Previously, we reported that fluid-phase endocytosis of native LDL by 
PMA-activated human monocytederived macrophages converted these macrophages into 
cholesterol-enriched foam cells (Kruth, H. S., Huang, W., Ishii, I., and Zhang, 
W. Y. (2002) J. Biol. Chem. 277, 34573-34580). Uptake of fluid by cells can 
occur either by micropinocytosis within vesicles (<0.1 microm diameter) or by 
macropinocytosis within vacuoles ( approximately 0.5-5.0 microm) named 
macropinosomes. The current investigation has identified macropinocytosis as the 
pathway for fluid-phase LDL endocytosis and determined signaling and 
cytoskeletal components involved in this LDL endocytosis. The 
phosphatidylinositol 3-kinase inhibitor, LY294002, which inhibits 
macropinocytosis but does not inhibit micropinocytosis, completely blocked 
PMA-activated macrophage uptake of fluid and LDL. Also, nystatin and filipin, 
inhibitors of micropinocytosis from lipid-raft plasma membrane domains, both 
failed to inhibit PMA-stimulated macrophage cholesterol accumulation. Time-lapse 
video phase-contrast microscopy and time-lapse digital confocal-fluorescence 
microscopy with fluorescent DiI-LDL showed that PMA-activated macrophages took 
up LDL in the fluid phase by macropinocytosis. Macropinocytosis of LDL depended 
on Rho GTPase signaling, actin, and microtubules. Bafilomycin A1, the vacuolar 
H+-ATPase inhibitor, inhibited degradation of LDL and caused accumulation of 
undegraded LDL within macropinosomes and multivesicular body endosomes. LDL in 
multivesicular body endosomes was concentrated >40-fold over its concentration 
in the culture medium consistent with macropinosome shrinkage by maturation into 
multivesicular body endosomes. Macropinocytosis of LDL taken up in the fluid 
phase without receptor-mediated binding of LDL is a novel endocytic pathway that 
generates macrophage foam cells. Macropinocytosis in macrophages and possibly 
other vascular cells is a new pathway to target for modulating foam cell 
formation in atherosclerosis.