Macrophage-conditioned medium containing very-low-density lipoproteins (VLDL) and its effects on smooth muscle cell triacylglycerol metabolism was investigated. Macrophages exposed to VLDL from normolipemic rats accumulated high levels of intracellular triacylglycerol, while similarly treated smooth muscle cells showed only slight changes. Medium, initially composed of VLDL and albumin, contained substantial levels of free fatty acids after exposure to macrophages. In the presence of albumin, the loss of VLDL triacylglycerol from the medium and the appearance of medium free fatty acids was consistent with a precursor-product relationship. The extent of medium fatty acid accumulation was dependent on the length of time of incubation with macrophages and was proportional to the concentration of VLDL and albumin added to the culture dish. This macrophage-conditioned medium, when given to smooth muscle cells, promoted a 6-12-fold increase in smooth muscle cell triacylglycerol levels over that produced by fresh VLDL and albumin. Similar increases in cell triacylglycerol levels could be produced using fresh medium approximating the oleate concentration and the fatty acid to albumin molar ratios found in macrophage-conditioned medium. In macrophage-conditioned medium with VLDL but without albumin, little free fatty acid was found. Other factors produced by macrophages did not seem to affect the metabolism of VLDL by smooth muscle cells since, in the absence of albumin, media with VLDL caused comparable responses in smooth muscle cell triacylglycerol accumulation whether or not the medium was previously exposed to macrophages. Thus, the minor changes in triacylglycerol content in smooth muscle cells promoted by medium containing VLDL and albumin were substantially enhanced by a prior exposure of the medium to macrophages, primarily due to the free fatty acids present in the macrophage-conditioned medium.