Augmented induction of tumor-specific resistance by priming with Mycobacterium tuberculosis (TBC) and subsequent immunization with PPD-coupled syngeneic tumor cells

The present study investigates the augmenting effect of tuberculin- (PPD) reactive amplifier T cells on the induction of syngeneic tumor immunity. PPD-reactive helper (amplifier) T cell activity was generated in C3H/HeJ mice by appropriate immunization with heat-killed Mycobacterium (Tbc). Immunization of these Tbc-primed mice with PPD-coupled syngeneic X5563 tumor cells led to augmented generation of in vivo tumor-neutralizing activity contingent on the presence of PPD-reactive amplifier T cell activity. Splenic T cells from these mice exhibited potent tumor-neutralizing activity using Winn's assay, whereas spleen cells from mice not primed with Tbc before PPD-X5563 immunization failed to neutralize viable X5563 tumor cells. After establishing that the neutralizing activity was tumor specific and mediated by T cells, the applicability of this augmentation of tumor-specific immunity to an immunotherapy model was explored. Immunization with PPD-X5563 in the early stages of the tumor-bearing state induced potent anti-tumor activity sufficient to reject the growing tumor. Pretreatment of mice with cyclophosphamide or light x-irradiation (250 R), procedures that eliminate suppressor cell activity nonspecifically, before priming with Tbc further potentiated the anti-tumor activity under these conditions. Thus, the present study elucidates the augmenting effect of PPD-reactive amplifier T cells in the induction of tumor-specific immunity and provides an effective method of immunotherapy in tumor-bearing animals.     

Reaginic antibody formation in the mouse. X. Possible role of suppressor T cells in transient IgE antibody responses.

The kinetics of IgE antibody response to alum-absorbed dinitrophenyl derivatives of ovalbumin (DNP-OA) was dependent on the dose of immunogen. A persistent IgE antibody response was obtained when high responder BDF1 mice were immunized with a minimum (0.05 microgram) dose. An increase of the immunogen to 10 microgram depressed IgE antibody responses but enhanced IgG antibody responses of both hapten and carrier specificities. Determination of T helper cell activity and B memory cells after immunization with different doses of antigen indicated that minimum immunogen was favorable for developing helper activity, whereas 1 to 10 microgram immunogen were more favorable than a 0.05-microgram dose for developing both IgE and IgG B memory cells. Nevertheless, neither helper T cells nor B memory cells in the spleen explains a transient IgE antibody response to a high (10 microgram) dose of DNP-OA. Evidence was obtained that immunization with 10 microgram OA induced generation of antigen-specific suppressor T cells, which were not detectable after immunization with 0.05 microgram OA. Transfer of suppressor T cells to DNP-OA-primed mice depressed both anti-hapten and anti-carrier IgE antibody responses. The results suggested strongly that suppressor T cells are involved in a transient IgE antibody response to a high-dose immunogen.     

Generation of specific helper cells and suppressor cells in vitro for the IgE and IgG antibody responses.

Normal splenic lymphocytes from BDF1 mice were cultured on ovalbumin (OA)-bearing syngeneic peritoneal adherent cells for 5 days and their subsequent helper function was tested by an adoptive transfer technique. Lymphocytes harvested from the culture were mixed with DNP-KLH-primed spleen cells and transferred into irradiated syngeneic mice followed by challenge with DNP-OA. The results showed that the cultured lymphocytes has helper function for both IgE and IgG anti-DNP antibody responses. Depletion of mast cells and T cells in the peritoneal adherent cell preparations did not affect the generation of helper cells in the culture. The helper function of the cultured lymphocytes was abolished by the treatment with anti-theta antiserum and complement and was specific for ovalbumin. The OA-specific helper T cells were generated in vitro by the culture of a T cell-rich fraction of normal spleen on T cell-depleted OA-bearing peritoneal cells. If the normal splenic lymphocytes or T cell-rich fraction were cultured with 10 mug/ml of OA in the absence of macrophages, cultured lymphocytes lacked helper function. The transfer of splenic lymphocytes or splenic T cells cultured with soluble OA to normal non-irradiated mice, however, suppressed both IgG and IgE antibody responses of the recipients to subsequent immunization with DNP-OA. The suppressor cells were sensitive to anti-theta antiserum and complement and their activity was specific for OA. The cultured cells transferred into normal mice did not suppress anti-hapten antibody response to DNP-KLH. Normal lymphocytes cultured on OA-bearing macrophages and had helper function in adoptive transfer experiments failed to suppress antibody response of non-irradiated recipients to DNP-OA. The results indicate that OA-bearing macrophages primed T cells and generated helper T cells, whereas the culture of normal lymphocytes with soluble OA in the absence of macrophages generated suppressor T cells.     

Reaginic antibody formation in the mouse. VII. Depression of the ongoing IgE antibody formation by suppressor T cells.

The ongoing IgE antibody formation against ovalbumin (OA) in high responder mice was depressed by i.v. injections of either native or urea-denatured ovalbumin (UD-OA). Adoptive transfer experiments to determine the helper function of spleen cells from the treated animals showed that helper function for both IgE and IgG antibody responses diminished after treatment. Evidence was obtained that treatment suppressed the expansion of IgE-G memory cells. When the same treatment with OA or UD-OA was given to OA-primed mice before the appearance of IgE antibody in their serum, OA-specific splenic suppressor T cells were demonstrable. Thus, the transfer of splenic T cells from treated mice into normal mice suppressed the primary IgE and IgG antibody responses of the recipeints to DNP-OA. It was also found that the transfer of the splenic T cells from UD-OA-treated mice into OA-primed mice depressed ongoing IgE antibody formation in the recipients. The results suggested strongly that the decrease of helper function and the depression of ongoing IgE antibody formation by repeated injections of UD-OA was caused by generation of antigen (OA)-specific suppressor T cells.     

Hapten-specific IgE antibody responses in mice. VII. Conversion of IgE "non-responder" strains to IgE "responders" by elimination of suppressor T cell activity.

Mice of the inbred strains SJL (H-2s) and AKR (H-2k) are "non-responders" and "low-responders," respectively, in terms of their capacity to develop antibody responses of the IgE class when immunized with conventional proteins and hapten-protein conjugates under conditions optimal for eliciting IgE responses in "high-responder" mice, such as BALB/c (H-2d), to these same antigens. For example, BALB/c mice preimmunized with ASC and then challenged 7 days later with DNP-ASC develop peak augmented primary IgE anti-DNP antibody responses of 320 PCA units, whereas SJL and AKR mice develop responses which are 16-fold and 4-fold lower, respectively. However, pretreatment of the latter two strains with appropriate doses of either x-irradiation (150 R), cyclophosphamide (100 mg/kg) or ALS (150 mul) before carrier-preimmunization strikingly enhances the magnitude of IgE antibody responses in such mice to levels as high as 64-fold above those of untreated control mice of the same strains. Evidence obtained in these experiments indicates that the capacity of such maneuvers to to convert poor IgE responders to high responder status reflects elimination of nonantigen-specific suppressor T lymphocytes which are naturally present and normally function to suppress or "dampen" the IgE antibody response in a relatively selective manner. It appears that these cells modulate IgE responses by acting at least at two distinct points: 1) The most effective activity seems to be at the level of induction of carrier-specific helper T cells; 2) A second locus of inhibitory activity is more distal in the response, either impeding helper T cell-B cell cooperative interactions or suppressing B cell differentiation and/or function directly. Taken collectively, these observations demonstrate that the state of poor responsiveness of the SJL and AKR strains for the IgE antibody class is not a reflection of a genetic inability to develop IgE responses but rather a manifestation of a genetic capability to actively inhibit IgE antibody synthesis.     

Reaginic antibody formation in the mouse. VII. Induction of suppressor T cells for IgE and IgG antibody responses.

Intravenous injections of urea-denatured ovalbumin (UD-OA) into OA-primed high responder mice suppressed the antibody response not only to the priming antigen but also to subsequent immunization with dinitrophenyl derivatives of OA (DNP-OA). The transfer of normal spleen cells or OA-primed spleen cells into UD-OA-treated animals did not restore the capacity of responding to DNP-OA to form anti-DNP IgE and IgG antibodies. The transfer of splenic T cell fraction from the UD-OA-treated animals into normal syngeneic mice diminished both IgE and IgG antibody responses of the recipients to DNP-OA. The B cell-rich fraction from the same donors failed to affect the anti-hapten antibody response and enhanced anti-cancer (OA) IgG antibody response of the recipients. It was also found that the transfer of T cell-rich fraction of OA-primed spleen cells failed to suppress antibody response of the recipients to DNP-OA. The results indicated that spleen cells of UD-OA-treated mice contained suppressor T cells which are distinct from helper cells. Suppressive activity of T cells in the UD-OA treated animals was specific for OA. The transfer of the T cell-rich fraction failed to suppress anti-DNP antibody response of the recipients to DNP-KLH.     

Activation through CD3 molecule leads a number of human T cell clones to induce IgE synthesis in vitro by B cells from allergic and nonallergic individuals

Seventy-eight clones established from tonsillar T lymphocytes of two nonallergic children were tested under different experimental conditions for their ability to induce in vitro IgE synthesis by B cells from allergic or nonallergic donors. After 24 hr preactivation with phytohemagglutinin (PHA), 11 out of 32 CD4+ clones from the first and 17 out of 36 CD4+ clones from the second tonsil donor showed the ability to induce IgE synthesis in vitro by B cells from both allergic and nonallergic individuals, whereas none of 10 CD8+ clones nor T blasts of PHA-induced cell lines obtained from unfractionated T cell suspensions of the same tonsils had such an effect. Seven of the 11 T cell clones from the first tonsil donor active on IgE production after pre-activation with PHA also induced IgE synthesis in vitro by nonallergic and allergic B cells upon stimulation with anti-CD3 monoclonal antibody. Under the same experimental conditions, virtually all of the T cell clones able to induce IgE synthesis in vitro by target B cells showed the ability to stimulate IgG and IgM production as well. T cell clones were also established from the peripheral blood of a nonallergic donor and were tested for their ability to induce IgE synthesis in autologous B cells. After preactivation with PHA, seven out of 35 CD4+ clones induced the production of detectable amounts of both IgE and IgG in autologous B cells. The addition to the cultures of PHA-stimulated unfractionated T cells inhibited in a dose-dependent manner the IgE but not the IgG synthesis induced by an autologous helper T cell clone in autologous B cells. Taken together, these data indicate that a remarkable proportion of human T cell clones upon triggering of the CD3 molecular complex were able to provide help for the synthesis of IgE in B cells from both allergic and nonallergic individuals. The successful induction of IgE synthesis by single T cell clones was apparently related to the lack of concomitant suppressor activity to which IgE-producing cells appeared to be exquisitely sensitive.     

Serologic and immunochemical characterization of HLA-A9 xenoantisera

There was a pronounced quantitative difference between the helper activities of B6C3F1 splenic T cells sensitized with unmodified vs modified antigens of SRBC. Modified SRBC induced the greater helper activity which was measured by the magnitude of an anti-TNP response (IgM and IgG) elicited in vivo by virgin B lymphocytes. Antigen modification was produced by conjugating SRBC with the hapten or simply by incubating SRBC in cacodylate buffer. There were restrictions with respect to both erythrocyte species and mouse strains for this differential priming to occur. The relatively poor performance of SRBC-primed T lymphocytes was apparently not due to suppressor T cells, but rather to activation of only one of two identified T cell subpopulations required for full helper activity. Unmodified SRBC activated a subpopulation of "helper" cells characterized as sensitive to elimination by ATS and long-lived after ATx, but failed to activate in B6C3F1 mice a second subpopulation of "amplifier" cells resistant to elimination by ATS and short-lived after ATx. In contrast, modified SRBC activated both helper and amplifier cells. Under appropriate conditions these subsets of T cells were strongly synergistic in promoting antihapten antibody formation especially of the IgG class. The involvement of two distinct types of T lymphocytes in the positive regulation of antibody responses raises interesting and novel questions concerning the sequence of events in the triggering of B cells and the subsequent development of the response.     

Human B cell differentiation. II. Suppression by T cells of T-dependent and T-independent plasma cell maturation.

In vitro human plasma cell generation induced by both T-dependent (PWM) and T-independent (NWSM) mitogens was found to be suppressed by peripheral blood lymphocytes preincubated with human aggregated IgG. T cells, but not B lymphocytes, were able to mediate the suppressive activity; since aggregated (Fab)'2 fragments were found unable to generate suppressor cells, it was concluded that the suppressor cell was a T lymphocyte bearing Fcgamma receptors. These cells appeared to be largely radiosensitive. In most cases the proliferative responses remained unchanged. Since NWSM-induced activation is not dependent on the presence of T cells, these results show that, at least in this case, T cells exert their suppressor function directly on B lymphocytes. Whether PWM-induced B cell differentiation is suppressed by the same mechanism or/and by inactivation of T helper lymphocytes remains under investigation.     

Genetic regulation of the antibody response to H-2Db alloantigens in mice. IV. Antigens that activate helper T cells for IgG, coded for by the non-H-2 genome

When B10.A(5R) mice are immunized with congenic C57BL/10 cells only 2-ME-sensitive antibodies (IgM type) are found directed against H-2Db. To obtain 2-ME-resistant antibodies (IgG type) 5R mice must be immunized with noncongenic cells (e.g., A.BY); in this case non-H-2 cell surface antigens will activate helper T cells to induce anti-Db IgG antibody production by B cells. An attempt was made to define helper antigens that activate helper T cells. Neither N-2 antigens of seven H-2Db recombinant strains nor a limited set of non-H-2 cell surface antigens were able to serve as helper antigens. By using individual backcross mice as antigen, one helper antigen was found on the background of strain A under the conditions used, whereas other backgrounds may carry more than one antigen. The helper antigen is dominantly expressed in F1 mice and has to be presented on the same cell as H-2Db to induce the switch from IgM to IgG.     

Induction of plasma cell differentiation of human fetal lymphocytes: evidence for functional immaturity of T and B cells.

Resembling the in vitro antibody response of the newborn cultures of cord blood lymphocytes stimulated with pokeweed mitogen (PWM) generated fewer plasma cells (PC) than comparable adult lymphocyte cultures and the response was almost exclusively of the IgM class. We investigated the cellular basis of this difference by preparing mixed cultures of newborn T or B lymphocytes with adult B or T cells. Substitution of adult for newborn T cells enhanced the response of newborn B cells, particularly of the IgG and IgA classes. The response of second trimester fetal spleen cells was also increased by adult T cells, although no IgA PC appeared. Conversely, adult B cells generated fewer PC particularly of the IgG and IgA classes when cultured with newborn T cells. The relatively poor IgA and IgG responses of newborn cells seems partially but not entirely due to deficiency of T cell helper function. Suppressor activity of newborn T cells was investigated by adding excess unrelated newborn or adult T cells to adult T + B cells: adult T cells improved the response whereas newborn T cells were variably suppressive. The results indicate that newborn T cells, although capable of helper function, are balanced toward suppression.     

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.     

Mechanisms of T and B cell collaboration in the in vitro production of anti-DNA antibodies in the NZB/NZW F1 murine SLE model

B/W mice spontaneously develop IgG antibodies to DNA that cause lethal immune nephritis. T and B cell interactions in the in vitro anti-DNA antibody response of B/W mice were investigated, and two distinct families of helper T cells that drive these responses were defined. First, the anti-DNA antibody-forming cell (AFC) response was found to be increased in B/W mice with nephritis and was inhibited with the monoclonal antibody anti-L3T4, suggesting a major role for helper T cells. Purified splenic T cells from mice with nephritis were able to augment both the IgG and the IgM anti-DNA AFC response of young B/W B cells. T helper cells were cloned from spleens of NZB/W F female mice with high titer anti-DNA antibodies and nephritis. The cloned T cells augmented both IgG and IgM anti-DNA AFC responses of young B/W B cells. Four clones--27.9, 30.7, 30.8, and 30.10--were selected for further study. These cells proliferated, in the context of syngeneic (H2d/z) antigen-presenting cells (APC) but not to allogeneic APC. Analysis of the mechanism of T helper cell clone-mediated augmentation of anti-DNA AFC revealed two populations: "cognate" T helper cells, which specifically augment anti-DNA AFC (30.7 and 30.10), and non-antigen-specific T helper cells (27.9 and 30.8), which augment the response of B cells of differing specificity by a bystander mechanism, probably through increased release of B cell growth and differentiation factors.     

PPD-induced immunoglobulin production in human peripheral blood lymphocytes. II. Separation of PPD-reactive helper T cells from PWM-reactive helper T cells in polyclonal immunoglobulin production of human peripheral blood lymphocytes.

We investigated the relationship bewteen PPD-reactive helper T cells and PWM-reactive helper T cells in polyclonal Ig production of human PBL. Elimination of PPD-reactive T cells by BUdR + light treatment resulted in a loss of helper function in PPD-induced Ig production, but had no effect on helper function in PWM-induced Ig production. On the other hand, elimination of PWM-reactive T cells resulted in a loss of helper function in both PPD-induced Ig production and PWM-induced Ig production. A blast cell-enriched fraction that was generated by PPD and separated by the velocity sedimentation method contained helper function in both responses. On the other hand, blast cell-depleted fractions did not contain PPD-reactive helper function, although the PWM-reactive helper function was evident. These results strongly suggest that PPD-reactive helper T cells are included in PWM-reactive helper T cells.     

Differences in the regulation of humoral responses between mice infected with Trypanosoma cruzi and mice administered T. cruzi-induced suppressor substance

In the present study, the effects of Trypanosoma cruzi and the T. cruzi-induced serum suppressor substance (SSS) on antibody responses were compared. Although infection with T. cruzi led to an alteration in T cell helper activity and a reduced specific B cell precursor frequency, SSS did not have a similar effect on either of these cell populations. The characteristics of the altered T cell helper activity was further investigated, and it was found that helper activity appeared earlier in infected mice than in normal or SSS-suppressed mice, and less antigen was required for optimal elicitation of T helper cells in infected mice. The potency of T cell helper activity also was shown to differ, and in the order T. cruzi-infected 6E normal 6E SSS-suppressed mice. It was found that spleen cells from T. cruzi-infected mice elaborated more potent specific helper factors than spleen cells from normal or SSS-suppressed mice, but did not produce a detectable nonspecific helper factor in vitro. Finally, the addition of B cells from low-dose primed, T. cruzi-infected mice to cultures of normal spleen cells resulted in subnormal responses to the priming antigen (sheep erythrocytes) but not to another unrelated antigen (trinitrophenyl-haptenated Brucella abortus), whereas similarly sensitized B cells from normal or SSS-suppressed mice caused no such effect.     

Induction of IgG and IgE synthesis in normal B cells by autoreactive T cell clones

During the course of generating tetanus toxoid (TT)-specific T cell clones frm an HLA-DR2,7 donor, four clones were obtained which proliferated in the presence of autologous monocytes alone without the addition of TT antigen. This proliferation was specifically inhibited by anti-HLA-DR framework mouse monoclonal antibody, and appeared to be HLA-DR-restricted. Two of the clones proliferated in response to HLA-DR2-bearing monocytes, and the other two clones proliferated in response to HLA-DR7-bearing monocytes. The capacity of these four autoreactive human T cell helper clones to induce IgE synthesis in B cells was studied. All four clones stimulated autologous peripheral blood B cells to synthesize IgE and IgG antibody. Induction of IgE synthesis in B cells by the autoreactive T cell clones followed the same pattern of HLA-DR restriction which governed the proliferative response of these clones. These results suggest that the interaction of autoreactive helper T cells with B cell HLA-DR antigens may be important in the activation of IgE immune responses in humans.     

The isolation and characterization of the human helper inducer T cell subset

Monoclonal antibody anti-4B4 was produced by fusing NS1 myeloma with spleen cells of a mouse immunized with Saguinus oedipus lymphocyte. This anti-4B4 antibody defines a 135-KD cell surface protein that is widely distributed throughout the hematopoietic system. More importantly, anti-4B4 is reactive with functionally unique human T cell subsets. Anti-4B4 antibody was reactive with approximately 41% of unfractionated T cells, 41% of T4+ inducer cells, and approximately 43% of T8+ cytotoxic/suppressor population. This antibody subdivided peripheral blood T4+ cells into two functionally distinct populations. The T4+4B4+ subset proliferates relatively poorly upon stimulation with Con A and autologous cell antigens (AMLR) but well on exposure to soluble antigens, and it provides a good helper signal for PWM-induced Ig synthesis. The T4+4B4- subset, in contrast, proliferates well to Con A stimulation and autologous cell antigen (AMLR) but relatively poorly to soluble antigen stimulation, and provides little help to B cells for PWM-induced Ig synthesis. The T4+4B4- subset is largely 2H4+ and functions as the inducer of the T8+ suppressor cells. Thus, the present results suggest that one can divide the human T4 population into two major subsets that are phenotypically and functionally distinct, the human helper inducer subset (T4+4B4+/H.I.) and its reciprocal population defined by anti-2H4, the suppressor inducer subset (T4+2H4+/S.I.).     

Regulation of antibody response in different immunoglobulin classes. II. Induction of in vitro IgE antibody response in murine spleen cells and demonstration of a possible involvement of distinct T-helper cells in IgE and IgG antibody responses.

In vitro induction of anti-DNP IgE as well as IgG1, IgG2a antibody responses was shown in murine spleen cell culture. Spleen cells primed three times with 1 mug of DNP-OA or DNP-Asc produced significant amounts of anti-DNP IgE as well as IgG antibodies by the in vitro stimulation with DNP-OA or DNP-Asc, respectively. Collaboration between DNP-primed B cells and carrier-primed T cells was required for the induction of both IgE and IgG antibodies with DNP-coupled T-dependent antigen. Carrier-specific T cells induced with a low dose of Asc (0.01 mug) showed helper function only on IgE antibody response, whereas T cells primed with a higher dose of Asc (10 mug) cooperated only with IgG-B cells. T cells primed with Asc in CFA showed helper function mainly on IgG antibody response but not on IgE antibody response. The result indicated the presence of a distinct population of T helper cells for IgE and IgG antibody responses. T-independent antigen (DNP-Ficoll) induced both anti-DNP IgE and IgG antibody responses in DNP-primed spleen cell population without the requirement of the collaboration of helper T cells.     

Augmentation of auto-anti-idiotypic antibody production by hapten-reactive helper T lymphocytes

Augmented auto-anti-idiotypic antibody production was effectively achieved by immunization of mice with haptenated myeloma protein in the presence of hapten-reactive helper T lymphocytes. Hapten-reactive helper T-lymphocyte activities were raised in BALB/c mice by immunization with para-azobenzoate (PAB)-derived mouse gamma globulin (MGG) prepared by amidination reaction (PAB_im-MGG). Helper T cell activity was effectively enhanced by pretreatment of mice with a PAB-derived nonimmunogenic copolymer of D-glutamic acid and D-lysine (D-GL) (PAB-D-GL) 3 days before priming with PAB_im-MGG; PAB-D-GL is a potent tolerogen of both PAB-specific suppressor T lymphocytes and PAB-specific B cells. After induction of these enhanced PAB-reactive helper T lymphocytes, mice were immunized with PAB-coupled TEPC-15 myeloma protein (PAB_im-T-15), which was also prepared by amidination reaction. Mice immunized in this way manifested strikingly enhanced titers of auto-anti-idiotypic antibodies, specific for the T-15 idiotype, as compared to control mice which had not been preimmunized with PAB_im-MGG. The ability of PAB_im-MGG preimmunization to facilitate auto-anti-idiotypic antibody production was due to the activity of PAB-reactive helper T cells since PAB-specific B cells had been abolished by prior treatment with PAB-D-GL. The implications of this model for future studies on immunological engineering the analysis of idiotype network phenomena are discussed.     

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.