Activated macrophage conditioned medium: identification of the soluble factors inducing cytotoxicity and the L-arginine dependent effector mechanism
| Autor: | Charles J. Parker, Read R. Taintor, Ina J. Amber, Zdenek Vavrin, Barbara B. Johnson, John B. Hibbs |
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| Rok vydání: | 1991 |
| Předmět: |
Lipopolysaccharides
medicine.medical_treatment Immunology Blotting Western Biology Adenocarcinoma Arginine Nitric oxide chemistry.chemical_compound Interferon-gamma Mice Interferon medicine Tumor Cells Cultured Immunology and Allergy Animals Interferon gamma Cytotoxicity Nitrites Aconitate Hydratase Immunity Cellular Mice Inbred BALB C Dose-Response Relationship Drug Tumor Necrosis Factor-alpha Macrophages Mammary Neoplasms Experimental Cell Biology DNA Macrophage Activation Molecular biology Cytostasis Culture Media Blot Cytokine Biochemistry chemistry Chromatography Gel Tumor necrosis factor alpha Female medicine.drug |
| Zdroj: | Journal of leukocyte biology. 49(6) |
| ISSN: | 0741-5400 |
| Popis: | Conditioned medium (CM) from cultures of cytotoxic activated macrophages causes inhibition of mitochondrial respiration, DNA synthesis, and aconitase activity in murine EMT-6 mammary adenocarcinoma cells by an l-arginine dependent effector mechanism. CM induces cytotoxicity and nitrite synthesis in EMT-6 cells in a dose dependent manner. We have identified the soluble factors in CM that induce cytotoxicity and synthesis of inorganic nitrogen oxides from l-arginine by EMT-6 cells. Using functional inhibition experiments, the activity of lipopolysaccharide (LPS), tumor necrosis factor alpha (TNFα), and interferon gamma (IFNγ) in CM was investigated. The LPS inhibitor polymyxin B and TNFα antibody produced a modest decrease in nitrite production, while IFNγ antibody markedly inhibited both nitrite production and cytostasis. Simultaneous treatment with polymyxin B, TNFα antibody, and IFNγ antibody reduced EMT-6 cell nitrite production by 81%, and cytostasis by 74%. By Western blot, IFNγ and TNFα were shown to be present in CM. When CM was subjected to hydrophobic interaction chromatography, a single peak of activity was eluted, and Western blot showed that the active fractions contained IFNγ. Furthermore, IFNγ antibody neutralized the activity in these chromatographic fractions. We conclude that induction of inorganic nitrogen oxide synthesis from l-arginine by the synergistic combination of IFNγ, TNFα, and LPS accounts for most of the biologic activity of CM, and that IFNγ is the major priming factor. |
| Databáze: | OpenAIRE |
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