Cholesterol in atherosclerosis. The images show proximal aorta atherosclerotic lesions from the atherosclerosis-susceptible apoE knockout mice fed a cholesterol- and saturated fat–enriched diet for 25 weeks. (a and b) Adjacent sections from the same lesion stained with hematoxylin (a) or oil red O (b). The asterisk in a marks an area filled with cholesterol crystals. The bright red-orange staining in b shows areas rich in neutral lipids, most likely cholesteryl ester inclusions in macrophage foam cells and cholesteryl ester-rich emulsions in extracellular regions of the lesion. Areas of the lesion that stain poorly with oil red O are often rich in free (i.e., unesterified) cholesterol. (c) Fluorescence microscopy image of another atherosclerotic lesion stained with the fluorescent dye filipin (shown as blue in this image), which binds specifically to areas enriched in free cholesterol. The bright filipin staining in almost the entire diseased intima indicates that advanced atherosclerotic lesions in this model are very rich in free cholesterol.