Characterization and genetic restriction of suppressor-effector cells induced by staphylococcal enterotoxin B

The capacity of the staphylococcal enterotoxins to stimulate all T cells bearing certain (but not all) TCR has generated a great deal of interest. This stimulation appears to involve specific binding of the toxin to class II Ags and subsequent stimulation via the TCR. Previous studies from this laboratory have demonstrated that staphylococcal enterotoxin B (SEB) induces multiple T suppressor cell populations that inhibit both primary and secondary plaque-forming cell responses. Efforts to characterize these suppressor cell populations have demonstrated that the suppressor population active early in the antibody response expresses the Lyt-1-2+ cell surface phenotype, whereas depletion analysis suggests that the population active late in an ongoing response bears the Lyt-1+2+ cell-surface markers. In the present study, enrichment for this late acting effector population with the use of sequential panning with anti-Lyt mAb reveals significant suppressive activity at both the initiation and effector phases of a 5-day Mishell-Dutton coculture. Additional experiments using I-J disparate strains of mice have demonstrated a genetic restriction at the "I-J" gene locus between the cells mediating SEB-induced suppression and their target. Depletion of SEB-primed splenocytes with anti-I-J mAb suggests that both the early and late effector cells bear I-J molecules on their surface. Taken together, these results show that SEB induces suppressor cell populations with properties similar to those exhibited by Ag-specific cell circuits.     

Suppressor T lymphocyte induction by a factor released from cultured blastocysts

For the analysis of immunologic escape mechanisms of embryos during the implantation period in mice, the effects of culture supernatant of blastocysts on in vitro responsiveness to alloantigen of mice was investigated. Blastocyst-cultured conditioned medium was prepared by culturing late blastocysts of outbred ICR mice for 5 days. The addition of culture supernatant containing four or eight blastocysts to allogeneic mixed lymphocyte culture inhibited both the MLR responses and the generation of cytotoxic T lymphocytes (CTL). Preincubation of the culture supernatant with lymphocytes syngeneic to the responder cells of MLR induced potent suppressor cell activity in the MLR. The supernatant did not inhibit the activity of CTL at the effector phase, but preinduced suppressor cells obtained by incubation of splenocytes with the supernatant showed almost complete suppression of CTL activity at the effector phase. Both of the suppressor cells, active on MLR and at the generation phase of CTL as well as active at the effector phase, had a surface phenotype of Thy-1+ and Ig-. The suppressive material could be extracted from the eight-cell stage of fertilized ova or blastocysts but not from unfertilized ova, indicating that the production of the factor(s) is dependent on the stages of early embryogenesis. These results suggest that the active induction of suppressor T lymphocytes by the factor(s) released from implanted embryos is one of the protective mechanisms from maternal immunologic attack.     

Lymphocyte subsets involved in tumor-activated nonspecific feedback suppression

Cells from the spleen, lymph nodes, and peritoneum of DBA/2 mice bearing a subcutaneous tumor mediate nonspecific suppression of an in vitro antibody response to sheep red blood cells (SRBC) when cocultured with a normal T-cell subset(s). The spleen cells from the tumor-bearing mouse required for the suppression bear the Lyt 1 and Ala 1 surface markers characteristic of "inducer" T cells and activated cells, respectively. The activity of this cell population is also sensitive to irradiation. The normal T-cell subset which cooperates in the suppression bears the Qa-1 surface antigen which has been associated with suppressor cell precursors in several systems but lacks detectable surface Lyt 1 and 2 markers. Suppression of antibody responses in spleen cell cultures from tumor-bearing mice alone could also be elicited, but only when increased numbers of cells were cultured. These data are consistent with the theory that a tumor-activated, Lyt 1+ T-cell subset has the capacity to nonspecifically suppress immune responses by activating a Qa-1+ subset(s) of T suppressor cells, perhaps via feedback signals.     

Suppressor T cells, distinct from "veto cells," are induced by alloantigen priming and mediate transferable suppression of cytotoxic T lymphocyte responses in vivo

Primary and secondary cytotoxic T lymphocyte responses to minor alloantigens can be suppressed by priming host mice with a high dose (10(8) cells) of alloantigenic donor spleen cells (SC). Such suppression is antigen specific and transferable into secondary hosts with T cells. One interpretation of this is that antigen-specific host suppressor T cells (Ts) are activated. Alternatively, donor Lyt-2+ T cells, introduced in the priming inoculum, may inactivate host CTL precursors (CTLp) that recognize the priming (donor) alloantigens. Donor cells that act in this way are termed veto T cells. The experiments described here exclude veto T cell participation in transferable alloantigen-specific suppression, and demonstrate the operation of an alloantigen-specific host-derived T suppressor (Ts) cell. The origin of the Ts has been studied directly by using Thy-1-disparate BALB/c mice. The cell responsible for the transfer of suppression of a secondary CTL response to B10 minors was of the host Thy-1 allotype, and so originated in the host spleen and was not introduced in the priming inoculum. Secondly, antigen-specific Ts generated in CBA female mice against B10 minors could act on CTL responses to an unequivocally non-cross-reactive-third party antigen (H-Y), provided the two antigens were expressed on the same cell membrane. Such third-party suppression is incompatible with the operation of veto T cells. Depletion of Thy-1.2+ or Lyt-2+ cells from the suppression-inducing donor SC inoculum did not abrogate suppression induction in BALB/c mice; instead, suppression was enhanced. The demonstration of veto cell activity in similarly primed mice by other groups of investigators indicates that both types of suppression may operate. However, our results show that only antigen-specific Ts can mediate the transferable suppression of CTL responses to alloantigens.     

The role of adherent accessory cells in the generation of effector suppressor T cells

The role of accessory cell populations in the generation of effector suppressor (Ts3) cells was studied. By using an in vitro culture system, it was previously determined that the induction of NP-specific effector suppressor activity requires T cells, antigen, and an anti-idiotypic B cell population. We now demonstrate that the generation of Ts3 cells in this system also requires accessory cells. The accessory population appears to play a role in the processing and presentation of antigen. These antigen-presenting accessory cells are required early in the induction phase of Ts3 generation. These accessory cells can present NP coupled to immunogenic or non-immunogenic polypeptide carriers, including polymers of L-amino acids. However, NP coupled to polymers of poorly metabolized D-amino acids fail to induce suppressor T cell generation. Furthermore, the data demonstrate that an H-2 homology must exist between the Ts3 precursors and the antigen-presenting cell population if suppressor activity is to be generated. We also characterize the differential genetic restrictions that govern the induction of Ts3 cells that control suppression of either T cell or B cell responses. The data suggest that although I-J region encoded gene products control the induction and effector phases of suppressor cell activity as measured on T cell responses, the suppression of B cell responses appear to be controlled by I-A gene products. Possible cellular mechanisms that might explain these findings are discussed.     

Lymphocyte function in experimental African trypanosomiasis. III. Loss of lymph node cell responsiveness

The relationship between immunosuppression and suppressor cell activity in the lymphoid organs of animals with experimental African trypanosomiasis has been examined further. In the present study we measure the primary in vitro PFC response to SRBC by spleen and lymph node cells from Trypanosoma rhodesiense infected or drug-cured C57BL/6 mice. Passive transfer experiments with this culture system tested for the presence or absence of suppressor cells. We demonstrate that infected mice exhibit immunosuppression in the spleen cell population several weeks before becoming suppressed at the level of the lymph node cell populations. Although suppressor cells are present in immunosuppressed spleen cell populations, suppression of lymph node cell responsiveness was not attributable to suppressor cells detectable withi, lymph nodes. After Berenil treatment of terminally infected mice immunocompetence was restored gradually, first to the lymph node cells and subsequently to the spleen cell population. Recovery of spleen cell responsiveness was attributable to the loss of detectable suppressor cell activity within spleens. These results demonstrate that there is anatomical restriction of the suppressor cell population to trypanosome-infected mouse spleen and that loss of immunocompetence in the lymph nodes may be due to factors unrelated to suppressor cell effects.     

Induction of T15-specific suppressor T cells in mice with the CBA/N defect by neonatal injection with anti-T15 idiotype antibody

Mice with the CBA/N defect (xid) are unresponsive to phosphorylcholine (PC), To determine whether idiotype-specific suppressor T cells can also be generated in these defective mice, defective (CBA/N X BALB/c)F1 male and nondefective (CBA/N X BALB/c)F1 female or (BALB/c X CBA/N)F1 male mice were neonatally injected with antibodies specific for the major idiotype of anti-PC antibody, i.e., anti-TEPC-15 idiotype (T15id) antibody. Suppressor cell activity was examined by co-culturing spleen cells from neonatally treated F1 mice with spleen cells of normal nondefective F1 mice in the presence of antigen. Spleen cells from defective (CBA/NM X BALB/c)F1 mice treated with anti-T15id antibody demonstrated a level of suppressor activity (greater than 83% suppression) comparable to that of similarly treated nondefective F1 mice. This suppression was specific for the T15id of anti-PC response, and a Lyt-1-2+-bearing T cell population appeared to be responsible for the active suppression. These suppressor T cells recognized T15 but not PC, based on a functional absorption test. These results indicate that the CBA/N defects, including the deficiency in the anti-PC response by B lymphocytes and a possible T cell defect, do not influence the generation of T15id-specific suppressor T cells by neonatal injection with anti-T15id antibody.     

Suppressor cell regulation of encephalitogenic T cell lines: generation of suppressor macrophages with cyclosporin A and myelin basic protein.

Chronic relapsing experimental allergic encephalomyelitis (CR-EAE) can be adoptively transferred using myelin basic protein (BP)-specific helper T cell lines, and suppressor cells may be important in recovery from EAE. In order to generate suppressor cells, spleen cells obtained from BP-complete Freund's adjuvant (CFA) inoculated SJL/J mice and from normal mice were cultured for 7 days with medium, with cyclosporin A (CsA), or with CsA and antigen (BP or purified protein derivative of mycobacterium (PPD)). Cultured spleen cells were assayed for suppressor activity in vitro by coculture with BP-specific and PPD-specific helper T cell lines derived from SJL/J mice. Immunized donor spleen cells cultured with cyclosporin A (CsA) and BP were potent inhibitors of T cell line proliferation, and suppressor activity was increased 17-fold compared with control splenocytes. The number of suppressor cells required to suppress PPD-specific line proliferation by 50% (I50) was significantly higher than the number required to suppress BP-specific line proliferation, suggesting an antigen-specific component to the suppression. The major effector cell required for suppression was a large granular Mac-1+ cell with the functional characteristics of a macrophage. Suppressor activity persisted after depletion of Thy 1.2+ cells, but suppression was no longer antigen-specific, suggesting that culture of spleen cells with CsA and BP may generate suppressor macrophages which are antigen-nonspecific and Thy 1.2+ suppressor cells which are antigen-specific. These suppressor cells may be important in the regulation of CR-EAE and the techniques described for their generation may prove useful for treatment and prevention of disease.     

Characterization of anti-idiotypic suppressor T cells (Tsid) induced after antigen priming

The induction and fine specificity of idiotype-specific suppressor T cells (Tsid) were studied. Spleen cells from C57BL/6 mice, immunized 4 wk previously with NP-KLH, failed to express NPb3 idiotype-bearing PFC when challenged in vitro with NP-Ficoll or NP-Brucella abortus. After treatment of NP-primed responder cultures with anti-Thy-1.2 anti-serum + C, NPb idiotype-bearing B cells could be detected. This B cell subset was preferentially suppressed by the addition of T cells from NP-primed mice. With this reconstitution protocol, it was determined that suppression of the NPb idiotype-bearing portion of the B cell response was mediated by a specifically induced T cell population (Tsid) that directly suppressed NPb-bearing B cells. As with a previously described suppressor population induced with hapten-modified syngeneic spleen cells (Ts2), the Tsid population bound and was lysed by NPb idiotype-bearing serum antibodies. However, the Tsid could be distinguished from the Ts2 population because it lacked I-J determinants and functioned as an effector T cell, not an intermediary suppressor cell. Furthermore, fine specificity studies with monoclonal NP-specific antibodies expressing various levels of serologically detectable NPb idiotypic determinants indicated that unlike the Ts2, the Tsid population reacts with conventional, serologically detected members of the NPb family. The combined idiotype binding studies for the Tsid and Ts2 populations demonstrate that the fine specificity of suppressor T cell populations reflects their independent mechanisms of regulation.     

Mlsa generated suppressor cells. I. Suppression is mediated by double-negative (CD3+CD5+CD4-CD8-) alpha/beta T cell receptor-bearing cells.

Grafting of cells from B10.D2 (H-2d) donors into H-2 compatible lethally irradiated (DBA/2 x B10.D2)F1 hosts results in a severe graft-vs-host reaction (GVHR), developed against DBA/2 non-H-2 Ag, with only 0 to 10% of animals surviving. This GVHR mortality rate is dramatically reduced (90 to 100% of animals survive) by donor preimmunization against Mlsa determinants. The protection against GVHR correlates with a decreased B10.D2 anti-DBA/2 proliferative response in vitro. Both in vivo and in vitro phenomena are associated with activation of CD5+ suppressor T cells in the spleens of immunized mice. The present work was designed to study the origin of these suppressor cells and to further characterize their phenotype. The results show that significant suppression is not inducible in "B" mice. In contrast, in mice that were only thymectomized or else pretreated in vivo with anti-CD4 or anti-CD8 mAb, the suppressor cells are activated as efficiently as in normal mice. The suppression of GVHR mortality and proliferative responses in vitro is lost after depletion from preimmunized splenocytes of CD5+ T cells and remains unaltered after depletion of CD4+ or CD8+ T cells or both. Depletion of asialo GM1+ cells removes all NK activity, whereas the suppression is decreased only slightly. FACS analysis showed that double-negative (DN) cells from normal and immunized mice contain both CD3+ and CD3- cells; the vast majority of the CD3+ DN T cells express the alpha/beta T cell receptor. Suppression of GVHR and of proliferative responses in vitro are abrogated after elimination of CD3+ cells. These results suggest that Mlsa generated suppressor cells: 1) are derived from post-thymic long-lived T cell precursors; 2) are low asialo GM-1+ but do not exhibit NK activity; 3) belong to a subset of peripheral CD5+ DN T cells bearing a CD3-associated alpha/beta-heterodimer.     

Activated suppressor cell dysfunction in progressive multiple sclerosis

Concanavalin A (Con A)-induced suppressor activity has previously been shown to be reduced in multiple sclerosis (MS) patients with active clinical disease. In this study, we demonstrate that OKT3, as well as Con A induced suppressor activity mediated by unfractionated peripheral blood mononuclear cells is reduced in patients with the progressive form of MS. By performing reconstitution experiments involving E+, T4+, or T8+ cells derived from either MS patients or controls, and normal allogeneic macrophages or E- cells, we sought to define the cellular basis for this suppressor defect. In both MS and control groups, E+ cells were required to obtain measurable levels of suppression. Suppressor levels induced by Con A-activated cultures containing E+ cells from MS patients were lower than those induced by those containing control donor E+ cells. Suppression mediated by T8+ cells from MS patients was also lower than for controls. In the control group, suppression mediated by T8+ cells exceeded that mediated by T4+ cells; such differences were not apparent in the MS group. These results suggest that although Con A-induced suppression can be mediated by a number of T and non-T cell subsets, the functional suppressor defect measured in the MS population does involve the T8+ cell subset.     

Regulation of the antibody response by the acute phase reactant: mouse serum amyloid P-component (SAP)

Serum amyloid P-component (SAP) is the major acute phase reactant (APR) of mice. Purified mouse SAP at 0.1 to 10.0 micrograms/ml selectively suppressed the secondary in vitro IgG antibody plaque-forming cell (PFC) response to the T-dependent antigen TNP-KLH but not to the T-independent antigens TNP-LPS and DNP-Lys-Ficoll. The suppression was antigen nonspecific. The mechanism of suppression occurred primarily through the activation of Lyt-1+, I-J+ suppressor-inducer cells, which in turn activated a Lyt-2+ suppressor T-cell population. The activity of preexisting, antigen-specific Lyt-2+ suppressor T cells was not influenced by SAP. The antigen-nonspecific suppressor T cells generated by SAP were sensitive to cyclophosphamide. Removal of SAP from the culture fluid with rabbit anti-Mo SAP antibody or agarose beads abrogated the suppression. Pentraxin proteins closely related to mouse SAP, such as human SAP and hamster female protein (FP), also displayed immunoregulatory activity of the antibody response by the same cellular mechanism. The results suggest that SAP regulates antibody responses by the activation of suppressor-inducer T cells and that the regulation of the antibody response during the acute stage of inflammation may occur via SAP.     

Cyclosporine A and peripheral tolerance. Inhibition of veto cell-mediated clonal deletion of postthymic precursor cytotoxic T lymphocytes.

Veto cell-mediated suppression of CTL responses has been proposed as one mechanism by which self tolerance is maintained in mature T cell populations. We have reported that murine bone marrow cells cultured in the presence of high-dose IL-2 (activated bone marrow cells) mediate strong veto suppressor function in vitro and in vivo, and that such veto activity is effected through clonal deletion of cytotoxic T cell precursors. In our studies, we have determined that bone marrow cell populations from athymic NCr-nu mice (H-2d) mediate strong veto cell activity without exposure to exogenous IL-2 in vitro. To examine mechanisms by which these naturally occurring veto cell populations in BM suppress precursor CTL (pCTL) responses, we used as a responding cell population in MLC, spleen cells of transgenic mice expressing at high frequency TCR specific for H-2 Ld encoded Ag with stimulation by H-2d-expressing cells in culture. Flow cytometric analysis was performed by staining the responding MLC cell population with the mAb 1B2 specific for the transgene-encoded TCR and determined changes of 1B2+ T cells. Such experiments demonstrated that the anti-H-2d cytotoxic response by these cell populations was specifically suppressed by NCr-nu (H-2d) bone marrow, and that 1B2+ pCTL were in fact specifically deleted from the responding cell population by incubation with such naturally occurring veto cell populations expressing the appropriate target Ag. In addition, to further understand the interactions of pCTL and veto cells and possible contributions by the latter to peripheral tolerance, we evaluated the effect of cyclosporine A (CsA) on veto cell-mediated suppression of pCTL of the transgenic mice. CsA inhibited veto cell-mediated suppression of cytotoxic T cell responses, and this inhibition correlated with a lack of clonal deletion of pCTL by veto cells in the presence of CsA. Furthermore, CsA exerted its effect through pCTL and not through veto cells, indicating that pCTL may play an active role in their own deletion by veto cells.     

Changes in the host natural killer cell population in mice during tumor development. 2. The mechanism of suppression of NK activity

Our earlier studies revealed that a rapid and progressive loss of splenic NK activity in mice during the development of a number of transplanted tumors as well as of spontaneous tumors was due to an inactivation of natural killer (NK) lineage cells rather than to their disappearance. The mechanism of this inactivation have now been explored in CBA/J mice receiving transplanted Ehrlich ascites tumors and in C3H/HeJ mice bearing spontaneous mammary tumors or receiving transplants of syngeneic mammary tumor lines of recent origin. A poor activation state or maturation arrest of NK lineage cells due to a low interferon level in vivo was ruled out, since the host NK activity could not be restored after administration of either an interferon inducer poly(I:C) or interferon-alpha, although such treatments enhanced the activity in tumor-free mice by four- to eightfold. Possible presence of host suppressor cells acting on the effector or preeffector stage of NK cells was explored by mixing spleen cells from tumor bearers with normal spleen cells either during the NK assay, or for a 20-hr period of in vitro short-term culture prior to the NK assay. Mixing during the NK assay led to a reduction of NK activity explicable by a simple dilution of active NK cell concentration rather a suppression of active NK cells. On the other hand, a 20-hr coculture of the mixed population at various ratios led to a complete abrogation of the NK activity, indicating that the suppressor cells acted on the preeffector stage of the NK Lineage. A further characterization of suppressor cells revealed that they were (1) contained in the adherent fraction of the spleen of tumor bearers, (2) of monocyte/macrophage morphology, (3) capable of phagocytosing latex particles, and (4) positive for surface Mac-1 antigen, as noted from a radioautographic binding of 125I-labeled monoclonal anti-Mac-1 antibody. The mechanism of the suppression was identified, at least in part, as being mediated by prostaglandin-like molecules, since the presence of indomethacin, a prostaglandin-inhibitor, during the 20-hr coculture period completely abrogated the suppression. Indomethacin exerted no direct effect on the recruitment or killer activity of NK lineage cells in vitro. NK cell suppression may be another normal immunoregulatory mechanism which alters the host-tumor balance in favor of the tumor rather than the host.     

Immunoregulation of lysozyme-specific suppression. III. Epitope-specific amplification of immunosuppression induced by monoclonal suppressor-T-cell products

The hen egg-white lysozyme (HEL)-specific suppression induced by soluble molecules produced by a monoclonal T-cell lymphoma line (LH8-105) obtained from HEL-specific suppressor T lymphocytes has been examined. Injection of I-J+ molecules from LH8-105 cell culture supernatant (TsFa) in HEL-primed mice during the afferent phase of the response induced Lyt-2+ second order suppressor T (Ts) cells which, upon transfer into HEL-CFA-primed syngeneic recipients, inhibit the delayed-type hypersensitivity (DTH) response to HEL. Transfer of spleen cells from TsFa-injected mice primed with HEL or human lysozyme suppresses the DTH response to HEL in recipient mice whereas this response is not affected by cell transfer from ring-necked pheasant egg-white lysozyme (REL)-primed and TsFa-injected mice, indicating that induction of second order Ts by TsFa is specific for a lysozyme epitope including phenylalanine at position 3. Fine antigenic specificity of second order Ts-cell induction is confirmed by similar results obtained upon injection of TsFa in mice primed with HEL N-terminal synthetic peptide or with an analog in which, as in REL, phenylalanine has been substituted by tyrosine at position 3. The same fine antigenic specificity observed in the induction of second order Ts cells is also present in the expression of TsFe suppressive activity. The similar antigenic specificity of Tsa and Tse suggests that Tse cells could result from amplification of the Tsa cell population or these two cell subsets could reflect different maturation stages of the same cell type rather than distinct T-cell populations activated in cascade.     

Immunosuppressive activity of staphylococcal enterotoxin B. II. Activation of suppressor-effector cells by a staphylococcal enterotoxin B-induced suppressor factor

The capacity of staphylococcal enterotoxins to stimulate all T cells bearing certain T cell receptors has recently generated a great deal of interest. These toxins are believed to bind directly both to the TCR:CD4 complex via its V beta domains and to class II MHC molecules on accessory cells prior to T cell activation. Previous studies from this laboratory have demonstrated that staphylococcal enterotoxin B (SEB) is capable of inducing multiple T suppressor cell populations which can inhibit in vitro antibody responses. Additional studies have demonstrated that the suppressive activity of these cells is mediated, at least in part, by an I-J-restricted suppressor factor. Efforts to characterize the inhibitory activity of this factor have demonstrated that the suppressive element is capable of activating both early and late acting suppressor cell populations in vitro. Analysis by both positive and negative selection shows that cells bearing the Lyt1-2+ surface marker phenotype are active early, whereas Lyt1+2+ cells are active both early and late in the antibody response. Additional experiments using various strains of mice as sources of suppressor factor and of naive splenocyte populations have demonstrated that activation of suppressor-effector cells by this suppressor factor is restricted at the I-J, but not Igh, gene locus. These studies suggest that this SEB-induced suppressor factor alone provides the signals necessary for the induction and activation of suppressor-effector cell activity.     

Isolation and characterization of novel suppressor T cell clones from murine fetal thymus

Murine fetal thymus from C57BL/6J (B6) and DBA/2J contains a cell population that suppresses CTL responses to alloantigens. This suppressor cell population was found to exist in high frequency in murine fetal thymus at the 14th day of gestation. The activity of this cell in the thymus declined rapidly with increasing time of gestation, and suppressor activity in the thymus was undetectable by the time of birth. On the other hand, suppressor activity could be detected in organ cultures of 14-day fetal thymus even after the organs were cultured for 14 or 21 days. Fetal thymocytes from B6 or DBA/2J mice were grown as long-term lines in interleukin 2 (IL 2)-containing medium. Clones of suppressor cells were derived from long-term cultures by micromanipulation. The clones had an average doubling time of 13 to 16 hr and were dependent on IL 2 for growth. The clones were 10- to 100-fold more efficient in suppressing CTL responses to alloantigens than day 15 fetal thymocytes. Analyses of cell surface molecules with the use of monoclonal antibodies and conventional anti-H-2 sera by radioactive binding assays showed that cloned suppressor cells from B6 fetal thymus were Thy-1 and Lyt-2+, and expressed little or no L3T4, Lyt-1, H-2K, H-2D, and class II molecules. The suppressor clones lacked the cytolytic activity of conventional CTL and also served as very poor target cells in CTL-mediated cytolysis. The suppressor function of the cloned cells was radiation-resistant, and this suppression could not be reversed by the addition of excess exogenous IL 2. The cloned cells suppressed CTL responses only when they were added within the first 48 hr of a 5-day culture period. Analyses of the antigen specificity of the suppressor cells showed that they suppressed CTL responses in a nonantigen-specific manner.     

Role of dextran-specific suppressor T cells in the regulation of the immune response by a reactive form of dextran

Reactive forms of antigens or haptens have been shown to induce a state of hyporesponsiveness mediated in part by suppressor T cells. Injection of Balb/c x C57B16 F1 (CB6F1) mice with a reactive form of dextran B1355S (periodate oxidized dextran, dex-P) specifically reduced responses to dextran immunization within 1 day after dex-P treatment. This unresponsiveness lasted at least 23 days and required a reactive form of dextran for its induction since native dextran and oxidized/reduced dextran failed to induce tolerance. Furthermore, hyporesponsiveness could be induced by iv injection of dextran-coupled cells, especially peripheral blood lymphocytes, a result which suggests that in vivo coupling to cellular antigens is involved in dex-P-induced hyporesponsiveness. Suppression of the anti-dextran response could be transferred to normal mice with T-cell-enriched spleen cell populations from dex-P-injected mice. Interestingly, the presence of B cells in the transferred cell preparations interfered with detection of suppression. Both Lyt 1+2- and Lyt 1-2+ cells were involved in the dex-P-induced suppression; indeed, mixtures of these types of T cells led to the most profound degree of suppression. The suppressive activity of spleen cells from dex-P-injected mice could be removed by passage over dextran-coated plates. Moreover, cells eluted from the plates specifically suppressed anti-dextran responses of normal mice, indicating that dex-P injection induces a population of antigen-binding suppressor cells. This system will allow the study of the suppressor-T-cell receptors in a well-defined idiotypic system.     

BCG-induced suppressor cells. I. Demonstration of a macrophage-like suppressor cell that inhibits cytotoxic T cell generation in vitro.

Spleen cells obtained from mice 5 to 40 days after infection with viable BCG organisms (BCG-spleens) were found to be unresponsive in vitro to both mitogenic and alloantigenic stimuli. Moreover, suppressor cells could be demonstrated in the spleens from these infected animals. When spleen cells from BCG-infected mice were added to either syngeneic or allogeneic normal spleen cells, the mixtures neither proliferated nor developed cytotoxic activity when cultured with alloantigen or with concanavalin A (Con A). The development of unresponsiveness post-infection paralleled the onset of suppressive activity. Spleen cells obtained from mice given heat-killed BCG were neither suppressive nor unresponsive. The suppressive activity of BCG-spleen cells was associated with an adherent, phagocytic cell that lacked membrane-associated Thy-1 antigen. Removal of this cell by passage through nylon wool columns resulted in a cell population that was no longer capable of suppression and that responded normally to alloantigen and to Con A. It would thus appear that BCG infection results in the development of a "suppressor" macrophage-like cell population within the spleen. The role of this cell type in regulation of the immune response in BCG-infected animals is as yet undefined.