell surface immunoglobulin showed both cell types to be present in numbers equal to fresh lymphocytes for up to 5 wk after culturing. However, the Fc receptor site on B cells was lost from cultured lymphocytes at the same time that MLC stimulation was lost. It is concluded that plant lectins can unmask new mitogenic sites on the cell surface as well as mask or delete existing sites, and that culturing lymphocytes for 1–6 wk will produce somewhat similar modulations. Modulation of surface membrane components by tissue culture or lectins may, therefore, have a profound effect in altering transplantation immunogenicity.     

Alloantigens expressed on activated human T cells different from HLA-A,B, C,and DR antigens

This study investigates alloantisera containing antibodies directed against antigens which are expressed on alloactivated human T lymphocytes but are absent on resting T and B cells. Among 39 defined anti-HLA-DR sera from multiparous women we found six sera giving positive reactions (more than 25 percent cytotoxicity) on in vitro alloactivated T cells, though negative reactions with resting B or T cells from the donors of either the responding or stimulating cell populations used for alloactivation. Two such sera were submitted to absorption and elution studies. Absorption of these sera with activated T cells did not remove the anti-HLA-DR activity. Furthermore, the antibodies eluted from activated T cells did not react with B cells but were positive only on activated T cells. In addition, we absorbed the sera with B cells and observed that they remained positive on activated T cells. The positive reactions do not seem to be due to either the passive acquisition of antigens from the stimulating population or to low levels of HLA-specific antibodies. As one of the sera we studied intensively gave clear positive and negative reactions on a panel of activated T lymphocytes, we believe it may recognize an antigen of an allogeneic system expressed on alloactivated human T cells.     

Human epidermal cells are more potent than peripheral blood mononuclear cells for the detection of weak allogeneic or virus-specific primary responses in vitro

Human epidermal cells (EC) and peripheral blood mononuclear cells (PBMC) have been used as antigen-presenting cells in allogeneic reactions or in self-restricted antiviral responses. Comparison of results from both cell types indicates that: (1) EC were better stimulators of primary proliferative responses in all the antigenic systems tested. (2) In secondary reactions, EC and PBMC functioned similarly for allogeneic responses, while a weak but significant difference could be observed in both (HSV1 or influenza A) virus-specific reactions. (3) By comparing pairs of HLA-identical mixed lymphocyte reaction (MLR)-negative siblings, positive responses were observed in several different families when lymphocytes of potential bone marrow donors were stimulated by EC of the recipient. This suggests that EC might be useful in detecting relatively weak proliferative responses in a number of antigenic systems, but especially in primary reactions against viral or putative minor histocompatibility antigens. (4) Despite this stronger antigen-presenting capacity in proliferative responses, EC induced lower levels of cytotoxic T lymphocyte (CTL) reactions than PBMC, not only in allogeneic responses but also in virus-specific self-restricted reactions.     

Antigen processing requirements for T cell activation: differential requirements for presentation of soluble conventional antigen vs cell surface MHC determinants

The present studies were undertaken to characterize the antigen-processing requirements involved in the responses to T cells to soluble antigen (antigen specific), to allogeneic cell surface MHC determinants (alloreactive), and to syngeneic MHC determinants (autoreactive). T cell clones were used that have dual cross-reactive specificities either 1) for self MHC plus soluble antigen and for allogeneic MHC products or 2) for syngeneic MHC and for allogeneic MHC, in order to permit comparison of the processing requirements for responses of the same T cell to distinct antigenic stimuli. The proliferative responses of antigen-specific, Ia-restricted T cell clones to soluble antigens were sensitive to treatment of antigen-presenting cells (APC) with 125 to 250 microM chloroquine, a lysosomotropic agent previously shown to inhibit the processing of soluble antigens. In contrast, the same T cell clones were only minimally affected in their ability to respond to similarly chloroquine-treated APC expressing allogeneic MHC products. The responses of autoreactive T cell clones to syngeneic stimulating cells and their cross-reactive responses to allogeneic cells were both resistant to chloroquine treatment of stimulating cells. The failure of chloroquine to inhibit antigen presentation to autoreactive T cell clones suggests that these clones are specific for self Ia not associated with in vitro processed foreign antigen. Thus, chloroquine sensitivity distinguishes the in vitro antigen-processing requirements for presentation of the soluble antigens tested from the requirements for presentation of syngeneic or allogeneic cell surface MHC determinants to the same T cells.     

Why do so many lymphocytes respond to major histocompatibility antigens?

A large fraction of T lymphocytes respond to allogeneic cells. Products of the major histocompatibility (H) complex influence the antigenicity of other cell surface components. We propose that lymphocytes responding to cells that differ genetically only at the major H locus recognize not only the major H difference, but also a multitude of other surface components in combination with the major H antigens. The large frequency of lymphocytes responding to allogeneic cells, then, becomes a function of the number of complex antigens they are recognizing on the foreign cells.     

Requirement of Ia-positive accessory cells in the MLR response against class II antigen on human B cell tumor line

In this report we have made a comparative study of the capacity of normal human stimulator cells and Epstein-Barr virus-transformed human B cell line Wa (EBV-Wa) cells to stimulate alloreactive T cells. Class II antigen (presumably HLA-DR4 determinant) on EBV-Wa cells was shown to act as a stimulating molecule in the mixed lymphocyte reaction (MLR) through a blocking study by using anti-Ia antibodies. Furthermore, it was found that HLA-DR-positive accessory cells in the responder population were required to elicit MLR responses against HLA-DR antigen on EBV-Wa cells. In contrast, HLA-DR-positive accessory cells in the responding cell population were not essential for elicitation of MLR responses against HLA-DR antigen on normal allogeneic peripheral blood mononuclear cells, as reported. The cell-cell interaction between responder HLA-DR-positive accessory cells and responding T cells in a major histocompatibility complex (MHC)-restricted manner was required for eliciting MLR responses against class II antigen on EBV-Wa cells such as antigen-presenting cell-T cell interaction in soluble antigen-specific T cell proliferative responses. The function of HLA-DR-positive accessory cells in the responder population could not be substituted for by the presence of interleukin 1. Furthermore, there was no obvious correlation between the degree of surface HLA-DR antigen expression on EBV-Wa cells and its stimulating ability. Thus, two distinct types of allo-class II, antigen-specific T cell activation between normal human stimulator cells and EBV-Wa cells were shown to exist.     

Expression of both I-A and I-E/C subregion antigens on accessory cells required for in vitro generation of cytotoxic T lymphocytes against alloantigens or TNBS-modified syngeneic cells

We have previously reported that radioresistant, Thy 1-negative accessory cells (SAC) are required for the in vitro generation of cytotoxic T-effector cells to allogeneic or trinitrophenyl-modified syngeneic cells. These SAC were found to provide accessory functions irrespective of whether they were syngeneic, semi-syngeneic, or allogeneic to the responding cells. To further characterize the accessory cells active in CML, the expression of Ia antigens on this functional population was assessed by pretreated SAC with anti-Ia reagents and complement and by testing the accessory cell function of these treated populations. The results of these studies demonstrated that the relevant accessory cells for allogeneic and TNP-self CTL express Ia determinants encoded by genes mapping in the I-A and I-E/C subregions. For the TNP-self CTL the accessory function of both SAC syngeneic or allogeneic to the responding and stimulating cells was specifically abrogated by treatment with anti-Ia reagents and complement.     

Generation of Human Alloantigen-specific T Cells from Peripheral Blood

The study of human T lymphocyte biology often involves examination of responses to activating ligands. T cells recognize and respond to processed peptide antigens presented by MHC (human ortholog HLA) molecules through the T cell receptor (TCR) in a highly sensitive and specific manner. While the primary function of T cells is to mediate protective immune responses to foreign antigens presented by self-MHC, T cells respond robustly to antigenic differences in allogeneic tissues. T cell responses to alloantigens can be described as either direct or indirect alloreactivity. In alloreactivity, the T cell responds through highly specific recognition of both the presented peptide and the MHC molecule. The robust oligoclonal response of T cells to allogeneic stimulation reflects the large number of potentially stimulatory alloantigens present in allogeneic tissues. While the breadth of alloreactive T cell responses is an important factor in initiating and mediating the pathology associated with biologically-relevant alloreactive responses such as graft versus host disease and allograft rejection, it can preclude analysis of T cell responses to allogeneic ligands. To this end, this protocol describes a method for generating alloreactive T cells from naive human peripheral blood leukocytes (PBL) that respond to known peptide-MHC (pMHC) alloantigens. The protocol applies pMHC multimer labeling, magnetic bead enrichment and flow cytometry to single cell in vitro culture methods for the generation of alloantigen-specific T cell clones. This enables studies of the biochemistry and function of T cells responding to allogeneic stimulation.     

Regulatory role of monomorphic and polymorphic determinants of HLA class I antigens in the proliferation of T cells stimulated by autologous and allogeneic B and T lymphoid cells

Monoclonal antibodies (mAb's) to monomorphic and polymorphic determinants of HLA Class I antigens were shown to inhibit proliferation of T cells stimulated with autologous and allogeneic B and T lymphocytes. Inhibition of proliferative responses was lower when T cells were used as stimulators than when B cells were used. The inhibitory activity was similar for mAb's to monomorphic and polymorphic determinants of HLA Class I antigens, suggesting that the density of antigen-antibody complexes on the cell membrane does not play a major role in the phenomenon. The anti-HLA Class I mAb's exerted their inhibitory effect at the level of both the responding and the stimulating cells. Addition of exogenous interleukin 2 to the mixed cultures did not affect the mAb-mediated inhibition.     

Thymocyte and macrophage interactions: separation of murine thymocyte subsets and enrichment of syngeneic cell-responding thymocytes by adsorption to macrophage monolayers

The spontaneous binding of murine thymocytes to macrophage monolayers was employed to separate thymocytes into macrophage-unbound and -bound subsets, and the functional reactivities of these two subpopulations were examined. Macrophage-unbound thymocytes were found to be enriched in functional subsets reactive to semi-allogeneic and allogeneic stimulating spleen cells by proliferation in mixed leukocyte culture (MLC). Furthermore, macrophage-unbound thymocytes were frequently found to respond to syngeneic spleen cells. This syngeneic proliferative response showed both memory and specificity upon subsequent restimulation and thus seems to represent a syngeneic mixed leukocyte reaction (SMLR). Syngeneic responding thymocytes were also found to produce interleukin 2 when cultured with syngeneic but not allogeneic stimulator cells. In contrast, macrophage-bound thymocytes showed greatly reduced proliferative responses to allogeneic stimulators and no responses to syngeneic stimulators. The macrophage-bound thymocyte subset was not enriched in detectable suppressive activity; proliferative responses of macrophage-unbound thymocytes to either allogeneic or syngeneic cells were neither suppressed nor enhanced when macrophage-unbound thymocytes were added to the cultures. Thus, the macrophage-unbound subset seems to be enriched in functionally mature thymocytes and the macrophage-bound subset appears to be enriched in functionally immature thymocytes. This functional separation of thymocytes by macrophage adherence also correlated well with thymocyte subpopulations separated by bovine serum albumin density gradients; the low density mature thymocytes showed enhanced responses to both allogeneic and syngeneic stimulators, whereas the high density immature cells were unresponsive. This correlation was supported further by binding studies in which T cell tumor lines derived from C57BL/6 mice were used. ERLD tumor cells, which are similar to cortical immature thymocytes in both enzymatic and surface antigenic markers, were found to bind readily to macrophages, whereas both EL-4 and E male G2 tumor cells, with characteristics of mature thymocytes, bound to macrophages poorly. The binding of thymocytes and ERLD tumor cells to macrophages was not genetically restricted. We speculate that thymocyte binding to macrophages may play a critical role during the functional maturation of thymocytes.     

Specific suppression of the in vitro parent anti-hybrid reaction. I. Characterization of a suppressor cell induced in hybrids by pretreatment with parent-strain spleen cells

Spleen cells from B6D2F1 hybrid mice pretreated with 5 X 10(7) B6 spleen cells iv 7 days earlier (B6-pretreated B6D2F1) exhibit a reduced capacity to stimulate the in vitro proliferative and anti-D2 CTL responses of B6 spleen cells. This inability of B6-pretreated B6D2F1 spleen cells to stimulate B6 spleen cells efficiently is due neither to the absence of stimulating cells bearing the D2 alloantigens nor to the destruction of B6 responding cells, but to the presence in the B6-pretreated B6D2F1 cell population of a suppressor mechanism, since the addition of B6-pretreated B6D2F1 spleen cells to a culture of normal B6 responding and irradiated B6D2F1-stimulating spleen cells can suppress the B6 anti-B6D2F1 response. This suppression is mediated by a nylon adherent, Thy-1-negative cell of parent-strain origin which is radioresistant at 2000 R. This suppressor cell is not induced by the injection to B6D2F1 hybrids of spleen cells from the other parent strain (D2) or an allogeneic strain (C3H). It does not suppress either the response of the other parent (D2) or an allogeneic strain (C3H) to B6D2F1 antigens, or the response of B6 cells to an allogeneic strain (C3H).     

Persistent effects of ouabain treatment on human lymphocytes: synthesis of DNA, RNA and protein in stimulated and unstimulated cells.

Pretreatment of human lymphocytes for 2 days in 2 X 10(-6)M ouabain caused irreversible loss of their subsequent capacity to stimulate in the mixed lymphocyte reaction (MLR). Pretreatment for the same period with 10(-7)M ouabain resulted in an enhanced incorporation of thymidine into DNA of the responding cells in the MLR; this effect was also on the stimulating cells, as previously reported by Christen et al. (Cell, Immunol. 19, 137-142 (1975)). Pretreatment of stimulating lymphocytes with 10(-7)M ouabain caused a persistent but reversible inhibition of the synthesis of RNA and protein in the MLR; peak incorporation of labelled uridine or alanine reached the same level as that of the control cultures, but 24 h later. Exactly the same persistent but reversible inhibition was found in the case of DNA syntheis of cells pretreated with 10(-7)M ouabain and then stimulated by antigens (streptolysin-O and varidase) or by mitogens (phytohemagglutinin and concanavalin A); the same level of incorporation of labelled thymidine occurred but 24-48 h later than in the case of the controls. Pretreatment with the cardiotonic steroid under these conditions also resulted in a pronounced inhibition of the basal, unstimulated levels of RNA and protein synthesis in the case of both control lymphocytes and those which had been treated with mitomycin C. The effects of ouabain pretreatment on basal RNA and protein synthesis were identical for both 2 X 10(-6)M and for 10(-7)M; the effect of pretreatment of stimulating cells with these two concentrations was completely opposite: irreversible inhibition of the proliferative response of allogeneic responding cells at the former concentration and delayed activation at the latter.     

Mouse T lymphocytes proliferative responses specific for human MHC products in mouse anti-human xenogeneic MLR

It has been reported that human T cells recognize the polymorphism of murine Ia antigens in the human anti-mouse xenogeneic mixed lymphocyte reactions (MLR). In this study, murine T cell recognition of human Class II antigens of the major histocompatibility complex (MHC) was analyzed in mouse anti-human xenogeneic MLR responses. The xenoreactive murine T cell proliferative response was blocked by adding anti-HLA-DR monoclonal antibody to the xenogeneic MLR culture. The specificity of xenoreactive murine T cells was examined with regard to the secondary and tertiary xenogeneic MLR system. The xenoreactive murine T cells were restimulated by distinct human stimulator cells that had no shared HLA antigens with the stimulator used in the primary MLR. The data presented here show that the murine xenoreactive T cells recognize the shared determinant(s) of HLA-DR antigen on non-T, non-B stimulator cells. The xenoreactive murine T cell proliferative responses were mediated by Thy-1+, Lyt-1+, and Lyt-2- cells. Furthermore, the xenoreactive T cell responses required Ia+ cells, and Ia antigen on accessory cells plays a crucial role in eliciting the xenoreactive responses against human stimulator cells, while Ia+ accessory cells in the responding cell population are not essential for the elicitation of allogeneic MLR responses, as reported previously.     

Generation of natural killer-like activity in mixed lymphocyte-tumor cell cultures. II. Correlation between expression of HLA-DR antigens on the activated T cells and their cytotoxic capability

In this study, we investigated the role of DR antigens in human mixed lymphocyte reactions (MLR) at the responder cell level. Upon stimulation by allogeneic lymphocytes or leukemic cell lines, a large proportion of T cells underwent blastogenesis and began expressing DR antigens. Analysis by a fluorescence-activated cell sorter revealed that both subpopulations of large activated T cell blasts and of small T lymphocytes became DR+ by synthesis and/or uptake. Depletion of DR+ responder cells from 6-day-old MLRs by treatment with anti-DR monoclonal antibodies (mAbs) plus complement (C) reduced but did not completely abrogate the natural killer (NK)-like activity of the responder lymphocytes, suggesting that the MLR-induced cytotoxic cells include both DR+ and Dr- populations. The expression of NK-like activity by the responder cells was also greatly reduced upon addition of anti-DR mAbs (without C) at the start of the mixed cultures. This effect was observed regardless of the presence of DR antigens on the stimulator cells, indicating that the anti-DR mAbs can interact with the antigens present on both the stimulator and responder populations. These data show that during an MLR, the continued presence of DR antigens on the responding population is essential for the expression and maintenance of the proliferative and cytotoxic capabilities of these cells.     

The pigeon cytochrome c-specific T cell response of low responder mice. I. Identification of antigenic determinants on fragment 1 to 65

An examination of the proliferative response to pigeon cytochrome c fragments 1 to 65 and 1 to 80 by T cells from mice that are low responders to the native molecule revealed that some of the strains could respond to antigenic determinants on these fragments. T cell clones derived from B10.A(3R) and B10.A(4R) mice were used to characterize the antigenic determinants on fragment 1 to 65. All of the clones recognized syngeneic A beta:A alpha Ia molecules as their restriction element. Three B10.A(3R) clones and six B10.A(4R) clones recognized fragment 39 to 65. Another four B10.A(4R) clones responded to fragment 1 to 38. By stimulating with a series of cytochrome c fragments from different species, as well as a synthetic peptide, it was possible to localize the antigenic determinant(s) recognized by the B10.A(3R) clones to residues 45 to 58. Each clone showed a unique pattern of responsiveness to the various fragments, suggesting a diversity of T cell receptors specific for the same peptide. One B10.A(3R) clone could be stimulated by many of the 1 to 65 fragments in association with allogeneic B10.SM presenting cells and by tuna fragment 1 to 65 in association with B10.M presenting cells, although the rank order of potency for several of the fragments was different than that observed with syngeneic antigen-presenting cells. In addition, the clone was poorly reactive to a synthetic peptide containing a conservative substitution, serine for threonine, at position 49. The implications of these results for subsite dissection (agretope and epitope) of the antigenic determinant recognized by this clone are discussed.     

Antigenic profile and functional characterization of human peritoneal macrophages

The antigenic profile and the functional properties of human peritoneal macrophages have been analyzed by using a panel of monoclonal antibodies (MoAb) and functional assays. All peritoneal macrophages were stained by the anti-class I HLA MoAb Q6/64. Between 40 and 100% of the cells were stained by the anti-HLA-DR + DP MoAb Q2/80, Q5/6, and Q5/13; approximately 80% of the cells were stained by the anti-HLA-DQ MoAb BT3.4, and about 95% were stained by the anti-macrophage MoAb OKM1. Peritoneal macrophages were not stained by the anti-dendritic cells MoAb Ki-M4 or by MoAb to T cell subsets, although all of the MoAb were reactive with the appropriate substrates. More than 60% of the cells expressed Fc receptors and C3 receptors, and displayed phagocytic activity. Peritoneal macrophages were effective in stimulating autologous and allogeneic lymphocytes and in presenting soluble antigens to T cells. These reactions were blocked by the anti-HLA-DR + DP MoAb Q5/13, but were not affected by the anti-dendritic cells MoAb Ki-M4 or by the anti-class I HLA MoAb Q6/64. These results suggest that human peritoneal macrophage preparations, without detectable contamination with dendritic cells, can induce proliferation of autologous and allogeneic T cells, and that class II HLA antigens play a significant role in these phenomena.     

Cellular requirements for lymphocyte restimulation after a first challenge in vitro.

Human peripheral blood lymphocytes from donors who were sensitized in vivo to bacterial antigens were stimulated by these antigens in vitro. When the cells from these first cultures were challenged with irradiated allogeneic lymphocytes, a proliferative response was obtained, the kinetics of which resembled those of a primary mixed lymphocyte reaction (MLR). On the other hand, the addition, under these conditions, of bacterial antigens never led to any second proliferative response. It was shown that: (1) the addition of irradiated autologous mononuclear cells, together with the bacterial antigens, led to a reconstitution of a proliferative response in second culture; (2) the cells capable of reconstituting the reactivity to tetanus toxoid could also be obtained from donors whose own cells did not respond to that antigen in primary cultures, and (3) the reconstituting activity in the second culture could not be provided by monocytes alone.     

Induction of interleukin 3 but not interleukin 2 or interferon production in the syngeneic mixed lymphocyte reaction

The syngeneic mixed lymphocyte reaction (SMLR) was assayed in the medium containing syngeneic normal mouse serum (NMS), by using nylon-adherent stimulator cells and nonadherent responder T cells, which were prepared from murine spleens in the absence of fetal calf serum (FCS) to avoid any sensitization to xenogeneic protein antigens. The responder cells in this SMLR, without definite background proliferation, generated specific proliferative response to the syngeneic stimulator cells in a dose-related fashion. The SMLR was accompanied by production of interleukin 3 (IL 3) but not interleukin 2 (IL 2) or interferon (IFN). No cytotoxicity against the syngeneic or allogeneic target cells was induced. Correlating with no production of IL 2 or IFN, no natural killer (NK) activity was detected. The proliferation was not inhibited by addition of specific antiserum for IFN-gamma. In contrast, proliferation in the responder cells when incubated with allogeneic stimulator cells was inhibited by anti-IFN-gamma serum and accompanied by production of IL 2 and IFN as well as IL 3, and by augmentation of NK activity and generation of cytotoxic T cells. Cell surface analysis revealed that the cells producing IL 3 in this SMLR system were Thy-1+ Lyt-1+2- helper T cells. Cells responding to the SMLR culture fluids with DNA replication were Thy-1-Lyt-1-2- asialo GM1- no-marker cells, which were the same as a population responsible for partially purified IL 3. On the other hand, when the responder cells were exposed to FCS before culture and assayed for SMLR in the FCS-free NMS medium, variable levels of IL 2 production were induced in response to the stimulator cells. The responder cells generated a high background DNA replication in the absence of syngeneic stimulators, suggesting that this IL 2 production may result from the stimulation of T cells by FCS as a foreign antigen. Overall, these results suggest that the SMLR may be a cellular interaction, in which non-T cells stimulate Lyt-1+2- helper T cells to produce IL 3 but not IL 2 or IFN. This IL 3 can, in turn, induce proliferation of IL 3 responding cells, which appear to be early precursors in lymphocyte differentiation, but no proliferative response or activation of IL 2- and IFN-dependent mature T cells or NK cells.     

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

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