Migration of natural suppressor cells from bone marrow to peritoneal cavity by live BCG

Delayed-type hypersensitivity (DTH) responses were suppressed in mice inoculated with bone marrow cells from mice that had been injected with 10(8) colony-forming units (CFU) of live BCG. Upon analysis of this DTH-suppression by the use of a macrophage migration inhibition (MI) assay, the in vitro correlate of DTH, suppressor macrophages in the peritoneal cavity were found to play an important role in DTH suppression. However, neither suppression of DTH nor production of suppressor macrophages was observed in mice inoculated with bone marrow cells from mice that had been injected with methotrexate (MTX), a folic acid antagonist, and 10(8) CFU of live BCG. Moreover, suppressor cells against the MI activity of peritoneal exudate cells from BCG cell wall-immunized mice existed in bone marrow cells from normal mice, natural suppressor (NS) cells, and they were sensitive to MTX. In addition, these NS cells phagocytized carbonyl iron particles, were adherent to Sephadex G-10, and had Fc receptors, but they had no B or T cell markers, suggesting that these cells belonged to a macrophage compartment. From this evidence, we hypothesized that the origin of suppressor macrophages in the peritoneal cavity induced by live BCG injection was MTX-sensitive NS cells in bone marrow, and that these NS cells were stimulated by a small dose of live BCG trapped in bone marrow after i.v. injection of a high dose of live BCG and migrated from bone marrow to the peritoneal cavity.     

Genetic and biological characterization of a T suppressor cell induced by anti-idiotypic antibody

The cellular and molecular characteristics of anti-idiotype-induced suppression have been investigated. We have shown that i.v. immunization of A/J or C.AL-20 mice with rabbit antibodies against the major cross-reactive idiotype on A/J anti-ABA antibodies induces splenic suppressor T cells (Ts) able to suppress T cell-mediated cytolytic and delayed-type hypersensitivity responses to ABA. In these studies, we compare the T suppressor activity manifested by anti-Id-induced suppressor cells with that described previously after conventional antigen priming. Results indicate that i.v. injection of anti-idiotypic antibodies primes for efferent level Ts; in contrast, i.v. administration of ABA-coupled cells induces afferent level suppressor cells. Soluble cell lysates, containing suppressor factor(s) derived from these anti-idiotype-induced Ts, can also mediate suppression of T cell immune responses in an efferent manner. Factor-mediated suppression is MHC-unrestricted and is also observed in mice pretreated with cyclophosphamide, suggesting that this activity is analogous to third-order suppression. Furthermore, this factor suppresses the T cell-mediated DTH and CTL responses in an antigen-nonspecific but Igh-restricted manner. These latter results suggest that the cellular elements conferring antigen specificity and Igh restriction are separate. The implications of these findings to the relationship between idiotypic elements, antigen-binding structures, and Igh restriction elements on immunoregulatory T cells are discussed.     

Suppression of the delayed-type hypersensitivity response to hen egg white lysozyme (HEL) by HEL peptides in a genetically high-responder mouse strain: evidence for requirement of the loop structure for induction of suppressor T cells

The delayed-type hypersensitivity (DTH) response of C3H/HeN mice to hen egg white lysozyme (HEL) can be blocked by a single iv injection of a solution of HEL in buffered saline 7 days before sensitization of animals with HEL in complete Freund's adjuvant (CFA). The minimal structure of HEL required for the suppression was examined by determining the abilities of various HEL-derivative peptides to inhibit HEL-DTH. Treatment of normal mice with Ploop I X II, sequence 57-107 (Cys64-Cys80, Cys76-Cys94), or P17 (sequences 1-27 and 123-129 linked by Cys6-Cys127) 7 days before immunization with HEL resulted in marked suppression of the DTH response. This inhibition of DTH involved generation of suppressor T cells (Ts). The results suggested that two suppressor pathways are involved. These data, together with another recent finding (1) that an entirely different portion of HEL is a suppressor determinant (SD) in A/J mice, indicate that different epitopes act as SDs in different strains of mice. Of the loop region peptides tested, Plc (intact loop I joined to a linear peptide, residues 84-97) was found to be the minimum structure capable of suppressing the HEL-DTH response; loop I or II alone did not cause suppression. Activation of Ts cells by the loop peptide depended on its conformational structure; completely reduced and carboxymethylated Ploop I X II did not cause suppression.     

Characterization of the suppressor cell(s) responsible for anterior chamber-associated immune deviation (ACAID) induced in BALB/c mice by P815 cells

Anterior chamber-associated immune deviation (ACAID) is a complex set of immune responses induced by the inoculation of antigens into the anterior chamber of the eye. Histocompatibility antigens, tumor-specific antigens, reactive haptens, and viral antigens have been shown to induce this phenomenon, which comprises the following specific host responses: high titer humoral antibodies, primed cytotoxic T cells, but specifically, impaired skin graft rejection and delayed-type hypersensitivity (DTH). Using the model system of ACAID induced by inoculation of P815 mastocytoma cells into the anterior chambers of H-2-compatible, but minor H-incompatible, BALB/c mice, we demonstrate that the impaired capacity of these animals to develop and express DTH is due to the activation of suppressor T cells. Generation of these cells requires an intact spleen, is not inhibited by cyclophosphamide pretreatment, and is abrogated by systemic treatment of the host with anti-I-J monoclonal antibodies. This splenic suppressor cell(s) can transfer suppression of DTH adoptively to naive syngeneic mice. One suppressor cell is Thy-1.2, Lyt-2.2, and I-Jd positive. A minority of these cells (or a second population of suppressor cells) also expresses the L3T4 surface marker. Suppression is exerted on the efferent limb of DTH expression, although afferent suppression is not excluded. P815-induced ACAID suppressor cells resemble similar cells induced by haptenated spleen cells inoculated into the anterior chamber of the eye. We propose that induction of these suppressor cells, whose target of action is selective for T DTH cells, but not for other types of T cells, is responsible for the phenomenon of immune privilege in the anterior chamber of the eye.     

Characterization of the accessory cells involved in suppressor T cell induction

The ability of UV-treated splenic adherent cells (SAC) to induce T cell-mediated immunity and suppressor T cells was analyzed in the 4-hydroxy-3-nitrophenyl acetyl (NP) system. UV irradiation of 0.88 KJ/m2 decreased the capacity of NP-coupled SAC to induce delayed-type hypersensitivity (DTH) responses by about 50%. The ability of uncoupled UV-treated SAC to induce allogeneic DTH response was also imparied, indicating that UV-treated SAC are inefficient at inducing DTH in these systems. TS1 induction by UV-treated NP-SAC was evaluated TS1 induction by UV-treated NP-SAC was evaluated by using adherent cells that were subjected to the same dose of UV irradiation that impaired DTH induction. Intravenous administration of 10(3) or 10(4) UV-treated NP-coupled SAC induced TS1 cells with the same efficiency as non-UV-irradiated cells. The TS1 cells induced in this fashion were antigen specific. Furthermore, to establish that the antigen was not reprocessed by the host, I-J-mismatched, UV-treated NP-SAC were unable to induce TS1 cells. The population of antigen-presenting cells responsible for TS1 induction appear to express both I-A and I-J determinants. TS2 induction by UV-treated accessory cells was also analyzed. TSF1 inducer suppressor factor was pulsed onto graded numbers of either normal or UV-treated adherent cells. The same levels of antigen-specific suppression were induced with normal and UV-treated cells. Finally, TS3 induction by UV-treated NP-SAC was analyzed. UV-treated and normal NP-SAC (3 X 10(3] induced antigen-specific suppression of NP DTH responses. I-J-mismatched, UV-treated NP-SAC failed to induce suppression, suggesting that the hapten was not reprocessed by the host under these experimental conditions. The accessory cell population responsible for TS3 induction appears to express both I-A and I-J determinants. Thus, there are at least two functional distinctions between the antigen-presenting cells that induce immunity vs those that induce suppressor cells. First, UV treatment selectively impairs the antigen-presenting cells, which activate the positive limb of the immune response. Second, I-J determinants appear to be specifically associated with the SAC, which induce suppressor T cells. Although these criteria can be used to distinguish the accessory cells involved in suppressor cell pathways from those controlling helper T cell induction, there were no discernible phenotypic differences among the accessory cells that induce the TS1, TS2, and TS3 subsets.     

Suppression of delayed-type hypersensitivity to third party "bystander" alloantigens by antigen-specific suppressor T cells

Delayed-type hypersensitivity (DTH) against alloantigens can be induced by sc immunization with allogeneic cells. The induction of DTH can be suppressed by iv preimmunization of the mice with similar allogeneic spleen cells, provided the cells are irradiated before injection. This suppression is mediated by T cells. The suppressor activity can be induced not only by H-2-and non-H-2-coded antigens, but also by H-2 subregion-coded antigens. Suppression induced by K, I, or D subregion-coded antigens is specific for that particular subregion as well as for its haplotype. I-J-coded alloantigens were found to not be necessary for the induction of antigen-specific suppressor T cells. After restimulation of suppressor T cells by the "specific" alloantigens, the DTH to simultaneously administered third-party alloantigens becomes suppressed as well. This nonspecific suppression of DTH to third party "bystander" alloantigens also occurs when the specific and the third-party antigens are presented on separate cells, provided that both cell types are administered together at the same site. The simultaneous presentation of both sets of alloantigens during the induction phase of DTH only is sufficient to prevent the normal development of DTH to the third-party antigens.     

Immune amplification of murine CD8 suppressor T cells induced via an immune-privileged site: quantifying suppressor T cells functionally

Background

CD8⁺ suppressor T cells exert antigen-specific suppression of the expression of hypersensitivity by activated T cells. Therefore, CD8⁺ suppressor T cells serve a major regulatory role for the control of active immunity. Accordingly, the number and/or activity of CD8⁺ suppressor T cells should be influenced by an immune response to the antigen. To test this hypothesis we used an adoptive transfer assay that measures the suppression of the expression of delayed-type hypersensitivity (DTH) by CD8⁺ suppressor T cells to quantify the antigen-specific suppression of DTH by these suppressor T cells.

Methods

Suppressor T cells were induced in the spleens of mice by the injection of antigen into the anterior chamber of an eye. Following this injection, the mice were immunized by the same antigen injected into the anterior chamber. Spleen cells recovered from these mice (AC-SPL cells) were titrated in an adoptive transfer assay to determine the number of AC-SPL cells required to effect a 50% reduction of antigen-induced swelling (Sw50) in the footpad of immunized mice challenged by antigen.

Results

Suppression of the expression of DTH is proportional to the number of AC-SPL cells injected into the site challenged by antigen. The number of AC-SPL cells required for a 50% reduction in DTH-induced swelling is reduced by injecting a cell population enriched for CD8⁺ AC-SPL cells. Immunizing the mice receiving intracameral antigen to the same antigen decreases the RSw50 of AC-SPL cells required to inhibit the expression of DTH.

Conclusions

The results provide the first quantitative demonstration that the numbers of antigen-specific splenic CD8⁺ suppressor T cells are specifically amplified by antigen during an immune response.     

Regulation of cell-mediated immunity in cryptococcosis. II. Characterization of first-order T suppressor cells (Ts1) and induction of second-order suppressor cells

Cryptococcosis patients frequently have high levels of cryptococcal antigen in their body fluids, and the levels of circulating antigen can generally be used to predict the patient's recovery, with high or rising antigen titers indicating a poor prognosis and low or decreasing levels a good prognosis. In a previous study, we reported on a murine model for studying the effects of cryptococcal antigen on host defense mechanisms. In that work, we demonstrated that an i.v. injection of cryptococcal antigen (CneF) into CBA/J mice, to simulate the antigenemia known to occur in human cryptococcosis, induced a population of T suppressor cells (Ts1) in the lymph nodes (LN). Upon adoptive transfer, the Ts1 cells specifically suppressed the afferent limb of the delayed-type hypersensitivity (DTH) response to cryptococcal antigen. In the present study, we show that the precursors of the Ts1 cells are sensitive to low-dose cyclophosphamide treatment and that the phenotype of the Ts1 cells is Lyt-1+, Ia+ (I-J+). LN cells from CneF-injected mice or a soluble factor derived therefrom can induce in the spleens of recipient mice a second-order suppressor cell population that suppresses the efferent limb of the DTH response. The cells that induce the second-order or efferent suppressor cells have the same phenotype as the cells that appear to suppress the afferent limb of the DTH response. The findings in this study indicate that a complex regulatory mechanism is responsible for the observed suppression of the DTH response in this infectious disease model. Furthermore, the suppressive circuit thus far defined for cryptococcal antigen is similar to the antigen-specific suppressor cell pathway outlined for certain chemically defined haptenic systems.     

Generation of suppressor T cells after local immunization with histocompatibility antigens

Subcutaneous (sc) hind-foot immunization (HFI) of mice with allogeneic spleen cells can induce a state of delayed-type hypersensitivity (DTH) as well as a state of suppression of DTH. This paper deals with the suppression induced by HFI. The state of suppression could be adoptively transferred by spleen cells and lymph node cells between Days 3 and 7 after HFI only. However, in the hind-foot-immunized mice the state of suppression lasted at least 25 days. The suppressor cells expressed the Thy-1+, Lyt-1-2+ phenotype and suppressed DTH antigen-specifically. The suppressor cells, however, also suppressed DTH responses to unrelated third-party alloantigens, provided the latter were administered during the induction of DTH together with the same alloantigens that were used for HFI. The HFI-induced T-suppressor cells suppressed the induction phase of DTH (i.e., the proliferative activity of the draining lymph node cells after secondary sc immunization), but not the expression phase of DTH (i.e., the activity of previously activated DTH effector T cells). H-2D compatibility between the donors of the HFI-induced T-suppressor cells and the recipients was required for the adoptive transfer of suppression. The differences in effect of local immunization versus systemic immunization on the induction and functional activity of T-suppressor cells are discussed.     

Inhibition of the delayed-type hypersensitivity response by staphylococcal enterotoxin B-induced suppressor T cells

Staphylococcal enterotoxin B (SEB) is a member of a family of gram-positive bacterial exotoxins which act as superantigens in both mouse and man. The administration of this toxin has been shown to inhibit antibody responses in vivo. We have previously shown that SEB is a potent inducer in vitro of multiple T suppressor cell populations. The present studies show that administration of microgram quantities of this toxin result in a reduced capacity to manifest a delayed-type hypersensitivity (DTH) response. In addition, we find that the failure to generate a normal DTH response appears to be due to the generation of a T suppressor cell population following SEB administration. Adoptive transfer studies show that the suppressor cells bear the CD5+ I-J+ CD4- CD8- Thy 1+ surface phenotype. The relationship of these cells to suppressor T cell populations generated following in vitro activation by SEB is discussed.     

Hapten-specific responses to the phenyltrimethylamino hapten. IV. Occurrence of mechanistically distinct idiotypic suppressor T cells before the appearance of anti-idiotypic suppressor T cells induced by the monovalent antigen L-tyrosine-p-azophenyltrimethylammonium

We have previously shown that a single i.p. injection of the monovalent synthetic antigen, L-tyrosine-p-azophenyltrimethylammonium [tyr(TMA)] in complete Freund's adjuvant induces an anti-idiotypic T suppressor cell (Ts2) population that can be detected 6 wk later by its ability to shut down delayed-type hypersensitivity (DTH) specific for the TMA hapten. In this paper we present evidence that 2 wk after tyr(TMA) administration, a subset of Ts, termed Ts1, appears that is both functionally and phenotypically distinct from the late appearing Ts2 population. The early occurring Ts1 act only at the induction phase of the DTH response and can also suppress this response intrinsically. This latter point is in marked contrast to our previous observation that the tyr(TMA)-induced anti-idiotypic Ts2 fail to function intrinsically and can only be detected upon adoptive transfer into naive mice. Ts1 bear idiotypic receptors and are Ly-1+,2- in contrast to the anti-idiotypic Ly-1-,2+ Ts2 population. In addition, unlike the Ts2 population, Ts1 are comparatively nylon wool-adherent. Adsorption of Ts1 on either antigen- or idiotype-coated petri dishes indicate that the suppressor activity can be transferred only by antigen-binding cells. Cellfree factors prepared from spleens containing the Ts1 population can suppress DTH only if administered at the induction phase of the response, in contrast to the factors derived from the Ts2 population that act both at induction as well as effector phases, suggesting that Ts1 and Ts2 can function via soluble mediators. Finally, we show that when Ts1-bearing mice are primed and boosted for anti-TMA antibody formation, the resulting response was overall reduced with respect to the idiotype-positive and negative plaque-forming cells that differs from the Ts2-bearing hosts wherein the idiotypic component is preferentially suppressed. The appearance of Ts1 before the detection of Ts2 in the same experimental animals is discussed with reference to a normal physiologic sequence of events involved in suppressor pathways.     

B and T lymphocytes regulated by idiotype anti-idiotype interactions inhibit delayed-type hypersensitivity to BCG in mice

Mice infected subcutaneously with 2 X 10(7) CFU of Mycobacterium bovis strain BCG (BCG) were able to mount a specific DTH response, whereas mice infected intravenously with the same dose of microorganisms were not. The suppression turned out to be mediated by id+ anti-PPD B lymphocytes, which arose very early during the infectious process and induced anti-id B lymphocytes. These cells were found at Day 4 after infection and exerted their effect by activating antigen-specific suppressor T lymphocytes, which affected the efferent phase of the DTH response. These results clearly indicate that the activation of a complex immunosuppressive circuit represents a mechanism by which BCG may interfere with the host's immune response already during the very early phases of infection.     

Delayed-type hypersensitivity (DTH) in BCG-sensitized mice I. Lack of suppressor T cell activity on DTH to sheep red blood cells.

Mice pretreated with an intravenous (i.v.) injection of BCG (BCG-sensitized mice) and then immunized intravenously with a high dose (10(8)--10(9)) of sheep red blood cells (SRBC) 2 weeks later developed strong delayed-type hypersensitivity (DTH) to SRBC, as in mice pretreated with cyclophosphamide (CY) (CY-treated mice) and then immunized with SRBC 2 days later; normal mice given the same dose of SRBC did not show such DTH. The mechanism of this strong DTH to SRBC which developed in BCG-sensitized mice was studied, by comparing it with that in CY-treated mice. The transfer of either whole spleen cells or thymus cells, but not serum, obtained from mice immunized with i.v. injections of 10(9) SRBC 4 days previously (hyperimmune mice) did not suppress either the induction or the expression of DTH to SRBC in BCG-sensitized mice, but suppressed those in CY-treated mice. The suppressor cells were SRBC-specific T cells. Adoptive transfer of DTH to SRBC by spleen cells from either BCG-sensitized mice of CY-treated mice to hyperimmune recipients failed. The adoptive transfer of DTH from BCG-sensitized mice to normal recipients also failed if the spleen cells from hyperimmune mice were cotransferred. Whole body irradiation (600 rad) of mice 2 hr before or after the time of immunization with SRBC reduced significantly DTH to SRBC in both BCG-sensitized and CY-treated mice. It was noticed that the total number of spleen cells in BCG-sensitized mice was 3--4 times larger than that in CY-treated mice. From these results, we conclude that the entity of effector T cells of DTH to SRBC induced in BCG-sensitized mice and in CY-treated mice was not different in terms of susceptibility to suppressor T cells and irradiation, but that the total numbers of effector T cells generated in these mice differed remarkably, resulting in the above-described different responsiveness to suppressor T cells transferred passively.     

Influence of 2'-deoxyguanosine upon the development of DTH effector T cells and suppressor T cells in vivo

Subcutaneous (s.c.) immunization of mice with allogeneic spleen cells can induce delayed-type hypersensitivity (DTH) to both major and minor histocompatibility antigens. Intravenous immunization with allogeneic spleen cells, however, induces a poor state of DTH. Furthermore, i.v. immunization with allogeneic spleen cells, especially if they have been irradiated, induces suppressor T lymphocytes. These suppressor T cells are capable of suppressing the host-vs-graft (HvG) DTH reactivity that normally arises after s.c. immunization. Moreover, they can suppress the development of anti-host DTH effector T cells during graft-vs-host (GvH) reactions. These models for HvG and GvH DTH reactivity were used to study the influence of 2'-deoxyguanosine (dGuo) and guanosine (Guo) on the generation of DTH-reactive T cells and suppressor T cells in vivo. It was found that daily i.p. administration of 0.01 mg dGuo to mice immunized i.v. partially prevented the generation of suppressor T cell activity, whereas daily administration of 0.1 or 1 mg dGuo resulted in a complete abolition. Administration of dGuo has no effect on the anti-host DTH reactivity by spleen cells from nonsuppressed donors except for when a daily dose of 10 mg is administered. This dose proved to be toxic for precursors of DTH effector T cells. Daily i.p. injection of Guo had no effect on the generation of suppressor T cells nor on the generation of DTH effector T cells. The effect of dGuo was found to be due to a direct effect on suppressor T cells and not to the induction of contrasuppressor cells. These data suggest a differential sensitivity of DTH-reactive T cells and suppressor T cells for dGuo. Because suppressor T cells and DTH-reactive T cells require proliferation for expressing maximal functional activity in the systems used, both cell types probably have different enzyme activities involved in the purine metabolism and similar deoxycytidine kinase activities, but have different nucleotidase (5'NT) activities, those in suppressor T cells being the lowest. If so, suppressor T cells will accumulate deoxyguanosine triphosphate, which causes an inhibition of the ribonucleotide reductase activity and thus of the DNA synthesis by these cells.     

T cell regulation of B cell activation: antigen-specific and antigen-nonspecific suppressor pathways are mediated by distinct T cell subpopulations

The present studies were carried out to characterize the cellular events involved in the induction and function of carrier-specific Ts cells, which selectively regulate the generation of IgG responses by Lyb-5- B cells. It was demonstrated that this regulation is in fact mediated by two distinct suppressor pathways. In one pathway, carrier-primed Lyt-1 + 2 - Ts cells are specifically activated by in vitro reexposure to the priming antigen. After this specific activation, these Lyt-1 + 2 - Ts cells are able to suppress IgG responses in an antigen-nonspecific manner. This suppression requires the participation of unprimed Lyt-1 - 2 + T cells, and is effective in both the early and the late phases of antibody responses. A second suppressor pathway requires the antigen-specific activation of primed Lyt-1 - 2 + Ts cells. Suppression of antibody responses by activated Lyt-1 - 2 + Ts cells is highly carrier specific, in contrast to the nonspecific effector function of Lyt-1 + 2 - Ts cells, appears to act without requirement for additional T cell populations; and is effective only early in the course of the antibody response. Thus, it appears that two Ts cell populations may function through distinct mechanisms to regulate the generation of IgG Lyb-5- B cell responses.     

Suppressor T cells and suppressor macrophages induced by Corynebacterium parvum

Corynebacterium parvum, injected intravenously into C57B1/6 mice (H-2^b) previously alloimmunized with P815 (H-2^d) mastocytoma cells, generated splenic suppressor cells that inhibited the development of primary cytotoxic lymphocytes in vitro. These suppressor cells differed from those generated by intravenous C. parvum injection of naive C57B1/6 mice. The former suppressor cells were effectively induced by administration of 700 μg of C. parvum whereas the latter suppressor cells were dependent upon higher doses (1400 μg) of adjuvant for their activation. Furthermore, suppressor cells generated in alloimmunized mice could only suppress C57B1/6 anti-P815 in vitro cytotoxic responses whereas suppressor cells generated in naive mice could suppress C57B1/6 anti-CBA (H-2^k) responses as well. Suppressor cells were not H-2 restricted in their action. Fractionation of spleens from alloimmunized, C. parvum-treated mice revealed the presence of suppressor T cells and suppressor macrophages. We were unable, however, to determine which cell was responsible for “antigen specificity” of suppression since the fractionation procedures seemed to trigger both suppressor cell types prior to adding them to the primary culture.     

Suppressor T cell circuits in contact sensitivity. III. A monoclonal T cell hybrid-derived suppressor factor specifically suppresses local DTH transfer by a DNP-specific T cell clone

Herein we described the direct suppressive effects of a monoclonal T cell hybridoma-derived, DNP-specific suppressor T cell factor (26.10.2 TsF) on the local transfer of delayed-type hypersensitivity (DTH) by a DNP-specific BALB/c T cell clone (dD1.9). The L3T4+, Lyt-2- dD1.9 T cell clone proliferated in response to DNP-OVA and DNBS, but not TNP-OVA or TNBS, in association with I-Ed determinants present on antigen-presenting cells. Similarly, local injection of histopaque-purified dD1.9 cell blasts resulted in DNP-specific, radioresistant, I-Ed-restricted, mononuclear cell-rich ear swelling responses. Incubation in 26.10.2 TsF specifically suppressed local transfer of DNP-specific DTH by dD1.9, but not local DTH responses transferred by BALB/c T cell clones specific for TNP or GAT. The suppressive effect of 26.10.2 TsF correlated with targeting on DNP-major histocompatibility complex determinants associated with the DTH T cell (TDH) targets. 26.10.2 TsF-mediated suppression was most pronounced after exposure of dD1.9 target cells to antigen (after the stimulation phase of the T cell clone maintenance procedure), and greatly reduced when dD1.9 was cultured for long periods in the absence of DNP (after the rest phase of clone maintenance). In additional support of this hypothesis, GAT-specific TDH, normally resistant to 26.10.2 TsF-mediated suppression, were rendered susceptible to suppression after surface DNPylation. The results demonstrate a direct, antigen-specific, effector phase regulatory effect of a monoclonal TsF on a cloned, antigen-specific T cell target, and strongly suggest that suppression is mediated via targeting on DNP determinants associated with the TDH target. Simplification of complex Ts circuitry operating in suppression of the efferent limb of DTH by the use of monoclonal TsF and cloned T cell targets should provide a basis for the future study of the molecular mechanisms of immune suppression.     

Induction of suppressor cells by a tumor-derived suppressor factor

Murine fibrosarcomas produce a factor that activates suppressor cells to inhibit expression of delayed-type hypersensitivity (DTH) responses to dinitrochlorobenzene (DNCB). This tumor-derived suppressor factor (TDSF) was partially purified by preparative isoelectric focusing of spent medium and 3 M KCl extracts of cultured methylcholanthrene-induced and spontaneous fibrosarcomas of C3H/He mice. Incubation of 1 micrograms/ml of a fraction, isoelectric pH less than 2.9, with normal syngeneic spleen cells for 1-6 hr at 37 degrees C induced suppressor cells that inhibited the primary DTH response to DNCB upon intraperitoneal transfer to normal C3H/HeJ mice. TDSF was not present in extracts of either syngeneic embryonic fibroblasts or normal spleen cells or in medium conditioned by normal peritoneal exudate cells but was present in 3 M KCl extracts of and the spent medium from four different cultured murine fibrosarcomas. TDSF activity was not restricted at the major histocompatibility complex. The suppressor cells inhibited the efferent limb of the DTH response because (1) hyporesponsive recipients of TDSF-treated spleen cells had splenic effector T cells capable of transferring DTH to DNCB into naive secondary recipients and (2) the ability of Lyt 1+,2- effector Tdth cells to transfer a secondary DTH response to DNCB was inhibited by co-incubation with macrophages or Lyt 1-,2+ T cells treated with TDSF. Preliminary biochemical analysis suggested that TDSF was an RNA- protein complex. Thus, several murine fibrosarcomas produced a soluble factor that activated splenic suppressor cells to depress the immune response to nonneoplastic antigens. These suppressor factors represent a novel group of regulatory molecules which may be ribonucleoprotein complexes.     

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.