Studies on the relationships between the immunogenicity and catabolism of antigens and their binding to the surface of macrophages (original) (raw)
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European Journal of Immunology, 1978
We have investigated the sequential interaction of macrophages, initiator T lymphocytes (ITL) and recruited T lymphocytes (RTL) in the development of a T cell-mediated response t o soluble antigens. Macrophages were pulsed with soluble antigens and used t o sensitize ITL in vitro. The ITL were irradiated t o prevent their proliferation and then injected into the footpads of syngeneic mice. Sensitized ITL were found t o recruit immunospecific RTL in the draining lymph nodes, as determined by a thymidine uptake assay of the lymph node cells in vitro. The richest source of lymphocytes with ITL activity was the thymus, and progressively less activity was detectable among spleen or lymph node lymphocytes. The magnitude of the subsequent RTL response could be modified by genetic differences between the ITL and the antigen-pulsed macrophages that were used t o sensitize them. Thus, ITL conveyed an immunogenic signal t o RTL whose magnitude reflected the genotype of the macrophages, but whose specificity was directed by determinants of the soluble antigen.
Cellular Immunology, 1985
MA 158.2, a rat monoclonal antibody with binding specificity for cells of the monocyte-macrophage lineage, reacts with an antigen (I 58.2) whose expression is enhanced on mononuclear cells activated to the tumoricidal phenotype by treatment with lymphokine supematant containing macrophage activating factor (MAF). The functional relevance of enhanced expression of this antigen has been examined in mouse peritoneal macrophages treated with a variety of immunomodulatory agents and assayed for augmented macrophage-mediated defense reactions, including 0; production, microbicidal, and tumoricidal activity. An interferon-y (IFN-7) preparation produced by recombinant DNA technology induced a dose-dependent increase in expression of the 158.2 antigen in inflammatory macrophages which was accompanied by acquisition of microbicidal activity against Listeria monocytogenes. However, these cells did not express tumoricidal activity and induction of this property required concomitant exposure to lipopolysaccaharide (LPS). Similar results were obtained using macrophages elicited with pyran copolymer. Exposure to LPS alone induced enhanced expression of antigen 158.2 but did not elicit microbicidal activity. Macrophages challenged with IFN-a, IFN-& MDP, and bestatin did not exhibit increased 158.2 and also failed to acquire tumoricidal activity when treated concomitantly with LPS. Collectively, these data indicate that the MA 158.2 antibody recognizes an antigen expressed by macrophage populations displaying the so-called primed phenotype in which microbicidal activity is expressed but in which induction of tumoricidal activity requires the addition of a second signal such as LPS. o 1985 Academic PBS. Inc.
Activation of a galactosidase-specific murine T hybridoma clone and of a human tetanus toxoid-specific T clone by antigen-presenting cells (APC) was used to evaluate the regulatory function ofantibodies complexed with the relevant antigen . Complexed antigen, in fact, is taken up with high efficiency thanks to Fc receptors borne by APC . Antibody/antigen ratio in the complexes proved to be a critical parameter in enhancing antigen presentation . Complexes in moderate antibody excess provided optimal T cell activation independently of the physical state of the complexes {precipitated by a second antibody or solubilized by complement) . Complexes in extreme antibody excess, on the contrary, did not yield T cell activation although taken up by APC efficiently. The effect of antibodies at extreme excess was observed with substimulatory dose of antigen (loss of potentiation) and with optimal dose of antigen (loss of stimulation). An excess of specific polyclonal antibodies hampers proteolytic degradation of antigen in vitro, supporting the view that a similar mechanism may operate within the APC that have internalized immune complexes in extreme antibody excess. The possibility that immune complex forming in extreme antibody excess may turn off the T cell response is proposed as a regulatory mechanism .
Journal of Experimental Medicine, 1975
Culture fluids rich in mononuclear phagocytes have a powerful effect on various in vitro reactions of lymphocytes (1-9). The culture fluids contain a mitogen for thymocytes and mature lymphocytes . Also found are active principles which differentiate memory B cells into antibody-secreting cells and which increase helper activities of T lymphocytes (2, 3). In a previous report we obtained from experiments in which lymphocytes were depleted from the cell preparations, strong evidence that the cellular source of the active component (s) was the macrophage (3). The activity secreted into the medium, however, was variable from experiment to experiment, perhaps related to the state of macrophage activation. In this paper we present results of experiments in which we studied the relation between macrophage activation and/or stimulation with the secretion of the various activities. We have found two ways in which to generate high levels of lymphostimulatory activities in macrophage cultures: one is to add a series of materials that are readily taken up by the macrophages; the second is to add a small number of activated T cells to the macrophage-rich cultures.
Journal of Experimental Medicine, 1976
Cultures of peritoneal exudate cells rich in macrophages were studied for the secretion of lymphostimulatory molecules. Two conditions produced increased secretion: (a) addition to the cultures of various agents that readily interacted with macrophages, such as latex particles, antibody-coated red cells, endotoxin, Listeria organisms, or Be salt; or (b) addition of activated lymphocytes. In the first case the increased activity was found during the first 24 or 48 h after uptake of the stimuli. Increased activity was found in normal or peptone-stimulated macrophages but not in macrophages after injection of endotoxin or thioglycollate. The addition of T lymphocytes from Listeria-infected mice to macrophage cultures increased greatly the activities. This increase was also produced by addition to antigen-primed T cells together with antigen. The lymphocytes by themselves did not secrete active factors. The lymphostimulatory activities were tested on thymocyte DNA synthesis and on antib...
Immunology, 1978
The mode of interaction with macrophages of two ordered synthetic polypeptides (Tyr-Tyr-Glu-Glu)-poly(DLAla)--poly(Lys), (T-T-G-G)-A--L, and (Tyr-Glu-Tyr-Glu)-poly(DLAla)--poly(Lys), (T-G-T-G)-A--L, which differ in their requirements for T-B cell co-operation in the process of antibody production, was compared. The binding of the two radiolabelled antigens to the surface of peritoneal adherent cells, their uptake by the cells and the rate of their degradation were investigated. Macrophages were found to be capable of degrading both poly-peptides with the same efficiency. (T-G-T-G)-A--L, the antigen which is less T-dependent, was bound to macrophage surfaces more readily than (T-T-G-G)-A--L, the T-dependent antigen, however, its uptake by the cells was found to be lower. Thus, (T-G-T-G)-A--L remains for a longer period in the form of a membrane bound polyvalent antigen.
The combined use of in vitro culture techniques together with efficient cell separation methods have revealed that macrophages are essential participants in the initiation of some antibody responses (1-4). The experimental design used was to obtain "purified" (macrophage-depleted) lymphocytes by a cell separation procedure and to demonstrate that these cells could not respond to a particular antigen unless macrophages were also present. Several mechanisms of macrophage function during the induction of the humoral immune response have been proposed. Since it was found that in vitro responses to particulate antigens, such as whole erythrocytes, required the participation of macorphages, whereas responses to smaller sized antigens, such as "soluble sheep red blood cell antigen" (3), or polymeric flagellin of Salmonella adelaide (POL) 1 did not, it was proposed that the role of macrophages may therefore be merely to reduce antigenic particles to a smaller and more immunogenic size. Experiments demonstrating that macrophage supernatants enhance responses to erythrocytes (5, 6) are consistent with this simple mechanism of macrophage function. Recently it was found that the immune response to thymus-dependent antigens, even those of small size, such as monomeric flagellin (MON), dinitrophenylated MON (DNP MON), or DNP fowl gamma globulin (DNP F~G), all required the presence of macrophages, whereas responses to the same antigenic determinants (DNP) on polymeric carriers (i.e. POL or DNP POL) which are thymus independent were also found to be
Analysis of Antibody Induction by Macrophages Treated Ex Vivo with Human Proteins in Mice
Reports of Biochemistry and Molecular Biology
Background: Macrophages are essential cellular components in various body tissues and tumor microenvironments. The high infiltration of macrophages into the tumor microenvironment determines the importance of ex vivo treatment of personalized macrophages with recombinant cytotoxic T-lymphocyteassociated protein 4 (rCTLA-4), programmed death-ligand 1 (rPD-L1), and programmed cell death protein 1 (rPD-1) proteins to block immune checkpoints. Methods: We investigated the development of humoral immunity against CTLA-4, PD-L1, and PD-1 receptors by introducing macrophages treated ex vivo with the corresponding proteins into mice. Peritoneal macrophages from BALB/c mice were cultured in medium containing recombinant human CTLA-4, PD-L1, and PD-1 proteins. Macrophages processing recombinant proteins were analyzed via immunofluorescence staining using antibodies against CTLA-4, PD-L1, and PD-1. The treated macrophages were administered intraperitoneally to mice to induce anti-CTLA-4, anti-PD-L1, and anti-PD-1 antibodies. The antibody titer in vaccinated mice was determined via enzyme-linked immunosorbent assays, followed by statistical analysis of the results. The specificity of the antibodies was determined via immunofluorescence staining in MCF7 cells. Results: The ex vivo treatment of macrophages with rCTLA-4, rPD-L1, and rPD-1 induced the formation of specific antibodies in vaccinated mice. The various rPD-L1 and rPD-1 concentrations used to treat macrophages had no significant effect on the specific antibody titers, while the anti-rCTLA-4 titer was dependent on the protein concentration in the culture medium. Immunofluorescence analysis revealed that anti-CTLA-4 and PD-L1 antibodies reacted with MCF7 cells. Conclusions: The ex vivo treatment of macrophages with rCTLA-4, rPD-L1, and rPD-1 can help induce humoral immunity and develop new approaches for cancer immunotherapy.