The purpose of the experiments reported here was to determine chemically the character and quantity of lipid in re-endothelialized and de-endothelialized areas of rabbit aortas. The aortas of 22 rabbits, Groups I and II, were de-endothelialized with a balloon catheter, and the rabbits were maintained on a lipid-poor diet for 4 weeks. Thirteen rabbits, Group II, were then fed an egg-supplemented diet for 6 weeks. Nine rabbits, Group I, were continued on the lipid-poor diet for an additional 6 weeks. Control rabbits with uninjured aortas were fed a lipid-poor diet for 10 weeks (Group III) or an egg-supplemented diet for 6 weeks (Group IV). Nonesterified cholesterol and fatty acids, cholesteryl esters, triacylglycerols, and squalene were quantitated in re-endothelialized and de-endothelialized aorta by thin-layer chromatography and fluorometric analysis. The results indicate 1) that there was approximately three times as much nonesterified cholesterol and cholesteryl ester in re-endothelialized aorta of Groups I and II as compared with adjacent de-endothelialized aorta and 2) that in re-endothelialized aorta of Group II the amount of total cholesterol correlated with serum cholesterol concentration in contrast to adjacent de-endothelialized aorta, with no correlation over a range of nearly 900 mg/100 ml. These studies indicate that the presence of endothelium favors accumulation of aortic cholesteryl esters. The results suggest that arterial lipid accumulation is not simply a result of passive filtration but may result from metabolic differences in the re-endothelialized neointima.