Suppression Of Antibody Production Research Articles

Cyclosporin A prevents the anti-murine antibody response to a monoclonal anti-tumour antibody in rabbits.

Repeated therapy of cancer with mouse monoclonal antibodies frequently produces antibodies directed against the administered antibody. We have investigated the ability of cyclosporin A (CsA) and the use of ultracentrifuged antibody to prevent the formation of anti-antibodies in rabbits. Courses of CsA, 20 mgkg-1 day-1, were given intramuscularly for 6 days to rabbits and a mouse monoclonal anti-human chorionic gonadotrophin antibody was given intravenously on day 2 with or without ultracentrifugation to remove microaggregates. The whole course was repeated after an interval of two weeks. Rabbit anti-mouse antibody production was prevented in all 8 animals that were given CsA and ultracentrifuged preparation (non UC-W14). Anti-mouse antibody was produced by all the animals that were not given CsA. A further dose of mouse antibody eight weeks after CsA demonstrated that immunological recovery had occurred in all four animals re-challenged with non UC-W14 but only 3/7 animals given an UC-W14 preparation. This suggests that CsA and ultracentrifugation facilitate the induction of immunological tolerance. The complete suppression of antibody production which could be achieved justifies a clinical trial of CsA and ultracentrifugation of antibody.

Specificity of the T cell immune response to acetylcholine receptor in experimental autoimmune myasthenia gravis. Response to subunits and synthetic peptides.

Myasthenia gravis (MG) and its animal model, experimental autoimmune MG (EAMG), are T cell-dependent diseases mediated by antibodies against acetylcholine receptor (AChR) on skeletal muscle. Most of the antibodies are directed toward conformation-dependent epitopes on the AChR, whereas T cells recognize denatured AChR. In search of T cell epitopes in EAMG, we tested 24 synthetic peptides covering 62% of the alpha-subunit sequence of Torpedo californica electric organ AChR in the T cell proliferation assay with lymph node cells from rats immunized with AChR. In Lewis rats, 2 of these peptides, [Tyr 100]alpha 100-116 and [Gly 89, Tyr 90]alpha 73-90, strongly stimulated T cells and, of these, [Tyr 100]alpha 100-116 was much more potent; 4 other peptides were weakly mitogenic and 18 were ineffective. None of the 24 synthetic peptides alone stimulated anti-AChR production and, when added to cultures along with AChR, [Tyr 100]alpha 100-116 and [Gly 89, Tyr 90]alpha 73-90 suppressed antibody production. Of twelve cloned T cell lines specific to AChR, 4 responded to [Tyr 100]alpha 100-116, indicating the importance of the epitope in alpha 101-116 in Lewis rats. In three other strains of rats whose responses to AChR and its subunits were similar to those in the Lewis rat, neither [Tyr 100]alpha 100-116 nor [Gly 89, Tyr 90]alpha 73-90 was stimulatory. Instead, completely different sets of peptides stimulated their T cells. When peptides were used as immunogens, each strain (except Lewis rats) responded only to the peptides that stimulated AChR-immune T cells from the same strain. Genetically restricted T cell recognition of AChR peptides in rats suggests that T cells from MG patients with different major histocompatibility haplotypes may recognize different AChR peptides.

Regulation of antibody production by hybridoma cultures: I. Anti-idiotype antibody-mediated down-regulation of anti-DNA antibody production by hybridoma cells

A series of anti-DNA antibody-producing hybridomas were obtained by fusing spleen cells from 6-month-old MRL/ lpr, autoimmune prone, mice with P3X63-Ag8 myeloma cells. Rabbit anti-idiotype (id) antibodies specific for several of the hybridoma proteins were prepared. It was shown that the anti-id antibody inhibited immunoglobulinin secretion by the hybridoma cells in an id-specific manner. Inhibition of antibody production was not due to a cytotoxic effect, since the anti-id, in fact, stimulated proliferation of the hybridoma cells.

Antibody-mediated graft versus host reactions in renal transplantation

Graft versus host reactions (GvHR) are frequent and severe complications of bone marrow transplantation. Although the major symptoms of GvHR in these patients are thought to be mediated through T lymphocytes, both clinical and experimental observations indicate that antibody production also is stimulated by GvHR. In animal models of GvHR, Gleichmann and colleagues 1 have demonstrated a range of antibody-mediated lesions, which become more pronounced when the antigenic differences between donor and host favor stimulation of helper T lymphocytes and a chronic disease process. Cnly limited attention has been given to the possibility that 'passenger lymphocytes' in solid tissue transplants may cause GvHR. In 1953 Simonsen suggested that the plasma cells he observed in the interstitium of transplanted canine kidneys were possibly of donor origin, but he abandoned this hypothesis when radiolabelling experiments demonstrated that cells of host origin infiltrated skin and kidney allografts (reviewed in Ref. 2). Yet, sporadic cases have been reported of unexpected hemolytic episodes following the transplantation of kidney, liver or lung from a donor who is Rh negative or O blood group to a patient whose erythrocytes bear Rh, A, B0 or AB antigens 3-1°. One report suggested that irradiation of the donor kidney prevented antibody production to the recipient's erythrocytes 5, and in a few cases antibodies that were eluted from the recipients' erythrocytes have been shown to be of donor a!!otype9-11; thus substantiating the hypothesis that the hemolytic episodes reflect GvHR. Some investigators 5.6.11 have argued that this phenomenon may be more frequently encountered when cyclosporine A (CyA) is used as an immunosuppressive agent, because selective effects of CyA on T lymphocytes might prevent rejection of the donor Blymphocytes but not suppress antibody production by these lymphocytes. Several recent reports have documented the Ionyitudinal titers of circulating antibodies to recipient ~, B or Rh antigens in CyA treated renal transplant recipients. These antibodies have been detected within 1 to 3 weeks after transplantation and they can persist for months 6, even after immunosuppression is discontinued 8. Both IgG and IgM antibodies have been detected in the serum and in eluates from the recipients' erythrocytes by the indirect antiglobulin test

Steroid-sensitive mechanism of soluble immune response suppressor production in steroid-responsive nephrotic syndrome.

Soluble immune response suppressor (SIRS), a lymphokine that suppresses antibody production and delayed type hypersensitivity in vivo, has been detected in urine and serum from certain patients with nephrotic syndrome. In the present paper, the relationship between SIRS production and nephrotic syndrome is further characterized. A striking correlation was found between detection of SIRS and the presence of steroid-responsive nephrotic syndrome (SRNS). A potential mechanism of SIRS production in SRNS patients was identified, in that lymphocytes from patients produced SIRS without requiring activation by exogenous agents, and incubation of normal lymphocytes with serum from patients activated the cells to secrete SIRS in culture. Although SIRS disappears rapidly from urine or serum after initiation of corticosteroid therapy, hydrocortisone (10(-6)-10(-7) M) did not block secretion of SIRS by activated suppressor cells. It did, however, inhibit in vitro activation of lymphocytes to produce SIRS by concanavalin A, interferon, or SRNS patient serum. The association of suppressor cell activation with SRNS and the sensitivity of both to steroids suggest that the pathogeneses of albuminuria and SIRS production are related.

Bovine milk IgG, but not serum IgG, inhibits pokeweed mitogen-induced antibody secretion by human peripheral blood mononuclear cells.

Bovine milk IgG markedly inhibits the pokeweed mitogen (PWM)-induced secretion of immunoglobulins from human peripheral blood mononuclear cells. Heat-aggregated bovine milk IgG is even more inhibitory, demonstrating significant inhibition when levels as low as 5-9 micrograms/ml are continuously present in the in vitro 14-day culture system. However, bovine serum IgG, regardless of its state of aggregation, and control proteins have little effect on PWM-induced secretion of human IgG, IgA, and IgM. In a similar fashion, goat milk IgG, especially when aggregated, inhibits human antibody secretion whereas goat serum IgG does not. Inhibition appears to be mediated by Fc gamma receptors since F(ab')2 fragments of milk-derived bovine IgG do not inhibit PWM-induced antibody secretion. The continuous presence of bovine milk IgG is not essential since preincubation of milk IgG with PWM and human mononuclear cells for 24 hr also results in inhibition of human immunoglobulin secretion. In examining potential mechanisms of inhibition, it was found that bovine milk IgG, bovine serum IgG, and another chitin-containing protein, bovine thyroglobulin, each caused a small and equal inhibition of the binding of 125I-labeled PWM to human mononuclear cells, yet only the milk IgG inhibited antibody production. These studies raise the question of whether bovine milk IgG might modulate the human immune system in vivo.

Suppression of cytolytic T-cell activity by staphylococcal enterotoxin B-induced suppressor cells: role of interleukin 2.

We have previously shown that staphylococcal enterotoxin B (SEB) is a potent inducer of suppressor T cells which function to inhibit antibody production in vitro. In the present paper we extend these studies and show that the SEB-induced suppressor cells also inhibit the development of cytotoxic lymphocytes in mixed-lymphocyte reaction (MLR) cultures. Since further analysis also showed that the level of interleukin 2 (IL-2) in cultures of SEB-primed cells was significantly reduced, experiments were carried out to determine the role of IL-2 in the inhibition of cytotoxic cell activity. Attempts to neutralize the suppression by supplementing MLR cocultures with delectinated supernatants from concanavalin A (Con A)-stimulated rat splenocytes were not successful. In addition, MLR cocultures supplemented on Day 0 with 50 units of affinity-purified IL-2 were also suppressed. Further analysis showed that the IL-2 activity in the supplemented MLR cocultures containing suppressor cells were significantly reduced by Day 3. However, repeated supplementation of the MLR cocultures with exogenous IL-2 was successful in achieving (and maintaining) "normal" levels of IL-2. The cytotoxic cell activity in these MLR cocultures remained suppressed. These results suggest that the inhibition of cytotoxic cell activity by SEB-induced suppressor cells is independent of IL-2 levels in the MLR coculture.

Inhibition of antibody production from the plasmacytoma cell line MOPC-315 by staphylococcal enterotoxin B-induced T-suppressor cells

Our previous results have shown that Staphylococcal enterotoxin B (SEB) induces a population (s) of T cells which has the capacity to suppress the antibody response of splenocytes in vitro. In the present report we have attempted to investigate the effect of SEB-primed cells on the secretion of antibody by the plasmacytoma cell line MOPC-315. We have found that the secretion of antibody by MOPC-315 is significantly reduced in as little as 24 hr of coculture with the suppressor cells. The suppressive activity is not antigen- or isotype-specific, since the antibody secretion by both MPC-11 and HOPC1 plasmacytomas are also inhibited by the SEB-primed cells. In addition, we have found that the SEB-primed cell population which inhibits the antibody production by the MOPC-315 cell line expresses the Lyt-1 +,2 − and Thy-1 + cell surface markers. The apparent relationship between the SEB-primed suppressor cell population and the population which inhibits a conventional antibody response is discussed.

Transient immune deficiency in patients with acute Epstein-Barr virus infection

To study the effect of primary Epstein-Barr virus (EBV) infection on antigen-specific antibody production, we immunized 17 college students who had developed acute infectious mononucleosis with the T-cell dependent neoantigen bacteriophage φX174. During the early phase of infectious mononucleosis, the proportion of peripheral blood lymphocytes displaying la and T8 (CD8) phenotypes was increased and the T helper/suppressor ( T4 T8 ) ratio was decreased (<1). These abnormalities disappeared during the convalescent phase. Correlating with EBV-induced changes in T lymphocytes, we demonstrated depressed humoral immune responses to bacteriophage φX174 both in vivo and in vitro, In vitro coculture experiments indicated that the la + suppressor T cells could inhibit antibody production and isotype switch. Removal of T8 + lymphocytes from patient T cells normalized in vitro antibody synthesis. In addition, impaired B-cell function was shown to be in part responsible for deficient antibody production. These studies demonstrate that infection with EBV affects both B and T lymphocytes and causes a broad-based transient immune deficiency in patients with uncomplicated infections mononucleosis.

Suppression of polyclonal immunoglobulin production by a soluble factor produced by a human thymus hybridoma

A human thymus cell hybridoma was established using thymus cells obtained from a patient with common variable hypogammaglobulinemia and associated thymoma. This hybridoma secreted a suppressor factor for polyclonal antibody synthesis. Supernatants of this hybrid showed 40–80% suppression of both IgM and IgG synthesis by pokeweed mitogen-stimulated human peripheral blood lymphocytes. Hybridoma supernatants were suppressive for immunoglobulin production only if added within the initial 48 h of the seven-day culture period. Suppression of antibody production by the hybridoma supernatant was prevented by preabsorption with T lymphocytes. Further, the suppressor factor was shown to inhibit antibody production in reconstructed cultures containing T4+ cells and B cells, yet the suppression could be abrogated by increasing the number of T4+ cells. The hybrid supernatant had no affect on the proliferation of human mononuclear cells in response to pokeweed mitogen, lipopolysaccharide, concanavalin A or alloantigen but inhibited phytohemagglutinin-induced proliferation. The target cell population for the inhibition of phytohemagglutinin responsiveness was shown to be a T4+ lymphocyte (helper inducer T cell). These results suggest that thymus hybridoma cells can produce immunoregulatory products that act through the modulation of T4+ lymphocyte function. To our knowledge this is the first human thymus cell hybridoma to be reported. Studies on such cell lines may provide important information on immunoregulatory thymic factors.

Paradoxical effects of cadmium exposure on antibacterial antibody responses in two fish species: Inhibition in cunners ( Tautogolabrus adspersus) and enhancement in striped bass ( Morone saxatilis)

Previous work in a marine fish, the cunner ( Tautogolabrus adspersus), showed that endocytosis of bacteria by cells in the liver and spleen was affected by 96-h exposure of the fish to cadmium at a concentration of 12 μg/mρ; however, antibody response to sheep erythrocytes was not affected. Since the latter finding was questionable because of short immunization times, and data from more than a single fish species were desirable, both the cunner and an anadromous fish, the striped bass ( Morone saxatilis), were examined for antibody responses against the bacterium Bacillus cereus in Freund's complete adjuvant during a 6- to 8-wk time period. Exposure to 12 μg/mρ cadmium caused significant inhibition of serum antibody titers (P<0.007) in cunners. Paradoxically, antibody response in striped bass exposed to 10 μg/mρ cadmium was enhanced sixfold. This enhancement was weaker but still evident when the antigen was injected 48 d after cadmium exposure. Peritoneal exudate cells from cadmium-exposed striped bass also showed more active migration through microporous filters than cells from non-exposed fish. Since the 96-h cadmium LC 50 was 26 μg/mρ for cunners and 20 μg/mρ for striped bass, the differences in antibody response could not be explained on the basis of differences in cadmium toxicity. Although geometric mean liver cadmium levels after exposure at 15°C were higher in cunners (163.8 ± 1.2μg/gm) than in striped bass (64.4 ± 1.2μg/g), cunners exposed to cadmium at 2°C had lower cadmium levels (46.4 ± 1.2μg/g) which were still effective in inhibiting antibody production at 8°C. On the other hand, striped bass not exposed to cadmium showed a strong, exponential rise in serum antibody when the temperature at immunization, 14°C, was reduced to 9°C; whereas cunners held in the same tanks exhibited a weaker, biphasic serum antibody response. Regardless of the cause (innate differences in cellular response, or better stress adaptability in an anadromous fish), the data show that chemical-stress effects on the immune system of one fish species cannot be extrapolated to another species.

Immunization and culture of rainbow trout organ sections in vitro

Splenic and anterior kidney sections or whole organs were excised from large (1 kg) or small (200 g) rainbow trout ( Salmo gairdneri) and placed in sterile 60 mm plastic plates containing 10 ml of Eagle's minimal essential medium (EMEM) supplemented with normal or fetal calf serum for in vitro culture. The organ samples were immunized in vitro by direct injection or by mixing in the medium Yersinia ruckeri O-antigen or dinitrophenyl-Ficoll. The medium was changed once during the 10-day incubation at 15 C. The passive hemolytic plaque assay demonstrated antibody production from the plaque-forming cells (PFC); passive hemagglutination was used to measure antibody titers in the media. High numbers of PFC occurred in cultures of either kidney or spleen, demonstrating that these organs can function independently for antibody production. Splenic sections from large fish produced more PFC than comparable whole organs from small fish. EMEM supplemented with 2% normal calf serum was a satisfactory culture medium. 2-hydroxyethyl-mercaptan an ingredient used in mammalian cell culture, inhibited antibody production in trout cells. These techniques are being used in the culture of organs and cells to elucidate pathways and sequences of antigen uptake and delivery of the immunopoietic tissues in trout.

Effect of cysteine ethylester hydrochloride (Cystanin) on host defense mechanisms (II): Restorative effects on the suppression of antibody production

The production of hemolytic plaque forming cells (HPFC) in the spleen of BALB/c mice immunized with sheep red blood cells was significantly inhibited by carrageenan treatment (0.3 mg/kg, i.p.; on days -3 and -1). Cysteine ethylester hydrochloride (ethylcysteine) restored the inhibition of the HPFC production by carrageenan treatment in a dose-dependent manner (10-100 mg/kg, p.o.). Ia positive cells (antigen-presenting cells) increased in the spleen adherent cells (SAC) obtained from immunized mice, whereas they decreased in the SAC obtained from carrageenan-treated mice. An increase of Ia positive cells occurred in the SAC of carrageenan-treated mice given ethylcysteine. Ethylcysteine (10-100 mg/kg, p.o.; on days -2 and -1) prevented both the suppression of the HPFC production and the decrease of the number of thymus lymphocytes and peripheral leukocytes induced by cyclophosphamide treatment (30 mg/kg, i.p.; on days -1 and 0). Lyt 1.2 positive cells (helper T cells) decreased in the spleen T cells of cyclophosphamide-treated mice, but increased in the spleen T cells from cyclophosphamide-treated mice give ethylcysteine. On the other hand, Thy 1.2 negative cells (B cells) did not increase in the spleen cells of cyclophosphamide-treated mice with or without ethylcysteine. These results suggest that ethylcysteine restores the immune response in immunosuppressed mice through the functions of macrophages and/or helper T cells.

Antigen-specific suppression of human antibody responses by allogeneic T cells. II. Cell interactions involved in the generation of suppression.

Specific antibody responses to influenza virus were obtained in vitro from human blood mononuclear cells (PBMC). Antibody production in these cultures was profoundly suppressed by the addition of allogeneic T cells with the surface phenotype Leu2a+ (CD8+), Leu8-. Suppression by allogeneic T suppressor (Ts) cells required interactions only between T-depleted B (E-) cells and allogeneic Leu2a+. No evidence was obtained for T-T cell interactions, or for Ts inducer cells similar to those described for nonspecific antibody responses to pokeweed mitogen. Moreover, allogeneic E+, or allogeneic Leu2a+ cells were able to suppress specific antibody responses by E- cells when help was provided by T cell-replacing factor showing that the target of suppression was the responding E- cells, and not T helper cells. In contrast to allogeneic T cells, allogeneic E- cells did not suppress antibody production when added to cultures of unfractionated PBMC (E- + E+). That is, Ts cells activated to allogeneic E- were unable to suppress antibody production by the syngeneic E- cells present in the same culture tube. This result shows that alloactivated Ts cells were specific for the allogeneic E- target cells, and that suppression was not mediated by nonspecific allogeneic effects. Allogeneic Ts cells therefore differ from Ts cells in pokeweed mitogen responses by their specificity, and by their activation in the absence of Ts inducer cells.

Human monocyte cell line (U937) releases suppressive IgG-binding factor(s).

Fc gamma receptor-bearing U937 cells, when incubated in serum-free buffer, were found to release spontaneously a suppressor material for pokeweed mitogen-drigen IgG synthesis which could be retained on Sepharose 4B-IgG immunosorbents. Immunosorbents coupled with IgM or F (ab')2 fragments of IgG were unable to retain the inhibitory activity of U937-derived material, suggesting a binding specificity for the Fc gamma fragment of IgG. This suppressor material corresponds therefore by definition to an IgG-binding factor (IgG-BF). The mechanism for in vitro suppression of the antibody response by U937-derived IgG-BF was investigated. It did not interfere with cell proliferation and displayed maximum effect when added at day 3 of the culture period. Tested for its effect on IgG, IgM and IgA synthesis, IgG-BF suppressed antibody production following a pattern specific for IgG. Finally gel filtration of the suppressor material gave rise to two peaks of inhibitory activity with an apparent molecular mass comprised between 30 to 40 kDa and 60 to 73 kDa, respectively.

Effects of antitumor polysaccharides fromAlbizza julibrissin on immune function

Polysaccharide fractions were prepared fromAlbizzia julibrissin by different extraction schedules. The fractions obtained were designated as PS-I and PS-II, respectively. Further purification of PS-I by Sephadex G-200 column chromatography gave two subfractions, Alju A and Alju B. Each fraction showed marked antitumor activity against sarcoma 180 solid form but not ascite form. PS-I and PS-II increased delayed hypersensitivity in normal and tumor bearing mice. PS-I treatment led to moderate restoration of the suppressed antibody production in tumor bearing mice.

Induction of suppression by a murine nonspecific suppressor-inducer cell line (M1-A5)

A murine nonspecific suppressor-inducer cell line (M1-A5) was established by the limiting dilution method from the spleen cells of a mouse bearing an advanced methylcholanthrene-induced fibrosarcoma. Indirect immunofluorescence studies demonstrated that the M1-A5 cells were Thy-1 −, sIg −, Ly-5 +, MAC −, and 45% asialo GM1 +. The M1-A5 cells were able to activate suppressor cells from unprimed, syngeneic normal spleen cells. These activated cells inhibited antibody production by cocultured syngeneic lymphoid cells. Induction of suppression by the M1-A5 cells was via the release of a suppressor-inducing factor, which was found to be protein in nature. Kinetic studies showed that when M1-A5 cells were separated from NSC by a dialysis tubing in Marbrook vessels, the M1-A5 cells required a minimum of 8 hr incubation period before suppressor cell activity could be demonstrated in precursor cells. On the other hand, induction of suppression by the suppressor-inducing factor required a minimum of 3 hr exposure of the precursor cells to the factor.

Inhibition of antibody production by 2-chloroadenosine

The ability of 2-chloroadenosine (2Cl Ado) to modulate lymphocyte function was examined in culture and in vivo . Mitogenic stimulation of B cell DNA synthesis was antagonized by 2Cl Ado while adenosine produced both stimulation and inhibitions. In culture, 2Cl Ado was found to suppress antibody production to sheep erythrocytes (SRBC) regardless of whether the nucleoside was added at the initiation of culture or 48 hours after sensitization. Inhibiting adenosine deaminase (ADA) did not affect the response to 2Cl Ado, and 1-homocysteine thiolactone was found to potentiate the inhibition suggesting formation of S-adenosylhomocysteine. Similar responses were found with adenosine provided ADA was inhibited. When 2Cl Ado was administered to mice 3–4 days after SRBC, a concentration-dependent decrease in antibody producing cells was observed. These data suggest that nucleosides can inhibit antibody production by inhibiting transmethylation reactions. 2Cl Ado appears to be an effective immunosuppressant without concomitant cytotoxicity both in culture and in vivo .

Kinetics of immune depression in burned rats: the sixth day phenomenon.

The data generated in a rat burn model supports the following conclusions: (1) There is a period of normal immune responsiveness surrounding day 6, in spite of peaks of immunosuppression before and after that time. (2) The spleen in burned animals is hypertrophic. This is due to its role as a filter (necrotic particles seen in macrophagesl and as a “germ reservoir” (spleen culture studies). Impairment of immune function in spleen cells is important since before day 6 there is an inhibition of antibody production with no alteration in lymphocyte transformation; after day 6, lymphocyte transformation and antibody production are both impaired. (3) Most important to the clinical treatment of burns, all of these phenomena are diminished or abolished by early removal of the burned tissue. The profile of immune depression in burned rats therefore seems to show two stages: before and after day 6. During the first, stress and prostaglandins from burned skin are probably the cause of immune depression; during the second, burn toxins and prostaglandins from the spleen cells are probably the cause.

Effects of Wy-18,251 (3-p-chlorophenyl)thiazolo[3,2-a]benzimidazole- 2-acetic acid), levamisole and indomethacin on the generation of murine T suppressor cells in vitro.

In vitro culture of normal BALB/c spleen cells with staphylococcal enterotoxin B (SEB) activates antigen non-specific suppressor T cells (Ts) which can be assayed by their ability to suppress antibody production in a plaque assay. Addition of the experimental immunomodulatory drug Wy-18,251 (10-100 microM) to cultures of spleen cells plus SEB significantly increased Ts activity relative to cultures without the drug. Similar results were obtained with levamisole, but, in contrast, indomethacin (0.1-10 microM) inhibited SEB-induced suppressor cell activity. The ability of Wy-18,251 to augment Ts activity could be therapeutically useful in the treatment of those autoimmune diseases, such as rheumatoid arthritis and systemic lupus erythematosus, in which hyperactive B cell function is a characteristic feature.