Cytolysis of Actinomycin D-Treated Target Cells by Cell-Free Supernatants from Human Monocytes

The lytic mechanism of human peripheral blood monocytes was studied by using as targets actinomycin D-treated WEHI-164, an NK-insensitive murine fibrosarcoma cell lineMonocytes, but not lymphocytes, lysed WEHI-164 target cells pre-treated with actinomycin D within 6 h in ⁵¹ Cr-release assaysBecause cytolysis could not be inhibited competitively by unlabeled WEHI/D target cells, contact-independent mechanisms of cytolysis were investigatedCell-free supernatants collected from monocytes cultured for 4-6 h at 37°C lysed target cells as effectively as effector cell preparations of monocytesSupernatants from lymphocytes cultured in parallel were not cytolyticCytolytic activity was not detected in supernatants from preparations of monocytes that were held on iceHowever, monocytes produced cytolytic activity whether they were isolated by adherence or remained unseparated in suspensions of mononuclear cells. The cytolytic activity in cell-free supernatants (CFS) from monocytes was unaffected by incubation with protease inhibitorsCFS activity was destroyed by heatStorage of CFS at 37°, 22°, 4°, or -20°C for 24 h decreased cytolytic activity; however, loss of cytotoxicity was minimized by storage at 4'C. The cytolytic substance detected in 4-h CFS from monocytes appeared to be a protein(s) based on the sensitivity of the cytolytic activity to proteasesCytolytic activity of CFS eluted from Sephacryl 200 in a single peak with an apparent molecular weight between 25,000 and 45,000 daltons.