The role of vitamin E on the anti-atherosclerotic effect of fish oil in diet-induced hypercholesterolemic rabbits was studied in male New Zealand white rabbits. The animals were randomly divided into six groups of 14 each according to the chow given them. Group I, control, rabbits were fed regular laboratory rabbit chow. In addition to the regular chow, Group II rabbits were fed a high (1%) cholesterol-enriched diet. Group III had the same diet as Group II plus 450 mg vitamin E/1000 g chow. Group IV also had the same diet as Group II plus 10% fish oil, but without vitamin E. Group V's diet contained 1% cholesterol, 10% fish oil, and 450 mg vitamin E/1000 g chow. Group VI had the same diet as Group V, but with 150 mg vitamin E/1000 g chow. At the end of a 6-week feeding, the ascending aorta from seven rabbits from each group was harvested for the analysis of prostanoid production, thiobarbituric acid reactive substances (TBARS), superoxide dismutase activity, and cholesterol and vitamin E levels; the same tissue from the other seven rabbits from each group was obtained for the delineation of atherosclerotic lesions by planimetry after the Sudan IV stain. The high cholesterol diet-treated rabbits had worse prostanoid synthesis and higher TBARS levels, which paralleled the severity of the atherosclerosis. Vitamin E or fish oil supplementation in a high cholesterol diet had beneficial prostanoid production, reduced aortic TBARS levels, and attenuated atherosclerotic lesions; these effects were potentiated when vitamin E and fish oil were combined together. The atherosclerotic lesion reduction in rabbits treated with high cholesterol plus fish oil and 450 mg vitamin E/1000 g chow reached statistical significance (P < 0.05) compared to high cholesterol and the same dose of vitamin E-treated rabbits. The rabbits treated with high cholesterol plus fish oil, but without vitamin E or with 150 mg vitamin E/1000 g chow showed an increased plasma creatine kinase activity at 4 (P < 0.01 and 0.05, respectively) and 6 (P < 0.001 and 0.01, respectively) week of feeding. These results suggest that vitamin E and/or fish oil attenuate atherosclerosis in high cholesterol-fed rabbits; vitamin E and fish oil potentiated the effect of each other. Furthermore, without sufficient vitamin E supplementation, rabbits treated with high cholesterol plus fish oil will show an elevation of plasma creatine kinase activity.