Soluble Suppressor Factor Research Articles

Human NK cells: From surface receptors to clinical applications

Natural killer (NK) cells play a major role in innate defenses against pathogens, primarily viruses, and are also thought to be part of the immunosurveillance against tumors. They express an array of surface receptors that mediate NK cell function. The human leukocytes antigen (HLA) class I-specific inhibitory receptors allow NK cells to detect and kill cells that have lost or under-express HLA class I antigens, a typical feature of tumor or virally infected cells. However, NK cell activation and induction of cytolytic activity and cytokine production depends on another important checkpoint, namely the expression on target cells of ligands recognized by activating NK receptors. Despite their potent cytolytic activity, NK cells frequently fail to eliminate tumors. This is due to mechanisms of tumor escape, determined by the tumor cells themselves or by tumor-associated cells (i.e. the tumor microenvironment) via the release of soluble suppressive factors or the induction of inhibitory loops involving induction of regulatory T cells, M2-polarized macrophages and myeloid-derived suppressor cells. The most important clinical application involving NK cells is the cure of high-risk leukemias in the haplo-identical hematopoietic stem cell transplant (HSCT) setting. NK cells originated from hematopoietic stem cells (HSC) of HLA-haploidentical donors may express Killer Immunoglobulin-like receptors (KIRs) that are mismatched with the HLA class I alleles of the recipient. This allows NK cells to kill leukemia blasts residual after the conditioning regimen, while sparing normal cells (that do not express ligands for activating NK receptors). More recent approaches based on the specific removal of TCR α/β+ T cells and of CD19+ B cells, allow the infusion, together with CD34+ HSC, of mature KIR+ NK cells and of TCR γ/δ+ T cells, both characterized by a potent anti-leukemia activity. This greatly reduces the time interval necessary to obtain alloreactive, KIR+ NK cells derived from donor HSC. Another promising approach is based on the use of anti-KIR blocking monoclonal antibodies (mAbs), rendering alloreactive any KIR+ NK cells.

The CD8-Derived Chemokine XCL1/Lymphotactin Is a Conformation-Dependent, Broad-Spectrum Inhibitor of HIV-1

CD8+ T cells play a key role in the in vivo control of HIV-1 replication via their cytolytic activity as well as their ability to secrete non-lytic soluble suppressive factors. Although the chemokines that naturally bind CCR5 (CCL3/MIP-1α, CCL4/MIP- 1β, CCL5/RANTES) are major components of the CD8-derived anti-HIV activity, evidence indicates the existence of additional, still undefined, CD8-derived HIV-suppressive factors. Here, we report the characterization of a novel anti-HIV chemokine, XCL1/lymphotactin, a member of the C-chemokine family that is produced primarily by activated CD8+ T cells and behaves as a metamorphic protein, interconverting between two structurally distinct conformations (classic and alternative). We found that XCL1 inhibits a broad spectrum of HIV-1 isolates, irrespective of their coreceptor-usage phenotype. Experiments with stabilized variants of XCL1 demonstrated that HIV-1 inhibition requires access to the alternative, all-β conformation, which interacts with proteoglycans but does not bind/activate the specific XCR1 receptor, while the classic XCL1 conformation is inactive. HIV-1 inhibition by XCL1 was shown to occur at an early stage of infection, via blockade of viral attachment and entry into host cells. Analogous to the recently described anti-HIV effect of the CXC chemokine CXCL4/PF4, XCL1-mediated inhibition is associated with direct interaction of the chemokine with the HIV-1 envelope. These results may open new perspectives for understanding the mechanisms of HIV-1 control and reveal new molecular targets for the design of effective therapeutic and preventive strategies against HIV-1.

Porcine regulatory T cells: Mechanisms and T-cell targets of suppression

Tregs are known for their suppressive capacity on various immune reactions. In swine, existence as well as suppressive activity of Foxp3 + Tregs could be demonstrated but detailed functional investigations are lacking. Therefore, we analysed the functional properties of porcine Tregs. We observed that besides TCR stimulation Tregs require IL-2 for activation. Furthermore, we investigated the following mechanisms of suppression: (i) cell–cell contact dependency, (ii) production of soluble suppressive factors and (iii) competition for growth factors. Our experiments revealed that suppression by porcine Tregs is abrogated by blocking cell–cell contact or by supplementing excessive amounts of IL-2. Additionally it could be shown that porcine Tregs produce immunosuppressive IL-10. Thereby, we demonstrated that porcine Tregs can use all main mechanisms of suppression mentioned above. Further investigations on the suppressive activity of Tregs using CFSE proliferation assays demonstrated that suppression affects T-helper cells as well as cytotoxic T lymphocytes and TCR-γδ T cells.

CSF-1–Dependent Red Pulp Macrophages Regulate CD4 T Cell Responses

The balance between immune activation and suppression must be regulated to maintain immune homeostasis. Tissue macrophages (MΦs) constitute the major cellular subsets of APCs within the body; however, how and what types of resident MΦs are involved in the regulation of immune homeostasis in the peripheral lymphoid tissues are poorly understood. Splenic red pulp MΦ (RPMs) remove self-Ags, such as blood-borne particulates and aged erythrocytes, from the blood. Although many scattered T cells exist in the red pulp of the spleen, little attention has been given to how RPMs prevent harmful T cell immune responses against self-Ags. In this study, we found that murine splenic F4/80(hi)Mac-1(low) MΦs residing in the red pulp showed different expression patterns of surface markers compared with F4/80(+)Mac-1(hi) monocytes/MΦs. Studies with purified cell populations demonstrated that F4/80(hi)Mac-1(low) MΦs regulated CD4(+) T cell responses by producing soluble suppressive factors, including TGF-β and IL-10. Moreover, F4/80(hi)Mac-1(low) MΦs induced the differentiation of naive CD4(+) T cells into functional Foxp3(+) regulatory T cells. Additionally, we found that the differentiation of F4/80(hi)Mac-1(low) MΦs was critically regulated by CSF-1, and in vitro-generated bone marrow-derived MΦs induced by CSF-1 suppressed CD4(+) T cell responses and induced the generation of Foxp3(+) regulatory T cells in vivo. These results suggested that splenic CSF-1-dependent F4/80(hi)Mac-1(low) MΦs are a subpopulation of RPMs and regulate peripheral immune homeostasis.

Generation and expansion of regulatory human CD4 + T-cell clones specific for pancreatic islet autoantigens

Autoantigen-specific regulatory T cells (Treg) are a potential cell therapy for human autoimmune disease, provided they could be generated in adequate numbers and with stable function. To this end, we determined the feasibility of cloning and expanding human CD4 + Treg specific for the type 1 diabetes autoantigens, GAD65 and proinsulin. Blood CD4 + cells stimulated to divide in response to GAD65 (in three healthy individuals) or proinsulin (in one type 1 diabetic) were flow sorted into single cells and cultured on feeder cells in the presence of anti-CD3 monoclonal antibody, IL-2 and IL-4. Clones were expanded over 4–6 weeks and screened for autoantigen-dependent suppression of tetanus toxoid-specific T-cell proliferation. Suppression by Treg clones was then confirmed against autoantigen-specific non-Treg clones. Of a total of 447 clones generated, 98 (21.9%) had autoantigen-dependent suppressor function. Treg clones were anergic but proliferated to autoantigen after addition of IL-2 or in co-culture with stimulated bulk T cells, without loss of suppressor function. Treg clones were stored over liquid N 2, thawed and further expanded over 12 days, whereupon they exhibited decreased suppressor function. Expansion of Treg clones overall was in the order 10 7–10 8-fold. Treg clones were not distinguished by markers of conventional CD4 +CD25 + Treg and suppressed independently of cell–cell contact but not via known soluble suppressor factors. This study demonstrates that autoantigen-specific CD4 + Treg clones with potential application as a cell therapy for autoimmune disease can be generated and expanded from human blood.

187 Autoresistance to X4 HIV Infection by soluble suppressor factors secreted from CD4+ T cells

187 Autoresistance to X4 HIV Infection by soluble suppressor factors secreted from CD4+ T cells

IFN-γ and TNF-α differentially regulate immunomodulation by murine mesenchymal stem cells

Murine mesenchymal stem cells (MSC) have the ability to inhibit allogeneic immune responses. Two different mechanisms, either cell contact-dependent or independent, have been proposed to account for this immunosuppression. The focus of this study was to elucidate the involvement of soluble suppressive factors secreted by murine MSC in an inflammatory setting, and their role in MSC immunomodulation. In a non-inflammatory environment, bone marrow derived murine MSC constitutively expressed low levels of COX-2, PGE-2, TGF-β1 and HGF, but not IL-10, PD-1, PD-L1 or PD-L2. These MSC were able to significantly reduce alloantigen driven proliferation in mixed lymphocyte reactions as well as mitogen driven proliferation. The pro-inflammatory cytokines IFN-γ and TNF-α did not ablate MSC mediated immunosuppression. MSC expression of PGE-2, IDO and PD-L1 was differentially regulated by these cytokines. COX-2 and PGE-2 expression by MSC were upregulated by both IFN-γ and TNF-α, and using a biochemical inhibitor this was shown to have an essential, non-redundant role in modulating alloantigen-driven proliferation. However, the surface expression of PD-L1 was induced by IFN-γ but not TNF-α and similarly functional IDO expression was only induced by IFN-γ stimulation. Blocking studies using neutralising antibodies and biochemical antagonists revealed that while PD-L1 induction was not essential, IDO expression was a prerequisite for IFN-γ mediated MSC immunomodulation. These data demonstrate that murine MSC expression of immunomodulatory factors dramatically changes in a pro-inflammatory environment and that IFN-γ in particular has an important role in regulating MSC immunomodulatory factor expression.

Autoactivation of matriptase in vitro: requirement for biomembrane and LDL receptor domain

In live cells, autoactivation of matriptase, a membrane-bound serine protease, can be induced by lysophospholipids, androgens, and the polyanionic compound suramin. These structurally distinct chemicals induce different signaling pathways and cellular events that somehow, in a cell type-specific manner, lead to activation of matriptase immediately followed by inhibition of matriptase by hepatocyte growth factor activator inhibitor 1 (HAI-1). In the current study, we established an analogous matriptase autoactivation system in an in vitro cell-free setting and showed that a burst of matriptase activation and HAI-1-mediated inhibition spontaneously occurred in the insoluble fractions of cell homogenates and that this in vitro activation could be attenuated by a soluble suppressive factor(s) in cytosolic fractions. Immunofluorescence staining and subcellular fractionation studies revealed that matriptase activation occurred in the perinuclear regions. Solubilization of matriptase from cell homogenates by Triton X-100 or sonication of cell homogenates completely inhibited the effect, suggesting that matriptase activation requires proper lipid bilayer microenvironments, potentially allowing appropriate interactions of matriptase zymogens with HAI-1 and other components. Matriptase activation occurred in a narrow pH range (from pH 5.2 to 7.2), with a sharp increase in activation at the transition from pH 5.2 to 5.4, and could be completely suppressed by moderately increased ionic strength. Protease inhibitors only modestly affected activation, whereas 30 nM (5 microg/ml) of anti-matriptase LDL receptor domain 3 monoclonal antibodies completely blocked activation. These atypical biochemical features are consistent with a mechanism for autoactivation of matriptase that requires protein-protein interactions but not active proteases.

Age-Related Induction of CD4+ T-Cell Unresponsiveness Is Reversible and Dependant of the Host's Environment.

A large number of CD4+ T-cells from either aged mice or humans display surface markers associated with an activated/memory phenotype. In spite of these changes however, these T-cells have a markedly decreased ability to proliferate and produce IL-2 in response to antigen stimulation in vitro. The cellular and molecular mechanisms involved in this age-related unresponsiveness of the CD4+ T-cell compartment remain poorly understood. Utilizing a well-established experimental system in which transgenic CD4+ T cells specific for a MHC class II restricted epitope of influenza hemagglutinin (HA) are adoptively transferred into non-transgenic recipients, we have previously elucidated important mechanisms involved in the induction and maintenance of CD4+ T-cell tolerance. Our studies were however limited to the analysis of T-cell function in lymphoma bearing young mice (4 to 10 weeks old). Here, we assessed the influence of the aged microenvironment in determining the phenotype and function of antigen-specific T-cells. CD4+ T-cells from young TCR transgenic mice (2 months old) were adoptively transferred into either old (20–24 months) or young (2 months old) non-transgenic mice. Two weeks later, clonotypic and non-clonotypic CD4+ T-cells were isolated from the spleens of these animals and their phenotype and function were determined in vitro. Reminiscent of the age-related changes observed within the normal CD4+ T-cell repertoire, young transgenic T-cells transferred into aged hosts have acquired an activated/memory phenotype but displayed a significant impairment in antigen-specific proliferation and IL-2 production in response to cognate antigen in vitro. These changes were not due to homeostatic proliferation of the transferred T-cells into the relatively lymphopenic aged host. To determine whether the changes observed in “aged” T-cells were reversible or not, we adoptively transfer old T-cells back into young hosts or into control old mice. While old transgenic T-cells transferred into an old environment remained fully unresponsive, the adoptive transfer of the same old T-cells into a young host restored their ability to proliferate and produce IL-2. Surprisingly, these “old” T-cells were able to produce significantly higher levels of IFN-gamma indicative of their memory/effector phenotype. Furthermore, young animals adoptively transferred with “aged” antigen-specific T-cells were now capable of rejecting A20 B-cell lymphomas expressing HA as a model tumor antigen (A20HA). Taking together, factor(s) present in the aged microenvironment are responsible for limiting the effector function of CD4+ T-cells that seem otherwise well equipped to become fully activated if the proper environment is provided (young microenvironment). The potential role of soluble suppressive factors as well as regulatory T-cells (Tregs) in the unresponsiveness observed in the T-cell compartment of aged hosts will be discussed.

HIV-1-suppressive factors are secreted by CD4+ T cells during primary immune responses.

CD4+ T cells are required for immunity against many viral infections, including HIV-1 where a positive correlation has been observed between strong recall responses and low HIV-1 viral loads. Some HIV-1-specific CD4+ T cells are preferentially infected with HIV-1, whereas others escape infection by unknown mechanisms. One possibility is that some CD4+ T cells are protected from infection by the secretion of soluble HIV-suppressive factors, although it is not known whether these factors are produced during primary antigen-specific responses. Here, we show that soluble suppressive factors are produced against CXCR4 and CCR5 isolates of HIV-1 during the primary immune response of human CD4+ T cells. This activity requires antigenic stimulation of naïve CD4+ T cells. One anti-CXCR4 factor is macrophage-derived chemokine (chemokine ligand 22, CCL22), and anti-CCR5 factors include macrophage inflammatory protein-1 alpha (CCL3), macrophage inflammatory protein-1 beta (CCL4), and RANTES (regulated upon activation of normal T cells expressed and secreted) (CCL5). Intracellular staining confirms that CD3+CD4+ T cells are the source of the prototype HIV-1-inhibiting chemokines CCL22 and CCL4. These results show that CD4+ T cells secrete an evolving HIV-1-suppressive activity during the primary immune response and that this activity is comprised primarily of CC chemokines. The data also suggest that production of such factors should be considered in the design of vaccines against HIV-1 and as a mechanism whereby the host can control infections with this virus.

Impairment of natural killer (NK) cytotoxic activity in hepatitis C virus (HCV) infection.

In the present study, we evaluated the NK cell cytotoxic activity in a group of HCV-infected individuals. Although the number of NK cells present in the peripheral blood of the HCV-infected patients was comparable to non-infected individuals, spontaneous NK cytotoxicity was four-fold lower (P < 0.001) than in normal donors. This functional impairment was not overcome by depletion of adherent or B cells, and it was partially restored by short-term (18 h) stimulation with IL-2. However, long-term stimulation (72 h) with this lymphokine induced activated killer cell (LAK) activity comparable to normal controls. The reduction in NK cytotoxic response does not seem to be due to soluble suppressive factors, since incubation of normal peripheral blood mononuclear cells (PBMC) with infectious HCV serums for a 4-h period does not affect NK spontaneous cytotoxic activity. Successful in vitro infection of PBMC with HCV infectious serum also resulted in an impairment of NK cytotoxicity, suggesting that altered NK function is associated with HCV infection and may be responsible, at least in part, for the chronicity of the infection.

Suppressor cell activity of ovine caruncular and intercaruncular tissues during the placentation period

We assessed the suppression of lymphocyte proliferation ovine endometrial cells recovered during each trimester (Days 45, 90, and 135) of pregnancy. We evaluated fractionated and unfractionated caruncular (C) and intercaruncular (IC) cells for suppression of cocultured peripheral blood lymphocytes (PBL) in phytohemagglutinin (PHA)-treated cultures. We also evaluated cells for the release of the suppressor factor into the culture medium and tested the factor for transforming growth factor-β (TGF-β) activity. Suppression of PHA-induced proliferation of PBL was evident for C and IC cells recovered from each day of pregnancy, and the activity was predominantly attributed to the population(s) of low-density (1.006–1.054 g/ml) cells. The activity was greater for unfractionated C than for IC cells on Day 45, whereas the pattern was reversed by Day 135 of pregnancy. For the C cells, the activity was greatest on Day 45 and least by Day 135. Although suppressor factor was released into the medium from cultured C and IC cells, its activity was not apparently mediated by TGF-β. In conclusion, we observed a temporal pattern in suppressor activity for unfractionated endometrial cells during pregnancy. Suppression was predominately mediated by a population(s) of low-density cells. In addition, the cells released a soluble suppressor factor that seems to be unrelated to TGF-β. The suppressor cells may provide immunological protection to the fetoplacental unit by suppressing specific lymphocyte responses directed toward conceptus tissues.

IL-12 Reverses Established Tolerance Mediated by Tcrαlβ+ but not by Tcrγδ+ Suppressor T Cells

Topical cutaneous painting with chemically reactive haptens induces the ability to subsequently elicit contact sensitivity (CS) responses in the skin. These CS responses are in vivo examples of acquired, antigen (Ag)-specific T cell immunity, and are a form of delayed-type hypersensitivity (DTH). In contrast, high dose i.v. administration of the hapten can induce Ag-specific tolerance. In some instances this specific immune hyporeactivity is due to suppressor T cells. We investigated the effect of IL-12 on reversal of tolerance due to suppressor T cells that were induced by i.v. administration of hapten in either normal TCRα+/+, or in immunodeficient TCRα−/- mice. In the TCRα+/+ mice, tolerance is mediated by TCRαβ+ suppressor T cells, while in the TCRα−/- mice the tolerance is due to suppressive TCRγδ+ cells. Treatment with IL-12 reversed suppressor mediated by the TCRαβ+ cells, but did not affect tolerance due to TCRγδ+ suppressor cells. Another difference was that the αβTCR+ suppressor cells produced a soluble suppressor factor that could replace the surppressor cells, while γδTCR+ suppressor cells did not. We hypothesized that IL-12 may strengthen responses of target CS-effector T cells influenced by the hapten-MHC-specificity of αβ suppresssor cells, or suppressor factor. On the other hand, γδTCR+ suppressive cells likely have specificity for the hapten alone, and are not MHC-restricted, and therefore probably do not operate via peptide-MHC interactions, that could be strengthened by IL-12. The ability of IL-12 to strengthen the resistance of CS-effector T cells to αβ TCR suppressor cells, may be due to the ability of IL-12 to increase T cell costimulation mediated by signaling mechanisms acting via B7.1 and B7.2. In contrast, αβTCR+ suppressor cells, that are largely hapten-specific, probably do not interact with peptide/MHC complexes on APC, and thus are not affected by IL-12 strengthening of co-stimulation.

Implication of macrophage inflammatory protein-1alpha in the inhibition of human haematopoietic progenitor growth by dengue virus.

The mechanisms were investigated of haematopoietic progenitor growth inhibition, observed after in vitro infection of cord blood mononuclear cells (CBMNC) by a clinical isolate of dengue 3 (29-56DSS). The level of virus replication was not different when CBMNC were inoculated with 29-56DSS compared with a prototype strain of dengue 3 (H-87) which had no inhibitory effect. An inhibitory effect was also observed when cell-free and heat-inactivated supernatants from 29-56DSS cultures, but not from H-87 cultures, were added to cultures of normal CBMNC, suggesting an indirect mechanism via the release of soluble suppressive factor(s). Macrophage inflammatory protein-1alpha (MIP-1alpha) was detected at a significantly higher level in 29-56DSS cultures than in controls. Blocking experiments with anti-MIP-1alpha antibody demonstrated that the inhibitory effect was related at least partly to high MIP-1alpha levels. To our knowledge, this is the first report suggesting an indirect effect of dengue infection on haematopoiesis mediated by a suppressive cytokine.

CD4 T suppressor cells mediate interferon tau protection against experimental allergic encephalomyelitis

Interferon tau is a type I IFN that was originally identified as a pregnancy recognition hormone produced by trophoblast cells. It is as potent an antiviral agent as IFN α and IFN β, but lacks the toxicity associated with high concentrations of these IFNs in tissue culture and in animal studies. We recently showed that IFN τ, like IFN β, can prevent the development of experimental allergic encephalomyelitis (EAE). We report here that IFN τ prevents EAE in mice by induction of suppressor cells and suppressor factors. Suppressor cells can be induced by IFN τ in tissue culture, and in vivo by either intraperitoneal injection or by oral administration to mice. Incubation of suppressor cells with myelin basic protein (MBP)-sensitized T cells blocked or delayed the MBP-induced proliferation. Further, intraperitoneal injection of suppressor cells into mice blocked induction of EAE by MBP. Suppressor cells possessed the CD4 T cell phenotype, and produced soluble suppressor factors that inhibited MBP activation of T cells from EAE mice. The suppressor factors were found to be IL-10 and TGF β, which acted synergistically to inhibit the MBP activation of T cells from EAE mice. These findings are important for understanding the mechanism(s) by which type I IFNs protect against autoimmune disease.

Analysis of T-cell receptor alpha beta chains of CD8+ suppressor T cells induced by tolerogenic conjugates of antigen and monomethoxypolyethylene glycol. Involvement of TCR alpha-CDR3 domain in immunosuppression.

A number of investigators have demonstrated that there exists a relationship between Ag receptors of Ts cells (TCR) and their soluble suppressor factors. With a view to elucidating this relationship, the primary structures of receptors of Ts cells, induced in mice by tolerogenic conjugates of Ag and monomethoxypolyethylene glycol, were characterized in this study. The cDNA encoding the alpha and beta chains of TCR of cloned Ts cells specific for (i) OVA and (ii) human monoclonal (myeloma) IgG (HIgG) were produced by polymerase chain reaction. From the analysis of the V alpha genes of TCR of Ts cells it was deduced that these receptors utilized a new member of the V alpha 15 gene family, which was productively joined to the J alpha genes that differed for each of the Ts cells of the two distinct specificities. Similarly, sequence analysis of the beta chain cDNA of the two Ts cell clones revealed that both clones utilized the V beta 8.2 gene, and that their J beta gene differed from each other. It is inferred that the Ts cells generated in response to the different tolerogenic Ag(mPEG)n conjugates belonged to a subset of T cells utilizing similar TCR alpha beta chains and differed only in their J alpha/J beta regions. Most importantly, pretreatment of mice with a mixture of pentadecapeptides comprising the TCR alpha chain of the OVA-Ts cells, down-regulated the immune response specific to OVA, but not to HIgG. Moreover, injection of mice with a pentadecapeptide corresponding to the CDR3 region of the TCR alpha chain of either OVA-Ts or HIgG-Ts suppressed specifically the Ab response to OVA or HIgG, respectively. On the basis of all these results, it is concluded that the CDR3 of the TCR-alpha chain of Ts cells plays a pivotal role in the Ts network underlying the specific down-regulation of the immune responses induced by tolerogenic Ag(mPEG)n conjugates.

Altered cytokine production and accessory cell function after HIV-1 infection.

We investigated cytokine production and accessory cell function in human macrophage hybridoma cell lines and primary monocytes after infection with HIV-1. HIV-1 infection induced IL-10 production in the macrophage hybridoma cell line with loss of IL-12 1 wk after infection. There were also significant increases in production of IL-10 (537 +/- 521 vs 687 +/- 625 pg/ml) while there was a reduction in IL-12 (6.3 +/- 3.1 vs 1.2 +/- 1.0 pg/ml, p = 0.021) in the primary monocytes 5 days after HIV-1 infection. In addition, the hybridoma cell lines and primary monocytes failed to support PHA, Con A, PWM, or anti-CD3- induced T cell proliferation 1 wk after infection. The viability of the T cells cocultured with the HIV-1-infected macrophage cell lines or the primary monocytes as determined by propidium iodide staining was unaltered and there was no increase in apoptosis-specific DNA strand breaks or increased expression of Bcl-2 in the T cells. No soluble suppressor factor was present, since UV-inactivated supernatants from the hybridoma cell line and primary monocytes failed to inhibit mitogen- and anti-CD3-induced T cell proliferation. Early events in T cell activation, including calcium flux and phosphotyrosine kinase activity, were intact in the T cells cocultured with the HIV-1- infected hybridomas and monocytes but there was reduced IL-2 production. Addition of exogenous IL-2 restored the proliferative responses. Taken together, these data suggest that alteration of cytokine production and accessory cell function for mitogens and anti-CD3-induced T cell proliferation independent of induction of apoptosis, suppressor factor production, or inhibition of T cell signaling occurs very early after HIV-1 infection and may contribute to the global immunosuppression observed in AIDS.

Soluble HIV suppressive factors: more than one Holy Grail?

Soluble HIV suppressive factors: more than one Holy Grail?

Establishment of a natural suppressor cell line producing soluble suppressor factor other than transforming growth factor-beta.

Natural suppressor (NS) cell line (Clone 59) was established from the bone marrow of adult C3H/Hej mice in the presence of WEHI-3 conditioned media. Clone 59 cells suppressed the generation of cytotoxic T lymphocytes from normal mouse spleen. This suppression was seen at a responder-to-suppressor cell ratio of 1000:1 and lacked antigen specificity or MHC restriction. Clone 59 cells expressed the 'null' surface phenotype (Thy1.2-, CD3-, Lyt-2-, L3T4-, surface Ig-, MAC-1-) by immunofluorescent staining. Clone 59 cells exhibited no cytolytic activity against NK cell-sensitive YAC-1 and natural cytotoxic L929 target cell lines. Non-specific suppression, with a cell-free supernatant from the Clone 59-NS cells, also was observed. The supernatant did not inhibit [3H]-thymidine uptake by CTLL-2 cells which were proliferating in response to IL-2. Anti-transforming growth factor-beta (TGF-beta) monoclonal antibody had no effect on suppression, suggesting that the non-specific suppression is mediated by some soluble factors other than TGF-beta. Clone 59 cells may be useful in identifying non-specific suppressor cells in adult bone marrow and studying their functional role in the regulation of tolerance and self-reactivity.

Macrophage colony stimulatory factor-activated bone marrow macrophages suppress lymphocytic responses through phagocytosis: a tentative in vitro model of Rosai-Dorfman disease.

NBXFO hybridoma cells produced macrophage colony-stimulating factor (M-CSF), which stimulated the growth of murine bone marrow-derived macrophages with potent suppressor activity. These macrophages suppressed lymphocyte responses to mitogens and antigens in a dose-dependent manner. Using a transwell chamber, we demonstrated that macrophages needed physical contact with the lymphocytes to suppress lymphocyte proliferation on day 1 in the concanavalin A mitogen reaction. In addition, no soluble suppressor factor was detected at that time. The number of lymphocytes disappeared with time when they were cocultured with the macrophages. Electron microscopy revealed that the macrophage phagocytosized the lymphocytes after 7 1/2 h. Dextran sulfate, heparan, and fucoidan prevented the macrophages from suppressing the lymphocytes. This phenomenon resembles the human disease sinus histiocytosis, also called Rosai-Dorfman disease, in which macrophages (histiocytes) phagocytosize autologous lymphocytes; occasionally, this disease is associated with immunological abnormalities. Thus we believed that macrophage-activating cytokines, such as M-CSF, may stimulate macrophages to phagocytose lymphocytes in vivo.