Previous investigations have shown that macrophages play a pivotal role in the induction of angiogenesis in both physiological and pathological settings. This investigation examines the relative production of the angiogenic modulator thrombospondin-1 (TSP1) by activated and nonactivated monocytes and macrophages. TSP1, a multifunctional extracellular matrix molecule, has been reported recently to inhibit angiogenesis both in vitro and in vivo. To examine the relationship between the level of TSP1 production by macrophages and expression of the angiogenic phenotype, murine monocytelike cells (WEHI-3) and human peripheral blood monocytes were each activated in vitro and examined for TSP1 production and angiogenic activity in rat corneal bioassay. Nonangiogenic monocytes produced low levels of TSP1 messenger RNA. Surprisingly, activated, potently angiogenic monocytes and macrophages exhibited as much as a sixfold increase in steady state TSP1 messenger RNA over unstimulated levels. Biosynthetic labeling studies demonstrated that TSP1 protein secretion increased in conjunction with increased TSP1 messenger RNA levels in angiogenic macrophages. The results demonstrate that activated monocytes and macrophages actively produce the angiogenic modulator TSP1 and suggest that TSP1 production may be a component of the angiogenic phenotype. In addition, the data suggest that the ability of macrophages to mediate angiogenesis results from a complex interplay of positive and negative regulators.