Antibodies to the L3T4 and Lyt-2 molecules interfere with antigen receptor-driven activation of cloned murine T cells

When L3T4+ cloned murine helper T lymphocytes (HTL) are stimulated with antigen or immobilized anti-T cell receptor (TCR) monoclonal antibodies (mAb) at concentrations which are optimal for proliferation, anti-L3T4 mAb inhibits activation as measured by proliferation and lymphokine production. Under similar conditions, IL 2-independent proliferation of Lyt-2+ cloned murine cytolytic T lymphocytes (CTL) stimulated by anti-TCR mAb is inhibited by anti-Lyt-2 antibodies. Proliferation of cloned HTL and CTL cells stimulated by IL 2 is not affected by the anti-L3T4 and anti-Lyt-2 mAb. The inhibition of TCR-induced activation of the T cell clones is not due to interference with the binding of the anti-TCR mAb. Stimulation of the TCR has been proposed to induce lymphokine secretion and proliferation by T cells through a pathway involving the activation of protein kinase C and the stimulation of an increase in the concentration of intracellular free calcium. However, proliferation of T cells stimulated by PMA (which activates protein kinase C) plus the calcium ionophore A23187 (which increases the concentration of intracellular free calcium) is not affected by mAb reactive with the Lyt-2 or L3T4 structures. If TCR stimulation does indeed activate T cells by activating protein kinase and increasing intracellular free calcium, then our data suggest that anti-L3T4 and anti-Lyt-2 mAb inhibit TCR-driven proliferation at some step before the activation of protein kinase C and the stimulation of a rise in intracellular free calcium concentration. Our results suggest that anti-L3T4 and anti-Lyt-2 mAb interfere with early biochemical processes induced by stimulation of the TCR. In HTL, which proliferate via an autocrine pathway, anti-L3T4 mAb appears to inhibit proliferation by interfering with signaling events involved in lymphokine production. Inhibition of IL 2-independent proliferation of Lyt-2+ cells by anti-Lyt-2 mAb appears to occur by a different mechanism. The precise molecular basis for the interference of each cell type has not yet been characterized.     

Role of L3T4+ and LyT-2+ cells in experimental visceral leishmaniasis

In contrast to euthymic (nu/+) BALB/c mice, athymic nude (nu/nu) BALB/c mice fail to control the visceral intracellular replication of Leishmania donovani, do not generate the macrophage-activating lymphokine IFN-gamma, and show little or no granulomatous tissue response. To characterize the T cell requirement for successful defense against L. donovani, nude mice were first reconstituted with unfractionated nu/+ immune spleen cells, which readily conferred the capacity to control and eliminate visceral (hepatic) L. donovani. In reconstituted mice, acquired resistance was paralleled by the ability of spleen cells to generate high levels of leishmanial Ag-stimulated IFN-gamma and the development of well formed liver granulomas. In contrast, nude mice reconstituted with either L3T4+- or Lyt-2+-enriched immune spleen cells alone failed to control visceral parasite replication and did not develop effective granulomas despite the finding that transfer of L3T4+ cells largely and Lyt-2+ cells partially restored the capacity to secrete IFN-gamma. To determine whether both T cell subsets were also required in a normal host, nu/+ BALB/c mice were treated with cell-depleting anti-L3T4 and anti-Lyt-2 mAb. Depletion of either T cell subset inhibited the acquisition of resistance to L. donovani and impaired the tissue granulomatous response. Thus, successful T cell-dependent host defense towards intracellular L. donovani and the tissue expression (granulomas) of this mechanism appear to require both L3T4+ and Lyt-2+ cells. A primary role for the L3T4+ cell may be IFN-gamma production; the role of the Lyt-2+ cell and the precise interaction of the two T cell subsets remain to be identified.     

Effects of anti-Lyt-2 and anti-L3T4 monoclonal antibodies on the function of cytotoxic T lymphocyte/helper T lymphocyte hybrid T cell clones

CTL/HTL hybrid clones provide a unique system that allows detailed analysis of the role of Lyt-2, L3T4, and other structures involved in T cell functions. We have demonstrated previously that the fusion of cloned murine CTL and helper T lymphocytes with defined specificity generated hybrid cells that expressed both Lyt-2 and L3T4 as well as two TCR. Data obtained with these hybrid clones demonstrated that cytolysis is closely linked to the CTL TCR. We have analyzed the effects of anti-Lyt-2 and anti-L3T4 as well as anti-TCR mAb on cytolysis, proliferation, and lymphokine release by a number of hybrid clones. We found that anti-Lyt-2 and anti-L3T4 mAb were able to inhibit both proliferation and lymphokine release by the hybrid clones in response to stimulation of either the CTL or helper T lymphocyte parent TCR. In contrast, only anti-Lyt-2 and anti-CTL TCR mAb were able to block cytolysis of target cells bearing the Ag recognized by the CTL TCR. These results provide further evidence that cytolysis is closely linked to the CTL TCR and that Lyt-2 and L3T4 have more than a passive role as accessory molecules on the surface of T lymphocytes.     

Dual regulation of resistance against Toxoplasma gondii infection by Lyt-2+ and Lyt-1+, L3T4+ T cells in mice

Studies were performed to attempt to define the T cell subset responsible for resistance to Toxoplasma gondii. A temperature-sensitive mutant (ts-4) strain of T. gondii was used for immunization because it causes infection but does not persist in the host. Immunization with this strain induced marked resistance against lethal challenge infection with virulent strains of T. gondii in mice. The resistance could be transferred to normal recipient mice by i.v. injection of spleen cells from ts-4-immunized mice. Marked inhibition of cyst formation in the recipient mice was also noted. The protective activity of immune spleen cells was removed by pretreatment of the spleen cells with anti-Thy-1.2 and C, indicating that T cells are responsible for the observed protection. Pretreatment of immune spleen cells with anti-Lyt-2.2 and C completely ablated their protective effect; pretreatment with anti-Lyt-1.2 or anti-L3T4 and C had lesser effects on their ability to transfer resistance. The effect of anti-Lyt-1.2 was the same as that obtained with anti-L3T4. This suggested that one T cell subset that is partially responsible for protection has both Lyt-1.2 and L3T4 markers on the cell surface. These results indicate that there are substantial roles for both the Lyt-2+ and Lyt-1+, L3T4 T cell subsets in dual regulation of resistance against toxoplasma infection and that Lyt-2+ T cells are the principal mediator of the resistance.     

The role of L3T4+ and Lyt-2+ T cells in the IgE response and immunity to Nippostrongylus brasiliensis

The role of L3T4+ and Lyt-2+ T cells in protective immunity to Nippostrongylus brasiliensis (Nb) was studied in BALB/c mice that were depleted of either the L3T4+ or Lyt-2+ T cell population by injection with rat mAb specific for the appropriate determinant. Host responses to Nb infection including spontaneous elimination of adult worms, development of intestinal mucosal mast cell hyperplasia and the generation of a polyclonal IgE response were all completely blocked by 0.5 mg anti-L3T4 antibody administered simultaneously with Nb inoculation. However, administration of 0.5 mg of anti-Lyt-2 antibody at the same time and 7 days after inoculation with Nb had no effect on any of these responses. Injection of anti-L3T4 antibody as late as 9 days after Nb inoculation interfered with spontaneous cure of Nb infection and anti-L3T4 antibody injection 11 days after Nb inoculation inhibited serum IgE levels measured on day 13 by 50%. In addition, administration of anti-L3T4 antibody at the time of the peak serum IgE response, 13 days after Nb inoculation, accelerated the decline in serum IgE levels. Injection of previously Nb-infected mice with anti-L3T4 antibody at the time of a second Nb inoculation prevented the development of a secondary IgE response but did not affect immunity to Nb infection based on finding no adult worms in the intestines of these mice. These data indicate that 1) L3T4+ T cells are required for spontaneous cure of Nb infection, development of intestinal mucosal mast cell hyperplasia, and the generation and persistence of an IgE response during primary infection with Nb and 2) L3T4+ T cells are required for a considerable time after inoculation for optimal development of these responses. However, L3T4+ T cells are not required for all protective responses in immune mice. In contrast, our data indicate that considerable depletion of the Lyt-2+ T cell population has no significant effect on either worm expulsion or the generation of serum IgE responses.     

Role of L3T4+ and Lyt-2+ T cell subsets in protective immune responses of mice against infection with a low or high virulent strain of Toxoplasma gondii

In order to elucidate the role of T cell subsets in protective immunity against infection with high virulent and low virulent strains of Toxoplasma gondii, monoclonal antibodies specific for T cell subsets were injected into mice before immunization or challenge infection. Treatment of mice with monoclonal antibody to either L3T4+ or Lyt-2+ T cells before they were immunized with Toxoplasma cell homogenate prepared from high virulent RH strain tachyzoites markedly reduced survival after mice were challenged with low virulent bradyzoites of the Beverley strain. Thus, induction of protective immunity against bradyzoites of the Beverley strain requires the presence of both L3T4+ and Lyt-2+ T cells. In contrast, mice injected with living bradyzoites of the low virulent Beverley strain after immunization with Toxoplasma cell homogenate acquired protective immunity against high virulent tachyzoites of the RH strain. Lyt-2+ T cells alone appear to be final effector cells for protection against the challenge with high virulent RH strain tachyzoites, since treatment of the bradyzoite-immune mice with anti-Lyt-2 antibody, but not anti-L3T4 antibody, before challenge significantly increased mortality.     

Impact of genetically regulated T cell proliferation on acquired resistance to Listeria monocytogenes

Two lines of mice genetically selected for high and low in vitro responses to PHA were used to evaluate the impact of T cell polyclonal expansion on acquired resistance to Listeria monocytogenes. The selective breeding induced two major consequences in low responder mice: (1) a reduction of the number of L3T4+ cells and (2) a restriction of T cell expansion upon PHA stimulation, predominantly affecting the Lyt-2+ subset, and associated with an abridgment of IL-2 production. In vivo PHA stimulation induced anti-Listeria protection in high responder mice, but was much less effective in low responder mice. Flow cytometer analysis revealed that T cell proliferation was also reduced in low responder mice during the course of Listeria infection, implying both L3T4+ and Lyt-2+ subsets. This defect did not apparently influence the kinetics of bacterial elimination in host tissues, which was similar in both lines during primary Listeria infection. In contrast, the expression of delayed-type hypersensitivity to Listeria antigens and the level of immunologic memory were significantly reduced in low responder mice. In vivo selective T cell depletion by anti-L3T4 or anti-Lyt-2 mAb allowed us to demonstrate the predominant role of Lyt-2+ cells in protection and that of L3T4+ cells in the expression of delayed-type hypersensitivity.     

Surface markers of T cells causing lethal graft-vs-host disease to class I vs class II H-2 differences

Information was sought on the phenotype of lymphoid cells causing lethal graft-vs-host disease (GVHD) in irradiated mice expressing whole or partial H-2 differences. In all strain combinations tested, pretreating donor lymph node (LN) cells with anti-Thy-1 monoclonal antibodies (MAb) plus complement (C) abolished mortality. With GVHD directed to class I H-2 differences, pretreating LN cells with anti-Lyt-2 MAb prevented mortality, whereas MAb specific for Ly-1 or L3T4 cell surface determinants caused severe mortality. These data imply that lethal GVHD directed to class I H-2 differences is mediated by L3T4-, Lyt-2+ cells; this subset of T cells was shown previously to control GVHD directed to multiple minor histocompatibility antigens, i.e., antigens seen in the context of self-class I molecules. With whole H-2 differences, GVHD appeared to be controlled largely but not exclusively by L3T4+, Lyt-2-T cells. This T cell subset was also the predominant cause of GVHD directed to class II differences. With class II incompatibilities, depleting donor cells of L3T4+ T cells, either by pretreatment with anti-L3T4 MAb + C or by fluorescence activated cell sorter selection, greatly reduced but did not completely abolish GVHD. These data might imply that L3T4-, Lyt-2+ cells have some capacity to elicit anti-class II GVHD. A more likely possibility, however, is that the residual GVHD to class II differences observed with Lyt-2+-enriched cells reflected minor contamination with L3T4+ cells.     

Importance of L3T4+ and Lyt-2+ cells in the immunologic control of infection with Mycobacterium bovis strain bacillus Calmette-Guérin in mice. Assessment by elimination of T cell subsets in vivo

The course of infection after injection of small doses of bacillus Calmette-Guérin (BCG) was studied in mice which were depleted in vivo of T cell subsets by administration of either anti-L3T4 or anti-Lyt-2 mAb. The results presented herein strongly suggest that the L3T4+ subpopulation play a pivotal role in the immunologic control of BCG infection because the depletion of L3T4+ cells led to a dramatic increase in the number of viable bacteria. Depletion of Lyt-2+ cells had no significant effect on the course of infection. These results were confirmed by using adoptive transfer experiments which showed that protective immunity was mediated by L3T4+ cells generated in the spleen as a result of infection. Moreover, T cells capable of controlling the recurrence of BCG multiplication from residual bacteria remaining in organs after the recovery from infection were shown to belong to the L3T4+ subpopulation.     

Functional analysis of T lymphocyte subsets in antiviral host defense

The role of different T cell subsets in antiviral host defense was investigated by treating thymectomized C57BL/6 and CBA/J mice with monoclonal rat anti-Lyt-2 or anti-L3/T4 IgG 2b antibodies 14 and 10 days before infection. This treatment depleted the respective T cell subsets to undetectable levels in peripheral blood when assayed by immunofluorescence. In mice treated with anti-Lyt-2, induction of cytotoxic T cells was reduced to less than 1 to 2% after intravenous infection with Armstrong strain of lymphocytic choriomeningitis virus (LCMV). In addition, no primary swelling of the footpad could be detected following local inoculation of the virus. In animals treated with anti-L3/T4, antiviral cytotoxic T lymphocyte responses were reduced by a factor of 10. These L3/T4+ cell-depleted mice showed delayed footpad swelling after local injection of LCMV Armstrong. After intracerebral infection with LCMV, anti-Lyt-2-treated mice were resistant and those injected with anti-L3/T4 were totally susceptible to LCMV Armstrong-triggered immunopathologic disease. Virus could be detected in the blood of antibody-treated mice 7 days after inoculation; however, no virus could be measured in the blood of surviving anti-Lyt-2-treated animals 15 days after intracerebral infection. Serum titers of interferon-alpha,beta induced by viral infection remained unaffected by depletion of T cell subsets. Anti-L3/T4 antibody-treated C57BL/6 mice failed to generate IgG antibodies against the New Jersey strain of vesicular stomatitis virus, whereas Lyt-2+ cell-depleted mice had normal antivesicular stomatitis virus (New Jersey strain) IgG antibody titers.     

Cloned Listeria monocytogenes specific non-MHC-restricted Lyt-2+ T cells with cytolytic and protective activity

Mice were infected with Listeria monocytogenes and Lyt-2+ T cell clones capable of lysing Ag-primed bone marrow macrophages were established. In accordance with earlier findings obtained at the population level, some T cell clones were identified which lysed bone marrow macrophages of different MHC type provided the relevant Ag was present. This unusual target cell recognition was further analyzed using a T3+, L3T4-, Lyt-2+, F23+, KJ16+ T cell clone, designated L-28. Target cell lysis by this clone was Ag specific, apparently non-MHC restricted. In contrast, YAC cells and P815 cells were not lysed by clone L-28. However, lysis of irrelevant targets could be induced by anti-T3, F23, or KJ16 mAb. Furthermore, Ag-specific lysis was blocked by anti-Lyt-2 mAb and by F(ab)2 fragments of F23 mAb. In addition to its cytolytic activity, clone L-28 produced IFN-gamma after co-stimulation with accessory cells, Ag, and rIL-2 and conferred significant protection on recipient mice when given together with rIL-2. These data suggest that non-MHC-restricted Lyt-2+ killer cells generated during listeriosis are cytolytic T lymphocytes that interact with their target Ag via the T cell receptor/T3 complex and the Lyt-2 molecule and, furthermore, that these cells play a role in anti-listerial resistance. The possible relevance of IFN-gamma secretion and target cell lysis for antibacterial protection is discussed.     

Role of L3T4 antigen in the activation of various functions of Lyt-1+2- T cells against vaccinia virus

The present study defines assay systems for vaccinia virus-reactive Lyt-1+2- T cells mediating various functions and investigates the positivity of L3T4 antigen on these Lyt-1+2- T cells as well as the role of L3T4 antigen in the activation of these T cells with respect to their functions. C3H/He mice were immunized against vaccinia virus by inoculating viable virus intraperitoneally (i.p.). Anti-vaccinia virus reactivity in lymphoid cells from these immunized mice was assessed by proliferative response, helper T cell activities involved in cytotoxic T lymphocyte (CTL) and B cell (antibody) responses, delayed type-hypersensitivity (DTH) response, and production of lymphokines such as interleukin 2 (IL2) and macrophage-activating factor (MAF). The results demonstrate that all of the above anti-vaccinia virus responses were mediated by Lyt-1+2- T cells and that these Lyt-1+2- T cells expressed L3T4 antigens on their cell surfaces. Moreover, such anti-vaccinia Lyt-1+2- T cell responses were inhibited in the presence of anti-L3T4 antigen antibody. These results indicate that there is a reciprocal relationship between Lyt-2 and L3T4 markers, and that L3T4 antigen is closely related to the activation of various functions of anti-vaccinia virus Lyt-1+2- T cells.     

A role for Lyt-2+ T cells in resistance to cutaneous leishmaniasis in immunized mice

The role of Lyt-2+ T cells in immunologic resistance to cutaneous leishmaniasis was analyzed by comparing infection patterns in resistant C57BL/6 mice and susceptible BALB/c mice induced to heal their infections after sub-lethal irradiation or i.v. immunization, with similar mice treated in vivo with anti-Lyt-2 antibodies. Administration of anti-Lyt-2 mAb resulted in a dramatic reduction in the number of lymphoid cells expressing the Lyt-2+ phenotype. Such treatment led to enhanced disease in both resistant C57BL/6 and irradiated BALB/c mice, as assessed by lesion size, but did not affect the capacity of these mice to ultimately resolve their infections. In contrast, anti-Lyt-2 treatment totally blocked the induction of resistance in i.v. immunized mice. These results suggest, that Lyt-2+ T cells may play a role in immunity to a Leishmania major infection and that their relative importance to resistance may depend on how resistance is induced.     

Characterization of the inflammatory response in the central nervous system of mice susceptible or resistant to demyelination by Theiler's virus

Intracerebral inoculation of mice with Theiler's murine encephalomyelitis virus results in an intense inflammatory response of mononuclear leukocytes which infiltrate into the central nervous system. Resistant strains of mice have the ability to clear virus whereas susceptible strains become infected persistently and are associated with chronic demyelination which is proposed to be immune-mediated. In an attempt to better understand the role of the immune response during demyelination, mononuclear leukocytes were isolated from the central nervous system of infected mice and stained by an immunoperoxidase technique with anti-Thy-1.2, anti-L3T4, anti-Lyt-2 and anti-MAC-1 mAb. Infection of susceptible SJL/J mice resulted in a biphasic immune response which peaked on days 7 and 27 post-infection. In contrast, a single peak (day 7) was observed in resistant C57BL/10SNJ mice. The presence of Thy-1.2, L3T4, and MAC-1+ cells was similar between the two strains. However, although the number of Lyt-2+ cells peaked on day 7 in C57BL/10SNJ mice, they were not detected in SJL/J mice until 14 days post-infection and gradually increased in number over the course of infection. To further study the role of T cells in demyelination, serial frozen sections of brain and spinal cord were stained for the presence of Lyt-2 and L3T4+ cells in the lesions of chronically infected SJL/J mice. L3T4+ cells were observed predominantly in perivascular regions while Lyt-2+ cells were observed infiltrating the parenchyma. These results provide further evidence that Lyt-2+ lymphocytes are important in the mechanism of susceptibility/resistance to Theiler's murine encephalomyelitis virus-induced demyelination.     

Suppression of Ia-unrestricted primary anti-Qa-1 cytotoxic T lymphocyte responses by class II major histocompatibility complex-restricted cellular interactions

A model has been established for investigating the cellular interactions for the generation and regulation of primary cytotoxic T lymphocyte (CTL) responses to Qa-1 alloantigens. Although NZB anti-BALB/c one-way mixed leukocyte cultures (MLC) generate anti-Qa-1b CTL, anti-Qa-1 CTL responses are not generated during BALB/c anti-NZB one-way MLC or during two-way MLC with NZB and BALB/c spleen cells. However, depletion of L3T4+ cells from the spleens of BALB/c mice before two-way MLC with NZB spleen cells resulted in anti-Qa-1b CTL responses. Likewise, the addition of anti-L3T4 monoclonal antibody (mAb) or anti-I-Ad mAb to two-way MLC with NZB and BALB/c spleen cells resulted in the generation of anti-Qa-1b CTL. Conversely, anti-Lyt-2 mAb inhibited the generation of anti-Qa-1 CTL. These data indicate that class II major histocompatibility complex-restricted cellular interactions are capable of suppressing the generation of Ia-unrestricted anti-Qa-1 CTL responses by Lyt-2+ responder cells. This model provides a novel opportunity to both characterize the cellular interactions responsible for regulating primary CTL responses to the Qa/Tla-encoded class I molecule Qa-1, and determine the contribution of this L3T4+ Ts-dependent defect in NZB mice to the pathogenesis of autoimmunity.     

Resistance in murine schistosomiasis is contingent on activated IL-2 receptor-bearing L3T4+ lymphocytes, negatively regulated by Lyt-2+ cells, and uninfluenced by the presence of IL-4

These studies assess the roles of subpopulations of T lymphocytes in inducing and modulating resistance to Schistosoma mansoni. C57BL/6 mice were depleted in vivo of L3T4+, Lyt-1+, Lyt-2+, IL-2R+ cells, or IL-4 by administration of appropriate mAb. Resistance and various correlative parameters of the immune response were studied in normal, depleted, and congenitally athymic mice. Depletion of T lymphocytes by anti-L3T4 or anti-IL-2R mAb reduced the development and expression of resistance, IgG2a and IgE antibody formation, and delayed type hypersensitivity reactivity against schistosome Ag. Depletion with anti-IL-4 antibody led to profound suppression of IgE-eosinophil-mediated antibody-dependent cell-mediated cytotoxicity and passive cutaneous anaphylaxis responses against the parasite and no effect on IgG2a antibody, Ag-mediated blast transformation, or resistance. Depletion of Lyt-2+ cells produced augmented development and expression of resistance and an increase in the immunological parameters of anti-schistosome reactivity. These studies suggest that protective immunity to S. mansoni in mice, induced by irradiated cercariae, is dependent on L3T4+, IL-2R+ lymphocytes and negatively regulated by Lyt-2+ cells. IL-4 does not appear to be essential for the development of resistance but is essential for the IgE response to the parasite.     

Conditions of anti-Lyt-2-mediated inhibition of TcR/CD3-induced IFN-gamma secretion by a CTL clone

The relationship between the T cell receptor (TcR) for antigen (Ag) and the Lyt-2/3 molecule during T cell activation was studied using the T cell clone KB5.C20, which is dependent upon Lyt-2 for target cell killing. This cytolytic T cell clone can be activated to secrete IFN-gamma by stimulation with H-2Kb expressing cells or with monoclonal antibodies directed against a clonotypic structure of the TcR or against associated CD3 molecules. IFN-gamma production induced by H-2Kb can be inhibited by anti-Lyt-2mAb. In addition, TcR-mediated activation using the anticlonotypic mAb Désiré-1 in soluble form can be inhibited by anti-Lyt-2 mAb in soluble form either as a divalent IgG or as its monovalent Fab fragment. Anti-Lyt-2 mAb immobilized on plastic wells was also inhibitory. Stimulation induced by the anti-TcR mAb or by anti-CD3 mAb immobilized on plastic can be inhibited only with plastic immobilized and not with soluble anti-Lyt-2mAb, however. These results are discussed in terms of local interactions between TcR and Lyt-2 molecules.     

L3T4+ T cells regulate Abelson virus-induced lymphomagenesis

To evaluate the role of T cells in regulation of lymphomagenesis, experiments were performed using Abelson murine leukemia virus (AMuLV). In vitro transformation of bone marrow target cells by this B lymphotropic retrovirus was inhibited by peripheral lymph node cells from naive mice. The inhibitory activity depended on Thy-1+ L3T4+ cells but did not require Lyt-2+ cells. In vivo depletion of L3T4+ T cells with a mAb (GK1.5) altered the course of AMuLV-induced lymphoma. L3T4 depletion of naturally resistant C57BL/6 mice resulted in dramatic susceptibility to lymphoma induction. Lymphoma cells from anti-L3T4-treated C57BL/6 mice infected with AMuLV displayed the B lineage transformation marker P1606C3. These studies reveal an important immunologic component of Abelson disease resistance involving L3T4+ T cells.     

Treatment of murine lupus with F(ab')2 fragments of monoclonal antibody to L3T4. Suppression of autoimmunity does not depend on T helper cell depletion

Treatment with mAb to the L3T4 Ag on Th cells can inhibit autoimmunity in mice. However, the mechanism by which anti-L3T4 inhibits autoimmunity is not known. In these studies, lupus-prone NZB/NZW F1 (B/W) mice were treated with F(ab')2 fragments of mAb to L3T4 to determine whether Th cell depletion is required for the beneficial effects of anti-L3T4. We first showed that treatment of female B/W mice with F(ab')2 anti-L3T4 from age 5 to 9 mo significantly reduced autoantibody production without depleting L3T4+ cells. However, treatment was complicated by the development of a host immune response to the rat mAb fragments. To circumvent this problem, female B/W mice were treated with a single high-dose of intact rat mAb to L3T4 (GK1.5) at age two mo. to induce immune tolerance to the mAb. Then, after recovery of L3T4+ cells, the mice were treated from age four to 14 mo with either F(ab')2 anti-L3T4 (0.5 mg 3 times per wk), intact anti-L3T4, or saline. In mice tolerized by this regimen, neither the F(ab')2 rat mAb nor the intact rat mAb elicited a host response. The mAb fragments bound target Ag but did not deplete the Th cells, whereas intact mAb to L3T4 profoundly depleted the L3T4+ cells. Despite this difference, both therapies had the same substantial beneficial effects on autoimmunity. They significantly decreased anti-DNA Ab production, improved renal function and prolonged survival. The initial tolerizing dose, by itself, did not inhibit autoimmunity. These findings show that anti-L3T4 suppresses autoimmunity by directly altering Th cell function through the L3T4 Ag, and not solely by depleting Th cells. They also document the detrimental effects of the host immune response to therapy with anti-L3T4 mAb, and they demonstrate a new strategy by which this response may be prevented.     

Depletion of L3T4+ and Lyt-2+ cells by rat monoclonal antibodies alters the development of adoptively transferred experimental autoimmune thyroiditis

To delineate the contribution of L3T4+ and Lyt-2+ cells in the pathogenesis of experimental autoimmune thyroiditis (EAT), synergistic pairs of monoclonal antibodies (mAb) to the T cell subsets were used in conjunction with the adoptive transfer of mouse thyroglobulin (MTg)-activated cells from immunized mice. Initial experiments verified the important role of L3T4+ cells in the transfer of EAT. Subsequent experiments pointed to the relative contribution of both L3T4+ and Lyt-2+ cells, depending on the stage and extent of disease development. Treatment during disease with L3T4, but not Lyt-2, mAb alone significantly reduced thyroiditis. However, in situ analysis of the cellular infiltrate in thyroid sections revealed that, after treatment with mAb, the appropriate subset was eliminated without altering the amount of the other subset in the remaining lesion. In addition, treatment during severe thyroiditis following the transfer of MTg-activated lymph node cells showed that Lyt-2 mAb alone also reduced thyroid infiltration. When the recipients were pretreated with either pair of mAb before transfer, disease development was only moderately affected. We conclude that (i) donor L3T4+ cells are the primary cells responsible for the initial transfer and development of thyroiditis; and (ii) previous in vitro cytotoxicity data, plus current monoclonal antibody treatment of disease and in situ analysis, further implicate a role for Lyt-2+ cells in EAT pathogenesis.