Macrophage-mediated suppression of immune responses in Toxoplasma-infected mice. II. Both H-2-linked and -nonlinked control of induction of suppressor macrophages

The suppressive effect of Toxoplasma infection on initiation of memory cells to dinitrophenylated keyhole limpet hemocyanin (DNP-KLH) was drastically different among inbred strains of mice. C57BL/6 (B6), C57BL/10 (B10), and SJL mice showed markedly suppressed secondary anti-DNP responses when infected. In contrast, the suppression did not occur in BALB/c mice. The infected DBA/2 and C3H/He mice produced moderately suppressed responses. In B6 mice, an injection with 1 X 10(2) organisms of T. gondii induced a suppressed elicitation of the memory cells to DNP-KLH. However, in BALB/c mice, the responses were not affected even by inoculation with 1 X 10(4) organisms. The difference in the suppressive effect of infection between B6 and BALB/c mice was also observed in the primary anti-DNP antibody responses to DNP-KLH. Both H-2-linked and -nonlinked genes appeared to be responsible for the regulation of the immunosuppression, since the suppressive effect of infection in B10.D2 mice, which have the B10 background and the same H-2 haplotype as BALB/c, was weaker than that of B10 mice, but stronger than in BALB/c mice. In vitro studies using a primary anti-sheep erythrocytes (SRBC) antibody response system demonstrated that the activation of plastic-adherent suppressor cells by Toxoplasma infection, in which suppressor macrophages have been proved to be the responsible cells for the suppressive activity, was controlled by both H-2-linked and -nonlinked genes.     

Macrophage-mediated suppression of immune responses in Toxoplasma-infected mice. III. Suppression of antibody responses to parasite itself

In acute Toxoplasma infection, anti-sheep erythrocytes (SRBC) antibody responses were strongly suppressed in the infected C57BL/6 mice, and the mice produced low titers of only 2-mercaptoethanol (2-ME)-sensitive antibodies but not 2-ME-resistant antibodies. By contrast, the infected BALB/c mice produced much higher titers of both 2-ME-sensitive and -resistant anti-SRBC antibodies than the infected C57BL/6 mice. In anti-Toxoplasma antibody responses, the 2-ME-resistant antibody titers were significantly lower in the infected C57BL/6 mice than in the BALB/c mice in the early phase of infection, suggesting that the suppressive effect of Toxoplasma infection affects antibody responses to Toxoplasma itself as well as to the unrelated antigen, SRBC. A histological study revealed that in the infected C57BL/6 mice, a large number of acid phosphatase-positive, macrophage-like cells infiltrated into the follicles of their spleens, and an involution of follicles occurred in the acute phase of infection. This histological change was not observed in the infected BALB/c mice. The infected C57BL/6 mice, which had the suppressed anti-Toxoplasma antibody responses, made five times as many as cysts in their brains as compared with the BALB/c mice at the fifth week of infection.     

Suppression of antibody responses in allogeneic mice by products of lymphoid tissue. I. Allogeneic suppressive factor (ASF) from spleens repopulated with thymus cells.

A cellfree extract prepared from the spleen cells of C3H mice is capable of suppressing antibody responses to SRBC when extract material is exposed to alloantigens. The observed immunosuppression was attributed to a soluble factor in the extract. This allogeneic suppressive factor (ASF) was detected in extracts prepared from the spleen cells of unirradiated mice as well as those of irradiated mice repopulated with thymocytes, provided that mice were previously immunized with SRBC. Donors of actively suppressive ASF preparations did not need to be previously exposed to alloantigens. Extracts from thymus and marrow cells of unirradiated mice and the spleen cells of irradiated mice repopulated with marrow cells (or no cells) did not contain ASF. C3H thymocytes stimulated with SRBC generated more ASF activity in spleens of C3BF1 hosts than in those of C3H hosts, indicating that alloantigenic stimulation enhances the production or activity of ASF. Once produced, C3H ASF was able to suppress antibody responses in cell transfer experiments only if exposed to C3BF alloantigens of either donor lymphoid cells or irradiated hosts. Once exposed to alloantigens, ASF appears to be capable of suppressing antibody responses of syngeneic C3H or semi-allogeneic C3BF cells. When both donor lymphoid cells and hosts were syngeneic with the donor of the ASF, there was enhancement of antibody formation in cell transfer experiments. C3H ASF did not interfere with education of C3BF thymocytes to SRBC or with the generation of precursors of anti-SRBC antibody-forming cells by C3BF1 marrow cells. ASF may interfere with cellular cooperative events necessary for humoral immune responses or with terminal differentiation of B cells. Production of ASF could partially account for the suppression of antibody responses observed during graft-vs-host reactions.     

Proliferation of natural suppressor cells in long-term cultures of spleen cells from normal adult mice.

Natural suppressor cells were induced by culturing spleen cells from normal adult mice for 2 to 3 wk. The suppressor cells were large in size, nonadherent and nonspecifically suppressed the plaque-forming cells response of fresh spleen cells to SRBC in vitro. The suppressive activity of the cells was not affected by treatment with indomethacin or anti-Thy-1, anti-Ig, anti-Ia, or anti-asialoGM1 plus complement. Phenotype analysis by FACS showed that Thy-1, L3T4, Ly-2, CD3-epsilon, TCR-alpha beta, Ig, B220, Ia, and asialoGM1 Ag were all absent in the suppressor cells, although they were wheat germ agglutinin receptor positive. The suppressor cells did not demonstrate cytotoxicity against either YAC-1 or P-815 cells. Enriched large cell populations from fresh normal spleens expressed the same phenotypes and also exhibited the suppressive activity. These findings suggest that a minor population of natural suppressor cells exist in the normal adult mouse spleen and they proliferate during the in vitro culture of spleen cells.     

Immunologic suppression after oral administration of antigen. I. Specific suppressor cells formed in rat Peyer's patches after oral administration of sheep erythrocytes and their systemic migration.

Rats given 10(10) sheep erythrocytes (SRBC) orally were found to contain specific suppressor cells to SRBC in their Peyer's patches (PP) and mesenteric lymph nodes (MLN) after 2 days of feeding. After 4 days of feeding, similar suppressor cells were found in the thymus and spleen, but they were missing in the PP or MLN. These suppressor cells effectively blocked IgM and IgG plaque-forming cell responses to SRBC in Mishell-Dutton cultures and delayed-type-hypersensitivity responses to SRBC when transferred to syngeneic recipients, but they did not affect responses to horse erythrocytes. The orally induced specific suppressor cells appeared to be T2 cells since their activity was eliminated by in vivo treatment of SRBC-fed rats with anti-rat lymphocyte serum but not by adult thymectomy. Because carrageenan partially relieved the suppression observed in culture, the actual suppressive mechanism may also involve a macrophage.     

Modulation of mouse anti-SRBC antibody responses by placental extracts. II. Antigen specificity and regulatory role of B and T cell populations affected by two distinct placental fractions

Previous results from our group had shown that when CBA mice are primed to sheep red blood cells (SRBC) in the presence of various doses of placental extract (PE) (or liver extract [LE] as control), their spleen cells injected into normal syngeneic recipients have important immunoregulatory properties. Low doses of PE (0.25 to 4 mg per mouse) induce a marked decrease of the PFC response against SRBC in recipient animals. In contrast, higher doses of PE (8 to 13 mg per mouse) have a potentiating effect on the same response. LE is not different from a saline injection, at any dose. The suppressive and enhancing effects can be reproduced with two distinct placental fractions (PF) of 40 KD and 60 KD, respectively. In the present report, we have studied the requirement for an antigenic stimulation at the same time as the injection of PE, and the antigenic specificity of the subsequent immunoregulatory effects. In addition, we have analyzed the functional properties of the spleen cell populations affected by PE or placental fractions: surface Ig- cells mediate the suppressive effect due to low doses of PE or the 40-KD fraction, whereas surface Ig+ cells are responsible for the enhancing effect due to high doses of PE or the 60-KD fraction. These immunoregulatory activities do not appear to be related to the presence of Ig fragments in PF, because our results have shown that no Ig fragments can be detected in either PF. Surface Ig- T cell populations from spleen cells treated with the 40-KD fraction and antigen have been further separated into Lyt-2- and Lyt-2+ subpopulations. Our results showed that Lyt-2+ cells alone suppress the PFC response anti-SRBC in both normal and irradiated syngeneic recipients. Thus, the injection of a 40-KD PF in the presence of antigen activates splenic T suppressor cells capable of specifically regulating a secondary antibody response in vivo.     

Suppressor T cells induced by soluble immune complexes can adoptively transfer inhibition of cytophilic antibody receptors on macrophages.

Immune complexes (soluble antigens of L1210 and antibody to L1210) when given to allogeneic C3H mice generated suppressor cells that inhibited receptors for cytophilic antibody on macrophages. Thymocytes or nylon-nonadherent splenic T cells (4 × 10⁷) from immune-complex-treated mice transferred this suppressive activity when injected into normal syngeneic mice. Maximal suppression of macrophages occurred 4 to 6 days after transfer. In contrast, even 5 × 10⁷ nylon-adherent, non-T spleen cells from immune-complex-treated (“suppressed”) mice failed to induce macrophage suppression in the syngeneic recipients. When T-cell-depleted “B” mice were used as recipients, neither thymocytes nor splenic T cells from suppressed mice were able to transfer suppressive activity. However, the admixture of 2 × 10⁷ normal syngeneic thymocytes with 4 × 10⁷ thymocytes from suppressed mice restored the latter's ability to elicit suppression of macrophages in T-cell-deprived recipients. Peritoneal monocytes from recipients of suppressor thymocytes (to L1210) could not attach cytophilic antibody to L1210 but could attach cytophilic antibody to EL-4 and sheep erythrocytes. Thus, suppressor T cells induced by immune complexes can transfer immunologically specific macrophage suppression (inhibition of cytophilic antibody receptors) to syngeneic recipients. The suppressor cells required the cooperation of normal T cells, suggesting either recruitment of suppressor cells from, or a helper effect by, the normal T cells, in order to produce their effect.     

Suppressor cells in mice infected with Trypanosoma brucei.

Within 2 to 3 days of infection with Trypanosoma brucei strain S42, the ability of spleen cells from infected CBA mice to mount a primary in vitro antibody response to sheep red blood cells (SRBC) is profoundly reduced, and suppressor cells are generated as detected by cell mixture experiments. Suppressor cell activity lies in the T and adherent cell compartments of spleens from infected mice, but not in the B cell compartment, although antibody responses to a thymus-independent antigen, DNP-Ficoll, are significantly reduced. Suppression of antibody responses of normal spleen cells depends on viable cells from infected mice. The trypanosome, itself, plays no direct role in suppression, and we have ruled out the possibility of antigenic competition as a mechanism of suppression. Our data is consistent with the model of suppressor T cells induced by concanavalin A mitogenesis. We hypothesize that trypanosome antigens may directly stimulate T cells with the concomitant release of factors with affinity for macrophage surfaces thus becoming suppressive for T and B cell responses.     

Lymphocyte function in experimental African trypanosomiasis. VIII. Loss of suppressor T cell function in lymph nodes

The immunosuppression that occurs in mice experimentally infected with African trypanosomiasis has been examined further. In the present study we have examined lymph node cells from Trypanosoma rhodesiense-infected C57Bl/6J mice for the ability to produce mitogen induced antigen-nonspecific suppressor T cells (Ts). Inguinal, mesenteric, and brachial lymph node cells were harvested from uninfected control mice and from mice at different periods of infection. These cells were cultured with or without concanavalin A (Con A) for 48 hr to induce Ts activity. After stimulation, the control and infected lymph node cells were passed over Sephadex G-10 columns to remove suppressor macrophages that arise during the infection from Con A-induced Ts. The column passed cells were then added to normal mouse responder spleen cells in a primary in vitro antibody response culture system with sheep erythrocytes (SRBC) as antigen. The resultant plaque-forming cell responses to SRBC indicated that Ts function was not induced in infected lymph node cell populations. However, early in the infection, a stimulatory signal was provided by both the untreated and Con A-treated infected lymph node cells, which was lost in the terminal stage. Determinations of T cell subpopulations revealed that the infected Lyt 2.2-bearing subpopulation was not significantly altered from normal controls. We conclude that T. rhodesense infected mice fail to mount normal lymph node cell antigen nonspecific Ts responses and that this loss of activity may be due to an intrinsic dysfunction in the suppressor T cell population.     

Suppressive effect of nonviable Mycobacterium paratuberculosis on the delayed-type hypersensitivity reaction to sheep erythrocytes in mice.

The effect of nonviable Mycobacterium paratuberculosis on the delayed-type hypersensitivity reaction to sheep erythrocytes (SRBC) in mice was evaluated by means of delayed-type footpad swelling. Intraperitoneal (i.p.) injection with nonviable M. paratuberculosis into mice from 28 days before to 1 day after immunization with SRBC resulted in a significant suppression of foot-pad swelling to SRBC. The suppressive effect could be transferred by i.p. injection of spleen cells or peritoneal exudate cells from mice which had been pre-treated with nonviable M. paratuberculosis into non-treated recipient mice. The suppressive effect of spleen cells was retained even after passing them through a nylon wool column. The suppressive effect of spleen cells was abolished by treatment with anti-Thy 1.2 monoclonal antibody plus complement or anti-Lyt 2.2 monoclonal antibody plus complement. However, treatment of spleen cells with anti-mouse gamma globulin antiserum plus complement or anti-Lyt 1.2 monoclonal antibody plus complement did not affect the suppressive effect of spleen cells. The suppression of footpad swelling to SRBC induced by pre-treatment with nonviable M. paratuberculosis could be reversed by i.p. administration of cyclophosphamide. Serum antibody response to SRBC in mice was not affected by pre-treatment with nonviable M. paratuberculosis. These findings indicate that T cells appear to be involved in the suppression of delayed-type hypersensitivity reaction to SRBC in mice by pre-treatment with nonviable M. paratuberculosis.     

Activation of distinct helper and suppressor T cells in experimental trypanosomiasis.

Spleen cells taken from mice soon after infection with Trypanosoma brucei S 42 enhance the primary in vitro antibody response of normal spleen cells to sheep red blood cells (SRBC), but do not affect their response to DNP-Ficoll. Spleen cells harvested later in the infection (day 6 onwards) suppress the antibody response of normal spleen cells to both SRBC and DNP-Ficoll. The enhancing and suppressive effects of "infected" spleen cells are sensitive to treatment with anti-Thy 1.2 anti-serum and complement, and can be mediated by nylon wool-purified populations of T cells. The enhancing T cell is sensitive to ALS, not lost within 4 weeks of adult thymectomy, and bears the Ly-1+, 23- phenotype. The suppressor T cell is insensitive to ALS, lost within 20 weeks of adult thymectomy, and bears the Ly-1+, 23+ phenotype. The significance of the activation of distinct helper and suppressor T cells is discussed in relation to the pathogenesis of trypanosomiasis.     

Helper cell factors restore antibody responses of allogeneic bone marrow chimeras: evidence for ineffective cellular interactions

We investigated the nature of deficient antibody responses to SRBC in stable, fully allogeneic bone marrow chimeras. No evidence for a suppressor cell-mediated mechanism was found. Chimera spleens possessed adequate numbers of antigen-reactive B cells to produce a normal antibody response. Using separated chimera cell populations and soluble helper factors, we assessed the functional capabilities of chimera B cells, T cells, and macrophages. Our data suggest that the failure of allogeneic chimeras to produce antibody is not the result of impaired B cell, T cell, or macrophage function, but rather that it is due in ineffective cellular interactions that normally result in the generation of helper factors. In vitro stimulation of chimera macrophages with LPS, and of chimera spleen cells with Con A, resulted in the release of soluble helper factors that were capable of fully restoring chimera B cell responses.     

Demonstration of active suppressor cells in spleens of young NZB mice

NZB mice, a strain prone to the development of autoimmune disease, have during the first 2 weeks of life suppressor cells in their spleens which can in coculture with adult spleen cells suppress the antibody response to sheep red blood cells (SRBC) generated in culture by the adult cells. The suppressive activity of spleen cells from NZB mice in the first week after birth is similar to that of spleen cells from 4-day-old C57BL/6 mice, a strain which does not spontaneously develop autoimmune disease. As in “normal” strains of mice, suppressor cell activity in NZB mice is diminished at 2 weeks and undetectable at 3 weeks of age. The data indicate that there is no defect inherent in the suppressor cells detected in the spleens of newborn and young NZB mice and suggest that the development of autoimmune responses does not result from a lack of suppressor cells in the young animals.     

Immunologic dysfunction during viral oncogenesis. I. Nonspecific immunosuppression caused by malignant rabbit fibroma virus

Malignant rabbit fibroma virus (MV) is a potent oncogenic poxvirus that produces a rapidly progressive syndrome of disseminated myxosarcoma, immunosuppression, and fatal gram-negative infection. MV is probably a recombinant between Shope fibroma virus (SFV) and rabbit myxoma virus, and is capable of preventing or aborting the in vitro proliferative responses of rabbit lymphocytes to B and T lymphocyte mitogens. Proliferative responses to sheep erythrocytes (SRBC) are similarly affected, although MV does not alter ongoing antibody responses to SRBC. Splenic lymphocytes from MV tumor-bearing rabbits suppress antibody and proliferative responses to SRBC when added to lymphocytes from SRBC-primed rabbits. Finally, lysates of cultured splenic lymphocytes from rabbits given MV suppress both proliferative and antibody-forming responses to SRBC. When MV is removed from these lysates by UV inactivation or by centrifugation, the suppressive activity remains. We therefore conclude that MV induces immunologic unresponsiveness in rabbits by at least two mechanisms. First, a direct suppressive effect of added virus on in vitro lymphocyte proliferation is seen. There is no effect in this situation if an antibody response is already in progress. Second, spleen cells exposed to MV in vivo produce one or more soluble factors capable of suppressing both proliferative and antibody responses of normal lymphocytes.     

Effects of antigen-feeding on intestinal and systemic immune responses. III. Antigen-specific serum-mediated suppression of humoral antibody responses after antigen feeding.

Feeding mice sheep erythrocytes (SRBC) caused a significant decrease in splenic IgM antibody responses to SRBC given ip. Reduced IgM responses were due to a suppressor factor in the serum of fed mice rather than due to a lack of IgM antibody-forming cell precursors or to the presence of suppressor T cells. Although feeding initially primed mice to produce greater IgA and IgG anti-SRBC responses after SRBC challenge, the initial primed state was transitory. Mice fed SRBC for longer than 8 weeks had significantly reduced splenic IgG and IgA responses after SRBC challenge.Suppression of IgM responses by serum from fed mice was antigen-specific and not H-2 restricted. Serum from fed mice inhibited the induction of IgM anti-SRBC responses but did not block the expression of already established responses. The size of the suppressor factor and the ability to remove suppressor activity from serum by anti-mouse immunoglobulin suggested that suppression was mediated by antibody. However, the determinants against which the antibody was directed appeared to differ among batches of suppressor sera. Suppressor activity did not appear to be mediated by immune complexes, or soluble antigen. Oral feeding of antigen can have a marked influence on host systemic immune responses when the antigen used for feeding is subsequently administered parenterally. Thus, oral antigen administration may provide a way for specifically manipulating systemic immune responses in vivo. In addition, antigen-feeding may provide a means for producing transferable factors that suppress humoral antibody responses.     

In vitro activation of suppressor cells from spleens of mice treated with radioactive strontium

Mice were treated with two 100-muCi injections of 89Sr to deplete marrow-dependent (M) cells. Mice so treated responded normally to immunization with sheep red blood cells (SRBC) in vivo; moreover, spleen cells from 89Sr-treated mice were able to respond to SRBC after infusion into irradiated recipient mice. However, spleen cells from mice treated with 89Sr did not respond to SRBC in vitro and mixtures of normal spleen cells with the latter were also not able to respond in vitro. The discrepancy between in vivo and in vitro responses was abolished by culturing spleen cells for 24 hr before testing their ability to respond to SRBC in the adoptive transfer in vivo. Pretreatment of spleen cells from 89Sr-treated mice with 1000 R of gamma-radiation lessened their suppressive activity. The suppressor cells were detected in spleens of athymic nude mice treated with 89Sr. The suppressive activity, after the 24-hr culture period, was not abolished by irradiation and was active in vivo as well as in vitro. Thus, depletion of M cells by 89Sr results in the appearance within the spleen of thymus-independent suppressor cells, which require a short period of in vitro cultivation before becoming functionally active.     

Soluble factors from BCG-induced suppressor cells inhibit in vitro PFC responses but not cytotoxic responses.

A macrophage-like suppressor cell is present in the spleens of BCG-infected C57BL/6 mice. This suppressor cell is capable of suppressing both in vitro cytotoxic and PFC responses of normal C57BL/6 spleen cells. Suppression was not caused by changes in the kinetics of the responses or in the quantities of antigen required for stimulation. Suppression of the in vitro cytotoxic response could not be linked to any soluble mediator. In contrast, supernatants obtained from BCG spleen cell cultures, which failed to inhibit alloantigen-induced cytotoxic responses, suppressed the in vitro PFC response to SRBC by normal C57BL/6 spleen cells. It is postulated that either BCG-induced macrophage-like suppressor cells inhibit these in vitro responses via different mechanism(s) or these responses are regulated by different suppressor cell subpopulations within the monocyte/macrophage compartment of BCG spleen cells.     

Heterologous antibody responses in mice with chronic T. cruzi infection: depressed T helper function restored with supernatants containing interleukin 2

Antibody production to sheep erythrocytes (SRBC) or hapten-conjugated SRBC (TNP-SRBC) was studied in mice with chronic Trypanosoma cruzi infections. Studies in vivo demonstrated that both IgM and IgG anti-SRBC responses were suppressed during chronic infection. Secondary IgG responses were suppressed regardless of whether the primary immunization was given before or after infection. The ability of cells from infected mice to provide help for antibody production was examined in vitro. Anti-SRBC responses were restored to cultures of whole spleen cells from infected mice by the addition of interleukin 2 (IL 2)-rich supernatants, indicating that these cells were capable of antibody production when sufficient help was provided. T cells from SRBC-primed infected mice were unable to provide significant help to normal B cell/M phi cultures for in vitro anti-TNP or anti-SRBC responses. The percentages of Thy-1+, Lyt-1+, and Lyt-2+ spleen cells were not significantly different between normal and infected mice. Anti-TNP and anti-SRBC responses were restored to cultures that contained T cells from infected mice and normal B cell/M phi by the addition of IL 2-rich spleen cell supernatants. The suppression of in vitro antibody responses in mice with chronic T. cruzi infections was associated with a lack of T cell help, which was provided by exogenous spleen cell supernatant.     

Suppression of Con A mitogen-induced proliferation of normal spleen cells by macrophages from chickens with hereditary muscular dystrophy

Spleen cells from chickens with hereditary muscular dystrophy (MD) give low blastogenic responses to the T cell mitogen concanavalin A (Con A) while exhibiting normal mitogen stimulated blastogenic responses to the T cell mitogen phytohemagglutinin (PHA). The addition of MD spleen cells to normal spleen cells caused a marked suppression of the Con A response of the normal cells while not affecting the PHA response of the normal cells. The suppressive activity by the MD spleen cells requires viable cells and is contact mediated. The suppressive activity is attributed to the presence in MD spleens of a population of suppressor cells with characteristics typical of macrophages. The suppressor cell activity was not removable by complement-mediated lysis using anti-T or anti-B sera, but it was reversible by treatment with carrageenan or carbonyl iron magnet, by passage through a Sephadex G-10 column, and by adherence to plastic petri dishes or glass beads. MD spleen cells depleted of the suppressor cell population remained unable to respond to Con A.     

A study of the mechanism of Con A-induced immunosuppression in vivo

The plaque-forming cell (PFC) response to sheep erythrocytes (SRBC) is suppressed in a dose-related manner when concanavalin A (Con A) is administered intravenously to mice prior to or after immunization with antigen. The magnitude of suppression as well as the duration of the Con A effect greatly depends on the concentration of antigen used for immunization. Although profound suppression of the anti-SRBC PFC response is observed in intact mice pretreated with Con A for 4-24 hr, spleen cells from these mice do not exhibit suppressive activity when transferred into normal recipients or when cotransferred with normal spleen cells into irradiated recipients. Moreover, the cells from Con A-treated mice respond as normal spleen cells to SRBC when transferred alone into irradiated hosts. Suppression of the anti-SRBC PFC is only observed when adoptive hosts of cells from Con A-treated mice are also injected with Con A within 48 hr (but not 72 hr) of cell transfer and immunization. This time course of responsiveness to the suppressive effects of Con A is similar to that observed in normal mice and in irradiated recipients of normal spleen cells. The immune response to SRBC is also suppressed in adoptive hosts of normal spleen cells that are pretreated with Con A 4-24 hr prior to irradiation and cell transfer. Although functionally inactive when transferred into adoptive hosts, spleen cells from mice pretreated with Con A for 4-24 hr can suppress a primary antibody response to SRBC in vitro. The suppressive activity, which cannot be detected in the spleens of mice when the interval between pretreatment and assay is longer than 24 hr, is present in a subpopulation that bears the Thy 1.2 and Lyt 2 phenotype. Taken together the results obtained in in vivo and in vitro functional assays suggest that a suppressor cell population is activated following in vivo treatment with Con A, but that the cells rapidly lose their state of activation when removed from a Con A environment. This phenomenon is in all probability responsible for the failure to demonstrate suppressive activity in the spleens of Con A-treated mice using in vivo functional assays.