Activation of antitumor cytotoxicity of human blood mononuclear cells by a basic factor from dialysable human-leukocyte extract.

Dialysable human leukocyte extract (10 kDa molecular weight cutoff) contained a basic factor stimulating natural killer (NK) cytotoxicity of human peripheral blood mononuclear cells (PBMC) against human K562 tumor cells when PBMC were pre-incubated with the factor for 72h prior to cytotoxicity assays. This cytotoxicity-stimulating factor (CySF-L2) could be enriched by adsorption to ion exchange resin Dowex 50WX2 (H-form) eluting at pH 9.0-9.6 when a pH gradient between pH 3 and pH 10 was used. Further purification was achieved by chromatography on DEAE-Sepharose. The factor has a molecular weight of approximately 1000 Da and is insensitive to protease and exopeptidase treatment but sensitive against treatment with endoglycosidase F. Using immunomagnetic cell sorting, complement-mediated cell depletion and depletion by planning, the cytotoxic effector cells activated during pre-incubation of PBMC with CySF-L2 could be identified as CD16+ CD14+ monocytes/macrophages and as Leu7+ Leu19+ CD16+ CD3- CD8- NK cells.

The CySF-L2 factor from dialysable human leucocyte extract activates natural killer cytotoxicity by induction of interferon gamma.

The mechanism of natural killer (NK) cytotoxicity activation of human peripheral blood mononuclear cells (PBMC) by CySF-L2 was elucidated. CySF-L2 is a cytotoxicity-stimulating factor isolated from dialysable human leucocyte extract, which activates NK cytotoxicity against NK-sensitive and insensitive tumour cells (K562; Daudi; Raji; MOLT4) when preincubated with effector cells for 72 h. CySF-L2-mediated activation was synergistic to interleukin-2(IL-2)-mediated activation of NK cytotoxicity. Induction of interferon gamma (IFN gamma) release was the crucial step during CySF-L2-mediated NK cytotoxicity activation since enhancement of NK activity was completely blocked when anti-IFN gamma antibodies were present during treatment of PBMC. Anti-IFN alpha, anti-TNF alpha (tumour necrosis factor alpha) anti-IL-1 and anti-IL-2 antibodies showed no blocking effect. Analysis of the supernatant culture medium after 72 h incubation of PBMC and their highly purified subpopulations demonstrated that CySF-L2 induced release of IFN gamma from CD3+T cells and CD56+CD3- NK cells and of TNF alpha and prostaglandin E2 from monocytes. CySF-L2 was also capable of activating NK cytotoxicity of highly purified (98%) CD56+CD3- NK cells as well as of monocytes (94% pure). Cell cooperation studies connected with analysis of cytokine release and enhancement of NK cytotoxicity indicated that CySF-L2 might play an essential role in the up and down regulation of NK cytotoxicity by the cytokine network.