ovalbumin and Mast-Cell-Sarcoma

IL-10 is a cytokine secreted by a wide variety of cells type that has pleiotropic stimulatory and suppressive activities on both lymphoid and myeloid cells in vitro. To analyze the consequences of high IL-10 secretion by APCs in immune responses, we produced transgenic mice expressing human IL-10 directed by the MHC class II Ea promoter. Despite alterations in the development of T and B cells, no gross abnormalities were detected in peripheral lymphocyte populations or serum Ig levels. However, when immunized using conditions that give either a Th2-type or a Th1-type response, IL-10 transgenic mice failed to mount a significant T or B cell immune response to OVA. IL-10 transgenic mice were also highly susceptible to infection with intracellular pathogens like Listeria monocytogenes or Leishmania major, in contrast to IL-10 transgenic mice, where the transgene was express in T cells. Finally, the recently described stimulatory effect of IL-10 on CD8+ T cells was confirmed by the ability of IL-10 transgenic mice to limit the growth of immunogenic tumors by a CTL-mediated mechanism. These results demonstrate, that, depending on the type of immune response, IL-10 can mediate immunosuppressive or immunostimulatory activities in vivo.

Topics: Animals; Antigen-Presenting Cells; B-Lymphocytes; Base Sequence; DNA Primers; Humans; Interferon-gamma; Interleukin-10; Leishmania major; Listeria monocytogenes; Lymphocyte Activation; Mast-Cell Sarcoma; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Transgenic; Monocytes; Ovalbumin; Polymerase Chain Reaction; Recombinant Proteins; T-Lymphocytes; Up-Regulation

Using a bipalmitoylated lipopeptide consisting of an ovalbumin helper T-cell epitope covalently linked to an influenza virus cytotoxic T-lymphocyte (CTL) epitope, we addressed possible factors that may be critical for CTL induction. Antigen processing of lipopeptide appears to be required for T-cell induction since there was virtually no in vitro binding of lipopeptide to purified MHC molecules. A major portion of lipopeptide immunogenicity was due to its particulate nature inasmuch as CTL induction in mice correlated with insoluble lipopeptide constructs, whereas more soluble analogs were significantly less immunogenic. Immunohistological analysis of tissue from immunized animals revealed that lipopeptide migration from the s.c. injection site to the spleen could be detected as early as 1 h after immunization and cell-associated lipopeptide was observed on macrophages and dendritic cells, implicating both cell populations in the processing and presentation of lipopeptide particles to CTLs.

Topics: Amino Acid Sequence; Animals; Epitopes, T-Lymphocyte; Female; H-2 Antigens; Histocompatibility Antigens Class II; Influenza Vaccines; Lipoproteins; Lymphocyte Activation; Mast-Cell Sarcoma; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Molecular Sequence Data; Nucleocapsid Proteins; Nucleoproteins; Ovalbumin; Particle Size; Peptide Fragments; RNA-Binding Proteins; T-Lymphocytes, Cytotoxic; Tumor Cells, Cultured; Viral Core Proteins

To determine whether the subretinal space can extend immune privilege to allogeneic tumor cell grafts that do not possess their own inherent immune privilege.. P815 tumor cells were injected into the anterior chamber (AC), the subretinal (SR) space, or subconjunctivally in eyes of BALB/c (allogeneic), SCID (immune incompetent), normal DBA/2 (syngeneic), or DBA/2 mice presensitized with P815 cells transfected with interleukin-12 and B7.1. Tumor growth was observed clinically and histologically for up to 50 days. BALB/c recipients were tested for suppression of DBA/2-specific delayed hypersensitivity and concomitant immunity. The SR space of tumor-containing eyes was assessed for its capacity to support ovalbumin (OVA)-specific anterior chamber associated immune deviation (ACAID).. P815 cells injected into the SR space of presensitized and normal DBA/2 and SCID mice grew progressively, resulting eventually in recipient death. Tumor cells injected into the SR space of eyes of BALB/c mice grew progressively until day 14, followed by tumor regression resulting in phthisis bulbi (14/35) or tumor elimination (19/35) with preserved ocular anatomy by day 35. Despite elimination of tumors from the SR space, BALB/c recipients exhibited DBA/2-specific ACAID and concomitant immunity. In addition, OVA injected into the SR space of eyes from which tumor has been eliminated induced ACAID.. Various parameters of immune privilege, originally described for the AC, are characteristic of immune privilege within the SR space. However, because P815 cells placed in the AC prove lethal for BALB/c recipients, but P815 cells placed in the SR space resolve without jeopardizing the host's life, immune privilege in the SR space can be distinguished from immune privilege in the AC, and this may have implications for grafts of retinal tissue placed within the SR space.

Topics: Animals; Anterior Chamber; Cell Survival; Eye Neoplasms; Female; Hypersensitivity, Delayed; Immunity; Immunization; Mast-Cell Sarcoma; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Mice, SCID; Neoplasm Transplantation; Ovalbumin; Retina; Transplantation, Homologous

To determine whether the subretinal space supports the induction of deviant immune responses to cell-associated and soluble antigens and to elucidate factors influencing the immunologic properties of the subretinal space.. P815 mastocytoma cells were used as cell-associated antigens and were inoculated into the anterior chamber (AC), the vitreous cavity (VC), the subretinal space, and subconjunctivally in H2-compatible, but minor H-incompatible, BALB/c mice. Ovalbumin, as a soluble antigen, was similarly injected into eyes, after which recipient animals were immunized with ovalbumin and complete Freund's adjuvant. Delayed-type hypersensitivity (DTH) was assessed by ear challenge. To alter the conditions in the subretinal space, the outer blood-retinal barrier was disrupted by compromising retinal pigment epithelial (RPE) cells with a systemic injection of sodium iodate. Immune privilege in the AC was abolished by mild corneal cauterization.. Antigen-specific DTH did not develop in mice in which alloantigenic tumor cells or ovalbumin was injected into the AC, the VC, or the subretinal space, and the mice's spleens contained lymphocytes capable of suppressing DTH when adoptively transferred into naive mice. When RPE cells were compromised with sodium iodate, tumor cells or ovalbumin injected into the subretinal space or the VC did not induce immune deviation, although the AC of these eyes still promoted AC-associated immune deviation. By contrast, when immune privilege in the AC was abolished by corneal cauterization, neither tumor cells nor ovalbumin injected into the subretinal space or the VC of eyes elicited immune deviation.. The subretinal space supports immune deviation for histoincompatible tumor cells and soluble protein antigens by actively suppressing antigen-specific DTH. Acute loss of immune privilege in the subretinal space and the VC does not cause loss of privilege in the AC, but abolition of immune privilege in the AC eliminates the capacity of the subretinal space and the VC to support immune deviation to antigens injected locally.

Topics: Adoptive Transfer; Animals; Anterior Chamber; Antigens, Neoplasm; Blood-Retinal Barrier; Extracellular Space; Female; Hypersensitivity, Delayed; Injections; Iodates; Mast-Cell Sarcoma; Mice; Mice, Inbred BALB C; Ovalbumin; Retina; Spleen; T-Lymphocytes; Vitreous Body

DNA immunization has been shown to elicit both antibody and CTL responses against antigens expressed by infectious organisms. Because CTL responses have been implicated in rejection of cancer, we investigated whether DNA immunization by particle bombardment using a gene gun could induce CTL responses that were capable of rejecting tumors in mice. DNA immunization by particle bombardment using genes encoding beta-galactosidase and ovalbumin primed mice to generate CTLs in two genetic backgrounds (DBA/2 and C57BL/6 strains, respectively). DNA immunization was more potent in inducing CTLs than immunization with an optimized regimen of ovalbumin peptide plus immune adjuvant. Immunity induced by DNA immunization protected mice against s.c. challenge with tumors expressing the beta-galactosidase antigen. Tumors were rejected even when DNA immunization was started 3 or 7 days after tumor challenge as tumors were becoming established. Tumor rejection required CD8(+) T cells, confirming a role for CTLs in vivo. These studies show that DNA immunization by particle bombardment can efficiently induce CTL responses that are capable of rejecting even established tumors.

Topics: Animals; beta-Galactosidase; Biolistics; Cancer Vaccines; Injections, Subcutaneous; Mast-Cell Sarcoma; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Ovalbumin; Recombinant Proteins; Spleen; T-Lymphocytes, Cytotoxic; Transfection; Tumor Cells, Cultured; Vaccines, DNA

Exogenous, nonreplicating protein antigens (Ags) are usually taken up by antigen-presenting cells (APCs) via endocytosis or pinocytosis and enter the major histocompatibility complex (MHC) class II processing and presentation pathway. Although exogenous Ags are not processed and presented in the class I pathway by most cells, soluble proteins can enter the class I processing and presentation pathway if they are introduced directly into the cytoplasm of APCs. The purpose of these studies was to determine whether exogenous proteins could be processed and presented to T cells if they were delivered into cells by electroporation. The conditions for electroporation were optimized so that the viability of the electroporated cells was high, and the majority of electroporated cells had protein incorporated. Electroporated B cells not only presented exogenous ovalbumin to CD8+, class I MHC-restricted T cells but also stimulated CD4+, class II MHC-restricted T cells. Electroporated cells also primed Ag-specific cytotoxic T lymphocytes (CTLs) in vivo, stimulated CTL precursors in vitro, and served as target cells for lysis by Ag-specific CTLs, indistinguishable from transfected cells. Thus, electropermeabilized cells were structurally intact, and the introduced exogenous protein was processed and presented in association with both class I and class II MHC molecules. This approach is as efficient and reproducible as other techniques of delivering exogenous proteins into the intracellular processing pathways. These studies suggest that electroporation could be employed for the study of cell-mediated immunity to various exogenous proteins.

Topics: Animals; Antigen Presentation; Antigen-Presenting Cells; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Electroporation; Epitopes; Female; Histocompatibility Antigens Class I; Histocompatibility Antigens Class II; Hybridomas; Interleukin-2; Mast-Cell Sarcoma; Mice; Mice, Inbred C57BL; Ovalbumin; T-Lymphocytes, Cytotoxic; Thymoma; Thymus Neoplasms; Tumor Cells, Cultured

Antigens in extracellular fluids can be processed and presented with major histocompatibility complex (MHC) class I molecules by a subset of antigen presenting cells (APCs). Chicken egg ovalbumin (Ova) linked to beads was presented with MHC class I molecules by these cells up to 10(4)-fold more efficiently than soluble Ova. This enhanced presentation was observed with covalently or noncovalently linked Ova and with beads of different compositions. A key parameter in the activity of these conjugates was the size of the beads. The APC that is responsible for this form of presentation is a macrophage. These cells internalize the antigen constructs through phagocytosis, since cytochalasin B inhibited presentation. Processing of the antigen and association with MHC class I molecules appears to occur intracellularly as presentation was observed under conditions where there was no detectable release of peptides into the extracellular fluids. When injected in vivo in C57BL/6 mice, Ova-beads, but not soluble Ova, primed CD4- CD8+ cytotoxic T lymphocytes (CTLs). Similar results were obtained in BALB/c mice immunized with beta-galactosidase-beads. The implications of these findings for development of nonliving vaccines that stimulate CTL immunity are discussed.

Topics: Animals; Antigen-Presenting Cells; Azides; Cell Line; Cytochalasin B; Deoxyglucose; Female; Histocompatibility Antigens Class I; Kinetics; Lymphoma, T-Cell; Macrophages; Mast-Cell Sarcoma; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Phagocytosis; Spleen; T-Lymphocytes, Cytotoxic; Tumor Cells, Cultured

Topics: Animals; Avidin; Cell Line; Horseradish Peroxidase; Humans; Lymph Nodes; Mast Cells; Mast-Cell Sarcoma; Mice; Mice, Inbred DBA; Ovalbumin; Peroxidases; Rats; Skin; Staining and Labeling

Topics: Animals; Antigen-Antibody Complex; Cattle; Cell Membrane; Cell Transformation, Neoplastic; Hot Temperature; Immunoglobulin G; Mast-Cell Sarcoma; Mice; Mice, Inbred DBA; Ovalbumin; Receptors, Fc; Rosette Formation; Serum Albumin, Bovine; Solubility

Cultured murine mastocytoma (AB-CBF1-MCT-1) cells were stimulated to release endogenous or incorporated histamine or serotonin by an IgE-mediated mechanisms without loss of viability. Stimulation was achieved by incubation of the cells with rat IgE-anti-IgE, rat IgE-anti-light chain, fluoresceinated rat IgE-anti-fluorescein, IgE-enriched mouse anti-ovalbumin-ovalbumin, or covalently linked dimers of rat IgE, at doses similar to those optimal for normal peritoneal mast cells. Active cell metabolism and Ca++ were required to obtain release. Despite the latter, no dose of the calcium ionophore, A23187, could be found which caused release without concomitant cytotoxicity. Phosphatidylserine did not enhance release.

Topics: Animals; Antibodies; Antibodies, Anti-Idiotypic; Calcimycin; Calcium; Cells, Cultured; Cytotoxicity, Immunologic; Fluoresceins; Histamine Release; Immunoglobulin E; Immunoglobulin Light Chains; In Vitro Techniques; Mast-Cell Sarcoma; Mice; Ovalbumin; Rats; Sarcoma, Experimental; Serotonin

Topics: Animals; Carcinoma, Hepatocellular; Cytotoxicity Tests, Immunologic; Guinea Pigs; Immune Sera; Immunity, Cellular; Liver Neoplasms; Lymphocytes; Lymphotoxin-alpha; Mast-Cell Sarcoma; Mice; Ovalbumin; Rabbits; Spleen

Topics: Absorption; Animals; Antibody Specificity; Antigens; Antilymphocyte Serum; B-Lymphocytes; Cell Adhesion; Cell Separation; Cells, Cultured; Chickens; Complement System Proteins; Cytotoxicity Tests, Immunologic; Dinitrophenols; Erythrocytes; gamma-Globulins; Immunization; Lymph Nodes; Lymphocytes; Macrophages; Mast-Cell Sarcoma; Mice; Ovalbumin; Serum Albumin; Solubility; T-Lymphocytes

Topics: Amines; Aniline Compounds; Animals; Antibody Specificity; Antigens; Arsenicals; Benzenesulfonates; Cell Membrane; Chickens; Chromium Radioisotopes; Cytotoxicity Tests, Immunologic; Dinitrophenols; Erythrocytes; gamma-Globulins; Hemagglutination Tests; Hemocyanins; Immune Sera; Lymph Nodes; Lymphocytes; Mast-Cell Sarcoma; Mice; Ovalbumin; Serum Albumin; Serum Albumin, Bovine; Solubility; Spleen; Tyrosine

Topics: Animals; Antibodies; Antigen-Antibody Complex; Binding Sites, Antibody; Cattle; Dinitrophenols; Erythrocytes; gamma-Globulins; Guinea Pigs; Immune Adherence Reaction; Immune Sera; Immunoglobulin A; Immunoglobulin E; Immunoglobulin G; Immunoglobulin M; Immunoglobulins; Mast Cells; Mast-Cell Sarcoma; Mice; Myeloma Proteins; Ovalbumin; Rabbits; Serum Albumin, Bovine

Topics: Absorption; Animals; Binding Sites, Antibody; Immune Sera; Immunoglobulin E; Mast-Cell Sarcoma; Mice; Microscopy, Electron; Ovalbumin; Passive Cutaneous Anaphylaxis

Topics: Animals; Carbon Isotopes; Cold Temperature; Drug Stability; Mast-Cell Sarcoma; Metabolism; Mice; Muramidase; Ovalbumin; Proteins; Serine; Stimulation, Chemical; Time Factors; Xylose