Effector mechanisms of cytolytically activated macrophages. II. Secretion of a cytolytic factor by activated macrophages and its relationship to secreted neutral proteases.

Abstract

Medium without serum conditioned by culture for 16 hr over macrophages, which had been cytolytically activated by bacillus Calmette-Guerin in vivo, effectively lysed murine MCA-I sarcoma targets but not murine embryo fibroblasts at dilutions of up to 1/60. Supernatants of inflammatory macrophages elicited by thiogy-collate broth were not lytic but became so when the macrophages were activated in vitro for direct cytolysis by endotoxin. Six other murine neoplastic cell lines, both adherent and nonadherent, were lysed by the supernatants of activated macrophages; two other types of non-neoplastic cells were not lysed by such supernatants. Lytic activity of the supernatants, which was not reduced by dialysis, was inhibited by both heat (56°C for 60 min) and by fetal calf serum (10% v/v). MCA-1 targets cultured in balanced salt solution were lysed within several hours (3 to 5) after exposure to lytic supernatant. Generation of lytic activity by the macrophages was inhibited by cycloheximide or by heating the macrophages to 56°C for 60 min. Lytic activity of the supernatants was inhibited by bovine pancreatic trypsin inhibitor (750 KIU/ml) and by diisopropylfluorophosphate (2 x 10-3M.) Supernatants of both types of activated macrophages, when partitioned by gel filtration, contained a major peak of cytolytic activity. This peak cochromatographed with a distinct peak of proteolytic activity not present in supernatants of the TG inflammatory macrophages. The cytolytic and proteolytic activities of this peak were inhibited by bovine pancreatic trypsin inhibitor. It is suggested that the lytic activity in supernatants of activated macrophages represents a potential mechanism by which activated macrophages can lyse neoplastic cells and that this lytic activity is related to a specific neutral protease secreted by activated macrophages.