Antigen-presenting cell-T cell interaction in the chicken is MHC class II antigen restricted

The involvement of the MHC in the recognition of Ag by avian T lymphocytes was analyzed. PBL from chickens primed with keyhole limpet hemocyanin in vivo were induced to synthesize DNA in an in vitro response to specific Ag. Responding cells were T cells as judged by immunofluorescence staining. In vivo Ag-primed PBL were stimulated in vitro with specific Ag and further propagated in the presence of IL-2. Subsequent Ag-specific T cell proliferation required the presence of Ag-pulsed peripheral blood adherent cells (APC). T cell responses were restricted by the MHC of the APC; Ag presented by allogeneic APC did not support T cell proliferation. By using MHC-recombinant chicken lines, the gene products controlled by MHC class II loci were shown to restrict the T cell-APC interaction. This conclusion was substantiated by the inhibition of the Ag-specific T cell response by a mAb against chicken MHC class II gene products but not by a mAb against chicken MHC class I gene products.     

Activation of human T4 cells by cross-linking class I MHC molecules

These studies examined whether cross-linking class I MHC molecules results in functional or biochemical responses in human T4 cells. The initial studies demonstrated that cross-linking class I MHC molecules either by culturing highly purified T4 cells with immobilized mAb to class I MHC Ag or reacting the T4 cells with mAb to class I MHC Ag and then cross-linking the mAb with goat antimouse Ig (GaMIg) enhanced T4 cell proliferation induced by an immobilized mAb to CD3, OKT3. More-over, immobilized but not soluble mAb to class I MHC Ag enhanced T4 cell proliferation induced by the combination of two mAb to CD2, OKT11, and D66.2. Finally, T4 cells reacted with mAb to CD3 and class I MHC Ag proliferated in the presence of IL-2 when cross-linked with GaMIg more vigorously than T4 cells reacted with either mAb alone. Cross-linking class I MHC molecules was also found to stimulate T4 cells directly. T4 cells reacted with mAb to class I MHC Ag or beta 2 microglobulin and cross-linked with GaMIg proliferated vigorously in the presence of IL-2 or PMA. In addition, it was demonstrated that cross-linking class I MHC molecules by culturing T4 cells with immobilized mAb to class I MHC Ag induced T4 cell proliferation in the presence of IL-2. T4 cell proliferation in the presence of IL-2 and PMA could also be induced by reacting the cells with specific mAb to polymorphic determinants on class I MHC molecules and cross-linking with GaMIg. Cross-linking mAb to CD4 or CD11a did not have a similar functional effect on T4 cells. Finally it was demonstrated that adding GaMIg to T4 cells reacted with mAb to class I MHC Ag but not CD11a resulted in an increase in intracellular calcium concentration. The data demonstrate that cross-linking class I MHC molecules results in the generation of at least one activation signal, a rise in intracellular calcium concentration, and, thereby, stimulates human T4 cells.     

Class II MHC/peptide complexes are released from APC and are acquired by T cell responders during specific antigen recognition.

T cell expression of class II MHC/peptide complexes may be important for maintenance of peripheral self-tolerance, but mechanisms underlying the genesis of class II MHC glycoproteins on T cells are not well resolved. T cell APC (T-APC) used herein were transformed IL-2-dependent clones that constitutively synthesized class II MHC glycoproteins. When pulsed with myelin basic protein (MBP) and injected into Lewis rats, these T-APC reduced the severity of experimental autoimmune encephalomyelitis, whereas unpulsed T-APC were without activity. Normal MBP-reactive clones cultured without APC did not express class II MHC even when activated with mitogens and exposed to IFN-gamma. However, during a 4-h culture with T-APC or macrophage APC, recognition of MBP or mitogenic activation of responder T cells elicited high levels of I-A and I-E expression on responders. Acquisition of class II MHC glycoproteins by responders was resistant to the protein synthesis inhibitor cycloheximide, coincided with transfer of a PKH26 lipophilic dye from APC to responders, and resulted in the expression of syngeneic and allogeneic MHC glycoproteins on responders. Unlike rested I-A- T cell clones, rat thymic and splenic T cells expressed readily detectable levels of class II MHC glycoproteins. When preactivated with mitogens, naive T cells acquired APC-derived MHC class II molecules and other membrane-associated proteins when cultured with xenogeneic APC in the absence of Ag. In conclusion, this study provides evidence that APC donate membrane-bound peptide/MHC complexes to Ag-specific T cell responders by a mechanism associated with the induction of tolerance.     

Proteolytically modified human beta 2-microglobulin augments the specific cytotoxic activity in murine mixed lymphocyte culture

A proteolytically modified form of beta 2-microglobulin (beta 2-m) present in the serum of patients suffering from autoimmune, immunodeficient diseases and cancer has been reported in the literature. In the present study we show that human beta 2-m as well as the proteolytically modified human form (M-beta 2-m) bind to murine lymphocytes expressing H-2 class I antigens; M-beta 2-m, when added at day 0 and 1 of culture in nanomolar concentrations to a one-way murine allogeneic mixed lymphocyte culture (MLC) augments the generation of specific cytotoxic T lymphocytes; M-beta 2-m increases the endogenous production of interleukin 2 in the MLC culture; monoclonal antibody which reacts with both the native beta 2-m and M-beta 2-m molecule blocks the augmentation of cytotoxic T lymphocyte production induced by M-beta 2-m; murine as well as human MLC responder cells can proteolytically modify native human beta 2-m; and the modifying activity of murine MLC responder cells was blocked in an intermediary step by an alloantibody, which reacts specifically with murine major histocompatibility complex, class I-associated beta 2-m. These findings suggest that the modification process is preceded by an association of human beta 2-m with the cell surface of the responder cells. Our data indicate that the modification of beta 2-m might reflect early events in allospecific responder cell activation.     

Glomerular mesangial cells in local inflammation. Induction of the expression of MHC class II antigens by IFN-gamma

Glomerular mesangial cells (MC) were isolated from rats and cultured for a prolonged period of time, resulting in a homogeneous cell population. MC were characterized as belonging to the smooth muscle type. They were negative for MHC class II expression. IFN-gamma and TNF alpha suppressed the proliferation of MC, demonstrating receptors for these cytokines on MC. IFN-gamma or TNF alpha, respectively, enhanced basal MHC class I Ag expression of proliferating cells in culture. The combination of the two cytokines yielded stronger effects. IL-1 beta was ineffective in enhancing MHC class I Ag expression, although MC possessed receptors for this cytokine. IFN-gamma dose dependently induced the expression of MHC class II Ag, while TNF alpha or IL-1 beta were ineffective alone. The combination of IFN-gamma with TNF alpha or IL-1 beta resulted in an enhanced induction of MHC class II Ag, compared to IFN-gamma administration alone. These findings suggest that proliferating mesangial cells of the smooth muscle type may participate in local inflammatory responses or substitute for macrophages by meeting the accessory cell requirement in the interaction with T lymphocytes. Furthermore, the data have important implications for the evaluation of the role of mesangial cells in autoimmune disease of the kidney.     

Some cloned murine CD4+ T cells recognize H-2Ld class I MHC determinants directly. Other cloned CD4+ T cells recognize H-2Ld class I MHC determinants in the context of class II MHC molecules.

Murine T lymphocytes recognize nominal Ag presented by class I or class II MHC molecules. Most CD8+ T cells recognize Ag presented in the context of class I molecules, whereas most CD4+ cells recognize Ag associated with class II molecules. However, it has been shown that a proportion of T cells recognizing class I alloantigens express CD4 surface molecules. Furthermore, CD4+ T cells are sufficient for the rejection of H-2Kbm10 and H-2Kbm11 class I disparate skin grafts. It has been suggested that the CD4 component of an anti-class I response can be ascribed to T cells recognizing class I determinants in the context of class II MHC products. To examine the specificity and effector functions of class I-specific HTL, CD4+ T cells were stimulated with APC that differed from them at a class I locus. Specifically, a MLC was prepared involving an allogeneic difference only at the Ld region. CD4+ clones were derived by limiting dilution of bulk MLC cells. Two clones have been studied in detail. The CD4+ clone 46.2 produced IL-2, IL-3, and IFN-gamma when stimulated with anti-CD3 mAb, whereas the CD4+ clone 93.1 secreted IL-4 in addition to IL-2, IL-3, and IFN-gamma. Cloned 46.2 cells recognized H-2Ld directly, whereas recognition of Ld by 93.1 apparently was restricted by class II MHC molecules. Furthermore, cytolysis by both clones 46.2 and 93.1 was inhibited by the anti-CD4 mAb GK1.5. These results demonstrate that CD4+ T cells can respond to a class I difference and that a proportion of CD4+ T cells can recognize class I MHC determinants directly as well as in the context of class II MHC molecules.     

Inhibition by anti-HLA class II monoclonal antibodies of monoclonal antibody OKT3-induced T cell proliferation. Studies at the mRNA level

mAb to monomorphic determinants of HLA class II Ag have been shown to inhibit monocyte-dependent OKT3-induced T cell proliferation, indicating that MHC class II molecules play a regulatory role also in Ag nonrestricted, CD3-induced T cell proliferation. This effect involves several steps in the process of T cell activation and proliferation, including IL-1 beta, IL-6, and IL-2 secretion and IL-2R alpha expression. In the present study, we analyzed the effect of an anti-HLA class II mAb (Q5/6) on the mRNA expression of genes related to monocyte and T cell activation. mRNA levels for early (early c-myc, c-fos) and late (late c-myc, N-ras, c-myb) genes involved in T cell activation were determined as well as mRNA levels for IL-1 beta, IL-6, IFN-gamma, IL-2, and IL-2R alpha. The kinetics of mRNA induction for ICAM-1 was also investigated. The results show that in T lymphocytes the expression of c-fos and early c-myc mRNA was unaffected by mAb Q5/6, whereas the c-myb and N-ras mRNA levels were strongly diminished as well as those of IL-2, IL-2R alpha, and IFN-gamma mRNA. An early increase of ICAM-1 mRNA was partially inhibited. In monocytes, a marked reduction of IL-1 beta and IL-6 mRNA was found. It is concluded that the HLA class II determinant involved in the inhibition mechanism can be engaged in the control of IL-1 beta and IL-6 mRNA levels and constitute an accessory signal up-regulating IL-2 and IL-2R alpha gene activation, through a pathway not affecting c-myc and c-fos expression.     

Heterologous cross-linking of Lyt-2 (CD8) to the alpha beta-T cell receptor is more effective in T cell activation than homologous alpha beta-T cell receptor cross-linking

Ag recognition of Lyt-2 (CD8)-positive T lymphocytes requires the presentation by APC of a suitably processed Ag in association with MHC class I molecules. In previous studies we have obtained evidence that, for optimal activation, both the alpha beta-TCR and Lyt-2 have to participate in this recognition process. In the current study we investigate the functional consequences of limited cross-linking of these cell surface molecules by using soluble, dimeric hetero- and homoconjugates of mAb to Lyt-2 and to the TCR beta-chain (F23.1). Heterologous cross-linking of Lyt-2 to the TCR induced a vigorous, selective Lyt-2+ T cell proliferative response. Functionally active cytotoxic cells were generated, and a high frequency of responding cells was observed in limiting dilution analyses. In contrast, homologous TCR cross-linking initiated a less pronounced proliferation with a relatively low frequency of response, whereas Lyt-2 cross-linking resulted in no cellular proliferation. Significant T cell activation occurred with exposure to anti-Lyt-2: F23.1 mAb dimers at concentrations an order of magnitude lower than those required for stimulation by F23.1:F23.1 mAb dimers. The induction of proliferation by mAb dimers occurred in the absence of Fc components and in rigorously APC depleted, purified T cell preparations. Effective stimulation of resting T cells could be induced also by heterodimers of monovalent Fab fragments. Heterologous cross-linking of Lyt-2 to the TCR was superior to homologous TCR cross-linking primarily with respect to proliferation in IL-2 containing media and to IL-2R expression, whereas proliferation in response to other lymphokines and the production of IL-2 itself were similar under both cross-linking regimens. Thus, when linked to the TCR, Lyt-2 contributed a strong, positive signal toward IL-2-dependent growth of resting T cells. We assume that in the case of Ag-driven T cell activation, the class I MHC molecule acts as the physiologic cross-linking ligand for Lyt-2 and the TCR.     

Inhibition of graft-versus-host disease by double-negative regulatory T cells

Pretransplant infusion of lymphocytes that express a single allogeneic MHC class I Ag has been shown to induce tolerance to skin and heart allografts that express the same alloantigens. In this study, we demonstrate that reconstitution of immunoincompetent mice with spleen cells from MHC class I L(d)-mismatched donors does not cause graft-vs-host disease (GVHD). Recipient mice become tolerant to skin allografts of lymphocyte donor origin while retaining immunity to third-party alloantigens. The mechanism involves donor-derived CD3(+)CD4(-)CD8(-) double-negative T regulatory (DN Treg) cells, which greatly increase and form the majority of T lymphocytes in the spleen of recipient mice. DN Treg cells isolated from tolerant recipient mice can suppress the proliferation of syngeneic antihost CD8(+) T cells in vitro. Furthermore, we demonstrate that DN Treg cells can be generated in vitro by stimulating them with MHC class I L(d)-mismatched lymphocytes. These in vitro generated L(d)-specific DN Treg cells are able to down-regulate the activity of antihost CD8(+) T cells in vitro by directly killing activated CD8(+) T cells. Moreover, infusing in vitro generated L(d)-mismatched DN Treg cells prevented the development of GVHD caused by allogeneic CD8(+) T cells. Together these data demonstrate that infusion of single MHC class I locus-mismatched lymphocytes may induce donor-specific transplantation tolerance through activation of DN Treg cells, which can suppress antihost CD8(+) T cells and prevent the development of GVHD. This finding indicates that using single class I locus-mismatched grafts may be a viable alternative to using fully matched grafts in bone marrow transplantation.     

Suppression of lymphocyte alloreactivity by early gestational human decidua. II. Characterization of the suppressor mechanisms

We have earlier shown that first trimester human decidual cells (typical decidual cells and decidual macrophages) suppress lymphocyte alloreactivity (MLR and CTL generation) in vitro in an MHC-unrestricted manner and that this suppression is mediated by PGE2. The present study explored the mechanisms underlying this suppression by noting the effects of decidual cells (+/- indomethacin or anti-PGE2 antibody) or chemically pure PGE2 on numerous T lymphocyte activation events following allogeneic stimulation in mixed lymphocyte culture (MLC) or polyclonal activation with Con A. The results revealed that this suppression was the net result of an action of PGE2 on at least two events during lymphocyte activation: (i) a down-regulation of IL-2 receptor development on lymphocytes, quantitated with a radioimmunoassay and radioautography; this was noted in MLC or Con A-stimulated lymphocyte cultures in the presence of decidual cells (reversible in the presence of indomethacin or anti-PGE2 antibody), or PGE2, but not PGF2 alpha; (ii) an inhibition of IL-2 production in the MLC, measured with a bioassay using an IL-2-dependent T cell line (CTLL-2) and a recombinant IL-2 standard. These effects blocked cell proliferation and eventual generation of killer cells in the MLC. Decidual cells or PGE2 did not interfere with IL-2-dependent proliferation of CTLL-2 cells, which require an interaction between IL-2 receptors on these cells and IL-2. Finally, neither agent interfered with the lytic function of CTL, once generated. These results indicate that the PGE2-mediated immunosuppressor function of early gestational human decidual cells is accomplished by an afferent blockade of the early events in T lymphocyte activation.     

Negligible Immunogenicity of Induced Pluripotent Stem Cells Derived from Human Skin Fibroblasts

Human induced pluripotent stem cells (hiPSCs) have potential applications in cell replacement therapy and regenerative medicine. However, limited information is available regarding the immunologic features of iPSCs. In this study, expression of MHC and T cell co-stimulatory molecules in hiPSCs, and the effects on activation, proliferation and cytokine production in allogeneic human peripheral blood mononuclear cells were examined. We found that no-integrate hiPSCs had no MHC-II and T cell co-stimulatory molecules expressions but had moderate level of MHC-I and HLA-G expressions. In contrast to human skin fibroblasts (HSFs) which significantly induced allogeneic T cell activation and proliferation, hiPSCs failed to induce allogeneic CD45⁺ lymphocyte and CD8⁺ T cell activation and proliferation but could induce a low level of allogeneic CD4⁺ T cell proliferation. Unlike HSFs which induced allogeneic lymphocytes to produce high levels of IFN-γ, TNF-α and IL-17, hiPSCs only induced allogeneic lymphocytes to produce IL-2 and IL-10, and promote IL-10-secreting regulatory T cell (Treg) generation. Our study suggests that the integration-free hiPSCs had low or negligible immunogenicity, which may result from their induction of IL-10-secreting Treg.     

Human anti-porcine T cell response: blocking with anti-class I antibody leads to hyporesponsiveness and a switch in cytokine production.

Intervention in the molecular interactions that lead to an immune response is possible at various stages of Ag recognition and T cell activation. Perturbation of the interaction of the TCR with the MHC/peptide ligand complex is one approach that has shown promise for autoimmunity and graft rejection in blocking T cell-activated responses. In this study, we investigated the effect of altering the target MHC class I molecule by blocking with Abs. We established a system that analyzed the human T cell response against MHC class I+/class II- porcine stimulatory cell targets. The primary human response against porcine smooth muscle cells was CD8+ T cell dependent. In the presence of F(ab')2 fragments of the MHC class I-reactive Ab, PT-85, the proliferative response was inhibited and production of IL-2 and IFN-gamma was blocked. Moreover, in a secondary response, proliferation was reduced and type 1 cytokine levels were inhibited. In contrast, levels of IL-10 and IL-4 were sustained or slightly increased. These findings indicate that Ab against MHC class I blocked the recognition of porcine cells by the human CD8+ T cells and altered the cytokine secretion profile. Thus, a single treatment with PT-85 F(ab')2 directed against the MHC class I molecule provides an attractive approach to the induction of T cell tolerance that may provide long-term graft survival in porcine-to-human cell transplantation.     

Role of class II histocompatibility antigens in Staphylococcus aureus protein A-induced activation of human T lymphocytes

The capacity of peripheral blood monocytes and B lymphocytes to support staphylococcal protein A (SpA)-induced proliferation of autologous and allogeneic T cells, as well as the role of major histocompatibility complex (MHC) class I and II molecules in this activation process, were investigated. Highly purified peripheral T lymphocytes did not proliferate in response to SpA, but their response was reconstituted by both irradiated (or mitomycin C-treated) monocytes and B lymphocytes. The effect of B cells on the SpA-induced T-cell response could not be explained by a contamination of residual accessory cells because long-term continuous B-cell lines restored SpA-induced T-cell DNA synthesis as effectively as did monocytes. Support of SpA responsiveness by B cells could not be accounted for by polyclonal binding of SpA to cell surface immunoglobulins, since the ability of SpA-unreactive and SpA-reactive B cells was comparable. The cells from two human leukemic lines--K562 and Raji--showed the same ability in supporting the pokeweed mitogen-induced T-cell response, but the class II-positive Raji cells were much more effective than class II-negative K562 cells in restoring the T-cell responsiveness to SpA. Monoclonal antibodies specific for monomorphic determinants of MHC class II antigens, as well as their F(ab')2 fragments, consistently inhibited the SpA-induced proliferative response, whereas antibodies specific for MHC class I antigens were without effect. The antibodies specific for class II antigens appeared to act at the level of accessory cell, since pretreatment with these antibodies inhibited the ability of SpA-pulsed monocytes or Raji cells to present SpA to autologous or allogeneic T lymphocytes, respectively. These data indicate that either monocytes or normal and lymphoblastoid B cells can act as accessory cells for the proliferative response of human T cells to soluble SpA and that monomorphic determinants of MHC class II molecules play an important role in this activation process.     

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.     

Prethymic nylon wool-passed bone marrow cells, substituting for helper T cells, can augment the generation of cytotoxic T lymphocytes from their precursors.

Nylon wool-passed bone marrow (NW-BM) cells treated with anti-Thy.1 monoclonal antibody and complement were added to a mixed lymphocyte culture which contained a limiting number of lymph node cells, as responder cells, and a sufficient number of mitomycin-c-treated allogeneic spleen cells as stimulator cells. NW-BM cells of the same MHC haplotype as responder cells enhanced the generation of allo-specific cytotoxic T lymphocytes (CTL) not only at a relatively high dose (3 x 10(3) cells/well) of responder cells, but also at an extremely dilute dose (1 x 10(3) cells/well). NW-BM cells which had a third-party MHC haplotype, a haplotype different from both responder and stimulator cells, also enhanced the generation of CTL at relatively high doses, but not at low doses, of responder cells. NW-BM cells which had MHC haplotypes identical with those of responder cells induced CTL from helper T cell-depleted responder cells, but NW-BM cells which had the third-party haplotype did not. These results showed that the enhancing effects of NW-BM cells of the same MHC haplotype as responder cells might be due to a specific helper effect and the enhancing effect of NW-BM cells of the third-party haplotype might be due to a nonspecific filler effect, which only conditioned the cultured cells. It was also found that, to exhibit the helper effect, NW-BM cells had to possess MHC class II, but not MHC class I, molecules in common with CTL precursors. This study showed that in the induction of CTL, prethymic NW-BM cells had a capability comparable to that of mature helper T cells.     

Induction of MHC class I presentation of exogenous antigen by dendritic cells is controlled by CD4+ T cells engaging class II molecules in cholesterol-rich domains.

We investigated interactions between CD4+ T cells and dendritic cells (DC) necessary for presentation of exogenous Ag by DC to CD8+ T cells. CD4+ T cells responding to their cognate Ag presented by MHC class II molecules of DC were necessary for induction of CD8+ T cell responses to MHC class I-associated Ag, but their ability to do so depended on the manner in which class II-peptide complexes were formed. DC derived from short-term mouse bone marrow culture efficiently took up Ag encapsulated in IgG FcR-targeted liposomes and stimulated CD4+ T cell responses to Ag-derived peptides associated with class II molecules. This CD4+ T cell-DC interaction resulted in expression by the DC of complexes of class I molecules and peptides from the Ag delivered in liposomes and permitted expression of the activation marker CD69 and cytotoxic responses by naive CD8+ T cells. However, while free peptides in solution loaded onto DC class II molecules could stimulate IL-2 production by CD4+ T cells as efficiently as peptides derived from endocytosed Ag, they could not stimulate induction of cytotoxic responses by CD8+ T cells to Ag delivered in liposomes into the same DC. Signals requiring class II molecules loaded with endocytosed Ag, but not free peptide, were inhibited by methyl-beta-cyclodextrin, which depletes cell membrane cholesterol. CD4+ T cell signals thus require class II molecules in cholesterol-rich domains of DC for induction of CD8+ T cell responses to exogenous Ag by inducing DC to process this Ag for class I presentation.     

Differential regulatory role of monomorphic and polymorphic determinants of histocompatibility leukocyte antigen class I antigens in monoclonal antibody OKT3-induced T cell proliferation

Monoclonal antibodies (mAb) to monomorphic and polymorphic determinants on the heavy chain of histocompatibility leukocyte antigen (HLA) class I antigens inhibit mAb OKT3-induced T cell proliferation, whereas the anti-beta 2-microglobulin mAb NAMB-1 does not affect it. The inhibitory effect of anti-HLA class I mAb is specific, is not an Fc-mediated phenomenon, does not require accessory cells, and does not involve early stages of T cell activation. Distinct determinants of HLA class I antigens regulate T cell proliferation by different mechanisms, because the anti-HLA-A2, A28 mAb CR11-351, and the mAb W6/32 to a framework determinant of HLA class I antigens block interleukin 2 (IL-2) secretion and IL-2 receptor expression, whereas the mAb CR10-215 to a monomorphic determinant blocks only IL-2 receptor expression. The mAb CR10-215 and W6/32 induced a 50% of maximal inhibition of T cell proliferation, when added after 27 and 12 hr, respectively, of incubation of peripheral blood mononuclear cells with mAb OKT3. On the other hand, the mAb CR11-351 inhibited T cell proliferation even when added after 38 hr of incubation of peripheral blood mononuclear cells with mAb OKT3 and was the only one to inhibit proliferation of cycling T lymphocytes. It is suggested that HLA class I antigens regulate T cell proliferation by interacting with cell-surface molecules involved in T cell activation. The differential inhibitory activity of the anti-HLA class I monoclonal antibodies tested may reflect the different ability of the corresponding determinants to interact with activation molecules.     

Lack of specificity in the mechanisms involved in the enhancement of the concanavalin A driven human T lymphocyte stimulation by beta-endorphin: studies on activation marker expression, cell cycle and interleukin release.

We report on the effects of a physiological concentration of Beta-Endorphin (BE) (10(-12)M) on Concanavalin A (ConA) stimulated human peripheral blood T-lymphocytes and monocytes. We evaluated the effect of timing of BE addition to the culture medium on thymidine uptake, the kinetics of expression of activation markers (CD69, CD25 and CD71) on CD4+ and CD8+ lymphocytes, and of class II MHC antigens on CD14+ cells (monocytes), the kinetics of interleukin-1 (IL-1), interleukin-2 (IL-2) and interferon gamma (IFN-gamma) release, and the cell cycle. Data show that BE is able to influence T lymphocyte only when added together with ConA at the beginning of culture, suggesting its major activity is on the early phases of the T cell response. BE did not increase the amount of class II MHC antigens on monocytes and did not preferentially stimulate CD69, CD25 and CD71 antigen expression on either CD4+ or CD8+ lymphocytes. After 24 hours, the relative proportions of CD4+ and CD8+ lymphocyte in S and G2-M phases were not affected by BE, although the opioid did augment the number of cells in the proliferative compartments of the cell cycle, S and G2-M, indicating an actual increase in the number of cells committed to proliferation. BE did not consistently influence the amount of IL-1, IL-2 and IFN-gamma found in the supernatant of ConA stimulated cultures. The mechanism of the enhancing effect on the proliferative response of normal human lymphocytes to ConA by BE, does not seem to be selective for or unique to specific lymphocyte subsets.     

IL-12 augments antigen-dependent proliferation of activated T lymphocytes.

Ag-dependent T cell activation requires multiple transmembrane signals including activation of Ag-specific T cell receptor in combination with signals delivered through cytokine receptors. IL-12 is a heterodimeric cytokine involved in the regulation of NK and T lymphocyte responses. In examining the role of IL-12 in T cell activation, we found a direct relationship between Ag stimulation and IL-12-induced proliferation. Unlike IL-2, which induced proliferation of CTL either in the presence or absence of a CD3/TCR co-signal, IL-12 mediated proliferation of CTL only when the cells were recently co-stimulated with alloantigen or solid-phase anti-CD3 antibody. After culture in the absence of alloantigen or anti-CD3 for 7 to 14 days, these CTL lost the ability to proliferate to IL-12 alone. Under these conditions, however, IL-12 synergized with low-dose IL-2 to induce CTL proliferation. Restimulation with alloantigen or solid-phase anti-CD3 restored the ability of the CTL to proliferate to IL-12 alone. Not all Ag signals resulted in IL-2 independent proliferation to IL-12. For example, CTL with specificity for influenza matrix peptide proliferated best when co-cultured with peptide Ag presented on self MHC and a combination of IL-2 and IL-12. This evidence suggests that IL-12 may be useful in expanding an Ag-specific T cell population, as the culture of CTL with IL-12 and low-dose IL-2 leads to proliferation only in response to an Ag co-signal.