The concept that macrophages can become foam cells as a result of a disturbed balance between the uptake of cholesterol from lipoproteins and cholesterol efflux is generally accepted. ABCA1 and ABCG1 are two cholesterol transporters that may act sequentially to remove cellular cholesterol, but currently their combined role in vivo is unknown. We report here that targeted disruption of both ABCA1 and ABCG1 in mice, despite severe plasma hypocholesterolemia, leads to massive lipid accumulation and foam cell formation of tissue macrophages. A complete ablation of cellular cholesterol efflux in vitro is observed, whereas in vivo macrophage-specific reverse cholesterol transport to the feces is markedly decreased. Despite the massive foam cell formation of tissue macrophages, no lipid accumulation was observed in the vascular wall, even in mice of 1 year old, indicating that the double knockout mice, possibly because of their hypocholesterolemia, lack the trigger to attract macrophages to the vessel wall. In conclusion, even under hypocholesterolemic conditions macrophages can be converted into foam cells, and ABCA1 and ABCG1 play an essential role in the prevention of foam cell formation.