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We studied the capacity of cultured mouse peritoneal macrophages to generate superoxide anion (O(2-)), the initial product of conversion of oxygen to microbicidal species, during phagocytosis of opsonized zymosan or upon contact with the membrane-active agent phorbel myristate acetate (PMA). Macrophages from mice infected with Bacille Calmette-Guerin (BCG) or injected intraperitoneally with thioglycollate broth or endotoxin, released up to 12 times more O(2-) than did resident peritoneal macrophages, depending upon the cell type and whether the stimulus was zymosan or PMA. There was little if any O(2-) release from resting (unstimulated) macrophages. The density of cells on culture dishes was an important variable since crowding of the dish markedly reduced the efficiency of O(2-) production. The enhanced O(2-) release of chemically elicited and infection-activated macrophages was noted after stimulation with a wide range of concentrations of PMA and zymosan, at all time points studied (up to 120 min), and with cells maintained for 140 rain to 16 days in culture. The O(2-) response of resident cells improved twofold to zymosan and ninefold to PMA during the first 3 days in culture. The capacity to release O~ appears to be limited to actively phagocytic cell types: murine macrophage-like tumor lines and cultured human monocytes released O(2-) when stimulated by PMA or zymosan, fibroblast and endothelial lines and embryo-derived cells did not. Activity of superoxide dismutase, which removes O(2-), was not detectable in culture supernates of any cell type, and thus, differences in detectable O(2-) could not be attributed to variations in the release of this enzyme. We conclude that the phagocytosis- associated respiratory burst is significantly enhanced in mononuclear phagocytes obtained ai~r chemical inflammation or BCG infection. Increased capacity to generate O(2-) and other oxygen radicals during phagocytosis could contribute to the improved microbicidal and tumoricidal activity of activated macrophages.
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