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O Leo

Publications and source records attributed to O Leo.

At least 91 records · Page 5Linked to original sources
PubMed1991

Cytokine release syndrome induced by the 145-2C11 anti-CD3 monoclonal antibody in mice: prevention by high doses of methylprednisolone.

The hamster mAb 145-2C11 specific for the CD3 complex of murine T lymphocytes shares many properties with OKT3, including the induction of T cell activation. In vivo, the injection of 145-2C11 entails a variety of pathologic changes in relation to the systemic release of cytokines. We tested the effects on this cytokine release syndrome of different doses of methylprednisolone (m-PDS) given at various intervals of time before the 145-2C11 mAb. The administration of high doses of m-PDS (50 mg/kg) 2 to 3 h before the mAb resulted in an almost complete inhibition of the systemic release of TNF-alpha, IL-2, and IL-6. As far as the pathologic changes are concerned, the hypothermia, the acute renal tubular necrosis, and the fatty infiltration of the liver were completely prevented whereas the hypoglycemia was only partially attenuated. The protective effect of m-PDS on the toxicity of 145-2C11 was confirmed by the reduction of the mortality rate among galactosamine-sensitized mice. The inhibition of the release of cytokines by m-PDS did not affect the immunosuppression triggered by 145-2C11 as assessed by the CTL activity against alloantigens measured 48 h after the injection of the mAb. We conclude that the administration of very high doses of glucocorticoids 2 to 3 h before 145-2C11 prevents the release of cytokines and attenuates the acute toxicity of the mAb. Similar protocols could allow mitigation of the cytokine-release syndrome induced by the OKT3 mAb in man.

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PubMed1991

Induction of long-term but reversible unresponsiveness after activation of murine T cell hybridomas.

T cell receptor (TCR)-mediated stimulation of murine T cell hybridomas induces cell activation and lymphokine production. We have observed that following productive activation, T cell hybridomas become refractory to a subsequent stimulation. Hyporesponsiveness is long-lasting but reversible, and is not due to reduced receptor expression. It is noteworthy that hyporesponsiveness is cell autonomous, persists in spite of cell division and does not require continuous exposure to ligands. Hyporesponsive cells retain the ability to produce lymphokines in response to pharmacological agents bypassing receptor triggering, indicating that they possess a functional enzymatic machinery for interleukin 2 synthesis and secretion. Reduced phosphatidylinositol hydrolysis was observed in these cells in response to TCR stimulation, suggesting that hyporesponsiveness result from defective transduction of activation signals. The development of unresponsiveness following receptor stimulation resembles desensitization, and may thus play an important role in the regulation of an immune response and in the induction of immune tolerance.

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PubMed1991

Evidence that pentoxifylline reduces anti-CD3 monoclonal antibody-induced cytokine release syndrome.

Pretreatment with pentoxifylline (PTX), a methylxanthine known for its beneficial effects on tissue lesions induced by the injection of endotoxin or recombinant cytokines, was shown to decrease the systemic release of tumor necrosis factor and interleukin 2 occurring after the administration of the anti-CD3 monoclonal antibody 145-2C11 in mice. In parallel, PTX attenuated the hypothermia and the rise in blood urea nitrogen observed in this model. The protective effect of PTX on the toxicity of 145-2C11 was confirmed by the reduction of the mortality among D-galactosamine-sensitized animals. The mitigation by PTX of the release of cytokines did not affect the immunosuppression entailed by 145-2C11 as assessed by the unmodified cytotoxic T lymphocytes (CTL) unresponsiveness against alloantigens measured 48 hr after the injection of the mAb. In vitro experiments on human peripheral blood leukocytes indicated that PTX alone or in synergy with methylprednisolone (m-PDS) also inhibited the release of TNF and IL-2 induced by OKT3. Finally, in a preliminary pilot trial conducted in kidney transplant recipients, we observed that pretreatment with PTX (20 mg/kg i.v.) in addition to m-PDS (2 g i.v.) reduced by half the amount of TNF released in the blood stream after the first injection of OKT3, while no further reduction of the low levels of IL-2 was found.

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PubMed1991

Hyper IgE in stimulatory graft-versus-host disease: role of interleukin-4.

Intravenous injection of 2 x 10(8) DBA/2 spleen cells into adult intact (C57BL/6 x DBA/2) F1 mice results in a stimulatory graft-versus-host reaction (GVHR) linked to the recognition by donor CD4+ T cells of Ia alloantigens on host B cells. In the experiments presented here, we found that this GVHR is associated with a major increase in IgE serum levels which was already present 7 days after the cell transfer. At 6 weeks, mean IgE levels were more than 200-fold above the control values. Host B cells were responsible for the hypersecretion of IgE in stimulatory GVHR since it was also observed when the DBA/2 donor inoculum was depleted of B cells but not when the F1 recipients were irradiated. The induction of IgE secretion required donor CD4+ T cells as treatment of the donor inoculum with lytic anti-CD4 monoclonal antibody (MoAb) completely prevented the occurrence of the hyper IgE whereas depletion of CD8+ cells had no influence on this parameter. The role played by interleukin-4 (IL-4) in this model was analysed in vivo by the administration of the 11B11 anti-IL-4 rat MoAb (total dose 36 mg) during the first 12 days following induction of stimulatory GVHR by 8 x 10(7) DBA/2 spleen cells. This treatment completely prevented the development of hyper IgE whereas the administration of a control rat MoAb had no significant effect. We conclude that stimulatory GVHR in mice is associated with a major increase in serum IgE which is mediated by IL-4.

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PubMed1991

Modulation of murine host-versus-graft disease by anti-CD3 monoclonal antibody.

BALB/c mice made tolerant to A/J alloantigens by neonatal injection of (A/J x BALB/c)F1 spleen cells develop a host-versus-graft (HVG) disease due to the activation of donor B cells by a subset of host alloreactive helper T cells. We have investigated the effects of a single neonatal injection of the 145-2C11 anti-mouse CD3 monoclonal antibody (MoAb) on the establishment of allotolerance and on the development of the immunopathological features of HVG disease. First, this treatment did not modify the specific anti-donor cytotoxic T lymphocyte (CTL) unresponsiveness or the persistence of circulating immunoglobulins bearing donor allotype. Second, the hyper IgE, the hyper IgG1 and the increased expression of Ia antigens on B cells found in untreated HVG mice were not observed after injection of the 145-2C11 MoAb. Likewise, treated mice displayed lower levels of anti-DNA IgG antibodies and less glomerular immune deposits as compared with untreated HVG mice. We conclude that the administration of the anti-CD3 MoAb did not interfere with the induction of allotolerance but exerts a pronounced inhibitory effect on the associated immunopathological syndrome.

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PubMed1990

Immune surveillance: both CD3+ CD4+ and CD3+ CD8+ T cells control in vivo growth of P815 mastocytoma.

The aim of this study was to examine whether a spontaneous immune response controls neoplastic growth in P815-bearing DBA/2 mice, and to characterize the cells involved in tumor resistance in vivo. Several cell lineages such as T-cell-receptor (TcR)-bearing T cells, NK cells and macrophages mediate some anti-tumor activity in vitro. P815 was chosen as a model because it is weakly immunogenic and is a good target both for tumor-specific, MHC-restricted CTL-mediated lysis and for MHC-unrestricted lysis exerted by long-term cultured lymphocytes or activated macrophages. Since most "NK-like activity" in freshly isolated populations appears to be associated with CD3- cells, whereas antigen-specific, MHC-restricted T cells mostly express CD3 determinants, CD3 was a good marker for evaluating the role of T cells and "NK" cells in tumor resistance in vivo. The survival of anti-CD3-treated animals that were inoculated with tumor cells was strongly reduced (mean survival time: 17 days vs. 40 days for the control group) and was associated with increased tumor growth rate. We followed the same approach to define the T-cell subset(s) that mediate(s) this immune response. Both CD4+ and CD8+ T cells were required for induction of immune control on neoplastic growth. The approach used has revealed the important role of CD4+ T cells in immune responses that control in vivo growth of a class-I-positive, class-II-negative tumor and suggests that these cells may play a central role in tumor resistance. Since CD4+ cells are activated by soluble, exogenous proteins, this finding may have important implications for immunotherapy.

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PubMed1990

Anti-CD3 antibodies induce T cells from unprimed animals to secrete IL-4 both in vitro and in vivo.

Recently, functional heterogeneity among Th cells has been recognized. Based on pattern of lymphokine secretion, two mutually exclusive subsets of CD4+ cells have been defined and designated Th1 (secreting IL-2 and IFN-gamma) and Th2 (secreting IL-4 and IL-5). Identification of these subsets was mostly based on the study of long term cultured T cell lines and clones, and little is known about the Th heterogeneity in vivo. In particular, it has been suggested that IL-4 producing cells cannot be detected in vivo or in primary stimulations in vitro unless responder cells had been previously primed. Our data however, indicate that anti-CD3 mediated stimulation can induce T cells isolated from unprimed animals to IL-4 production. An assay system based on the ability of IL-4 to increase Ia expression of B cells present in the environment of activated T cells was found to be more sensitive than detection of secreted IL-4 in the supernatant by conventional bioassays and was used to study IL-4 production by unprimed lymphocytes polyclonally stimulated in vivo and in vitro by anti-CD3 mAb. The results obtained indicate that CD4+ CD8- T cells able to produce IL-4 upon receptor-specific stimulation exist in the preimmune pool of adult animals. Remarkably, these cells can also be stimulated in vivo by treating animals with anti-CD3 mAb, as indicated by the in vivo induction of IL-4 specific mRNA and hyper-Ia expression on B cells. These results indicate that the inability to detect IL-4 in primary cultures is not due to different activation requirements of Th2 cells but may simply result from their lower frequency in unprimed animals.

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PubMed1990

Hypothermia and hypoglycemia induced by anti-CD3 monoclonal antibody in mice: role of tumor necrosis factor.

The possible involvement of tumor necrosis factor-alpha (TNF) in the metabolic disturbances induced by anti-CD3 monoclonal antibodies (mAb) was analyzed in DBA/2 mice injected with 50 micrograms of the anti-murine CD3 mAb 145-2C11. First, we found that 145-2C11 induces a profound hypothermia maximal between 3 h and 6 h after the injection (at 3 h: -3.0 +/- 0.1 degrees C) as well as hypoglycemia (blood glucose levels at 6 h and 24 h: 76 +/- 13 mg/100 ml and 92 +/- 22 mg/100 ml, respectively, p less than 0.001 as compared with control values). These metabolic changes are preceded by the release of TNF into the circulation (peak serum TNF levels at 2 h: 50 +/- 23 pg/ml, p less than 0.01 as compared with controls). The release of TNF induced by 145-2C11 depends on the effect of the mAb on T cells as it is not observed in athymic nude mice while lipopolysaccharide-resistant C3H/HeJ mice also display a significant rise in serum TNF (peak levels at 2 h: 59 +/- 44 pg/ml). Pretreatment of DBA/2 mice with 12 mg of rabbit anti-murine TNF antibodies completely prevents the hypothermia while the hypoglycemia is significantly attenuated. Finally, F(ab')2 fragments of 145-2C11 induce only a transient hypoglycemia (blood glucose levels at 6 h: 109 +/- 14, p less than 0.001 as compared with controls) but neither hypothermia nor significant TNF release. We conclude that TNF is a major mediator of the acute metabolic changes induced by the intact form of 145-2C11.

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PubMed1989

CD4-Ia interactions can occur in the absence of T-cell receptor/antigen-Ia recognition.

The T-cell differentiation antigen, CD4, is expressed by major histocompatibility (MHC) class II restricted T lymphocytes. CD4+CD8- T cells use their T-cell receptor to recognize foreign antigens in association with MHC class II products (Ia). The association between CD4 expression and restriction by MHC class II products has led to the hypothesis that CD4 may interact with monomorphic determinants of MHC class II molecules. A large body of experimental evidence suggests that CD4 interaction with MHC class II molecules leads to an increase in the binding avidity of T cell-stimulator cell interactions. A direct test for a functional CD4-MHC class II interaction in T-cell activation requires a separate evaluation of CD4-Ia interactions from T-cell receptor (TcR)-antigen (Ag)/Ia recognition. However, a separate evaluation proves difficult since the T-cell receptor and CD4 may interact with the same MHC class II molecule. In this report, we use a T-cell activation protocol where TcR-Ag/Ia recognition is replaced by TcR complex-anti-CD3 antibody interactions. Therefore, the affinity of the TcR complex for its ligand (the anti-CD3 mAb) is independent from MHC expression on target cells and allows a separate evaluation of the role of accessory molecules in T-cell activation. We have analysed the effects of monoclonal anti-MHC class II antibodies on the activation of a CD4+ T-cell hybridoma in the absence of its TcR restricting MHC class II molecule (I-Ek) but in the presence of unrelated MHC class II molecules (I-Ed, I-Ad). The data obtained indicate a functional interaction between the CD4 molecule and a non-polymorphic region of the MHC class II product in T-cell triggering.

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PubMed1988

Selective expression of the Ly-6C.2 epitope on Lyt-2+ T cells that can develop in the absence of thymic influence.

The present study focused on attempts to correlate the expression of T cell differentiation antigens and development. A series of mAb was developed that recognized an Ly-6.2-linked Ag, Ly-6C.2. These mAb reacted with a subset of peripheral Lyt-2+ T lymphocytes but not Lyt-2+ thymocytes. Functional analysis of the Lyt-2+, Ly-6C.2- and Lyt-2+, Ly-6C.2+ subsets demonstrated that both cell types responded in class I and class II allogeneic MLC as well as in virus-specific cell-mediated cytolysis assay, although precursor frequency analysis suggested that the majority of precursor CTL were Ly-6C.2+. The marker did not appear to represent an activation Ag because Ly-6C.2-, Lyt-2+ precursor CTL generated Ly-6C.2-, Lyt-2+ CTL. Finally, virtually all splenic Lyt-2+ T cells from athymic (nu/nu) mice were Ly-6C.2+ whereas all thymic CTL were Ly-6C.2-. These results suggested that Ly-6C.2+, Lyt-2+ T cells may represent a distinct lineage of Lyt-2+ T cells that can develop in the absence of thymic influence.

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PubMed1988

Activation properties of T cell receptor-gamma delta hybridomas expressing diversity in both gamma- and delta-chains.

To elucidate the structure, diversity, and activation properties of the murine T3-associated gamma delta-receptor, examination was made of the gamma delta and T3 components, T cell receptor (TCR) gene transcription, activation properties, and lymphokine production in a panel of four cloned T cell hybridomas expressing a TCR-gamma delta. Biochemical analysis of the gamma and delta proteins expressed on these hybridomas reveals new gamma and delta species not observed in whole populations of dLy-1 Lyt-2-L3T4-thymocytes from which these hybridomas were derived. Thus, analysis of expression of the TCR-gamma delta complex at the clonal level indicates that both gamma and delta appear to be expressed as multiple distinct gene products within a homozygous inbred mouse strain. Northern blot analysis reveals that, whereas all gamma delta hybridomas had mature 1.5-kb TCR-alpha-chain and mature TCR-gamma-chain transcripts, none had mature 1.3-kb TCR-beta-chain transcripts, thus indicating that the type of TCR heterodimer expressed reflects of the state of TCR gene transcription in these hybridomas. Our results also reveal striking similarities between TCR-gamma delta and TCR-alpha beta cells with respect to their activation properties. First, all five of the T3 components associated with the gamma delta-complex are of the same size and have the same glycosylation patterns as those associated with alpha beta-heterodimers. Second, induction of function in these gamma delta cells (assayed by lymphokine production) can be achieved with a variety of stimuli known to elicit activation signals in alpha beta cells as well: direct receptor-engagement (i.e., through anti-Thy-1), and phorbol 12-myristate 13-acetate-plus-ionomycin-mediated. Collectively, these findings suggest that gamma delta T cells express a receptor of at least limited diversity and use T3-mediated activation pathways very similar to those employed by TCR-alpha beta-bearing T cells.

Animals↗
PubMed1988

Mitogenic activation of EL-4 cells does not require surface Thy-1 expression.

The ability of monoclonal anti-Thy-1 antibodies to stimulate IL-2 production and T-cell proliferation has raised the possibility that Thy-1 may play an important role in T-cell activation. To examine this postulated role we have produced Thy-1-negative variants of the murine T lymphoma EL-4 by mutagenesis with ethyl methanesulfonate (EMS) and subsequent negative selection with anti-Thy-1 monoclonal antibodies (mAbs) and complement. Although the parental EL-4 cell line produced interleukin-2 (IL-2) in response to concanavalin A (Con A), phytohemagglutinin, anti-Thy-1 mAbs, and an anti-T3 mAb, as well as after exposure to phorbol myristate acetate (PMA), only PMA was capable of inducing IL-2 production by several Thy-1-negative cell lines. The loss of responsiveness to cell surface stimulatory ligands appeared to be correlated with loss of Thy-1 expression because mutagenized cells selected for high levels of Thy-1 expression all responded normally to Con A. However, when Thy-1 expression was reconstituted in the "nonresponder" (Thy-1-negative) cell lines either by transfection of a Thy-1.2 gene or by 5-azadeoxycytidine treatment, the revertant cell lines were still unable to produce IL-2 when stimulated with Con A, anti-Thy-1, or anti-T3. Furthermore, several other independently derived Thy-1-negative EL-4 cell lines responded normally to mitogens and mitogenic mAbs. Taken together, these results suggest that Thy-1 expression is not required for the T-cell activation process and that the EMS mutagenesis procedure resulted in an additional mutation(s) responsible for the inability of certain Thy-1-negative cell lines to be triggered by mitogens and mitogenic mAbs. These cell lines may prove to be valuable tools for further biochemical and molecular studies of the sequence of events associated with T-cell activation.

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PubMed1987

Role of accessory molecules in signal transduction of cytolytic T lymphocyte by anti-T cell receptor and anti-Ly-6.2C monoclonal antibodies.

Accessory molecules present on the cell surface of cytolytic T lymphocytes (CTL) play an important role in their activation. Antigen-specific recognition by CTL is inhibited by antibodies against Lyt-2, L3T4, or LFA-1 molecules. Presently it is not known whether these molecules function by binding a ligand such as class I or class II on the target cell or by delivering a signal that down-regulates T cell activation. In the present study we utilized anti-T cell antibodies including anti-T3 and anti-T cell receptor (alpha/beta) as well as an anti-Ly-6.2C monoclonal antibody to activate CTL clones to kill irrelevant targets or secrete BLT esterase. The redirected lysis assay system utilizes the fact that heteroconjugates between anti-T3, and anti-T cell receptor, or anti-Ly-6.2C and anti-trinitrophenyl can trigger CTL lysis of trinitrophenyl-coupled targets that did not express antigen. In this system anti-Lyt-2 antibodies as well as anti-LFA-1 antibodies inhibited triggering via T cell receptor-related molecules but not via the anti-Ly-6.2C heteroconjugate. In addition, the anti-Lyt-2 was shown to inhibit conjugate formation in the heteroaggregate assay system suggesting that the anti-Lyt-2 antibodies acted early in inhibiting CTL activity. Similar results were observed in a system in which the CTL clones were triggered to secrete a BLT-esterase-like activity in the absence of target cells. Anti-T3 coated on plastic was shown to activate BLT-esterase secretion. This secretion was inhibited by anti-Lyt-2 and anti-LFA-1. Thus, it would appear that both the Lyt-2 molecule and the LFA-1 molecule act as signal-transducing elements involved in CTL activation. In particular, the Lyt-2 molecule appears to preferentially function in receptor-mediated T cell activation.

Animals↗
PubMed1987

Activation of murine T lymphocytes with monoclonal antibodies: detection on Lyt-2+ cells of an antigen not associated with the T cell receptor complex but involved in T cell activation.

A new T cell molecule defined by the mAb 143-4-2 has been identified that is involved in T cell activation. The expression of the 143-4-2-defined epitope is linked to the previously characterized Ly-6 locus and restricted to bone marrow cells and to a subset of peripheral Lyt-2+ cells. In comparison to other anti-Ly-6.2 mAb, the 143-4-2 mAb appears to be directed at an allogeneic determinant of the Ly-6.2C molecule. The anti-Ly-6.2C antibody can promote the lysis of antigen-non-bearing target cells by alloreactive CTL clones, and in the presence of cofactors (PMA or IL 2) induces a subset of Lyt-2+ cells to proliferate, perhaps through an autocrine pathway. Although the antibody described has antigen-like effects as described for anti-TcR complex reagents, studies performed with a recently derived anti-murine T3 mAb suggest that the Ly-6.2C molecule is not associated on the cell surface with components of the TcR complex. Nevertheless, cell surface expression of the TcR complex is required for optimal triggering of T cells via the Ly-6.2C molecule. Because Ly-6.2C determinants are expressed in bone marrow and not in the thymus, the possibility is considered that expression of this molecule identifies a distinct subset of extrathymically derived T cells.

Animals↗
PubMed1987

Identification of a monoclonal antibody specific for a murine T3 polypeptide.

A monoclonal antibody (145-2C11) specific for the murine T3 complex was derived by immunizing Armenian hamsters with a murine cytolytic T-cell clone. The antibody is specific for a 25-kDa protein component (T3-epsilon) of the antigen-specific T-cell receptor. It reacts with all mature T cells and can both activate and inhibit T-cell function. These results identify T3-epsilon as a cell surface protein involved in the transduction of activation signals.

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PubMed1986

Identification of monoclonal antibodies specific for the T cell receptor complex by Fc receptor-mediated CTL lysis.

Monoclonal antibodies (mAb) directed at the T cell receptor complex (TcR) on cloned T cells have generally been identified by their ability to inhibit the clone's antigen-specific function. Because such inhibition is highly dependent on antibody concentration and affinity, detection of anti-clonotypic antibodies to murine alloreactive T cells has been very difficult. In this report, an alternative method is described on the basis of the ability of antibodies specific for the TcR complex to activate T cells in an antigen-independent manner. The assay is based upon the observation that soluble antibodies to human T3 promote lysis of irrelevant, Fc receptor-positive targets by a human CTL line. By using this approach, an anti-TcR mAb has been identified among a panel of murine mAb generated against an alloreactive CTL clone. Induction of lysis by soluble anti-TcR mAb has been shown to require both the expression of Fc receptors on the target cell and conjugate formation between the effector and the target cell. This assay provides a screening procedure that is much more sensitive than inhibition of function, and it preferentially detects antibodies specific for cell surface molecules involved in T cell activation.

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