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Initial characterization of a lymphokine pathway for the immunologic induction of tumor necrosis factor-alpha release from human peripheral blood mononuclear cells.

R S Kornbluth, S A Gregory and T S Edgington
J Immunol September 15, 1988, 141 (6) 2006-2015;
R S Kornbluth
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S A Gregory
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T S Edgington
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Abstract

Under endotoxin-free conditions, unstimulated human PBMC do not release TNF-alpha, as measured in a sensitive assay with 51Cr release in 6 h from actinomycin D-treated WEHI 164 cells. IFN-gamma alone at less than or equal to 10,000 U/ml is insufficient to elicit TNF-alpha release. Similarly, the lymphokine-rich supernatant of PBMC stimulated by allogeneic cells is also insufficient to induce TNF-alpha release in a short term assay. However, when PBMC are first primed with IFN-gamma for 48 h and then exposed to lymphokine supernatant for 6 h, effector cells within the PBMC population are triggered to express TNF-alpha-mediated cytotoxicity. All of the measured cytotoxicity is attributable to TNF-alpha because it could be abolished by a specific anti-TNF-alpha neutralizing mAb. Although IFN-gamma serves to prime PBMC in this assay system, it fails to trigger the release of TNF-alpha. Instead, a second lymphokine (provisionally termed "cytotoxicity triggering factor" (CTF) is required to induce TNF-alpha release from IFN-gamma-primed human PBMC. In kinetic studies, IFN-gamma priming was optimal when PBMC were exposed to IFN-gamma (150 U/ml) for 48 h. In contrast to the prolonged interval for priming, CTF need be present for 6 h or less for maximal induction of TNF-alpha-mediated cytotoxicity. In dose-response studies, IFN-gamma priming (48 h) required at least 4 U/ml and was complete with 20 to 100 U/ml. By using fully primed PBMC, the response to CTF followed a sigmoidal dose-response curve, which allowed the quantitation of CTF in half-maximal units. Activated Th lymphocytes constitute one cellular source for CTF. CTF is produced by cloned allorective T3+T4+T8-M1- Th cells after alloantigen stimulation, and also by nylon wool-purified T cells after stimulation with PMA and A23187 calcium ionophore. Unstimulated T cells do not release CTF. In physicochemical studies, CTF activity elutes from Sephadex G-100 as a major discrete peak of Mr 55 kDa and minor peaks of 14 kDa and greater than 150 kDa. On the basis of multiple criteria, CTF is distinguishable from several other cytokines: IFN-gamma, IL-1, IL-2, GM-CSF, MIF, CSF-1, TNF-alpha, and lymphotoxin (TNF-beta). We conclude that, by acting together, IFN-gamma and CTF provide a lymphokine pathway whereby Ag-responsive human Th cells induce the immunologic release of TNF-alpha from effector cells present in PBMC.

  • Copyright © 1988 by American Association of Immunologists

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The Journal of Immunology
Vol. 141, Issue 6
15 Sep 1988
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Initial characterization of a lymphokine pathway for the immunologic induction of tumor necrosis factor-alpha release from human peripheral blood mononuclear cells.
R S Kornbluth, S A Gregory, T S Edgington
The Journal of Immunology September 15, 1988, 141 (6) 2006-2015;

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Initial characterization of a lymphokine pathway for the immunologic induction of tumor necrosis factor-alpha release from human peripheral blood mononuclear cells.
R S Kornbluth, S A Gregory, T S Edgington
The Journal of Immunology September 15, 1988, 141 (6) 2006-2015;
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Print ISSN 0022-1767        Online ISSN 1550-6606