Evidence for accessory cell function by class II MHC antigen-expressing airway epithelial cells

Expression of major histocompatibility complex (MHC) class II antigens is a requirement for accessory cell function in antigen presentation. Recent reports have demonstrated the presence of class II antigens on human bronchial epithelial cells. In the present study, immunohistochemical staining revealed HLA-DR on human airway epithelial cells obtained from two different mucosal sites (lobar bronchus and nasal turbinates). To determine whether airway epithelial cells bear functional class II molecules that allow for their cognate interaction with T lymphocytes, cells isolated from these sites were used in mixed lymphocyte cultures (MLR), as an in vitro model of accessory cell function. Freshly isolated cells (11 bronchi/3 turbinates) stimulated allogeneic T lymphocytes (stimulation index [S.I.] = 9.3 [mean]; P less than 0.001 compared to T cells alone). In order to assess the potential role of contaminating conventional accessory cells, bronchial epithelial cell isolates were first preincubated in a serum-free, growth factor-supplemented medium that functionally eliminates potential non-epithelial stimulators prior to MLR culture. Conventional accessory cell-depleted epithelial cells were still capable of stimulating allogenic T lymphocytes in 18 of 23 MLR cultures (S.I. = 5.5 [mean]; P less than 0.0005 compared to T cells alone). The addition of an anti-class II monoclonal antibody (VG2.2) at the onset of culture completely inhibited the MLR response (n = 10). No shift in the CD4+/CD8+ ratio was detected between lymphocytes harvested from airway epithelial cell MLR (1.42 +/- 1.29) and the ratio from T lymphocytes cultured alone (1.3 +/- 0.75), suggesting that both CD4+ and CD8+ T lymphocytes were proliferating in response to stimulation from alloepitopes recognized on airway epithelial cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Stimulation with dendritic cells decreases or obviates the CD4+ helper cell requirement in cytotoxic T lymphocyte responses

We investigated the need for CD4+ helper T (Th) cells in the induction of murine cytotoxic T lymphocyte (Tc) responses across minor or major histocompatibility (MHC) antigenic differences with either normal spleen cells (NSC) or purified dendritic cells (DC) as antigen-presenting cells (APC). Generation of a secondary in vitro class II MHC-specific Tc response was totally CD4+ Th cell-dependent with both types of APC. Likewise, male antigen (H-Y)-primed class II mutant bm12 T cells, which do not respond to H-Y presented on NSC, do respond to H-Y presented on DC in a completely CD4+ Th cell-dependent fashion. All other Tc responses, including primary anti-class I MHC, primary anti-class I + II MHC plus anti-minor H, and secondary C57BL/6 (B6) anti-H-Y, although not completely CD4+ Th cell dependent, were greatly augmented in the presence of CD4+ Th cells, but only with NSC as APC. In contrast, with DC as APC these responses were entirely or largely CD4+ Th cell independent. Similarly, H-Y primed class I MHC mutant bm14 T cells, which do not respond to H-Y presented on NSC, do respond to H-Y presented on DC in a completely CD4+ Th cell-independent fashion. The combined results indicate that DC can directly present class I MHC alloantigen or class I MHC plus nominal antigen (e.g. minor H) to CD8+ cells and generate a Tc response by these cells without the requirement for CD4+ Th cells.     

Phenotype and function of peripheral blood and bone marrow T-cell colonies: identification of DC3-, 4-, 8-autoreactive T cells

The existence of immature autoreactive T cells has been examined in peripheral blood (PB) and bone marrow (BM) derived T-lymphocyte colonies (TLC) that have previously been shown to be potentially generated from CD3-negative BM-T cells. An extensive phenotypical analysis of total and T-depleted TLC showed that both PB- and BM-derived TLC contained a mean of 5% immature T lymphocytes (ITL), which were negative for the CD3, CD4, and CD8 antigens but displayed the CD2 and CD7 antigens. The only detectable immune functions of these isolated ITL were an allo- and an autoreactivity without cytolytic activity. The self-reactivity of ITL was not detected in bulk non T-depleted TLC cells and seemed to be actively suppressed by autologous mature T cells. In addition, the auto-MLR of ITL was totally inhibited by anti-HLA class II but not by anti-class I monoclonal antibody (MoAb) and only partially by anti-CD4 moAb, whereas the anti-CD3 and anti-CD2 MoAbs gave no inhibition. Once activated, ITL could acquire in culture a mature T cell CD3 + CD4 + phenotype. The CD3-, 4-, 8-auto-reactive T cells present in T colonies could represent pre- or post-thymic cells that have not yet undergo or that have escaped the thymic selection.     

Xenogeneic recognition of soluble and cell surface HLA class II antigens by proliferative murine T cells

In order to characterize the murine anti-human xenogeneic mixed lymphocyte reactions (MLR), we studied T cell proliferative responses against various human lymphoid cells by immunization of mice either with cellular or purified HLA-DR antigens. Data presented here indicated that small amounts of soluble HLA-DR antigen were able to prime mice, and that the xenogeneic MLR depends on the expression of HLA class II antigens on the stimulating cells. Experiments using a mutant cell line clearly showed that HLA-DP molecules were also sufficient in eliciting a primary or a secondary xenogeneic MLR while no secondary proliferative response was obtained with cells expressing only HLA class I molecules. Using a large panel of human cells with various haplotypes, our results also showed that (a) nonpolymorphic determinants of HLA class II antigens trigger dominantly the murine T cells and (b) the xenogeneic response required I-E and L3T4 accessory molecules and was not inhibited with anti I-A and monomorphic anti-HLA class II antigen monoclonal antibodies. Altogether these results suggest that HLA class II antigens act as nominal antigens in triggering a murine anti-human proliferative response.     

The response of gamma delta T cells to Plasmodium falciparum is dependent on activated CD4+ T cells and the recognition of MHC class I molecules.

Peripheral blood gamma delta T cells from non-exposed individuals respond to antigens of the malaria parasite, Plasmodium falciparum, in vitro. This response, largely caused by T cells bearing the V gamma 9+ chain of the T-cell receptor, is stimulated by components of the parasite expressed on the schizont stage and released at schizont rupture. The response of V gamma 9+ T cells to parasite components is inhibited by antibodies to major histocompatibility complex (MHC) class I and class II. However, the inhibition by anti-MHC class II antibodies can be overcome by the addition of interleukin-2 (IL-2) to the cultures, suggesting that gamma delta T cells themselves do not recognize MHC class II molecules but require an MHC class II-dependent response taking place in the culture. In contrast, the inhibition by anti-class I antibodies cannot be reversed by addition of IL-2. Since an accompanying CD4+ T-cell response occurred in peripheral blood mononuclear cells cultured with P falciparum antigens, it was considered that these cells provide the cytokines necessary for the subsequent activation and expansion of V gamma 9+ T cells recognizing components of the parasite and MHC class I molecules. This was confirmed by reconstituting the response of enriched gamma delta T cells to P falciparum schizont extract by addition of purified CD4+ T cells.     

CD8 is involved in both class I- and class II—induced proliferation of a cytolytic T-cell clone with dual specificity for HLA-B27 and HLA-DR2 antigens

A CD3⁺ CD4⁻ CD8⁺ cytolytic T-lymphocyte (CTL) clone, CTL 47, could be induced to proliferate in the presence of exogenous interleukin 2 by either HLA-B27.1⁺ or HLA-DR2⁺ cells. B27.1-induced proliferation was strongly and equally inhibited by an anti-B27 and by an anti-CD8 monoclonal antibody (MoAb). DR2-induced proliferation was inhibited by the same anti-CD8 MoAb less efficiently and with a different time course than anti-class II blocking, only being significant when the antibody was added ab initio or very early during the assay. These results indicate that CD8 is essential for class I—induced proliferation but that it also enhances class II—induced stimulation of this CTL clone. It is proposed that the necessary role of CD8 in class I—induced proliferation is related to its interaction with the same class I molecule bound by the T-cell receptor. The accessory role in class II—induced proliferation would be due to an additive effect on the avidity of cell adhesion, resulting from interaction of CD8 with the class I antigens on the stimulator cell, or perhaps to a regulatory role of CD8 as a transducer of early signals for T-cell activation.     

Establishment of stable CD8+ suppressor T cell clones and the analysis of their suppressive function.

Stable CD8+ suppressor T cell (Ts) clones were established by a relatively simple method. Keyhole limpet hemocyanin (KLH)-primed spleen cells from C3H mice were depleted of B cells and CD4+ T cells by panning and cytotoxic treatment, and the resulting CD8+ T cells were periodically stimulated with antigen and irradiated syngeneic spleen cells followed by manifestation in interleukin-2 (IL-2) containing medium. T cell clones with a definite suppressor function were established by limiting dilution. They were defined as classical effector type Ts of CD8+ phenotype as they had constant and definite suppressor functions in antigen-induced T cell proliferation and specific antibody response against T cell-dependent antigens without detectable cytotoxic activity against both antigen presenting cells (APC) and helper T cells (Th). They showed no helper activity for B cells and produced no detectable helper type lymphokines such as IL-2 and IL-4. CD8+ Ts clones were able to inhibit the antigen-induced IL-2 production of normal and cloned T cells. Their suppressive activity was antigen-nonspecific and major histocompatibility complex-unrestricted. CD8+ Ts clones were also able to suppress the proliferative response of Th clones induced by immobilized anti-T cell receptor (TcR) and anti-CD3 mAbs but not the response induced by concanavalin A (ConA) and IL-2. All the CD8+ T cell clones established independently utilized the TcR V beta 8 gene. Syngeneic antigen presenting cells could induce proliferation of these CD8+ clones, which was blocked by anti-CD8 and anti-I-Ak monoclonal antibody (mAb) but not by anti-class I mAbs. The stimulation of CD8+ Ts clones with immobilized anti-CD3 resulted in the release of a suppressor factor(s) that potently inhibited the antigen-induced proliferation of CD4+ Th clones and the in vitro secondary antibody formation.     

Analysis of the peripheral T-cell compartment in the MHC class II deficiency syndrome.

To analyse the impact of lack of MHC class II expression on the composition of the peripheral T-cell compartment in man, the expression characteristics of several membrane antigens were examined on peripheral blood lymphocytes (PBL) and cultured T cells derived from an MHC-class-II-deficient patient. No MHC class II expression could be detected on either PBL or activated T cells. Moreover, the expression of MHC class I was reduced both on PBL and in vitro activated T cells compared to the healthy control. However, the reduced expression of CD26 observed on the PBL of the patient was restored after in vitro expansion. Despite the presumably class-II-deficient thymic environment, a distinct but reduced single CD4+ T-cell population was observed in the PBL of the patient. After in vitro expansion, the percentage of CD4+ cells dropped even further, most likely due to a proliferative disadvantage, compared to the single CD8+ T-cell population. However, proliferation analysis showed that T-cell activation via the TcR/CD3 pathway is not affected by the MHC class II deficiency.     

Role of CD8+ T Cells in Mercury-Induced Autoimmunity or Immunosuppression in the Rat

In Brown-Norway (BN) rats mercuric chloride induces an autoimmune disease characterized by an increase in serum IgE concentration, and by the production of anti-glomerular basement membrane antibodies responsible for a glomerulonephritis with a heavy proteinuria. (i) This disease results from a B-cell polyclonal activation probably due to frequent anti-class II T cells. (ii) The self limitation observed in this model is associated with both a decrease in the frequency of anti-class II T cells and the emergence of CD8+ T cells able to suppress these autoreactive T cells. (iii) In Lewis (LEW) rats which do not develop autoimmunity, HgC12 provokes the appearance of non-antigen-specific CD8+ T cells responsible for a depression of T-cell functions. The aim of this work was to test the effect of treatment with an anti-CD8 monoclonal antibody (MoAb) in both BN and LEW rates, Anti-CD8 MoAb-treated rats were effectively depleted in CD8+ T cells. However, neither the induction nor regulation phases of mercury-induced autoimmunity were modified in BN rats. Mercury-induced immunosuppression in LEW rats was abrogated; however, depletion in CD8+ T cells did not allow the disease to occur in that strain. Finally, CD8 depletion induced in normal BN rats rats the appearance of rare anti-class II T cells showing that these cells are normally present in that strain but negatively controlled by suppressor T cells.     

Comparative Analysis of Syngeneic and Allogeneic Mixed Lymphocyte Reaction of ''Naturally'' Activated and Resting T Cells

'Naturally' activated (NA) or resting T lymphocytes obtained from the spleen of normal BALB/c mice were compared in their capacity to mount a syngeneic mixed lymphocyte reaction (SMLR). Both T-cell subsets were able to proliferate and secrete IL-3/GM-CSF in SMLR cultures. IL-2 was present in 'resting' T-cell SMLR supernatants, and barely detectable in NA T-cell SMLR supernatants. Both NA and 'resting' T-cell SMLRs were inhibited with anti-class II, anti-CD4, or anti-IL-2R MoAbs. NA T cells exhibited a background proliferative and secretory activity in the absence of syngeneic accessory cells. This autonomous activity was susceptible to anti-CD4, but poorly inhibited with anti-class II MoAbs. Both NA and 'resting' T lymphocytes displayed strong responsiveness to allogeneic stimuli. The analysis of the relative frequency of proliferating cells in the SMLR (BALB/c), or allo-MLR (B10, B10.A, B10.D2) from NA or 'resting' T cells indicated an enrichment for syngeneic reactivity among NA T lymphocytes. The meaning of these results for NA T-cell function and repertoire is discussed.     

MHC class II-dependent activation of CD4+ T cell hybridomas by human mast cells through superantigen presentation.

Human mast cells (MC) were examined for expression of MHC class II antigens and for their ability to activate CD4+ T cell hybridomas through presentation of superantigen (SAg). HMC-1, a leukemic immature MC line expressing class II Ags, was shown to efficiently present the staphylococcal enterotoxin B (SEB) SAg to responding T cell hybridoma on treatment with interferon-gamma (IFN-gamma), which up-regulated class II molecules. The study was then extended to human normal MC. Almost pure (>99%) cord blood-derived MC (CBMC) were shown to express class II Ags (HLA-DR and HLA-DQ) and CD80, which were up-regulated by IFN-gamma treatment and, to a lesser extent, by interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF). CBMC directly activated CD4+ T cell hybridomas through presentation of SEB and TSST1 SAgs. The production of IL-2 required a cell-to-cell contact between T cells and CBMC and it was inhibited by anti-class II antibodies. Furthermore, an additional pretreatment of CBMC by IFN-gamma or GM-CSF or IL-4 had no effect on their presenting efficiency. This previously unknown function of human MC, i.e., MHC class II-dependent activation of CD4+ T cells, may be critical in subsequent cellular activation events because colocalization of mast and T cells is frequently observed at sites of antigen entry.     

阻断型CD154抗体和CD80抗体生物学作用的研究

为比较阻断型抗人CD154单克隆抗体(1B1)和抗人CD80单克隆抗体(3H8)的单独及联合应用在调节CD4+T细胞对同种抗原的初次和再次免疫应答中的作用。采用免疫磁珠阴性选择法分离获得人外周血CD4+T细胞、将纯化的CD4+T细胞与刺激细胞体外共培养,通过检测培养上清IL-2、IFN-γ水平以及CD4+T细胞的增殖来评价1B1和3H8的生物学作用。结果显示,1B1和3H8单独或联合应用能不同程度地抑制混合淋巴细胞反应中CD4+T细胞对同种抗原刺激的增殖,下调CD4+T细胞分泌IL-2和IFN-γ,并且在再次反应中能有效诱导CD4+T细胞对同种抗原的免疫低反应性。因此,1B1和3H8的单独及联合应用在移植排斥的免疫干预及同种抗原免疫耐受的诱导中具有潜在的应用前景。     

Role of B70/B7-2 in CD4+ T-cell immune responses induced by dendritic cells.

Dendritic cells (DC) are potent antigen-presenting cells (APC). However, the molecular basis underlying this activity remains incompletely understood. To address this question, we generated murine monoclonal antibodies (mAb) against human peripheral blood-derived DC. One such antibody, designated IT209, stained differentiated DC and adherent monocytes, but failed to stain freshly isolated peripheral blood mononuclear cells (PBMC). The antigen recognized by IT209 was identified as B70 (B7-2; also recently identified as CD86). Using this mAb we studied the role of B70 in CD4+ T-cell activation by DC in vitro. IT209 partly inhibited the proliferative response of CD4+ T cells to allogeneic DC and to recall antigens, such as tetanus toxoid (TT) and purified protein derivative (PPD) of tuberculin, presented by autologous DC. More importantly, the mAb had a potent inhibitory effect on the primary response of CD4+ T cells to autologous DC pulsed with human immunodeficiency virus (HIV) gp160 or keyhole limpet haemocyanin (KLH). Adherent monocytes, despite their expression of B70, failed to induce T-cell responses to these antigens. IT209-mediated inhibition of CD4+ T-cell responses was equivalent to that produced by anti-CD25 mAb, whereas an anti-CD80 mAb was only marginally inhibitory and did not augment the effect of IT209. These findings indicate that the B70 antigen plays an important role in DC-dependent CD4+ T-cell activation, particularly in the induction of primary CD4+ T-cell responses to soluble antigens. However, since activated monocytes, despite their expression of B70, failed to prime naive T cells to these antigens, our results suggest that additional molecules contribute to the functions of DC in CD4+ T-cell activation.     

Synergistic T-T cell interaction present in alloreactivity: determination of 'MLR helper' T cell subsets

CD4 and CD8 T cell subsets involved in primary allo-MLR responses were re-evaluated in the present study. The slope analysis of B6 allo-MLR responses to MHC class I-disparate bm1 and class II-disparate bm12 revealed the presence of a T-T cell interaction between CD4 and CD8 T cell subsets. The allo-MLR response of B6 CD4 T cells to bm12 was completely blocked by anti-L3T4, indicating that the function of CD4 T cells is required for allo-MLR responses to bm12. It was further demonstrated that the function of CD8 T cells in class II-disparate allo-MLR is to augment the proliferative response of CD4 T cells. On the other hand, CD8 T cells primarily responded to class I-disparate allo-MLR, and CD4 T cells function to augment the proliferative response of CD8 T cells. It was further indicated that CD4 and CD8 T cells recognize both class I and class II MHC. The CD4 T cell recognition of allo-class I and CD8 T cell recognition of allo-class II induces insufficient signals for proliferation of these T cells. Thus, the class I-reactive CD4 and class II-reactive CD8 T cell subsets function as helper cells ('MLR helper' T cells) for proliferating T cells in MLR responses.     

Purified human NK cells do not function as accessory cells in T-cell proliferative responses.

In the present study we investigated the accessory function of natural killer (NK) cells in PHA, anti-CD3 (OKT3) and solid-phase-bound anti-CD3-induced T-cell proliferative responses, as well as in allogeneic mixed lymphocyte reaction (MLR). Although NK cell-enriched Percoll fraction cells showed accessory activity in all of the studied T-cell responses, the accessory function in anti-CD3 and allogenic responses were always found to be strictly dependent on the proportions of contaminating monocytes (Mo) and dendritic cells (DC). Purified CD16+ NK cells were totally incapable of inducing T-cell proliferation in anti-CD3, solid-phase-bound anti-CD3 and in allogeneic MLR responses, whereas these NK cells could support PHA-induced T-cell proliferation, although to a very low extent. Growing evidence suggests an inhibitory role of NK cells in various immune responses, but in the studied T-cell responses NK cells did not suppress either Mo or DC accessory ability.     

Thyroglobulin-induced T-cell in vitro proliferation in Hashimoto's thyroiditis: identification of the responsive subset and effect of monoclonal antibodies directed to Ia antigens

Recently it was reported that the peripheral blood and thyroid gland of patients with Hashimoto's thyroiditis contain activated (Ia+ and/or MLR4+) T cells and high levels of 5/9+ ("helper") T lymphocytes. In normal individuals the 5/9 monoclonal antibody recognizes a T-cell fraction that includes all T lymphocytes with inducer activities. Here, circulating 5/9+ and 5/9- T lymphocytes were isolated from patients with Hashimoto's disease, and the proliferative response induced by human thyroglobulin was investigated. The results show that the total thyroglobulin-induced lymphocyte DNA synthesis is confined to the 5/9+ T-cell fraction. Further subfractionation of 5/9+ into MLR4+ and MLR4- cells clearly indicates that no substantial differences exist in their proliferative capacities. Whether 5/9, MLR4, and Ia antigens, all expressed on the thyroglobulin-responsive T-cell subset, are involved in thyroglobulin-induced cell proliferation, was also analyzed. Although both 5/9 and MLR4 monoclonal antibodies had no effect, complete inhibition of antigen-induced blastogenesis was observed upon addition of monoclonal antibodies (D1/12 and BT2/9) directed to common determinants of Ia antigens. This inhibitory effect was also observed when T or non-T fractions were separately incubated with the monoclonal antibodies before culture. These results indicate that in humans, as in animals, the major histocompatibility complex may play a role in autoimmune thyroiditis. The data show that (a) the thyroglobulin-induced proliferative response is confined to a subset (5/9+) of T lymphocytes and (b) Ia antigens are involved in thyroglobulin-induced lymphocyte DNA synthesis in Hashimoto's disease.     

Differences between Primed Allogeneic T-Cell Responses and the Primary Mixed Leucocyte Reaction

Recent investigations have demonstrated that the primary mixed lymphocyte reaction (MLR) is dependent on certain accessory molecules, e.g. CD4 and LFA-1. We have compared the requirements of the primary MLR and the responses of alloreactive, primed lymphocytes (PL) by inhibition studies using monoclonal antibodies (MoAb) directed against (i) adhesion molecules belonging to the CD11 cluster of leucocyte antigens (CD11a, LFA-1; CD11b, MAC1 = CR3; and CD11c, p 150,95); (ii) various T cell-related antigens (CD2, CD4, CD5 and CD8); and (iii) recombinant IL-1 beta. The CD5-, CD11a- and CD11c-reactive MoAb significantly inhibited the primary MLR (inhibition = 25%, P less than or equal to 0.01; 48%, P less than or equal to 0.01 and 13%, P less than or equal to 0.05, respectively) but these MoAb did not inhibit the primed lymphocyte reaction (PLR). The CD11b-reactive MoAb had no significant influence on either of the responses. CD2- and CD4- reactive MoAb significantly inhibited both primary MLR (greater than 80%, P less than or equal to 0.01) and to a lesser extent the PLR (40-65%, P less than or equal to 0.01). A MoAb reactive with IL-1 beta inhibited the primary MLR (38%, P less than 0.01) and the purified protein derivative (PPD) induced lymphocyte transformation response (42%, P less than or equal to 0.01) of peripheral blood mononuclear cells (PBMC), whereas primed allogeneic responses to PBMC and Epstein-Barr virus (EBV) cell lines were unaffected by this MoAb. In addition, preliminary data indicated that PL seemed neither to bind exogenous IL-1 (as opposed to CD4+ PBMC) nor to possess membrane-bound IL-1. The differences between 'virgin' and primed, allogeneic T-cell responses indicate that profound changes in the functional capability of the responding T-cell population take place during the bulk expansion. The results indicate that during repeated priming with alloantigen and bulk expansion, the proliferative response of T lymphocytes becomes independent of (i) the interaction with the CD11 adhesion molecule(s), (ii) the CD5 molecule, and (iii) the cytokine IL-1 beta.     

HLA-Dw/LD directed cytotoxic T cell clones

T lymphocyte clones were derived by micromanipulation from an MLC between a stimulator and responder matched for class I but mismatched for class II HLA antigens; among the possible stimulating antigens were DR4 and Dw4. Two of the clones, when tested for cytotoxicity on a panel of DR4 positive and negative targets, appeared to recognize a determinant closely associated with Dw4, but did not lyse, with one exception, targets expressing other DR4 associated Dw specificities or DR4 negative targets. Blocking studies, using monoclonal antibodies directed against monomorphic epitopes on class I or class II molecules, revealed that the cytotoxic activity of these clones was strongly inhibited by an anti-class II (L-243) but not by an anti-class I (w6/32) monoclonal antibody. Both clones were T3⁺, T4⁺, T8^?. These findings show that cytotoxic T cell clones, (directed against class II antigens), may have specificity that correlates with a T lymphocyte-defined LD/Dw determinant. The blocking experiments using monoclonal antibodies give further support to the idea that these clones recognize determinant(s) on a class II molecule. The cell surface phenotyping results are in agreement with previous reports that most anti-class II cytotoxic clones have a T4⁺ T8^? phenotype.     

Human suppressor T cells induced in vitro with an autologous renal allograft-derived T cell line. I. Suppressor cell induction, function, and specificity

The functional characteristics of T suppressor (Ts) cells generated from the peripheral blood lymphocytes (PBL) of a kidney transplant recipient who had excellent graft function for 1 year were examined. Ts cells were induced by co-culture of PBL with an autologous alloreactive cytotoxic T lymphocyte (CTL) line (EE-1) previously grown from a routine renal allograft biopsy of this patient performed 10 days posttransplant. The EE-1 line included CD3+ T cells of CD8+ and CD4+ phenotypes with cytotoxic specificity for disparate class 1 (HLA-B8) and class II (HLA-DR1 and 3) antigens of the kidney donor (JC). The EE-1 induced Ts cell lines (designated TsEE) were found to significantly suppress (50%-95%) autologous fresh responder EE-PBL stimulation by donor EBV-transformed cells (JC-EBV) in mixed lymphocyte reaction (MLR) assay. TsEE cells were CD3+ (98%) and predominantly CD8+ (68-80%), showed no cytotoxic activity, and were suppressive only at the early phase of MLR stimulation. In three-party cell test MLR assays, TsEE-mediated suppression appeared restricted to responder cells sharing HLA-B7 with the suppressor line, and was not abrogated by the addition of exogenous interleukin-2 (IL-2). TsEE cells also showed restricted suppression of CTL generation but not mature CTL activity. The restricted suppressor activity of TsEE lines was dependent upon their induction and restimulation with the autologous EE-1 line.