Human Atherosclerotic Plaque Contains Both Oxidized Lipids and Relatively Large Amounts of α-Tocopherol and Ascorbate

Abstract

Abstract We assessed the antioxidant status and contents of unoxidized and oxidized lipids in freshly obtained, homogenized samples of both normal human iliac arteries and carotid and femoral atherosclerotic plaque. Optimal sample preparation involved homogenization of human atherosclerotic plaque for 5 minutes, which resulted in recovery of most of the unoxidized and oxidized lipids without substantial destruction of endogenous vitamins C and E and 87% and 43% recoveries of added standards of α-tocotrienol and isoascorbate, respectively. The total protein, lipid, and antioxidant levels obtained from human plaque varied among donors, although the reproducibility of replicates from a single sample was within 3%, except for ubiquinone-10 and ascorbate, which varied by 20% and 25%, respectively. Plaque samples contained significantly more ascorbate and urate than control arteries, with no discernible difference in the vitamin C redox status between plaque and control materials. The concentrations of α-tocopherol and ubiquinone-10 were comparable in plaque samples and control arteries. However, approximately 9 mol percent of plaque α-tocopherol was present as α-tocopherylquinone, whereas this oxidation product of vitamin E was not detectable in control arteries. Coenzyme Q ₁₀ in plaque and control arteries was only detected in the oxidized form ubiquinone-10, although coenzyme Q ₁₀ oxidation may have occurred during processing. The most abundant of all studied lipids in plaque samples was free cholesterol, followed by cholesteryl oleate and cholesteryl linoleate (Ch18:2). Approximately 30% of plaque Ch18:2 was oxidized, with 17%, 12%, and 1% present as fatty acyl hydroxides, ketones, and hydroperoxides, respectively. In comparison, 7-ketocholesterol was detected at an ≈75-fold lower concentration. Normal arteries contained similar levels of protein as atherosclerotic arteries, much less free cholesterol, and no detectable amounts of unoxidized or oxidized cholesteryl esters. Together, these results demonstrate the coexistence in human plaque of large amounts of oxidized cholesteryl esters with significant concentrations of ascorbate and vitamin E in their reduced, antioxidant-active form. We conclude that compared with healthy human arteries, advanced atherosclerotic plaques are not deficient in the antioxidant vitamins C and E, despite the occurrence of massive lipid oxidation.