Immunosuppressive effects of glycosylation inhibiting factor on the IgE and IgG antibody response

Glycosylation inhibiting factor (GIF) was purified from culture filtrates of a T cell hybridoma, 23A4, by affinity chromatography on anti-lipomodulin Sepharose. The factor exhibited phospholipase inhibitory activity upon dephosphorylation. Immunization of BDF1 mice with aluminum hydroxide gel (alum)-absorbed dinitrophenyl derivatives of ovalbumin (DNP-OA) resulted in persistent IgE and IgG antibody formation. However, repeated injections of the affinity-purified GIF into the DNP-OA-primed mice beginning on the day of priming prevented the primary anti-hapten antibody responses of both the IgE and the IgG1 isotypes. Treatment with GIF also diminished on-going IgE antibody formation in the DNP-OA-primed mice. The treatment changed the nature of IgE-binding factors formed by BDF1 spleen cells. Incubation of spleen cells from OA + alum-primed mice with OA resulted in the formation of IgE-potentiating factor, whereas spleen cells of OA-primed, GIF-treated mice formed IgE-suppressive factor upon antigenic stimulation. It was also found that Lyt-2+ T cells in the OA-primed, GIF-treated mouse spleen cells released GIF, which had affinity for OA and bore I-Jb determinant(s). Transfer of a Lyt-1+ cell-depleted fraction of the OA-primed, GIF-treated mouse spleen cells into naive syngeneic animals resulted in suppression of the primary anti-DNP IgE antibody response of the recipients to alum-absorbed DNP-OA, but failed to affect the anti-DNP antibody response to DNP-keyhole limpet hemocyanin. The results indicate that GIF treatment during the primary response to OA facilitated the generation of antigen-specific suppressor T cells.     

Immunoregulatory role of Ig isotypes. I. Induction of contrasuppressor T cells for contact sensitivity responses by antibodies of the IgM, IgG1, and IgG3 isotypes

A profound state of specific tolerance for the contact sensitivity reaction can be produced by i.v. exposure to hapten on the surface of syngeneic macrophages. When the same haptenated cells are incubated with specific antibody to form cell-bound Ag-antibody complexes, i.v. injection induces immunity rather than tolerance. We observe that such cell-bound Ag-antibody complexes induce not only effector cells for contact sensitivity but also hapten-specific contrasuppressor T (Tcs) cells, which are capable of rendering effector cells resistant to the inhibitory effects of Ts cells. Whereas the induction of the effector cells of contact sensitivity by cell-bound complexes required I region compatibility between the injected cells and the recipient, the induction of Tcs cells showed no genetic restriction. On the other hand, induction of contrasuppression required intact Fc on the complexed antibody, inasmuch as F(ab')2 fragments of specific antibody did not induce immunity. In addition, Tcs cells could also be induced by Ag-antibody complexes on opsonized TNP-mouse RBC treated with anti-TNP antibody. Immunity induced by cell-bound Ag-antibody complexes was observed only when antibodies of the IgM, IgG3, or IgG1 isotypes are used to generate the complexes. Further studies demonstrated that the Tcs cells induced in this way displayed the phenotype of Tcs cells described in other systems (Lyt-1+,2- I-J+, Vicia villosa lectin-adherent) and released a hapten-specific contrasuppressor factor. These studies indicate that Tcs cells can be induced independently of other T cells (such as the effector cells of contact sensitivity) and are likely to be responsible for some of the immunoregulatory effects of cell-bound Ag-antibody complexes. The role of antibody isotype in the induction of Tcs cells is discussed.     

Regulatory responses in contact sensitivity: afferent suppressor T cells inhibit the activation of efferent suppressor T cells.

Two types of suppressor cells regulate the contact sensitivity (CS) response to picryl chloride (PCL). Afferent suppressor T cells (Ts-aff) inhibit the generation of CS responses to PCL, while efferent suppressor T cells (Ts-eff) inhibit the activity of Th 1 cells that mediate CS reaction. Intravenous injection of mice with TNP-substituted peritoneal exudate cells (TNP-PEC) induces Ts-eff cells that block the adoptive transfer of contact sensitivity. The induction of Ts-eff cells is prevented by the presence of Ts-aff cells, which in turn are induced by the injection of TNP-PEC coupled with antibodies of the IgG2a and IgG2b isotype (TNP-PEC-Ab). If an animal is injected with TNP-PEC prior to or simultaneously with TNP-PEC-Ab, it generates only Ts-aff cells, while if it is injected with TNP-PEC alone or TNP-PEC prior to TNP-PEC-Ab, it generates Ts-eff cells. Ts-aff cells effect only the generation of Ts-eff cells, as the addition of Ts-eff cells to assays for Ts-eff cells has no inhibitory effect on the suppressive effects of Ts-eff cells in adoptive transfer. Our experiments show that Ts-aff cells induced by TNP-PEC-Ab are phenotypically either Lyt 1+2- or Lyt 1-2+, but only the latter inhibit the generation of Ts-eff cells in vivo. The Ts-aff cells that inhibit Ts-eff activity adhere to the lectin Vicia villosa (VV), while Ts-eff cells are VV nonadherent. In addition, Ts-aff cells can prevent the generation of Ts-eff to linked haptens presented on the same PEC. It appears that a cascade of Ts cell interactions are involved in the regulation of CS responses.     

The action of interleukin-4 on antigen-specific IgG1 and IgE production by interaction of in vivo primed B cells and carrier-specific cloned Th2 cells

The production of anti-trinitrophenyl (TNP) antibodies of different isotypes from in vivo primed B cells was studied using the plaque-forming cell method. It was shown that these B cells secrete anti-trinitrophenyl antibodies of different isotypes only in the presence of Th2 cells specific for keyhole limpet hemocyanin (KLH) and the hapten-carrier conjugate TNP-KLH. Lipopolysaccharide-stimulated primed B cells without cells from the Th2 clone did not produce anti-TNP-specific IgG1 or IgE antibodies even in the presence of the hapten-carrier antigen TNP-KLH. Supernatants from these Th2 clones cultured with antigen-presenting cells and the complete antigen were unable to activate primed B cells for antibody secretion. Cognate interaction between primed B cells and carrier-specific Th2 cells is a prerequisite for hapten-specific IgG1 or IgE production. Anti-IL-4 antibody inhibited secretion of anti-hapten IgE antibody. Therefore, for production of anti-hapten antibody of the IgE isotype IL-4 is also necessary.     

Prevention of cerebral malaria by adoptive transfer of malaria-specific cultured T cells into mice infected with Plasmodium berghei

Murine T cell populations specific for Plasmodium berghei parasites were generated in vitro from BALB/c immune lymph node cells. The malaria-specific T lymphocytes were shown: a) to proliferate specifically in vitro in response to stimulation with P. berghei-infected red blood cells; b) to exhibit the Thy-1+, Lyt-1+2- cell surface phenotype; c) to provide specific helper activity for an in vitro anti-hapten (TNP) plaque-forming cell antibody response; and d) to protect P. berghei-infected mice from early mortality due to cerebral malaria.     

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

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

T cell subsets regulating antibody responses to L-glutamic acid60-L-alanine30-L-tyrosine10 (GAT) in virgin and immunized nonresponder mice

T cell subsets from virgin and immunized mice, which are Ir gene controlled nonresponders to GAT, which regulate antibody responses to GAT have been characterized. Virgin nonresponder B10.Q B cells develop GAT-specific antibody responses to GAT, B10.Q GAT-M phi, and GAT-MBSA when cultured with virgin or GAT-primed Lyt-1+, I-J-, Qa1- B10.Q helper T cells. Virgin T cells are radiosensitive, whereas immune T cells are radioresistant (750 R); qualitatively identical helper activity is obtained with T cells from mice immunized with soluble GAT, B10.Q GAT-M phi, and GAT-MBSA. Responses to GAT and GAT-M phi are not observed when virgin or GAT-primed Lyt-1+, I-J+, Qal+ T cells are added to culture of virgin or GAT-primed Lyt-1+, I-J-, Qa1- helper T cells and virgin B cells; the GAT-specific response to GAT-MBSA is intact. The Lyt-1+, I-J+, Qa1+ T cells from mice primed with GAT, GAT-M phi, and GAT-MBSA were qualitatively identical in mediating this suppression. Virgin Lyt-2+ T cells have no suppressive activity alone or with virgin Lyt-1+, I-J+, Qa1+ T cells, whereas responses to GAT, GAT-M phi, and GAT-MBSA are suppressed in cultures of GAT-primed helper T cells containing GAT-primed Lyt-2+ T cells (with or without GAT-primed Lyt-1+, I-J+, Qa1+ T cells). Suppression of responses to GAT-MBSA in cultures of GAT-M phi-primed helper T cells requires both GAT-M phi-primed Lyt-1+, I-J+, Qa1+ T cells and Lyt-2+ T cells; the Lyt-1+, I-J+, Qa1+ T cells appear to function as inducer cells in this case. In cultures containing GAT-MBSA-primed helper T cells, either GAT-MBSA-primed Lyt-1+, I-J+, Qa1+ or Lyt-2+ T cells suppress responses to GAT and GAT-M phi; under no circumstances are responses to GAT-MBSA suppressed by GAT-MBSA-primed regulatory T cells. This regulation of antibody responses to GAT by suppressor T cells is discussed in the context of the involvement of suppressor T cells in responses to antigens under Ir control, and of the evidence that nonresponsiveness to GAT is not due to a defect in the T cell repertoire, but rather is due to an imbalance in the activation of suppressor vs helper T cells.     

Insulin-specific suppressor T cell factors

Murine antibody responses to heterologous insulins are controlled by MHC-linked immune response genes. Although nonresponder mice fail to make antibody when injected with nonimmunogenic variants of insulin, we have recently shown that nonimmunogenic variants stimulate radioresistant, Lyt- 1+2- helper T cells that support secondary antibody responses. However, the helper activity can not be detected unless dominant, radiosensitive Lyt-1-2+, I-J+ suppressor T cells are removed. In this paper we report that extracts of primed Lyt-2+ suppressor T cells contain insulin-specific suppressor factors (TsF) that are capable of replacing the activity of suppressor T cells in vitro. The activity of these factors is restricted by MHC-linked genes that map to the I-J region, and immunoadsorption studies indicated that they bind antigen and bear I-J-encoded determinants. Insulin-specific TsF consists of at least two chains, one-bearing I-J and the other the antigen-binding site. Furthermore, mixing of isolated chains from different strains of mice indicates that the antigenic specificity is determined by the antigen-binding chain and the MHC restriction by the H-2 haplotype of the source of the non-antigen-binding, I-J+ chain. Moreover, mixtures containing antigen-binding chain from allogeneic cell donors and I-J+ chain from responder cell donors have activity in cultures containing responder lymphocytes. This suggests that preferential activation of suppressor T cells, rather than differential sensitivity to suppression, results in the nonresponder phenotype to insulin.     

T cell-induced allotypic suppression. Origin of the CD8+ T cells maintaining IgG2ab suppression.

One of the problems raised by the T cell-induced allotypic suppression is the origin (donor or host) of the T cells responsible for the chronicity of the suppression. To address this point, we used T cells from Igha/a Thy-1.2 mice whose natural T cell activity against IgG2ab was enhanced in vivo. These T cells were injected into newborn Ighb/b Thy-1.1 mice where they induced complete suppression of IgG2ab expression in around 70% of these recipients. During a study that lasted more than 1 yr, we found that about 3% of the recipient splenocytes were T cells of the donor type. By means of suppression-transfer experiments, using either Thy-1.2+ or Thy-1.1+ cell-depleted splenocytes from mice suppressed in this manner we were able to unambiguously show that Thy-1.2+ cell-depleted splenocytes were incapable of transferring the suppression, whereas Thy-1.1+ cell-depleted splenocytes could. We thus demonstrated that suppression was maintained throughout the recipient's life by donor Thy-1.2+ T cells.     

Isotype-specific immunoregulation. Systemic antigen induces splenic T contrasuppressor cells which support IgM and IgG subclass but not IgA responses

In the present study, we have isolated and characterized the Lyt-1+, -2- T contrasuppressor (Tcs) cells from mice systemically primed with SRBC. Adoptive transfer of splenic Tcs cells from these mice abrogates oral tolerance and supports IgM and IgG anti-SRBC plaque-forming cell (PFC) responses; however, unlike the responses seen after transfer of Tcs cells derived from orally primed mice, low IgA responses were seen. Mice systemically primed with lower SRBC doses (0.01 to 1%) exhibited contrasuppression only within the L3T4- T cell subset, whereas mice primed with a high dose of SRBC (10%), harbored Lyt-1+, -2- Tcs cells in both the L3T4+ and L3T4- subsets. Both the L3T4- and L3T4+ Tcs cell subsets supported IgM and IgG responses when adoptively transferred to orally tolerized mice, and when added to tolerized spleen cell cultures. Splenic Tcs cells from systemically primed mice supported mainly IgG1 and IgG2b subclass anti-SRBC PFC responses, a pattern also seen with Tcs cells derived from orally primed mice. Both L3T4+ and L3T4- Tcs cells from systemically primed mice exhibited well established characteristics of contrasuppressor cells including binding to Vicia villosa lectin and expression of I-J. The splenic effector Tcs cells which support IgM, IgG1 and IgG2b anti-SRBC PFC responses are antigen-specific, since both L3T4- and L3T4+ Tcs cells from spleens of mice primed with 10% SRBC reverse tolerance to SRBC, but not to horse erythrocytes (HRBC). Further, both L3T4- and L3T4+ Tcs cells from HRBC-primed mice reverse tolerance to IgM and IgG anti-HRBC, but not to anti-SRBC responses. Isolation of T3-positive Lyt-1+, -2- and L3T4- Tcs cell subsets by flow cytometry followed by adoptive transfer, showed that effector Tcs cells express T3 and presumably contain an Ag-R (TCR-T3 complex). These studies show that systemic priming with heterologous RBC induces splenic Ag specific Tcs cells in a dose-dependent manner, which support IgM and IgG subclass responses, but not IgA responses.     

TMA-specific first-order T-suppressor hybridoma. I. Characterization of the hybridoma-derived single-chain inducer suppressor factor, TsF1

Experiments described in this report will characterize a monoclonal phenyltrimethylammonium (TMA) specific, first-order T-suppressor factor (TsF1) produced by a T-cell hybridoma, 8A.3. The hybridoma expressed the Thy-1, Lyt-1, Lyt-2 antigens as well as cross-reactive idiotypic (CRI) determinants but did not express I-J encoded epitopes. It was also found to bear determinants recognized by a monoclonal antibody raised against single-chain GAT-specific TsF1. The hybridoma-derived factor was capable of suppressing primary in vitro trinitrophenol (TNP)-specific responses induced with the Brucella abortus antigen, conjugated with TMA and TNP haptens (TMA-BA-TNP). In addition, in vivo administration of 8A.3 culture supernatant resulted in the specific suppression of TMA-specific delayed-type hypersensitivity (DTH) responses. Analysis of this factor revealed it to be an induction-phase, antigen-binding, CRI+, and I-J+ single chain polypeptide. Our results represent only the second such described single chain, antigen binding, I-J+ suppressor factor derived from a monoclonal T-cell hybridoma.     

In vitro T cell-mediated killing of Pseudomonas aeruginosa. IV. Nonresponsiveness in polysaccharide-immunized BALB/c mice is attributable to vinblastine-sensitive suppressor T cells

We reported previously that BALB/c mice immunized with a polysaccharide (PS) antigen isolated from immunotype 1 Pseudomonas aeruginosa and vinblastine sulfate develop T cell-mediated protective immunity, despite their failure to produce specific antibody. In vitro, Lyt-1-,2+, I-J+ T cells from vinblastine- and PS-immunized mice kill P. aeruginosa by secretion of a bactericidal lymphokine. BALB/c mice immunized with PS alone generate neither protective antibodies nor a protective T cell response. The current studies indicate that T cells from mice immunized with PS alone significantly suppress the bactericidal activity of T cells from mice immunized with vinblastine and PS. The suppressor T cells are of the same Lyt-1-,2+, I-J+ phenotype as the bactericidal T cells. Suppression is mediated by a soluble product of these suppressor T cells which both inhibits T cell proliferation and interferes with the production or release of the bactericidal lymphokine. Cyclophosphamide, used in other systems to remove suppressor T cells, fails to enhance bacterial killing and does not inhibit suppressor cell activity. These studies indicate that immunization with PS elicits responses in two functionally distinct subgroups of Lyt-1-,2+, I-J+ T cells, and that these cells are distinguishable by their sensitivity to vinblastine sulfate.     

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

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

Autoimmune thyroiditis induced in mice depleted of particular T cell subsets. I. Requirement of Lyt-1 dull L3T4 bright normal T cells for the induction of thyroiditis

T cell-depleted C3H/He or (C57BL/6xC3H/He)F1 (B6C3F1) mice were prepared by adult thymectomy and injection of antithymocyte serum, followed 3 wk later by lethal x-irradiation and bone marrow reconstitution. When these T cell-depleted mice were not injected or injected i.v. with normal spleen and lymph node cells treated with either anti-Thy-1, -L3T4 or -Lyt-2 antibody plus C or C alone, none of the groups of mice developed thyroiditis. In contrast, the adoptive transfer of normal cells treated with anti-Lyt-1 plus C resulted in high incidence of the production of antithyroglobulin antibody and the induction of typical thyroiditis lesion. The thyroid was the sole organ involved, because neither typical inflammatory lesion in other organs nor autoantibody such as anti-DNA antibody was detected in mice that exhibited thyroiditis. Analyses of surface phenotypes of cells required for inducing thyroiditis by the adoptive transfer revealed that an appreciable percentage of Lyt-1 dull T cells remained after the treatment of normal lymphoid cells with anti-Lyt-1 plus C. Almost all of these Lyt-1 dull T cells expressed magnitudes of L3T4 or Lyt-2 Ag comparable to those detected on Lyt-1 bright T cells. More important, the induction of thyroiditis was almost completely prevented by either in vitro or in vivo elimination of Lyt-1 dull L3T4+(bright) but not of Lyt-1 dull Lyt-2+(bright) T cells. These results indicate that Lyt-1 dull L3T4+ T cells existing in normal healthy individuals have potential to induce typical thyroiditis which is associated with the production of antithyroglobulin autoantibody, and that the activation and/or function of this T cell subset is regulated by the Lyt-1 bright T cell population coexisting in normal lymphoid cell population.     

NK cell-deficient mice develop a Th1-like response but fail to mount an efficient antigen-specific IgG2a antibody response.

NK cells have been shown to play a role in the modulation of B cell differentiation and Ab production. Using a novel murine model of NK cell deficiency, we analyzed the in vivo role of NK cells in the regulation of Ag-specific Ab production. After immunization with OVA or keyhole limpet hemocyanin in CFA, NK cell-deficient (NK-T+) mice developed an efficient Th1 response and produced significant levels of IFN-gamma but displayed markedly reduced or absent Ag-specific IgG2a production. There were no differences in the levels of Ag-specific IgG, IgG1, and IgG2b between NK-T+ and NK+T+ mice. Furthermore, NK cell-reconstituted, NK+T+ (tgepsilon26Y) mice produced significant amounts of Ag-specific IgG2a after immunization with OVA. These results indicate that NK cells are involved in the induction of Ag-specific IgG2a production in vivo. Moreover, they also demonstrate that the lack of Ag-specific IgG2a Ab production in NK-T+ mice is not associated with the impaired Th1 response and IFN-gamma production.     

Cellular subsets involved in cell-mediated immunity to murine Plasmodium yoelii 17X malaria

Cell mediated immunity to nonlethal Plasmodium yoelli 17X (PY17X-NL) was examined in the CBA/CaJ mouse by adoptive transfer of sensitized T lymphocyte subsets. In intact mice, PY17X-NL causes a self-limiting infection with parasitemia levels ranging from 10 to 25% of total red blood cells. Upon recovery, mice are refractory to subsequent challenge with the homologous parasite. In T cell-depleted mice, PY17X-NL infections are extremely virulent and result in death of the host after parasitemia levels reach 50% or higher. The transfer of either Lyt-1 T cells or Lyt-2 T cells from immune animals into normal, naive animals produced accelerated recovery to subsequent infection. However, this adoptive transfer of immunity by either subset was dependent upon the presence of an I-J+, Lyt-null cell in the immune population. T cell deprivation precluded the ability of animals to control blood-stage infections. When T cell-depleted mice were reconstituted with naive, Ig-negative (T cell-enriched) spleen cells, parasitemia levels were controlled and the parasites were eliminated. When T cell-deprived animals were reconstituted with naive Lyt-1+2-, Ig-negative spleen cells, they experienced twofold higher parasitemias of longer duration than mice receiving unfractionated cells. Two of six of these Lyt-1 mice died of fulminant infections, suggesting that the presence of naive Lyt-2 cells enhances the degree of protection. Immune Lyt-2 T cells were highly protective in T cell-depleted animals. Protection by sensitized Lyt-1 T cells correlated with the induction of a monocytosis. On the other hand, protection by Lyt-2T cells occurred in the absence of monocytosis. The possibility that the immunity induced by each T cell subset is mediated by a different effector mechanism is discussed.     

IL-1 as adjuvant. Role of T cells in the augmentation of specific antibody production by recombinant human IL-1 alpha

Human rIL-1 alpha significantly enhanced splenic plaque-forming cells (PFC) to SRBC in vitro and in vivo. A single i.p. injection was sufficient to produce a fivefold or greater increase in the generation of PFC in a primary response. IL-1 treatment resulted in an increased production of Ag-specific PFC, both in vitro and in vivo, in combination with suboptimal doses of Ag. When IL-1 was given with a primary dose of Ag in vivo, an enhanced IgG response occurred. IL-1 enhanced in vivo carrier priming for an anti-hapten PFC response, indicating increased Th activity. Furthermore, T cells from spleens of mice treated with IL-1 provided significantly more help in both carrier (SRBC)- and hapten (TNP)- specific PFC. The enhancement of PFC by IL-1 in vitro occurred even in the presence of an excess of neutralizing anti-IL-2 antibody. These results suggest that IL-1 may enhance T cell-dependent antibody production in part by increasing Th activity, and that the mechanism of IL-1 action in increasing antibody production involves pathways in addition to the induction of IL-2 secretion.     

Abnormal expression of surface immunoglobulin isotypes on antigen-binding B lymphocytes from mice tolerant to trinitrophenyl determinants

Temporary B-cell tolerance to the trinitrophenyl (TNP) hapten can be produced in BDF1 mice by intraperitoneal injection of trinitrobenzene sulfonic acid (TNBS). Antigen-binding cells (ABC) specific to TNP, measured as TNP donkey erythrocyte rosettes, are found in tolerant mice as well as in immune mice. We have studied the surface immunoglobulin isotype profile of these TNP-binding lymphocytes (TNP-ABC) in four groups of animals: nonimmune, immune, tolerant, and tolerant-challenged. Immune mice received intravenous TNP sheep erythrocytes (TNP-SRC), whereas tolerant-challenged mice received TNP-SRC and TNBS on Day 0. TNP-ABC from mice immunized with TNP-SRC exhibit increased expression of surface IgG and decreased expression of surface IgD, compared to the ABC from nonimmune mice. Tolerant mice have a higher proportion of ABC with surface IgG, and a lower proportion with surface IgD, than nonimmune mice. Tolerant-challenged mice have a lower proportion of ABC with surface IgG, and a higher proportion with surface IgD, than immune mice. Thus, B-cell tolerance in this model entails an attenuation of the surface immunoglobulin isotype switch (loss of IgD and gain of IgG) on ABC seen in the normal immune response. For most TNP-ABC, tolerogen exposure prevents the switch in surface isotypes normally induced by exposure to TNP antigen; i.e., the tolerance lesion precedes the surface isotype switch. However, a minority of the TNP-ABC appear to switch surface isotypes in response to the tolerogen itself.     

Role of antigen-specific T cell help in the generation of in vivo antibody responses. I. Antigen-specific T cell help is required to generate a polyclonal IgG1 response in anti-IgD antibody-injected mice.

A system in which injection of mice with an antibody to mouse IgD that they recognize as foreign stimulates a large, T cell-dependent IgG response was used to study whether Ag-specific T cell help is required to stimulate polyclonal (non-Ag-specific) IgG production in vivo. Igha x Ighb allotype heterozygous mice were injected with a conjugate of a foreign Ag coupled to a mAb specific for one of the two IgD allotypes expressed in these mice. This conjugate cross-links mIgD on B cells that express the recognized allotype. These cells process the conjugate and present the foreign Ag to Ag-specific T lymphocytes, which become activated. Thus, B cells of the recognized allotype can be stimulated by cross-linking of their mIgD, Ag-specific T cell help, non-Ag-specific cytokines, and non-Ag-specific contact with activated T cells. In contrast, B cells that express the Igh allotype not recognized by the Ag-anti-IgD antibody conjugate (bystander B cells) can be stimulated in this system only by non-Ag-specific cytokines and non-Ag-specific contact with activated T cells. Although both recognized and bystander B cells in conjugate-injected mice demonstrated substantial increases in size and Ia expression, only the recognized B cells were induced to synthesize DNA and to make a substantial polyclonal Ig response. Bystander B cells still failed to secrete IgG when mice were injected with an anti-IgD-Ag conjugate specific for the other Igh allotype as well as a mAb that cross-linked IgD of the bystander B cell allotype. These observations demonstrate that although non-Ag-specific cytokine and contact-mediated T cell help are sufficient to induce B cells to increase in size and Ia expression in anti-IgD antibody-injected mice, Ag-specific T cell help is required to stimulate the generation of an IgG response in these mice.     

Virus-neutralizing antibodies of immunoglobulin G (IgG) but not of IgM or IgA isotypes can cure influenza virus pneumonia in SCID mice.

The ability of monoclonal antibodies (MAbs) to passively cure an influenza virus pneumonia in the absence of endogenous T- and B-cell responses was investigated by treating C.B-17 mice, homozygous for the severe combined immunodeficiency (SCID) mutation, with individual monoclonal antiviral antibodies 1 day after pulmonary infection with influenza virus PR8 [A/PR/8/34 (H1N1)]. Less than 10% of untreated SCID mice survived the infection. By contrast, 100% of infected SCID mice that had been treated with a single intraperitoneal inoculation of at least 175 micrograms of a pool of virus-neutralizing (VN+) antihemagglutinin (anti-HA) MAbs survived, even if antibody treatment was delayed up to 7 days after infection. The use of individual MAbs showed that recovery could be achieved by VN+ anti-HA MAbs of the immunoglobulin G1 (IgG1), IgG2a, IgG2b, and IgG3 isotypes but not by VN+ anti-HA MAbs of the IgA and IgM isotypes, even if the latter were used in a chronic treatment protocol to compensate for their shorter half-lives in vivo. Both IgA and IgM, although ineffective therapeutically, protected against infection when given prophylactically, i.e., before exposure to virus. An Fc gamma-specific effector mechanism was not an absolute requirement for antibody-mediated recovery, as F(ab')2 preparations of IgGs could cure the disease, although with lesser efficacy, than intact IgG. An anti-M2 MAb of the IgG1 isotype, which was VN- but bound well to infected cells and inhibited virus growth in vitro, failed to cure. These observations are consistent with the idea that MAbs of the IgG isotype cure the disease by neutralizing all progeny virus until all productively infected host cells have died. VN+ MAbs of the IgA and IgM isotypes may be ineffective therapeutically because they do not have sufficient access to all tissue sites in which virus is produced during influenza virus pneumonia.