Rapid deletional peripheral CD8 T cell tolerance induced by allogeneic bone marrow: role of donor class II MHC and B cells

Mixed chimerism and donor-specific tolerance are achieved in mice receiving 3 Gy of total body irradiation and anti-CD154 mAb followed by allogeneic bone marrow (BM) transplantation. In this model, recipient CD4 cells are critically important for CD8 tolerance. To evaluate the role of CD4 cells recognizing donor MHC class II directly, we used class II-deficient donor marrow and were not able to achieve chimerism unless recipient CD8 cells were depleted, indicating that directly alloreactive CD4 cells were necessary for CD8 tolerance. To identify the MHC class II(+) donor cells promoting this tolerance, we used donor BM lacking certain cell populations or used positively selected cell populations. Neither donor CD11c(+) dendritic cells, B cells, T cells, nor donor-derived IL-10 were critical for chimerism induction. Purified donor B cells induced early chimerism and donor-specific cell-mediated lympholysis tolerance in both strain combinations tested. In contrast, positively selected CD11b(+) monocytes/myeloid cells did not induce early chimerism in either strain combination. Donor cell preparations containing B cells were able to induce early deletion of donor-reactive TCR-transgenic 2C CD8 T cells, whereas those devoid of B cells had reduced activity. Thus, induction of stable mixed chimerism depends on the expression of MHC class II on the donor marrow, but no requisite donor cell lineage was identified. Donor BM-derived B cells induced early chimerism, donor-specific cell-mediated lympholysis tolerance, and deletion of donor-reactive CD8 T cells, whereas CD11b(+) cells did not. Thus, BM-derived B cells are potent tolerogenic APCs for alloreactive CD8 cells.     

Defective MHC class II presentation by dendritic cells limits CD4 T cell help for antitumor CD8 T cell responses

Cancer immunosurveillance failure is largely attributed to insufficient activation signals and dominant inhibitory stimuli for tumor Ag (TAg)-specific CD8 T cells. CD4 T cells have been shown to license dendritic cells (DC), thereby having the potential for converting CD8 T cell responses from tolerance to activation. To understand the potential cooperation of TAg-specific CD4 and CD8 T cells, we have characterized the responses of naive TCR transgenic CD8 and CD4 T cells to poorly immunogenic murine tumors. We found that whereas CD8 T cells sensed TAg and were tolerized, the CD4 T cells remained ignorant throughout tumor growth and did not provide help. This disparity in responses was due to normal TAg MHC class I cross-presentation by immature CD8alpha+ DC in the draining lymph node, but poor MHC class II presentation on all DC subsets due to selective inhibition by the tumor microenvironment. Thus, these results reveal a novel mechanism of cancer immunosubversion, in which inhibition of MHC-II TAg presentation on DC prevents CD4 T cell priming, thereby blocking any potential for licensing CD8alpha+ DC and helping tolerized CD8 T cells.     

Modulation of CD8+ T cell response to antigen by the levels of self MHC class I

The response of H-Y-specific TCR-transgenic CD8(+) T cells to Ag is characterized by poor proliferation, cytolytic activity, and IFN-gamma secretion. IFN-gamma secretion, but not cytotoxic function, can be rescued by the B7.1 molecule, suggesting that costimulation can selectively enhance some, but not all, effector CD8(+) T cell responses. Although the H-Y epitope binds H-2D(b) relatively less well than some other epitopes, it can induce potent CTL responses in nontransgenic mice, suggesting that the observed poor responsiveness of transgenic CD8(+) T cells cannot be ascribed to the epitope itself. Previously reported reactivity of this TCR to H-2A(b) is also not the cause of the poor responsiveness of the H-Y-specific CD8(+) T cells, as H-Y-specific CD8(+) T cells obtained from genetic backgrounds lacking H-2A(b) also responded poorly. Rather, reducing the levels of H-2(b) class I molecules by breeding the mice to (C57BL/6 x B10.D2)F(1) or TAP1(+/-) backgrounds partially restored cytotoxic activity and enhanced proliferative responses. These findings demonstrate that the self MHC class I gene dosage may regulate the extent of CD8(+) T cell responsiveness to Ag.     

In vivo functional clonal deletion of recipient CD4+ T helper precursor cells that can recognize class II MHC on injected donor lymphoid cells.

Intravenous injection of semiallogeneic (C57BL/6XDBA/2)F1 lymphocytes into adult C57BL/6 recipient mice not only, as previously reported, reduces the recipients' cytotoxic T lymphocyte response in a subsequent in vitro mixed lymphocyte reaction against the injected cell type, but also reduces Th cell function in the same MLR. Thus lymphoid cells derived from the injected mice were greatly reduced in their ability to proliferate and to produce IL-2 in response to (C57BL/6XDBA/2)F1 stimulator cells in vitro, whereas third party responses were unaffected. This appears to be due to a reduction in the precursor frequency of IL-2-producing T lymphocytes specific for the injected cells as measured by limiting dilution analysis. Similar donor-specific reduction in the frequency of precursors of IL-2-producing cells was seen after i.v. injection of A.TL lymphocytes into A.TH recipients (differing at class II determinants I-A and I-E, but identical at K and D). Here there also appeared to be a functional clonal deletion of precursors of IL-2-producing Th cells, shown directly to be class II MHC reactive and CD4+. There is strong evidence that the reduction of class I-specific cytotoxic responses in the injected mice is a manifestation of donor cells that function as veto cells, i.e., that function as deletional APC that inactivate class I-reactive CTL precursors that recognize them. Our data in this study show that class II-specific Th responses are similarly reduced in the injected mice and suggest that CD4+ class II-reactive precursors of Th cells may be functionally inactivated in vivo by donor cells via a veto-like mechanism.     

Activation of human T cell clones and Jurkat cells by cross-linking class I MHC molecules

Cross-linking class I MHC molecules on human T cell clones by reacting them with various mAb directed at either monomorphic or polymorphic determinants on class I MHC molecules followed by cross-linking with GaMIg stimulated a rise in intracellular free calcium concentration ([Ca2+]i), and induced proliferation and IL-2 production. T cell clones varied in the mean density of class I MHC molecules and the capacity to respond to mAb to class I MHC molecules. However, the functional responses of the clones did not correlate with class I MHC density or the CD4/CD8 phenotype. mAb to polymorphic class I MHC determinants were less able to induce an increase in [Ca2+]i and a functional response in the T cell clones. Additive stimulatory effects were noted when mAb against both HLA-A and HLA-B determinants were employed. Cross-linking class I MHC molecules on Jurkat cells induced a rise by [Ca2+]i and induced IL-2 production upon co-stimulation with PMA. Cross-linking class I MHC molecules on mutant Jurkat cells that expressed diminished levels of CD3 and were unable to produce IL-2 in response to anti-CD3 stimulation triggered both a rise in [Ca2+]i and IL-2 production with PMA co-stimulation. In contrast, cross-linking class I MHC molecules on mutant Jurkat cells that were CD3- stimulated neither a rise in [Ca2+]i nor IL-2 production. The combination of mAb to CD28 or ionomycin and PMA, however, was able to induce IL-2 production by CD3- Jurkat cells. The data demonstrate that cross-linking class I MHC molecules delivers a functionally important signal to T cell clones and Jurkat cells and indicate that class I MHC molecules may function to transduce activation signals to T cells. In addition, the data demonstrate that transmission of an activation signal via class I MHC molecules requires CD3 expression. The data, therefore, support a central role for CD3 in the transduction of activation signals to T cells via class I MHC molecules.     

Induction of linked suppression in addition to the donor H-2 class I-specific unresponsiveness in recipient T cells by transfusing class I plus class II-disparate, but not class I alone-disparate, bone marrow cells

This study was undertaken to determine whether bone marrow (BM) cells contain a cell population with the capacity to induce an unresponsiveness of T cells specific to the BM self-H-2 class I antigens in vivo, i.e., veto cell population. Recombinant or congenic mice were infused intravenously with H-2-incompatible BM cells. One to several weeks later, donor H-2-and irrelevant H-2-specific responses in mixed lymphocyte reaction cultures of recipient T cells were assessed. Transfusion of H-2-incompatible BM of C57BL/10 (B10) recombinant strains caused a long-lasting cytotoxic T lymphocyte (CTL) unresponsiveness to the donor class I antigens in recipient lymph node cells. When class I plus class II-disparate BM cells were transfused, an anti-donor class I CTL response and a response against a third-party class I antigen, which was presented on the stimulator cells coexpressing the donor class I and class II, were significantly suppressed. This linked suppression lasted for less than 2 weeks after transfusion. Transfusion of class I-alone-disparate BM induced the donor class I-specific CTL unresponsiveness, but not the linked suppression. The induction of linked suppression was prevented considerably by transfusing nylon wool-nonadherent BM or by treating recipients with cyclophosphamide 2 days before transfusion. An anti-third-party class I CTL response, stimulated in vitro with fully allogeneic spleen cells, was not hampered by the BM transfusion. Coculturing the lymph node (LN) cells obtained from the class I plus class II-disparate BM recipient with normal LN cells interfered with the generation of both anti-donor class I and anti-linked third-party class I CTL, whereas, coculturing LN cells from the class I alone-disparate BM recipient inhibited neither specificity of CTL generation. Transfusion of class I plus class II-disparate BM resulted in a significant suppression of the donor class II-specific proliferative response. In contrast, transfusion of class I alone-disparate BM did not suppress any proliferative responses, including even a "linked" third-party class II-specific response. Transfusion of bm 1, (B6 X bm 1)F1, or (bm 1 X bm 12)F1 BM to B6 did not induce unresponsiveness in bm 1-specific CTL responses. However, the transfusion resulted in a significant suppression of bm 1-reactive proliferative response of recipient LN cells.(ABSTRACT TRUNCATED AT 400 WORDS)     

T cell activation. II. Activation of human T lymphoma cells by cross-linking of their MHC class I antigens

The present work demonstrates that antibody-induced cross-linking of MHC class I antigens on Jurkat T lymphoma cells leads to a rise in intracellular calcium (Cai2+) and, in the presence of phorbol ester (PMA), to IL-2 production and IL-2 receptor expression. The rise in Cai2+ exhibited a profile very different from that obtained after anti-CD3 antibody-induced activation suggesting that activation signals are transduced differently after binding of anti-CD3 antibody and class I cross-linking, respectively. However, when Cai2+ was examined in individual Jurkat cells by means of a digital image processing system no differences were observed after cross-linking with anti-CD3 and anti-MHC class I antibodies, respectively. Two CD3-negative mutant lymphoma lines were nearly totally refractory to class I cross-linking. Taken together our results may indicate the existence of a functional linkage between the T cell receptor complex and MHC class I molecules.     

CD27 instructs CD4+ T cells to provide help for the memory CD8+ T cell response after protein immunization

For optimal quality, memory CD8(+) T cells require CD4(+) T cell help. We have examined whether CD4(+) T cells require CD27 to deliver this help, in a model of intranasal OVA protein immunization. CD27 deficiency reduced the capacity of CD4(+) T cells to support Ag-specific CD8(+) T cell accumulation at the tissue site after primary and secondary immunization. CD27-dependent CD4(+) T cell help for the memory CD8(+) T cell response was delivered during priming. It did not detectably affect formation of CD8(+) memory T cells, but promoted their secondary expansion. CD27 improved survival of primed CD4(+) T cells, but its contribution to the memory CD8(+) T cell response relied on altered CD4(+) T cell quality rather than quantity. CD27 induced a Th1-diagnostic gene expression profile in CD4(+) T cells, which included the membrane molecule MS4A4B. Accordingly, CD27 increased the frequency of IFN-gamma- and IL-2-producing CD4(+) T cells. It did not affect CD40L expression. Strikingly, MS4A4B was also identified as a unique marker of CD8(+) memory T cells that had received CD27-proficient CD4(+) T cell help during the primary response. This apparent imprinting effect suggests a role for MS4A4B as a downstream effector in CD27-dependent help for CD8(+) T cell memory.     

Cutting edge: single-chain trimers of MHC class I molecules form stable structures that potently stimulate antigen-specific T cells and B cells

We report in this work the expression and characterization of class I molecules expressed as single-chain trimers consisting of an antigenic peptide-spacer-beta(2)-microglobulin-spacer H chain. Our results indicate that these single-chain constructs assemble efficiently, maintain their covalent structure, and are unusually stable at the cell surface. Consequently, these constructs are at least 1000-fold less accessible to exogenous peptide than class I molecules loaded with endogenous peptides, and they are potent simulators of peptide-specific CTL and Abs. Our combined findings suggest that single-chain trimers may have applications as DNA vaccines against virus infection or tumors.     

Genetic control of the immune response to ferredoxin: linkage and mapping of T cell proliferation and antibody production genes to the MHC of mice

The genetics of the immune response in the mouse were studied by using the antigenically simple, stable, naturally occurring protein ferredoxin (Fd) from Clostridium pasteurianum. The immune status of mice primed and boosted with Fd was assessed by using two parameters of immunity: T cell proliferation and serum antibody production with the ELISA method. In both assay systems, the response has been shown to be H-2 linked: k, b, and s haplotypes respond to Fd, and H-2d mice are nonresponders. It is apparent that different immunoregulatory events modulate the response in the responder strains; these factors become evident in the recombinant analysis of the response and to date an immunoregulatory gene(s) has been mapped to at least the K/I-A subregions. F1 analysis demonstrated a gene dose-dependent response of the strains studied.     

Molecular cloning of cDNA that encode MHC class I molecules from a New World primate (Saguinus oedipus). Natural selection acts at positions that may affect peptide presentation to T cells

To investigate the evolutionary pressures that drive the generation of polymorphism in primate MHC class I molecules, three cDNA that encode MHC class I alleles from a New World monkey, the cotton-top tamarin (Saguinus oedipus), were cloned and sequenced. These tamarin MHC class I alleles contained amino acid substitutions not found in any of the previously sequenced human MHC class I alleles. Moreover, the majority of these unique amino acid substitutions was located in the Ag recognition site at positions that have been shown to be critical in the presentation of viral peptides to T cells in mice and humans. These data suggest that selective pressures on MHC class I molecules preferentially act on the Ag recognition site and that the peptide binding or presenting functions of these molecules may drive the generation of MHC class I polymorphism. The novel Ag recognition sites of the tamarin MHC class I molecules, in addition to their restricted polymorphism, might account for the unusual susceptibility of the cotton-top tamarin to human pathogens.     

Inhibition by brefeldin A of the specific B cell antigen presentation to MHC class II-restricted T cells

We have shown previously that specific Ag presentation is prevented by the inhibition of protein synthesis but nonspecific presentation is not. In the present paper, Ag presentation by Ag-specific B cells was examined for sensitivity to brefeldin A (BFA), which blocks protein export from the endoplasmic reticulum. A20-HL B lymphoma expressing surface receptors specific for TNP was used as a B cell, and TNP-OVA was used as a specific Ag. The presence of BFA during pulsing of A20-HL cells with TNP-OVA inhibited the ability of the pulsed cells to stimulate 42-6A T cell clone, specific for OVA323-339 and Iad. The inhibition was not due to nonspecific toxicity of BFA, because the presence of BFA during pulsing of A20-HL cells with OVA323-339 did not affect their APC function. Ag binding to the receptor on A20-HL cells and internalization by the cells were observed in the presence of BFA. Thus, BFA might inhibit intracellular processing of specific Ag or intracellular complex formation of antigenic peptide from specific Ag with MHC class II molecules. Nonspecific Ag presentation by A20-HL cells, however, was resistant to BFA. A20-HL cells pulsed with OVA in the presence of BFA, even after fixation, could stimulate 42-6A cells to produce IL-2, although the IL-2 production was lower than that induced by A20-HL cells pulsed in the absence of BFA. These results suggest that the processing pathways for specific Ag and nonspecific Ag are different from each other, at least partly, in A20-HL cells.     

Evidence that multiple residues on both the alpha-helices of the class I MHC molecule are simultaneously recognized by the T cell receptor

Single amino acid substitutions at nine different positions on the H-2Kb molecules from in vitro-mutagenized, immunologically altered, somatic cell variants were correlated with their patterns of recognition by monoclonal antibodies (MAbs) and allogeneic cytotoxic T lymphocyte (CTL) clones. While MAbs were found to detect spatially discrete, domain-specific sites, CTLs interacted simultaneously with multiple residues on the alpha 1 and alpha 2 domains of the Kb molecule. The computer graphic three-dimensional Kb model structure showed that, of the seven CTL-specific residues analyzed, six residues were located on the alpha-helical regions of the two domains. Every CTL clone was found to interact with a distinct pattern of residues composed of a specific subset of the CTL-specific residues.     

The regulatory role of sialic acids in the response of class II reactive T cell hybridomas to allogeneic B cells

Two different kinds of alloreactive T cell hybridomas were established in previous experiments. One is reactive and the other is nonreactive to allogeneic I-A region-associated membrane antigen (mIa) on B cells. In the present experiments the difference between these hybridomas were analyzed by using representative clones, B cell mIa-reactive clone CB-11.4, and nonreactive clone HTB-9.3. Unresponsiveness of HTB-9.3 clone to allogeneic B cells could not be due to the inability of B cells in interleukin 1 production or the density of mIa molecules on B cells. HTB-9.3 clone could respond to C57BL/6 mouse B cells treated with neuraminidase (Nase), and Nase-treated HTB-9.3 clone could respond to normal B cells from C57BL/6 mouse, indicating that sialic acid on both B cells and HTB-9.3 clone plays a regulatory role in the alloreactivity of the clone. In response to B cells from C57BL/6 mouse, T cells from C3H/He mouse spleen showed similar reactivity to HTB-9.3 clone; that is, T cells could respond to Nase-treated B cells, and Nase-treated T cells to B cells, and T cells primed with C57BL/6 spleen cells in vitro showed similar reactivity to CB-11.4 clone. These results suggest that HTB-9.3 clone represents virgin T cells and CB-11.4 clone-primed T cells at least in alloreactivity. Anti-L3T4a was shown to block alloreactivities of both T cell hybridomas and splenic T cells against B cells more efficiently than against splenic adherent cells. These results suggest that L3T4a on T cell plays more important role in allogeneic response to B cells than to splenic adherent cells.     

CD8 T cell-mediated lung damage in response to the extracellular pathogen pneumocystis is dependent on MHC class I expression by radiation-resistant lung cells

Pneumocystis, a fungal, extracellular pathogen causes a life-threatening pneumonia in patients with severe immunodeficiencies. In the absence of CD4 T cells, Pneumocystis infection results in vigorous CD8 T cell influx into the alveolar and interstitial spaces of the lung. This response results in lung damage characterized by low pO2 and albumin leakage into the bronchoalveolar lavage fluid similar to other CD8 T cell-mediated interstitial lung diseases. How this extracellular pathogen elicits a CD8 T cell response is not clear, and it was the aim of our study to determine the Ag specificity of the recruited CD8 T cells and to determine whether MHC class I (MHC I) expression was necessary to initiate lung damage. Using an adoptive T cell-transfer model with either polyclonal wild-type CD8 T cells or transgenic influenza virus-specific CD8 T cells we found that CD8 T cell recruitment is Ag-specific and requires the continuous presence of the Pneumocystis pathogen. Bone marrow chimera experiments using Rag-1 and beta2-microglobulin-deficient mice as hosts demonstrated a requirement for MHC I expression on nonbone marrow-derived cells of the lung. This suggests either direct processing of Pneumocystis Ags by nonbone marrow-derived cells of the lung or the induction of lung damage triggered by a lung-specific autoantigen. Using perforin-, Fas-, and IFN-gamma-deficient animals, we showed that these molecules are not directly involved in the CD8-mediated lung damage. However, CD8 T cell-mediated lung damage is Ag-specific is induced by a MHC I-expressing nonbone marrow-derived cell in the lung and is dependent on the continued presence of live Pneumocystis.     

Sufficiency of the CD8+ T cell lineage to mount an effective tumoricidal response to syngeneic tumor-bearing novel class I MHC antigens

The origins of "help" in rejection of syngeneic tumors by the CD8 T cell lineage was examined with a model tumor inappropriately expressing novel class I MHC and subject to cytolytic T cell (CTL)-mediated rejection. The requirement for CD4+ Th cells to induce CD8+ CTL effectors in vivo was investigated by using C3H mice selectively depleted of either CD4+ or CD8+ T cells. Rejection of the tumor was vigorous and indistinguishable from normal mice after depletion of CD4+ T cells in vivo. In contrast, in CD8+ T cell-depleted mice tumors grew progressively, confirming that T cells of the CD8+ lineage are required for a tumoricidal immune response, and cells of this lineage are sufficient for a primary response. Taken together, these results demonstrate that, in the absence of CD4+ T cells in vivo, unprimed cells of the CD8+ lineage are fully competent to mount an effective CTL immune response to syngeneic cells expressing novel class I Ag, consistent with the concept that only T cells with class I recognition specificity may be required to satisfy the need for both help and effector functions in the response.     

Soluble class I MHC with beta2-microglobulin covalently linked peptides: specific binding to a T cell hybridoma

Soluble forms of the mouse MHC class I molecule, Dd, were produced in which the peptide binding groove was uniformly occupied by peptides attached via a covalent flexible peptide linker to the N terminus of the associated beta2-microglobulin. The MHC heavy chain and beta2-microglobulin were firmly associated, and the molecules displayed an Ab epitope requiring proper occupancy of the peptide binding groove. Soluble Dd containing a covalent version of a well-characterized Dd-binding peptide from HIV stimulated a T cell hybridoma specific for this combination. Furthermore, a tetravalent version of this molecule bound specifically with apparent high avidity to this hybridoma.     

Cell-sized, supported artificial membranes (pseudocytes): response of precursor cytotoxic T lymphocytes to class I MHC proteins

Novel cell-sized, supported artificial membranes bearing class I antigens have been prepared by a simple dialysis procedure and then used to study the requirements for antigen recognition by precursor cytotoxic T lymphocytes (CTL). The membranes were made by mixing lipid, H-2 antigen, and C18 alkylated 5 microns silica beads in deoxycholate, and dialyzing to remove the detergent. The H-2 antigen-bearing, cell-sized beads, termed pseudocytes (artificial cells), were able to simulate generation of secondary CTL responses with the same specificity as alloantigen-bearing spleen cells. Comparative analyses demonstrated that the size of an antigen-bearing structure, and thus its potential for multivalent interaction, was a critical determinant of effectiveness of antigen recognition, and showed that H-2 antigen was recognized as effectively on cell-sized beads as on allogeneic spleen cells. Generation of a response to antigen on the cell-sized beads was completely dependent on addition of lymphokines to the cultures. Thus, unlike liposomes, H-2 antigen on beads was not available to accessory cells for stimulation of Ia-dependent production of lymphokines by T helper cells. These results, as well as direct observations by microscopy, strongly indicate that antigen is recognized on the surface of the beads. Despite effective stimulation of secondary CTL responses, antigen on beads was completely inactive in stimulating a primary CTL response by naive spleen cells. The results of mixing experiments by using beads and alloantigen-bearing cells or plasma membrane vesicles indicate that the lack of a primary response may result from a requirement for a soluble factor(s) that is not needed for generation of secondary responses. The unique advantages of cell-size supported membranes for studying antigen recognition by T cells are discussed. The beads can be handled and used like antigen-bearing cells in functional assays, while possessing well-defined, readily varied, and easily quantitated composition.     

Rejection of allogeneic tumor is not determined by host responses to MHC class I molecules and is mediated by CD4-CD8+ T lymphocytes that are not lytic for the tumor

In previous studies, the murine SaI (A/J derived, KkDd) sarcoma was transfected with the allogeneic MHC class I H-2Kb gene, and expressed high levels of H-2Kb antigen. Contrary to expectations, the tumor cells expressing the alloantigen (SKB3.1M tumor cells) were not rejected by autologous A/J mice. Because these results contradict the laws of transplantation immunology, the present studies were undertaken to examine the immunogenicity of SKB3.1M and SaI cells in allogeneic hosts. Similar to SKB3.1M, SaI cells are lethal in some allogeneic strains, despite tumor-host MHC class I incompatibilities. Tumor challenges of SKB3.1M and SaI cells, however induce MHC class I-specific antibodies and CTL in both tumor-resistant and -susceptible hosts. Although the tumors induce specific CTL, tumor cells are not lysed in vitro by these CTL, suggesting that the tumor cells are resistant to CTL-mediated lysis. Since growth of these tumors does not follow the classical rules of allograft transplantation, and because the tumor is not susceptible to CTL-mediated lysis, we have used Winn assays to identify the effector lymphocyte(s) responsible for SaI rejection. Depletion studies demonstrate that the effector cell is a CD4-CD8+ T lymphocyte. Collectively these studies suggest that the host's response to MHC class I alloantigens of SKB3.1M and SaI cells does not determine tumor rejection, and that effector cells other than classically defined CTL, but with the CD4-CD8+ phenotype, can mediate tumor-specific immunity.     

Effects of monoclonal antibodies directed against murine T lymphocyte cell surface antigens on lymphokine production by cloned T lymphocytes reactive with class I MHC or Mls alloantigens

Monoclonal antibodies recognizing murine T lymphocyte cell surface structures implicated in T lymphocyte-mediated cytolysis, including Lyt-2, L3T4, LFA-1, and a cytolytic T lymphocyte (CTL) clonotypic determinant, were used as probes to investigate the role of these structures in lymphokine production by T cell clones induced by antigen or lectin. The clone-specific antibody 384.5 bound to and inhibited antigen-induced lymphokine production by the L3 CTL clone, but did not affect lymphokine production by other T cell clones. Antibodies against the T cell surface structures Lyt-2 or L3T4, which are expressed by mutually exclusive T cell subsets, inhibited antigen-induced lymphokine production by class I major histocompatibility complex (MHC) antigen-reactive CTL clones or an M1s-reactive helper T lymphocyte (HTL) clone, respectively. Antibody against the broadly distributed LFA-1 molecule inhibited antigen-induced lymphokine production by all of the clones tested. Lectin-induced lymphokine production by cloned T cells was not inhibited by the clonotypic antibody, anti-Lyt-2, or anti-LFA-1; slight inhibition of the HTL clone was observed with the anti-L3T4 antibody. None of these structures appear to be uniquely involved with a particular functional response. Our results suggest that each of these structures is involved with the interactions between the effector cell and the stimulating cell leading to lymphokine production.