We have previously demonstrated that rat resting macrophages have cytotoxicity against tumor cells. In the current study we have performed an in vitro experiment to explore the mechanism of Kupffer cells-mediated cytotoxicity against hepatomas. The coculture of tumor cells with Kupffer cells (derived from rat livers) stimulated syngeneic and allogeneic Kupffer cells to produce nitric oxide. Kupffer cell-mediated cytotoxicity was paralleled to the amount of nitric oxide and was abolished by the addition of N-G-monomethyl-L-arginine. The ability to stimulate Kupffer cells to produce nitric oxide resided on the membranous fragment of tumor cells. (C) 1994 Academic Press, Inc.