It has been suggested that oxidation of low-density lipoprotein (LDL) is a critical initiating step in human atherosclerosis, and that antioxidants can act as antiatherogens. Vitamin C (ascorbic acid, AA) is an antioxidant nutrient known to protect LDL against atherogenic modification in vitro more effectively than any other known natural or synthetic antioxidant. However, the mechanism of vitamin C's protective effect is unexplained, and it is not known whether vitamin C can act as antioxidant and antiatherogen in vivo. This proposal seeks to answer these questions. In the first part of the proposal, in vitro experiments will be conducted to identify the molecular mechanism of LDL protection by vitamin C. We have already shown that this mechanism is not explained by scavenging of free radicals by AA, but involves stable modification of LDL with decomposition product(s) of dehydroascorbic acid (DHA), the oxidation product of AA. Thus, the effects of AA, DHA, and selected DHA decomposition products on atherogenic modification of LDL will be investigated. LDL will be exposed to metal-ion-dependent (cupric ions or monocyte-macrophages) or metal-ion-independent (peroxyl radicals) oxidizing conditions. Progression of atherogenic modification of LDL will be assessed by measuring lipid peroxidation, electrophoretic mobility, and uptake of LDL by monocyte-macrophages. Possible mechanisms of action of vitamin C will be explored by investigating covalent modification of apolipoprotein B, chelation of metal ions, and peroxidase activity. We will also examine whether the reactions and mechanisms identified in vitro are operative in LDL in its natural plasma environment. Plasma will be incubated under oxidative stress to oxidize AA to DHA, or with DHA or its decomposition products. LDL will be isolated and examined for increased resistance to atherogenic modification. In the second part of the proposal, vitamin C's in vivo antioxidant and antiatherogenic effects will be investigated. Vitamin C-adequate and -marginally deficient guinea pigs on an atherogenic cholesterol diet will be exposed to hyperoxia, and compared to control animals on the same diets but without exposure to oxidative stress. In vivo lipid peroxidative damage will be assessed by measuring prostaglandin-like compounds formed by a free radical-catalyzed mechanism. LDL will be isolated from the animals and examined for antioxidant content and redox status as well as for susceptibility to atherogenic modification in vitro. Extent of atherosclerotic lesion development and tissue oxidative damage will be assessed at the conclusion of the study. In summary, in this proposal we will carry out a detailed analysis of vitamin C's antioxidant and antiatherogenic activities by studying its effects on three different levels, in vitro, in its natural plasma environment, and in vivo. The studies will elucidate fundamental mechanisms of antiatherogenesis and determine the importance of vitamin C as a dietary antioxidant and antiatherogen.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
5R29HL049954-03
Application #
2226021
Study Section
Nutrition Study Section (NTN)
Project Start
1993-06-01
Project End
1998-05-30
Budget Start
1995-06-01
Budget End
1996-05-31
Support Year
3
Fiscal Year
1995
Total Cost
Indirect Cost
Name
Boston University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
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Chen, K; Suh, J; Carr, A C et al. (2000) Vitamin C suppresses oxidative lipid damage in vivo, even in the presence of iron overload. Am J Physiol Endocrinol Metab 279:E1406-12
Carr, A; Frei, B (1999) Does vitamin C act as a pro-oxidant under physiological conditions? FASEB J 13:1007-24
Gokce, N; Keaney Jr, J F; Frei, B et al. (1999) Long-term ascorbic acid administration reverses endothelial vasomotor dysfunction in patients with coronary artery disease. Circulation 99:3234-40
Retsky, K L; Chen, K; Zeind, J et al. (1999) Inhibition of copper-induced LDL oxidation by vitamin C is associated with decreased copper-binding to LDL and 2-oxo-histidine formation. Free Radic Biol Med 26:90-8
Shwaery, G T; Samii, J M; Frei, B et al. (1999) Determination of phospholipid oxidation in cultured cells. Methods Enzymol 300:51-7
Carr, A C; Frei, B (1999) Toward a new recommended dietary allowance for vitamin C based on antioxidant and health effects in humans. Am J Clin Nutr 69:1086-107
McCall, M R; Frei, B (1999) Can antioxidant vitamins materially reduce oxidative damage in humans? Free Radic Biol Med 26:1034-53
Vita, J A; Keaney Jr, J F; Raby, K E et al. (1998) Low plasma ascorbic acid independently predicts the presence of an unstable coronary syndrome. J Am Coll Cardiol 31:980-6

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