Characterization of the precursors of the NK-like cytotoxic cells generated in the autologous mixed leukocyte reaction and by interleukin 2 activation

In vitro stimulation of E+ cells with autologous E- cells (AMLR) for 7 days results in the generation of NK-like killer cells with the same phenotype (E+, 4F+2, OKT-8, OKM-1) as the killer cells produced by activation of E+ cells with IL-2 for 3 days in our system. IL-2 activity was also detected in the course of AMLR culture in concordance with the generation of the killer cells. These data prompted us to characterize the progenitor cells in the 2 systems by using positive (panning) or negative (complement mediated cytolysis) selection techniques. The NK-like killer cells generated from both the AMLR responder population and by activation with IL-2 are derived from the same population of cells with an E+, OKM+1 but not Leu 7+ phenotype.     

Injury to autologous normal tissues and tumors mediated by lymphokine-activated killer (LAK) cells generated in vitro from peripheral blood mononuclear cells of glioblastoma patients

Activation of peripheral blood mononuclear cells (PBMC) with IL-2 generates lymphokine-activated killer (LAK) cells that show a broad target cell range. In adoptive immunotherapy using in vitro-generated LAK cells, the intensity and specificity of their cytotoxic activity affect the prognosis of cancer patients. The present study was designed to examine the tumor-specific spectrum of T lymphocytes generated from the PBMC of patients with recurrent glioblastoma by in vitro propagation with IL-2 plus either soluble or solid-phase anti-CD3 monoclonal antibody (MAb) in short-term or long-term cultures. Both short-term and long-term culturing with solid-phase anti-CD3 MAb plus IL-2 yielded broad-reactivity CD8+ alphabetaT and gammadeltaT lymphocytes, both of which were non-MHC restricted, as shown by the fact that they were able to lyse autologous glioblastoma cells, MHC class I+II- allogeneic glioblastoma cells, and MHC class I-II-NK-sensitive K562 target cells. More importantly, these cells from patients failed to lyse fresh autologous PBMC. These results demonstrate that cells generated using this approach are non-MHC-restricted LAK cells and exhibit marked tumor specificity. In contrast, incubation with soluble anti-CD3 MAb generated T lymphocytes that after long-term culture, were either CD4+ or CD8+. These caused significant lysis of both allogeneic and autologous glioblastoma target cells, the extent of lysis being greater than that using cells produced by culturing with the solid-phase MAb. However, both the CD4+ and CD8+ cells also caused greater lysis of autologous normal PBMC, indicating that cells generated using this approach may cause significant adverse reactions in cancer patients if used for immunotherapy.     

Production of a cytokine with interleukin 3-like properties and cytokine-dependent proliferation in human autologous mixed lymphocyte reaction

The autologous mixed lymphocyte reaction (AMLR) was assayed in a medium containing fresh autologous serum, by using nylon-adherent stimulator cells and nonadherent responder T cells, which were prepared from human peripheral blood mononuclear cells in the absence of fetal calf serum (FCS) to avoid any sensitization to xenogeneic protein antigens. DNA replication without a background proliferative response was induced by stimulator cells in the responder cells. The addition of monoclonal anti-HLA-DR antibody to the culture or treatment of the responder cells with complement plus anti-T4 but not anti-T8 monoclonal antibody suppressed the AMLR, suggesting that this specific AMLR involves an interaction between HLA-DR antigens and helper/inducer T cells. Regardless of this specific DNA replication, the AMLR generated no production of interleukin 2 (IL-2) and interferon gamma (IFN-gamma), both of which could be found in the allogeneic (allo) MLR. In addition, DNA replication in the AMLR was not inhibited by the addition of specific antisera for IL-2 and IFN-gamma, both of which significantly inhibited the DNA replication in allo-MLR. The AMLR was accompanied by production of a soluble factor, which could stimulate the proliferation of murine interleukin 3 (IL-3)-dependent cell line 32Dcl but not the proliferation of IL-2-dependent cell lines. This factor was also found to be responsible for proliferation of responder nonadherent cells in the AMLR. It strongly stimulated bone marrow cells, as did the murine IL-3. The factor had an Mr range, as determined by gel filtration, of 15,000-28,000, but it did not bind to fast protein liquid chromatography (FPLC)-MonoQ column. Thus, the factor is distinguishable from IL-2 in physicochemical or biological properties, but similar to murine IL-3. These results suggest that the human AMLR may be primarily a phenomenon in which non-T cells mediated by the HLA-DR antigens on the cell stimulate helper/inducer T cells to produce a lymphokine with IL-3-like properties, but no IL-2, which in turn stimulates the factor-dependent cells to proliferate.     

Autologous mixed lymphocyte reaction in man. X. T-T autologous mixed lymphocyte reaction in young and aging humans

T cells upon activation with mitogens or autologous non T cells express surface HLA-DR antigens and are capable of stimulating autologous T cells in the autologous mixed lymphocyte reaction (T-T AMLR). We have examined T-TA AMLR, using T-non T AMLR activated-(TA) T cells as stimulators in young (21-32 yr) and aging humans (62-84 yr). In aging subjects a significantly (p less than 0 . 01) higher proliferative response was observed in T-TA AMLR as compared to simultaneously studied young subjects. In allogeneic MLR, no significant difference was observed between young and aging subjects. The increased T-TA AMLR could be a mechanism responsible for deficient T-non T AMLR reported in aging humans.     

Enhancement of the impaired autologous mixed leukocyte reaction in patients with systemic lupus erythematosus.

The cellular basis of the impaired autologous mixed leukocyte reaction (AMLR) in patients with systemic lupus erythematosus (SLE) was investigated. Non-T cells from normal subjects and from SLE patients were fractionated into low and high density subpopulations. SLE patients were found to have an increased proportion of low density to high density non-T cells when compared to normal subjects. Although normal low-density non-T cells were highly enriched in AMLR stimulatory capacity, SLE low-density non-T cells induced minimal proliferation by autologous T cells. Brief incubation of SLE non-T cells with phorbol myristate acetate or formalin-treated Staphylococcus aureus resulted in marked augmentation of the capacity of those non-T cells to stimulate an AMLR, although the magnitude of the activated non-T cell-induced AMLR did not achieve that observed in normal subjects. No significant alterations in the expression of Ia molecules on the surface of the non-T cells were detected after in vitro activation. These experiments support the hypothesis that the impaired capacity of SLE T lymphocytes to proliferate in response to autologous non-T cells may in part represent a failure of SLE non-T cells to present an appropriate stimulus for the generation of a T cell response.     

Generation of T cells with lytic specificity for atypical antigens. III. Priming F1 animals with antigen-bearing cells also having reactivity for host alloantigens allows for potent lytic T cell responses

Here, we explore the conditions required for generating two different highly potent F1 antiparental killer cell populations to unusual antigens in rats. The first, L/DA anti-DA, has lytic specificity for two antigen systems: MTA, a mitochondrial antigen expressed on DA and DA Lewis (L) target cells restricted by RT1A class I molecules; and H, an antigen that maps to the class I-like RT1C region and is present only on parental target cells from donors homozygous at the major histocompatibility complex. The second killer population is generated in the reciprocal DA/L anti-DA combination and has lytic specificity only for the H antigen system. We show that the killer cells are T cells, and that generation of these F1 cytotoxic T lymphocytes (CTL) requires an in vivo priming step in which it is essential that the inoculated parental cells bear the relevant target antigens and possess alloreactivity for F1 host antigens. The requirement for alloreactivity and antigen on the same priming cell population suggests that these potent lytic responses depend on a situation akin to a hapten-carrier effect that bypasses otherwise ineffective helper responses by the host to these unusual antigens. Restimulation of F1 lymphocytes in culture is also necessary, requiring the presence of antigen on irradiated lymphoblast stimulator cells, but alloreactivity to responder cell antigens is not necessary; normal, nonactivated lymph node cells are completely ineffective as stimulators. For effective lysis, the target cells need not possess the potential for alloreactivity to responder F1 CTL. We also demonstrate in a preliminary way additional antigen systems defined by killer populations raised with other F1 antiparental strain combinations.     

Responder cells in the human autologous mixed lymphocyte reaction.

Isolated human T4+ cells proliferate in the autologous mixed lymphocyte reaction (AMLR), whereas isolated T8+ cells do not. However, in the presence of Interleukin 2 or T4+ cells, the T8+ cells demonstrated substantial proliferation. These studies suggest that T8+ cells recognize signals from autologous non-T cells, but require an additional factor for the subsequent proliferative response. Since this stimulus can be provided by T4+ cells, the AMLR appears to constitute an inducer circuit. Different defects in this circuit may be responsible for the common abnormality of the AMLR in different diseases.     

Antigen-reactive T cells can be activated buy autologous macrophages in the absence of added antigen

T cells responsive to macrophages (M phi) in the autologous mixed lymphocyte reaction (AMLR) contain those cells that can be induced to proliferate by soluble antigens. Negative solution (5-bromo-2-deoxyuridine and light) of T cells activated by autologous M phi also removed those cells required for reactivity to Candida albicans and purified protein derivative. Positive selection of T cells responsive to autologous M phi yields a population that is simultaneously enriched in antigen reactivity. Some patients demonstrating cutaneous anergy and diminished in vitro blast transformation in response to soluble antigen also lack T cells responsive to the AMLR to M phi. When considered in conjunction with previously reported data, these findings indicate the AMLR occurring between T cells and M phi in the absence of soluble antigen represents self recognition occurring between antigen-reactive T cells and antigen-presenting M phi.     

Human autologous mixed lymphocyte reactivity is primarily specific for xenoprotein determinants adsorbed to antigen-presenting cells during rosette formation with sheep erythrocytes

We present evidence that most T cells proliferating in response to autologous sheep erythrocyte (SRBC)-separated non-T cells (NT) cells are not specific for autoantigens but for antigens derived from xenogeneic sources. The conclusion was based on the following three observations. First, we found that NT cells isolated in the absence of xenoproteins by means of density gradient centrifugation on Percoll only weakly stimulated autologous T cells. Because this weak proliferation could not be expanded in restimulation experiments, its significance as an immune recognitive event remains questionable. NT cells isolated by the above method in the absence of xenogeneic determinants readily acquired stimulatory capacity after brief exposure to either SRBC or fetal calf serum. Second, restimulation of T memory cells generated in 1 degree autologous mixed lymphocyte reaction (AMLR) against SRBC-separated autologous NT cells was exclusively seen when NT cells exposed to or separated with xenoproteins were used for restimulation. Third, T memory cells generated against SRBC-separated autologous NT cells were specifically restimulated by autologous Percoll-separated NT cells that had been pulsed with a variety of xenogeneic mammalian sera. These xenogeneic determinants were preferentially recognized in context with autologous HLA-DR+ cells. From these findings and from our previous results that indicated an absolute requirement of HLA-DR+-adherent NT cells (8), we conclude that human AMLR primarily does not represent an autoantigen but a xenoantigen response that is genetically restricted by the HLA-DR type of the antigen-presenting cell.     

Impairment of the autologous mixed lymphocyte reaction in atopic dermatitis.

The T cell proliferative response to autologous non-T cells is termed the autologous mixed lymphocyte reaction (AMLR). Recent studies have suggested that the AMLR represents an inducer circuit for the activation of T8+ suppressor/cytotoxic effector cells. Since atopic dermatitis (AD) patients are deficient in T8+ cytolytic T cell function, we investigated the AMLR in AD. When sheep erythrocytes were used to separate T cells from non-T cells, the AMLR was found to be significantly decreased (P less than 0.001) in AD patients (n = 11; delta cpm = 1,550 +/- 393) when compared with normal control subjects (n = 13; delta cpm = 25,819 +/- 4,609). To exclude the possibility that these results were an artifact of the sheep erythrocyte separation, T cells were also separated on a fluorescence-activated cell sorter after treatment of peripheral blood lymphocytes with the OKT3 monoclonal antibody. AD T cells separated by the latter method were also found to have a significantly reduced AMLR response when compared with similarly treated normal T cells. Co-culture studies using cells from AD patients and their HLA identical siblings indicated that the defect resided at the responder T cell level rather than at the stimulator non-T cell level. Co-culture studies revealed no evidence for excessive suppressor cell activity resulting in the decreased AMLR. However, enumeration of T cells reactive with the monoclonal antibody T29, which recognizes a subset of T cells proliferating in the AMLR, demonstrated that AD patients (n = 8; % T29 = 2.5 +/- 0.7) had a significantly decreased (P less than 0.001) number of circulating T29+ T cells when compared with normal controls (n = 8; % T29 = 10.4 +/- 0.8). These studies suggest that a deficiency of T4+ T29+ cells contributes to the deficient AMLR in AD and possibly underlies the abnormalities of T8+ effector cells present in this disease.     

T cell contact with Ia antigens on nonhemopoietic cells in vivo can lead to immunity rather than tolerance.

Long-term H-2-heterozygous a----(a x b)F1 bone marrow (BM) chimeras prepared with supralethal irradiation (1,300 rad) are devoid of Ia+ host BM-derived antigen-presenting cells (APC), but show quite strong host Ia expression in germinal centers, probably on follicular dendritic cells (a class of nonhemopoietic stromal cells). To examine whether Ia expression on these non-BM-derived cells is capable of inducing post-thymic tolerance of T cells, thymectomized irradiated (a x b)F1 mice were reconstituted with parent alpha stem cells and then, 6 mo later, given parent alpha thymus grafts. As measured by primary mixed lymphocyte reactions and V beta expression, the CD4+ cells differentiating in the thymus-grafted mice showed no detectable tolerance to the H-2 (Ia) antigens of the host. To examine whether the thymus-grafted mice contained immunologically significant quantities of host Ia antigens, long-term alpha----(alpha x b)F1 chimeras were injected with normal strain alpha CD4+ cells; the donor cells were recovered from thoracic duct lymph of the chimeras and tested for host reactivity in vitro. The results showed that Ia expression in the chimeras was sufficient to cause selective trapping of a substantial proportion of host-Ia-reactive CD4+ cells soon after transfer and, at later stages, to induce strong priming. Tolerance was not seen. The data place constraints on the view that T cell recognition of antigen expressed on cells other than typical BM-derived APC leads to tolerance induction.     

Helper cells in the autologous mixed lymphocyte reaction. III. Production of helper factor(s) distinct from interleukin 2

In normal mice, the autologous mixed lymphocyte reaction (AMLR) can activate helper T cells that, in the presence of hapten-modified syngeneic cells, can induce a hapten-specific cytotoxic response. Supernatants from AMLR cultures contain a factor(s) that will mediate a cytotoxic T cell response to hapten-altered self. The AMLR factor is effective in facilitating the generation of cytotoxicity only in those cultures containing both T cells and hapten-altered, syngeneic, nonstimulatory cells. Factor production requires an interaction between Lyt-1+23- cells and non-T cells (the T cells synthesize it). The AMLR factor does not appear to be interleukin 2 (IL-2) because it does not activate thymocytes in the presence of antigen, nor does it maintain an IL-2-dependent cell line or function in co-stimulator assays. For the AMLR factor to facilitate the generation of cytotoxicity, thymic adherent cells are a necessary intermediate. These data suggest that the factor recoverable from AMLR cultures acts early in the cytotoxic pathway, before IL-1 production.     

Participation of the H-2 antigens of tumor cells in their lysis by syngeneic T cells

Cytotoxic T lymphocytes were generated in vitro against H-2 compatible or syngeneic tumor cells. In vitro cytotoxic activity was inhibited by specific anti-H2 sera, suggesting that H-2 antigens are involved in cell lysis. Two observations directly demonstrated the participation of the H-2 antigens on the tumor cells in their lysis by H-2-compatible T cells. First, coating of the H-2 antigens on the target tumor cell reduced the number of cells lysed on subsequent exposure to cytotoxic T cells. Second, when cytotoxic T cells were activated against an H-2 compatible tumor and assayed against an H-2-incompatible tumor, anti-H-2 serum that could bind to the target cell, but not to the cytotoxic lymphocyte, inhibited lysis. H-2 antigens were also shown to be present on the cytotoxic lymphocytes. Specific antisera reacting with these H-2 antigens, but not those of the target cell, failed to inhibit lysis when small numbers of effector cells were assayed against H-2-incompatible target cells or when effector cells of F1-hybrid origin and bearing two H-2 haplotypes were assayed against a tumor cell of one of the parental strains. These findings suggest that it is the H-2 antigens on the tumor cell and not those on the cytotoxic lymphocytes that are important in cell-mediated lysis of H-2-compatible tumor cells.     

Major histocompatibility complex-restricted self recognition. A monoclonal anti-I-Ak reagent blocks helper T cell recognition of self major histocompatibility complex determinants

The functional role of cell surface Ia antigens has been studied for in vitro antibody responses, using as a probe the ability of anti-Ia reagents to inhibit these responses. A hybridoma monoclonal anti-Ia reagent specific for a product of I-Ak (Ia.17) profoundly inhibited in vitro antibody responses to TNP-KLH by spleen cells of the I-Ak but not I-Ab haplotype. This inhibition by anti-I-Ak product, but not by interaction with T or B cell product, in spite of the fact that functional B cells as well as accessory cells could be shown to express the determinant detected by this hybridoma reagent. These results suggest that the Ia expressed by accessory cells in of unique functional importance in these responses. To further characterize the function of Ia antigens in this response system, the mechanism of anti- I-Ak inhibition was determined. The inhibition resulting from interaction of anti-I-Ak with accessory cell Ia was not mediated by nonspecific suppressor cells, nor was there nonspecific interference with accessory cell function as a result of the binding of anti-Ia antibody. The relationship between anti-Ia inhibition and T helper cell recognition of self determinations on accessory cells was analyzed using T cells from radiation bone marrow chimeras. It was demonstrated that (B10 X B10.A)F1 leads to B10 (F1 leads to B10) chimera T cells were able to cooperate with B10 (H-2b and I-Ab) but not B10.A (H-2a and I-Ak) accessory cells for responses to TNP-KLH; F1 leads to B10.A T cells were able to cooperate with B10.A but not B10 accessory cells; and both chimera populations were able to cooperate with (B10 X B10.A)F1 (F1) accessory cells. Monoclonal anti-I-Ak inhibited the cooperation of F1 leads to B10.A T cells with the same F1 accessory cells. Thus, inhibition by anti-I-Ak is dependent upon active helper T cell recognition of I-Ak-encoded determinants expressed on accessory cells. These findings demonstrate that T cells recognize self Ia determinants expressed on accessory cells, and that such recognition is required for the generation of T cell-dependent antibody responses.     

Dengue virus-specific human T cell clones. Serotype crossreactive proliferation, interferon gamma production, and cytotoxic activity

The severe complications of dengue virus infections, hemorrhagic manifestation and shock, are much more commonly observed during secondary infections caused by a different serotype of dengue virus than that which caused the primary infections. It has been speculated, therefore, that dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) are caused by serotype crossreactive immunopathological mechanisms. We analyzed clones of dengue serotype crossreactive T lymphocytes derived from the PBMC of a donor who had been infected with dengue 3 virus. These PBMC responded best to dengue 3 antigen, but also responded to dengue 1, 2, and 4 antigens, in bulk culture proliferation assays. 12 dengue antigen-specific clones were established using a limiting dilution technique. All of the clones had CD3+ CD4+ CD8 phenotypes. Eight clones responded to dengue 1, 2, 3, and 4 antigens and are crossreactive, while four other clones responded predominantly to dengue 3 antigen. These results indicate that the serotype crossreactive dengue-specific T lymphocyte proliferation observed in bulk cultures reflects the crossreactive responses detected at the clonal level. Serotype crossreactive clones produced high titers of IFN-gamma after stimulation with dengue 3 antigens, and also produced IFN-gamma to lower levels after stimulation with dengue 1, 2, and 4 antigens. The crossreactive clones lysed autologous lymphoblastoid cell line (LCL) pulsed with dengue antigens, and the crossreactivity of CTL lysis by T cell clones was consistent with the crossreactivity observed in proliferation assays. Epidemiological studies have shown that secondary infections with dengue 2 virus cause DHF/DSS at a higher rate than the other serotypes. We hypothesized that the lysis of dengue virus-infected cells by CTL may lead to DHF/DSS; therefore, the clones were examined for cytotoxic activity against dengue 2 virus-infected LCL. All but one of the serotype crossreactive clones lysed dengue 2 virus-infected autologous LCL, and they did not lyse uninfected autologous LCL. The lysis of dengue antigen-pulsed or virus-infected LCL by the crossreactive CTL clones that we have examined is restricted by HLA DP or DQ antigens. These results indicate that primary dengue virus infections induce predominantly crossreactive memory CD4+ T lymphocytes. These crossreactive T lymphocytes proliferate and produce IFN-gamma after stimulation with a virus strain of another serotype, and demonstrate crossreactive cyotoxic activity against autologous cells infected with heterologous dengue viruses.(ABSTRACT TRUNCATED AT 400 WORDS)     

Autologous rosette-forming T cells as the responding cells in human autologous mixed-lymphocyte reaction.

Autologous rosette-forming cells (Tar cells) have surface and functional characteristics of post-thymic precursors and among these characteristics there are some that have been identified in the responsive cell of the autologous mixed-lymphocyte reaction (AMLR). We therefore did AMLR with circulating mononuclear cells from normal subjects using as responding cells either total T cells, T cells depleted of Tar cells, or purified Tar cells. The response of Tar cells in AMLR was significantly greater than that of total T cells and these responded significantly more than Tar-depleted T cells. Conversely, Tar cells responded less than total T cells or T cells depleted of Tar cells in allogeneic mixed-lymphocyte reactions. Increasing numbers of Tar cells gave significantly greater AMLR responses both alone and when added to diminishing proportions of Tar-depleted T cells to keep the number of T cells constant in the system. Tar cells are the responding cells in AMLR but not in allogeneic mixed-lymphocyte reactions.     

Antigen recognition by a T cell clone outside the context of the major histocompatibility complex. A role for Mls in antigen presentation

A T cell clone isolated from antigen-primed CB6/F1 mice was shown to proliferate to keyhole limpet hemocyanin (KLH) in the presence of irradiated syngeneic F1 spleen cells, as well as spleen cells from either parental strain (BALB/c and C57BL/6). The genetic restriction involved in this antigen-specific proliferation was mapped using BXD (C57BL/6 X DBA/2) recombinant inbred strains of mice to the Mls gene on chromosome one. To exclude the role of Ia antigens as the restricting determinants, monoclonal anti-Ia antibodies were used to block the in vitro proliferative response of this clone. Although anti-Iab and anti- Iad blocked the proliferation of this clone to KLH in the presence of irradiated spleen cells from either parent, this effect was shown to be dependent on Ia molecules passively absorbed by the T cell clone from the irradiated filler cells. Since the T clone expressed Thy-1.2 and Lyt-1+ differentiation markers, its helper activity was compared with other KLH carrier-specific clones in an in vitro antibody synthesis assay. The Mls-KLH-restricted T cell clone, in contrast to other carrier-specific, major histocompatibility complex (MHC)-restricted T cell clones, was unable to cooperate with trinitrophenyl (TNP)-primed B cells in the presence of TNP-KLH to generate an anti-TNP response. These experiments suggest that non-MHC determinants, such as autologous Mls gene products, may play a role in genetically restricted antigen recognition by T lymphocytes.     

Reduction in susceptibility to natural killer cell-mediated lysis of human FO-1 melanoma cells after induction of HLA class I antigen expression by transfection with B2m gene

Induction of HLA class I antigens on cultured melanoma cells FO-1 after transfection with a human or a mouse B2m gene was associated with a statistically significant reduction in their susceptibility to natural killer (NK) cell-mediated lysis. These results indicate that the structural differences between human and mouse beta 2-mu do not abolish the ability of the HLA class I molecular complex to modulate NK cell-mediated lysis of melanoma cells FO-1. The role of HLA class I antigens in the phenomenon is corroborated by the ability of anti-HLA class I MAb to enhance, although to a different extent, the susceptibility of transfected FO-1 cells to NK cell-mediated lysis. Gamma interferon (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) significantly reduced the susceptibility to NK cell-mediated lysis of transfected FO-1 cells. Surprisingly, TNF-alpha reduced the extent of lysis more than IFN-gamma, although the latter cytokine enhanced HLA class I antigen expression more than the former one. This finding, in conjunction with a reduction in the susceptibility to NK cell-mediated lysis of untransfected FO-1 cells incubated with IFN-gamma or TNF-alpha, suggests that the two cytokines reduce NK cell-mediated lysis of transfected cells by modulating not only the expression of HLA class I antigens, but also that of other structures. Induction of HLA class I antigens and their modulation with IFN-gamma did not affect the susceptibility to lymphokine-activated killer (LAK) cell-mediated lysis of transfected FO-1 cells. Characterization of the molecular mechanism(s) underlying abnormalities in HLA class I antigen expression by melanoma cells and of the role of these molecules in the interactions of melanoma cells with various types of effector cells may suggest novel immunotherapeutic approaches to melanoma.     

Factors affecting the autologous mixed lymphocyte reaction in kidney transplantation.

In long-term well adapted kidney transplant recipients we have found a close correlation between the T helper (TH):T suppressor/cytotoxic (TS/C) subset ratios and the presence of T cells that respond in the autologous mixed lymphocyte reaction (AMLR). In 21 recipients with T cell E rosette levels ranging between 53 and 86% and TH:TS/C ratios between 0.15 to 2.10, ratios of greater than 0.8 correlated with AMLR responses (13/13), and ratios of less than 0.8 with AMLR nonreactivity (7/7). By contrast, the allogeneic MLR showed no apparent correlation with the TH:TS/C ratios or with the AMLR pre- or postoperatively. It was found that the AMLR in 22 of 23 normal individuals was markedly inhibited by autologous T cells obtained from peripheral blood lymphocytes, exposed to 3,000 rad (Tx) and added as a third component to the cultures. In contrast, 13 of 13 kidney transplant recipients failed to exhibit this Tx AMLR inhibitory cell population. The "naturally occurring" T inhibitory cells, fractionated by an affinity column chromatography procedure into x-irradiated TH and TS/C subsets, inhibited the AMLR to the same extent as unseparated Tx cells. In cell interchange studies performed in four of five HLA identical donor-recipient pairs the Tx cells of the (normal) donor inhibited the recipient AMLR (immunosuppressed), but recipient Tx cells failed to inhibit the donor AMLR. Finally T cells, primed in AMLR and allogeneic MLR for 10 d were tested for AMLR or allogeneic MLR inhibitory activity. Allogeneic MLR primed x-irradiated cells, inhibited both the AMLR and allogeneic MLR while AMLR x-irradiated primed cells inhibited neither reaction. The Tx AMLR inhibitor found in normal peripheral blood, appears to be a cell that is highly sensitive to the effects of biologic or pharmacologic immunosuppressive agents.     

Inhibition of T cell responses by activated human CD8+ T cells is mediated by interferon-gamma and is defective in chronic progressive multiple sclerosis.

The autologous mixed lymphocyte reaction (AMLR) involves the activation of T cells by autologous antigen presenting cells. Cells are generated during the course of the AMLR that have suppressive properties in vitro. In the present study we investigated the induction of CD8+ T cells in the AMLR with suppressive properties and the mechanism by which these cells downregulate in vitro proliferative responses. Purified CD8+ but not CD4+ T cells activated in the AMLR in conditioned medium inhibited proliferation of autologous T cells by anti-CD3 or PPD. Nonactivated CD8+ T cells did not suppress. The CD8+ T cells activated in the AMLR in the presence of conditioned medium (CD8+ Tact) were CD11b negative and were noncytotoxic. The inhibitory effect of CD8+ Tact cells was completely abrogated by anti-IFN-gamma antibody, but not by anti-IL-4, anti-IL-10, or anti-TGF-beta antibody. The induction of CD8+ Tact cells in the AMLR was blocked by anti-IL-2 or by anti-GM-CSF antibody and the combination of these two recombinant cytokines could support the induction of suppressive CD8+ Tact cells. CD8+ Tact cells were defective in patients with chronic progressive multiple sclerosis (MS) as compared to patients with relapsing-remitting MS or normal controls. Our studies provide a basis for understanding the mechanism of suppression by human CD8+ T cells in terms of specific cytokines, and demonstrate the potential importance of these cells in a human autoimmune disease as their function is defective in patients with progressive MS.