Suppressor cells in the spleens of tumor-bearing mice: enrichment by centrifugation on hypaque-ficoll and characterization of the suppressor population.

Spleen cells from mice bearing methylcholanthrene-induced sarcomas or a mammary adenocarcinoma suppressed the mitogen responses of normal spleen and lymph node cells. Lymph node cells from tumor bearers had no suppressive effects. Centrifugation of spleen cells layered on Hypaque-Ficoll (specific gravity of 1.08) produced a dense fraction which pelleted and a light fraction which was retained at the Hypaque-Ficoll-medium interface. Suppressive activity was not found in either fraction of normal spleen cells. In tumor-bearer spleen cells suppressor activity was greatly enriched in the light fraction. Treatment of the suppressor fraction with anti-theta or anti-Ig serum and complement did not remove suppressor activity. However, the suppressor cells were removed by passage through nylon wool or by carbonyl iron treatment. Also, the population which adhered to plastic Petri dishes contained the suppressor cell activity.     

Enterically induced immunologic tolerance. I. Induction of suppressor T lymphoyctes by intragastric administration of soluble proteins.

Specific immune unresponsiveness was induced in inbred mice (BDF1) by the administration of soluble ovalbumin (OVA) by gastric intubation. Anti-hapten (DNP) responses likewise were specifically diminished when animals were fed autologous carrier (OVA or keyhole limpet hemocyanin). Adoptive transfer of spleen cells demonstrated that the tolerant state could be maintained in irradiated recipient mice, and specific anergy could be transferred to normal recipient animals. Adoptive suppression was mediated by T lymphocytes, as demonstrated by nylon wool fractionation and susceptibility of the cells to anti-Thy 1.2 and complement. Transferred B cells had neither suppressive nor augmentative effects. Enteric administration of OVA also specifically diminished antigen-induced DNA synthesis of primed lymph node T cells, although suppressor cells were not identified in the lymph nodes per se.     

Suppression of antitumor immunity by macrophages in spleens of mice bearing a large MOPC-315 tumor

We had shown previously that progression of MOPC-315 plasmacytoma growth is associated with an increase in the percentage of macrophages in the spleen as well as a decrease in the ability of tumor-bearer spleen cells to mount an antitumor cytotoxic response upon in vitro immunization. Here we provide evidence that macrophages in the MOPC-315 tumor-bearer spleen are responsible at least in part for the suppression of the generation of antitumor cytotoxicity. Accordingly, removal of most macrophages by depletion of phagocytic cells or Sephadex G-10-adherent cells from spleens of mice bearing a large tumor resulted in augmented antitumor immune potential. Also, Sephadex G-10-adherent spleen cells from tumor-bearing (but not normal) mice drastically suppressed the in vitro generation of antitumor cytotoxicity by normal spleen cells. The suppressive activity of these adherent cells did not reside in contaminating suppressor T cells, since it was not reduced by treatment with monoclonal anti-Thy 1.2 antibody plus complement. The Sephadex G-10-adherent cell population from the tumor-bearer spleen suppressed the in vitro generation of antitumor cytotoxicity against autochthonous tumor cells but not against allogeneic EL4 tumor cells, and hence the suppression was apparently specific. The suppressive activity of the Sephadex G-10-adherent cell population from tumor-bearer spleens was overcome by treatment of the tumor-bearing mice with a low curative dose of cyclophosphamide. This immunomodulatory effect of a low dose of the drug in overcoming the suppression mediated by the Sephadex G-10-adherent cell population enables the effector arm of the immune system of tumor-bearing mice to cooperate effectively with the drug's tumoricidal activity in tumor eradication.     

Reversal of macrophage-augmented MLR reactivity by tumor-induced splenic suppressor T cells and their soluble factor(s)

Augmenting concentrations of macrophages or their supernatants failed to reverse T-cell hyporeactivity in tumor-bearing mice (TBM). Serial passaging over nylon wool columns depleted TBM spleen cells of a mildly adherent tumor-induced suppressor cell and restored mixed lymphocyte reaction (MLR) reactivity to the purified TBM T-cell population. The tumor-induced suppressor cell was extensively plated to remove macrophages and characterized as a T cell by its anti-Thy 1 serum sensitivity. This suppressor T cell, when added to normal T cells, abrogated all enhancing effects caused by addition of macrophages. Suppressor T-cell inhibition was non-contact dependent, since suppressor T-cell supernatants inhibited MLR activity in T cells treated with enhancing concentrations of macrophage supernatants. Thus it appears that tumor-induced T-cell debilitation is a reversible phenomenon, mediated not by macrophages but by soluble factor(s) from a nonphagocytic, mildly adherent, suppressor T cell.     

Adjuvant-induced nonspecific supressor cells: in vitro and in vivo studies

The in vitro mitogen responses of spleen cells from mice injected ip with the nonantigenic adjuvant, Al(OH)₃, are markedly depressed. This depressed reactivity was found to be mediated by a population of nylon wool adherent, Fc-receptor-bearing suppressor cells. Suppressor cells were detected only in the spleens of the adjuvant-treated mice, as the response of lymph node cells to mitogenic stimulation in vitro was found comparable to that of normal controls. Moreover, elevated levels of suppressive activity could be detected in sera of Al(OH)₃-treated mice during the first week after adjuvant administration, which, however, did not correlate with either the long-lasting presence of suppressor cells or the in vivo normal immune response of the adjuvant-treated animals. Studies designed to test the effect of suppressor cells on the generation of splenic PFC in vivo revealed that both the direct and indirect PFC responses against SRBC inoculated iv were enhanced rather than suppressed, as compared to those of the normal controls. Furthermore, the level of cytotoxic lymphocytes generated in spleens of Al(OH)₃-treated mice immunized with allogeneic tumor cells was equal to or higher than that of the normal controls. In view of the present results, we feel that the concept that splenic, nonspecific suppressor cells (macrophages) are immunosuppressive in vivo as well as the in vivo relevance of in vitro findings should be carefully reevaluated.     

Suppression of antitumor immunity by macrophages in spleens of mice bearing a large MOPC-315 tumor

Summary We had shown previously that progression of MOPC-315 plasmacytoma growth is associated with an increase in the percentage of macrophages in the spleen as well as a decrease in the ability of tumor-bearer spleen cells to mount an antitumor cytotoxic response upon in vitro immunization. Here we provide evidence that macrophages in the MOPC-315 tumor-bearer spleen are responsible at least in part for the suppression of the generation of antitumor cytotoxicity. Accordingly, removal of most macrophages by depletion of phagocytic cells or Sephadex G-10-adherent cells from spleens of mice bearing a large tumor resulted in augmented antitumor immune potential. Also, Sephadex G-10-adherent spleen cells from tumor-bearing (but not normal) mice drastically suppressed the in vitro generation of antitumor cytotoxicity by normal spleen cells. The suppressive activity of these adherent cells did not reside in contaminating suppressor T cells, since it was not reduced by treatment with monoclonal anti-Thy 1.2 antibody plus complement. The Sephadex G-10-adherent cell population from the tumor-bearer spleen suppressed the in vitro generation of antitumor cytotoxicity against autochthonous tumor cells but not against allogeneic EL4 tumor cells, and hence the suppression was apparently specific. The suppressive activity of the Sephadex G-10-adherent cell population from tumor-bearer spleens was overcome by treatment of the tumor-bearing mice with a low curative dose of cyclophosphamide. This immunomodulatory effect of a low dose of the drug in overcoming the suppression mediated by the Sephadex G-10-adherent cell population enables the effector arm of the immune system of tumor-bearing mice to cooperate effectively with the drug's tumoricidal activity in tumor eradication.This paper was presented in part at the annual meeting of the American Association of Immunologists, Chicago, Illinois, 10–15 April 1983     

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.     

Fractionation of lymphocyte subpopulations which regulate mixed lymphocyte reactions.

Four days after injection of allogeneic lymphocytes BALB/c splenic T cells suppress proliferation of syngeneic cells in mixed lymphocyte reactions (MLR). Conversely, lymph node cells from the same mice amplify MLR responses. To further characterize these functional subpopulations, alloantigen-primed lymphocyte suspensions from both organs were fractionated by velocity sedimentation at unit-gravity. After fractionation MLR suppressor cells from spleens localized exclusively in rapidlly sedimenting fractions of large cells. MLR suppressor activity of cells from these fractions, as well as that of unfractionated spleen cell suspensions, was abolished by treatment with anti-Thy-1.2 serum and complement. Spleen cell fractions of similar sedimentation velocity also secreted a soluble MLR suppressor into culture supernatants. Although inhibitory of MLR, spleen cells of rapid sedimentation velocity did not suppress responses to T cell mitogens. In marked contrast with the effects of spleen cells, large 4-day-alloantigen-primed lymph node cells had no suppressive activity in MLR. MLR amplifier cells of uncertain derivation were found in fractions of medium sedimentation velocity from both spleens and lymph nodes. Fractionation of alloantigen-primed lymph node cell suspensions did reveal, however, a subpopulation of small cells with MLR suppressor acitivty which was unaffected by treatment with anti-Thy-1 serum and complement. The data thus indicate that large alloantigen-activated lymphocytes are not intrinsically suppressive nor are cells which suppress MLR necessarily large. We consequently conclude that regulation of MLR responses by alloantigen-primed lymphocytes involves a complex interaction between distinct functional subpopulations of cells which are separable both by physical and biologic properties.     

Activation of suppressor T cells by low-molecular-weight factors secreted by spleen cells from tumor-bearing mice

The spleens of mice bearing large M-1 fibrosarcomas have been shown to contain several populations of cells which nonspecifically suppress antibody synthesis by cocultured normal spleen cells. It has now been shown that the spleens of tumor-bearing mice also contain inducer cells which secrete soluble factors capable of activating suppressor T cells from unprimed precursor cells. The activated suppressor cells are Thy 1+, Lyt 1+2+ and secrete a soluble suppressive factor. They inhibit the in vitro generation of antibody-forming cells by cocultured normal spleen cells stimulated by T-cell-dependent antigens. They do not, however, suppress the antibody response to T-cell-independent antigens and do not inhibit antibody synthesis by cocultured nude mouse spleen cells cultured with T-cell-dependent antigens and exogenous helper factors. In addition, suppression is blocked if conditioned medium containing T-cell growth factors is added to the suppressor cell assays. These data suggest that cells in the spleens of tumor-bearing mice secrete inducing factors which activate suppressor cells. These activated suppressor cells in turn secrete soluble suppressor factors which inhibit antibody synthesis, possibly by interfering with the synthesis or release of T-cell growth factors.     

Immunosuppression induced by attenuated Salmonella. Evidence for mediation by macrophage precursors

An attenuated aro A- strain of Salmonella typhimurium, SL3235, was previously shown to afford excellent protection in C3HeB/FeJ mice against challenge by virulent Salmonella and cross-protection against Listeria monocytogenes. In the present study, the immunologic status of immunized mice was evaluated by studying their ability to mount in vivo and in vitro plaque-forming cell responses to SRBC. SL3235-immunized mice exhibited marked suppression in their ability to generate anti-SRBC plaque-forming cell responses. Suppression was active and mediated by soluble factors as demonstrated by the capacity of immune cells to suppress the responses of normal cells in co-culture and across a membrane in transwell plates. Depletion of T cells from immune spleens did not alleviate the suppressive activity of the remaining cells. Depletion of splenic adherent cells resulted in partial alleviation of suppressive activity, demonstrating that mature macrophages are partly responsible for mediating the observed suppression. A sequential multi-step depletion procedure resulted in marked enrichment of a second suppressor cell population that was Mac1+, Thy-1.2-, surface Ig-, J11d-, non-adherent, non-phagocytic, and esterase negative. When cultured in vitro in the presence of L cell-conditioned medium, but not in the presence of Con A supernatant, these cells matured into typical macrophages within 72 h of culture. The cell population enriched for macrophage precursors (75%) retained the suppressive capacity of unfractionated splenocytes. Our data indicate that, in addition to the well-described involvement of mature macrophages in suppressing immune responses, bacterial infection may induce appearance of macrophage precursors that may also play an important regulatory role in the immune system.     

Adherent cell function in murine T lymphocyte antigen recognition. I. A. macrophage-dependent T cell proliferation assay in the mouse.

The data in this report describe a T cell proliferation assay with nylon wool column-purified murine lymph node lymphocyte from animals immunized by footpad injection of antigen in CFA. It was found that the in vitro immune response of sensitized T cells to soluble protein antigens was functionally dependent on the presence of adherent cells, more specifically macrophages, at all concentrations of in vitro antigen challenge. The response was due to T cells in that cytotoxic treatment of the immune lymphocyte cells with anti-Thy 1.2 serum and complement effectively eliminated the antigen-specific DNA synthetic responses. The antigen-specific proliferation of murine lymphocytes depleted of adhereent cells could not be reconstituted with either guinea pig macrophages nor murine fibroblasts, indicating the existence of species and cell type specificity. In contrast to previous observations in the guinea pig, soluble products of cultured adherent cells could at least partially replace the function of intact macrophages in the response to antigen.     

Effect of murine tumors upon delayed hypersensitivity to dinitrochlorobenzene. III. In vivo activity of the nonspecific suppressor cell

Tumor-induced immunosuppression was investigated in an in vivo model of delayed hypersensitivity (DH) to the chemical sensitizer, dinitrochlorobenzene (DNCB). DH to DNCB as measured in a footpad assay was decreased in C3H/HeJ mice bearing MCA-F, a 3-methylcholanthrene-induced syngeneic fibrosarcoma. Suppressor cells from the spleens of tumor-bearing mice inhibited the induction of DH to DNCB in otherwise normal syngeneic C3H/HeJ recipients. Ten million spleen cells (SpC) harvested from mice bearing MCA-F for 10 days and adoptively transferred to tumor-free mice at the time of sensitization with DNCB suppressed the response to the sensitizer. The suppressor cells were macrophages, since they were adherent to plastic, removed by treatment with a magnet after phagocytosis of carbonyl iron, resistant to exposure to gamma radiation and to treatment with anti-Thy 1.2 serum and complement. Further, the nonspecific suppressor cells were activated by progressive tumor growth rather than by induction of tumor-specific immunity using irradiated tumor cells. Titration studies revealed that suppression of DH occurred with the transfer of as few as 10(6) SpC. Thus, nonspecific suppressor cells are effective at inhibiting in vivo DH to DNCB and suggest that nonspecific suppression in the intact host occurs through mechanisms different from those involved in suppression in vitro.     

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.     

Cellular basis of tolerance to serum albumin in adult mice. I. characterization of T suppressor and T helper cells.

The surface markers and size of suppressor cells were determined in adult (BALB/c x C57BL/Ka)F1 mice which were tolerized with a single injection of deaggregated bovine serum albumin (BSA). Suppressor cells from the spleens of tolerazided donors were assayed in a cell transfer system in which graded numbers of cells were injected into irradiated syngeneic mice along with limiting numbers of T cells primed to BSA and an excess of B cells primed to DNP-BSA. Adoptive hosts were challenged with DNP-BSA in saline, and the anti-DNP response was measured. Suppressor cells were antigen specific as shown by the inhibitory activity of BSA-tolerant spleen cells on the response to DNP-BSA, but not to DNP-BGG. Suppressor cells were eliminated by in vitro treatment with anti-Thy 1.2, anti-Ly-2.2, anti-I-J subregion antisera and C, but not with anti-Ly-1 or anti-I-A subregion antisera. Neither unprimed nor primed helper T cells were detected in the spleen of tolerized donors after in vitro treatment with anti-Ly-2.2 antisera. Both helper and suppressor T cells from the spleens of primed or tolerized donors, respectively, showed a rapid sedimentation velocity (S greater than 3.7 mm/hr).     

Cyclophosphamide and abrogation of tumor-induced suppressor T cell activity

Summary Previously we have demonstrated that the in vitro generation of P815-specific anti-tumor cytotoxic T lymphocytes (CTL) was suppressed by splenic suppressor T cells from late tumor-bearing hosts (TBH). Suppression is not caused by in vitro growth of P815 from splenic metastases, since suppression was also seen with spleen cells from late TBH mice bearing a hypoxanthine/aminopterin/thymidine-sensitive subline (PHS-5) of P815 in the presence of HAT. Cyclophosphamide has been shown to inhibit theinduction of suppressor cells selectively in a number of immune responses, but evidence that it can inhibit active tumor-induced suppressor T cells is limited. We have found that suppressor T cells already induced by P815 in syngeneic late TBH are sensitive to low doses of cyclophosphamide (50 mg/kg) given 1 day before spleen harvest, but the in vitro CTL response of late TBH spleen cells could not be restored by pretreating the mice with cyclophosphamide, even when exogenous interleukin-2 was added to the cultures. Although 50 mg/kg cyclophosphamide did not inhibit the CTL response of spleen cells from mice immunized with P815 +Corynebacterium parvum, the same dose of cyclophosphamide eliminated the CTL response of spleen cells from early TBH. Interleukin-2 (IL-2) did not overcome this effect of cyclophosphamide, suggesting a direct effect on CTL. Ultra-low-dose cyclophosphamide (10 mg/kg) did not adversely effect early TBH CTL but was still able to eliminate suppressor T cell activity from late TBH. Nevertheless, late TBH CTL remained unresponsive after pretreatment of mice with ultra-low-dose cyclophosphamide, even when exogenous IL-2 was added in vitro. CTL precursor frequency analyses demonstrated that cyclophosphamide pretreatment had little or no effect on the numbers of CTL precursors from early TBH. Late TBH CTL precursor cells were not detectable in these studies, with or without suppressor T cell inhibition by cyclophosphamide pretreatment. Thus, it appears that most CTL precursor cells may be lost or irretrievably inactivated in the spleens of late TBH mice.This work was supported by grants CA42443, CA48075 and T32-CA09210 from the National Cancer Institute, Department of Health and Human Services, and an American Cancer Society Clinical Oncology Career Development Award (H. D. Bear)     

Suppression of the humoral immune response by plasmacytomas: mediation by adherent mononuclear cells.

Mice bearing plasmacytomas have a severely impaired ability to mount a primary immune response; T cells from these mice, however, appear by both in vivo and in vitro criteria to function normally. This unusual pattern of immunodeficiency appears to be mediated by a regulatory cell found in the spleens and peritoneal cavities but not in the lymph nodes or thymuses of mice bearing plasmacytomas. The number of cells with suppressor activity in the spleens of plasmacytoma-bearing mice is directly proportional to the size of the subcutaneous tumor borne by the host. These cells are capable of suppressing antibody production in in vitro cultures of normal spleen cells but have no demonstrable effect on the ability of normal spleen cells to proliferate in vitro in response to phytohemagglutinin or 8-Br-guanosine-3', 5'-monophosphate (T and B cell mitogens, respectively). Characterization of the suppressor cell population on the basis of its cell surface properties, its radioresistance, its morphology, and its ability to adhere to various solid matrices suggest that these cells are adherent mononuclear cells. These data support the concept that plasma cell tumors indirectly induce an impairment in the humoral immune response of their hosts by stimulating the expression of regulatory functions in a population of splenic and peritoneal macrophages.     

Adherent cells (macrophages?) in tumor-bearing mice suppress MLC responses

The spleens of mice bearing transplanted methylcholanthrene (MCA)-induced fibrosarcomas (MCA-1425 and MCA-1460) were shown to contain cells capable of suppressing the generation of cytolytic T lymphocytes (CTL) in mixed leukocyte cultures (MLC). The suppressive activity was first detected 21 days after tumor transplantation. No suppression was seen with lymph node cells taken at the same time as the spleen cells. The cells responsible for the suppressive activity were adherent to nylon wool and plastic dishes and they were not lysed by anti-T-cell serum plus complement. The suppressor cells were phagocytic and were resistant to irradiation (3000 rads) in vitro. Spleen cells from tumor-bearing nude mice were as suppressive as were spleen cells from tumor-bearing conventional mice. We conclude from these findings that T cells were not involved either as inducers or as effectors of the suppression observed, although the responsible adherent cells may have exerted their effect by interacting with a T-suppressor cell population in the MLC mixtures. While spleen cells of tumor-bearing mice were suppressive when added at any time during the first 4 days of a 5-day MLC, they showed no effect on the cytotoxicity of fully differentiated CTL. Indomethacin reversed suppression, suggesting that prostaglandins may have been involved.     

Graft-vs-host reactions (GVHR) across minor murine histocompatibility barriers. II. Development of natural suppressor cell activity

We explored the immunoincompetence of mice undergoing a chronic graft-vs-host reaction (GVHR) across minor histocompatibility barriers. BALB/c and B10.D2 mice are H-2d and mls b, and differ only with regard to minor histocompatibility antigens (MiHA). A large number of BALB/c mice were unirradiated or were irradiated with 300, 600, or 900 R. They then were injected with 5 X 10(7) spleen cells from either allogeneic B10.D2 or syngeneic BALB/c mice. The spleen cells from these recipient mice were assayed at various times post-irradiation/injection for their proliferative response to Con A and LPS, their ability to suppress the mitogen responses of normal spleen cells, and for the genetic specificity of this suppression. Spleen cells from BALB/c mice that had received 600 or 900 R (but not 0 or 300 R), and allogeneic B10.D2 lymphocytes, became very hyporesponsive to mitogens and became suppressive in vitro by days 7 to 10 post-irradiation/injection. These phenomena persisted for the entire 49 days of the experiment. After an initial period of splenomegaly, the spleens of these mice gradually became depleted of viable lymphocytes. Initial characterization of suppressor cells found in the spleens of GVH mice showed that they were not removed by treatment with anti-Thy-1.2 plus complement. GVH suppressors also were not adherent to plates coated with antiserum directed towards murine Ig. In addition, these cells did not adhere to plastic plates. Thus, we believe that the suppressor cells found in mice undergoing GVHD across MiHA are not mature T cells, B cells, or macrophages, but belong to a class of suppressor cells termed natural suppressor (NS). Genetic analysis of NS cell activity showed that as early as 10 days post-irradiation/injection, NS cells inhibited mitogen responses of all mouse strains tested, the exception being the relative difficulty in suppressing the LPS response of B10.D2 (syngeneic with donor cells). By day 42, this had developed into an almost complete inability to suppress a B10.D2 LPS response, although at this time NS cells were still capable of inhibiting all the other mitogen responses of all strains tested, including the Con A response of B10.D2 spleen cells. Moderate amounts of mitogen unresponsiveness and suppressor activity were seen in the syngeneic groups (BALB/c----BALB/c) but only if recipients received 600 or 900 R. This was a transient phenomenon that was maximal at day 14, and which we believe to be a similar but less severe degree of immunoincompetence when compared with that seen with allogeneic stimulation in the B10.D2----BALB/c GVH model.(ABSTRACT TRUNCATED AT 400 WORDS)     

Tumor modulation of autoreactivity: decreased macrophage and autoreactive T cell interactions

The autologous mixed lymphocyte reaction (AMLR) is an in vitro measure of autoreactivity, a key mechanism in immune homeostasis. In this system, macrophages (M phi) act as accessory cells to autoreactive L3T4+ T cells by presenting self-Ia and releasing soluble modulators. During tumor growth, changes occur in M phi and T cells. Tumor-bearing host (TBH) M phi have a reduced ability to act as accessory cells. In fact, TBH M phi suppressed autoreactivity by 60-70%. The decrease in TBH M phi or T-cell abilities was not due to differences in cell numbers or incubation time. Because tumor growth causes increased prostaglandin E2 (PGE2) production by M phi, indomethacin was used to assess the contribution of prostaglandins. Normal and TBH T-cell reactivity increased nearly 50% when stimulated by normal host M phi, while normal and TBH T-cell reactivity increased nearly 100% when stimulated by TBH M phi. Thus increased prostaglandin production is partly responsible for the increased TBH suppressor M phi activity and in the normal host, suppressor M phi may be responsible for maintaining immune regulation. To assess the direct role of prostaglandins in T-cell hyporesponsiveness, PGE2 was titrated into the cultures. PGE2 suppressed normal and TBH T-cell responsiveness in a dose-dependent manner. Normal host T cells were suppressed to a greater extent than TBH T cells by PGE2 (66% versus 42% suppression, respectively). Reduced Ia expression and active suppressor mechanisms are not the only mechanisms mediating hypoautoreactivity during tumor growth. TBH autoreactive L3T4+ T cells were less responsive to self-Ia; they were only 60-80% as reactive as their normal counterparts. To address whether the helper T (TH)-cell defect involved cytokines, T cells were treated with interleukin (IL)-1, IL-2, and IL-4. In all cases, the TBH T-cell response to the factors was decreased (only 60-75% as reactive as normal T cells). Because TBH M phi-mediated suppression can override the addition of IL-1, IL-2, and IL-4, indomethacin was also added with the exogenous interleukins. This coaddition significantly enhanced normal host autoreactivity above control levels while TBH autoreactivity (the combination of TBH T cells and TBH M phi) only returned to normal host unstimulated levels. Tumor growth modulates the immune response at least by (i) decreasing the accessory cell abilities of TBH M phi through decreased Ia expression and increased production of suppressive molecules such as prostaglandins; and (ii) decreasing the responsiveness to immune enhancing factors by TH cells.     

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.