A rapid assay for cytotoxicity of unstimulated human monocytes

Abstract

Human peripheral blood mononuclear cells were used as effectors against the Wehi 164 mouse fibrosarcoma cell line grown in suspension culture. In a standard 7-hour 51Cr release assay, specific release usually was below 10%. In contrast, pretreatment of Wehi 164 for 3 hours with dactinomycin (Act D), while leaving the tumor cells intact and viable, resulted in a drastic increase in its susceptibility to lysis, which reached 60% specific release. In terms of lytic units, this reflects up to a fiftyfold enhancement. Cell-separation experiments revealed that the effector cells were plastic-adherent and iron-phagocytic. Adherent cell fractions with 85-98% naphthol AS acetate-esterase (NAS)-positive cells were enriched in cytotoxicity, while nonadherent cells with less than 4% NAS-positive cells were almost devoid of activity. Depletion of phagocytic cells with iron and magnet resulted in a strong reduction of cytotoxicity by 82-95% compared to the cytotoxicity seen in control treated effector cells. In both kinds of analyses, natural killer cell activity showed a reciprocal behavior. The evidence indicates that the cytotoxic effector cells directed against Act D-treated Wehi 164 cells belong to the monocyte lineage. The system described should be useful in analyzing the cytotoxic function of unstimulated monocytes in a short-term assay without prior purification of the effector cells.