Distinction of partially purified human natural killer cytotoxic factor from recombinant human tumor necrosis factor and recombinant human lymphotoxin

Natural killer cell cytotoxic factor (NKCF), a cytotoxic factor contributing to human natural killer cell-mediated cytotoxicity, was generated from lymphocyte-conditioned medium using various stimuli. Crude NKCF activity was concentrated, and partially purified by ammonium sulfate precipitation and gel filtration. NKCF activities eluted as two molecular weight peaks, corresponding to Mr 33,000-43,000 (pool I) and approximately Mr 5,000 (pool II). The cytotoxic activity and target specificity of the partially purified NKCFs were found to be different from both recombinant human TNF and recombinant human lymphotoxin. In the NKCF assay, up to 10(6) units/ml of TNF and lymphotoxin had virtually no effect, whereas both NKCFs lysed 22% (range 17-33%) of the NK-sensitive target K562. In contrast, TNF and lymphotoxin were active in a standard assay against the sensitive murine L929 fibroblast cell line in all concentrations tested (10(-1)-10(6) units/ml). In addition, the effect of these cytotoxic factors in a short-term (4-h) chromium-release assay using peripheral blood mononuclear cells as effector cells was tested: only NKCF (pool I), but not TNF, lymphotoxin, or low molecular weight NKCF (pool II), enhanced NK and lymphokine-activated killer cell cytolysis, both against the NK-sensitive target K562 and the NK-resistant melanoma cell line SK-MEL 30. Results were not affected in the presence of neutralizing antibodies against TNF. NKCF could, therefore, be distinguished from TNF and lymphotoxin with respect to their biological activities.