Author + information
- Received December 1, 1996
- Revision received April 14, 1997
- Accepted April 24, 1997
- Published online August 1, 1997.
- Lori Mosca, MD, MPH, PhDA,*,
- Melvyn Rubenfire, MD, FACCA,
- Caroline Mandel, MS, RDA,
- Cheryl Rock, PhD, RDB,
- Tom Tarshis, MPHA,
- Alan Tsai, PhDC and
- Thomas Pearson, MD, MPH, PhD, FACCD
- ↵*Dr. Lori Mosca, Preventive Cardiology Program, The University of Michigan, 24 Frank Lloyd Wright Drive, P.O. Box 363, Ann Arbor, Michigan 48106-0363.
Objectives. This study sought to determine the effect of antioxidant supplementation on the susceptibility of low density lipoprotein (LDL) to oxidation in patients with established cardiovascular disease (CVD).
Background. Data are inconsistent regarding the role of antioxidant nutrients in the prevention of CVD.
Methods. The study design was a 12-week, double-blind, placebo-controlled clinical trial. Patients with CVD (n = 45) were randomized to 1) placebo control; 2) 400 IU of vitamin E, 500 mg of vitamin C, 12 mg of beta-carotene (mid-dose); or 3) 800 IU of vitamin E, 1,000 mg of vitamin C, 24 mg of beta-carotene (high dose) daily. Reduced susceptibility of LDL to oxidation was estimated by an increase in lag phase (minutes). Baseline and 6- and 12-week measurements of lipoproteins and lag phase were obtained. Plasma levels of antioxidants were measured at baseline and 12 weeks.
Results. Concentrations of alpha-tocopherol, vitamin C and beta-carotene significantly increased in the mid- and high dose groups during the trial. Lag phase significantly increased from baseline (190.1 ± 63.8 min [mean ± SD]) to 12 weeks (391.1 ± 153.0 min) in the high dose group (p < 0.01). A nonsignificant increase in lag phase in the mid-dose group was observed during the same time interval. A dose response was found for mean percent change from baseline to 12 weeks for lag phase for the placebo, mid- and high dose groups (p = 0.004 for trend).
Conclusions. A high dose combination of antioxidant nutrients reduces the susceptibility of LDL to oxidation in patients with CVD and may be useful in secondary prevention.
Despite significant reductions in risk factor levels and advances in medical and surgical intervention, cardiovascular disease (CVD) remains the leading cause of death in the United States . The risk of future cardiovascular events is substantially increased in subjects with established CVD compared with those without previous manifestations of clinical disease . Therefore, it is important to identify effective strategies for the secondary prevention of coronary artery disease (CAD).
Recent data suggest that antioxidant nutrients may play a role in the prevention of CAD. The protective effect of antioxidant nutrients may be mediated through inhibition of the oxidative modification of low density lipoprotein cholesterol (LDL), which is believed to be a key step in the pathogenesis of atherosclerosis [3, 4]. In addition, antioxidants may reduce platelet adhesiveness , preserve endothelial function and stabilize plaque . These mechanisms lend support for a role of antioxidants in the prevention of atherosclerosis and thrombosis and in the regulation of vasomotor tone.
Epidemiologic data have shown associations between antioxidant intake and reduced CVD in several prospective studies [8–15]. Data from randomized clinical trials have not supported the findings from observational studies. A Finnish trial , designed to test the effect of low dose alpha-tocopherol (50 mg) or beta-carotene (20 mg/day), or both, on the incidence of lung cancer in 29,133 male smokers, failed to show a protective effect of antioxidants on death from CVD. Recent results from large, randomized clinical trials showed no effect of beta-carotene alone (Physician’s Health Study , Skin Cancer Prevention Study ) or in combination with retinol (Carotene and Retinol Efficacy Trial ) in the primary prevention of CVD.
Few data are available to evaluate the role of antioxidant nutrients in secondary prevention. A subgroup analysis of the Cholesterol Lowering Atherosclerosis Study (CLAS) study demonstrated an association between supplemental vitamin E intake and a reduction in coronary lesion progression by angiography. The Cambridge Heart Antioxidant Study (CHAOS) , a secondary prevention trial of 2,002 patients with CAD followed up for a median of 510 days demonstrated a 77% reduction in nonfatal myocardial infarction in patients randomized to receive either 400 or 800 IU of vitamin E versus those receiving placebo.
Experimental evidence [21–30]demonstrating favorable alterations in indexes of oxidized LDL in subjects treated with antioxidant supplements provides biologic plausibility for a role of dietary micronutrients in CVD prevention. These data are limited to patients free of disease, and the generalizability of results to patients with CVD remains unclear. Patients with CVD have unfavorable indexes of oxidized LDL compared with healthy control subjects, and it may not be appropriate to extrapolate doses of antioxidant vitamins to be used in secondary prevention trials on the basis of data obtained from subjects without CVD [31–33].
This purpose of the present study was to test the effect of two doses of a combination of vitamin E, vitamin C and beta-carotene on lag phase, a surrogate measure of the susceptibility of LDL to oxidation, compared with placebo in a randomized clinical trial in subjects with established CVD.
Forty-five nonsmoking, nondiabetic subjects (39 men, 6 women; 39 to 80 years old) with established CAD were recruited from the cardiac catheterization laboratory and the preventive cardiology program at the University of Michigan Medical Center between January 1994 and May 1995. CAD was determined by angiography (at least one lesion >70% lumen stenosis), previous angioplasty, coronary artery bypass graft surgery, myocardial infarction by history validated by electrocardiographic changes or classic angina with an ischemic response on a treadmill test. Subjects had to have been on a stable diet and exercise program for 6 months and have normal hepatic and renal function, no evidence of malabsorption, pancreatic or biliary disease and no acute medical condition for at least 3 months before study entry. Patients currently receiving resin therapy or using vitamin supplements above the recommended dietary allowance (RDA) levels within the previous 6 months were excluded from the study. There were no exclusion criteria based on serum lipid status. All subjects gave informed consent. The study was approved by the Institutional Review Board of The University of Michigan Medical Center.
1.2 Experimental protocol.
After baseline measurements, subjects were entered into a 12-week double-blind, placebo-controlled clinical trial. Each subject was randomly assigned to receive 1) placebo (control group, n = 15); 2) 400 IU of vitamin E, 500 mg of vitamin C, and 12 mg of beta-carotene daily in two divided doses (mid-dose group, n = 15); or 3) 800 IU of vitamin E, 1,000 mg of vitamin C and 24 mg of beta-carotene daily in two divided doses (high dose group, n = 15). Supplements were given in the form of a capsule (vitamin E [200 IU] as dl-alpha-tocopheryl acetate, vitamin C [250 mg] and beta-carotene [6 mg]) obtained as a gift from Hoffmann-La Roche Inc. Each subject took two capsules (supplement or placebo, or both) twice a day to maintain blinding. Pill counts were done at 6 and 12 weeks to assess adherence to study protocol. Four subjects did not complete the trial (one assigned to the placebo group who had an adverse effect of headache within 2 weeks of the trial; one assigned to the high dose group who had an adverse effect of stomach discomfort after 1 week of therapy; two who did not return after the baseline visits because of scheduling difficulties). All 41 subjects who completed the trial were 100% compliant with the study medication.
1.3 Laboratory protocol.
Two lipid, lipoprotein and lag phase measurements were obtained 1 week apart immediately before study entry and were averaged to obtain baseline values. One laboratory measurement was done at study midpoint (6 weeks). Two ending measurements were done at 11 and 12 weeks and were averaged to obtain the 3-month follow-up laboratory values. Plasma tocopherol, carotenoid and vitamin C levels were measured once at baseline and again at 12 weeks in subdued light at the time of other laboratory measurements.
After a 12-h fast and a 15-min supine rest period, one 20-ml tube without anticoagulant and two 12-ml heparinized tubes of blood were obtained with the tourniquet deflated. Whole blood without anticoagulant was centrifuged at 2,000 rpm for 30 min to separate serum. Aliquots of serum samples were frozen at −15°C for up to 4 weeks for serum lipid and lipoprotein lipid analysis, and fresh serum was used for LDL lag phase assay.
Heparinized blood was separated by refrigerated centrifugation at 2,300 × gfor 10 min. Plasma aliquots designated for tocopherol and carotenoid quantification were preserved in sodium ascorbate and stored at −70°C in cryogenic tubes for up to 6 months. One-milliliter aliquots designated for vitamin C analysis were processed with 10% trichloroacetic acid before freezing to enhance stability during short-term storage.
1.4 Measurement of lipids, lipoproteins and lag phase.
For measurement of lag phase, LDL was isolated immediately from fresh serum according to the method of Weiland and Seidel . This method highly correlates (r = 0.9) with results obtained from ultracentrifugation to isolate LDL.
The oxidation of polyunsaturated fatty acids in a buffer system was continuously monitored at 234 nm with a Beckman DU-6 spectrophotometer up to 10 h according to the method of Esterbauer et al. . Lag phaseis expressed in minutes and is the time interval from addition of Cu2+ions to the onset of the propagation phase of LDL oxidation, which is the maximal rate of conjugated diene production. Lag phase is determined by extrapolating a tangent drawn from the slope of the most rapid rise in the absorbance curve to the horizontal time axis. Lag phase is indicative of the resistance of LDL to oxidation; the greater the resistance of LDL to oxidation, the longer the lag phase.
Cholesterol and triglyceride concentrations were determined enzymatically with commercially available reagents (Sigma) by the methods of Allain et al. for total cholesterol (TC), LDL cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C) and McGowan et al. for TC, LDL-triglycerides. The isolation of LDL-C was according to the method of Weiland and Seidel . HDL-C was isolated by precipitation with phosphotungstic/MgCl2according to the method of Assmann et al. .
1.5 Plasma antioxidant measurements.
Plasma carotenoids were separated and quantified according to the high performance liquid chromatographic (HPLC) method of Bieri et al. , as modified by Craft et al. and Craft with further modifications to reduce oxidative loss and improve recovery of compounds during analysis. This analytic method measures 90% of the total plasma carotenoids present and permits quantification of the predominant serum carotenoids, including beta-carotene.
Plasma tocopherols were separated and quantified according to the HPLC method of Bieri et al. . This analytic method permits separation and quantification of alpha- and gamma-tocopherol, using tocopherol acetate (Sigma) as an internal standard. Plasma vitamin C was measured by the derivative spectrophotometric method of Omaye et al. .
1.6 Measurement of covariates.
Demographic variables, lifestyle habits and medication usage were assessed by standardized questionnaire. Anthropometric and physiologic variables were evaluated based on a standard examination. Body fat was calculated from three gender-specific skinfold measurements using the generalized equations of Jackson and Pollock and Jackson et al. for predicting body density.
Detailed dietary histories based on 3-day food records were obtained by a nutritionist 1 week before study entry and during the final week of the study. Diet records were analyzed using Nutritionist IV software (version 2.1 1995 N-Squared computing). Average daily total energy intake, percent energy from fat, percent energy from saturated fat, percent energy from carbohydrate, percent energy from protein and grams of alcohol/day and intake of antioxidants were determined at the beginning and end of the study.
1.7 Statistical analysis.
Statistical analysis was performed based on intention to treat. Continuous variables are expressed as mean value ± SD. Normal probability plots of all continuous variables were examined, and the Shapiro-Wilk statistic was used to test for normal distribution. Data that were not normally distributed, including lag phase, was log transformed when appropriate. Within-group change for lipids, plasma antioxidants and lag phase during the treatment period were evaluated using paired ttests. Between-group differences at baseline and 6 and 12 weeks for lipids, plasma antioxidants and lag phase were evaluated by analysis of variance (ANOVA) with a Tukey test to control for multiple pairwise comparisons. The relations between continuous variables were evaluated using Pearson correlation coefficients or Spearman correlations when data were not normally distributed. Differences in mean values for dichotomous variables were examined by Student ttests or Wilcoxon rank sum for nonnormally distributed variables. Significant independent predictors of lag phase were incorporated into a multiple linear regression model to assess the independence of antioxidant supplementation on resistance of LDL-C to oxidation. All tests were two-tailed, and statistical significance was set at p < 0.05. All data were double entered, discrepancies were corrected, and analysis was performed using SAS for Windows version 6.08 (1992 SAS Institute, Inc.). Blinding was maintained until the analysis was complete. The study was designed to detect a 10% difference in LDL oxidation with 80% power at p = 0.05.
Descriptive information for the 45 subjects with CAD is shown in Table 1. The only statistically significant difference between groups for any of the variables was the use of beta-adrenergic blocking agents. In the placebo group, seven subjects reported current use of beta-blockers use compared with four in the mid-dose group and one in the high dose group.
2.2 Lipid profiles.
Mean baseline and 6- and 12-week lipid profiles, according to group assignment, are presented in Table 2. There were no significant differences in baseline TC, LDL-C, HDL-C or triglyceride levels between groups. In the placebo group, HDL-C increased from 0.90 mmol/liter at baseline to 1.02 mmol/liter (p < 0.05) from baseline to 6 weeks. In the high dose group, TC and LDL-C significantly increased between 6 and 12 weeks, from 5.64 to 6.08 mmol/liter and from 4.09 to 4.56 mmol/liter, respectively. HDL-C increased between 6 and 12 weeks, from 0.99 to 1.08 mmol/liter (p < 0.05) in the high dose group. LDL-C and triglycerides increased significantly from baseline to 12 weeks in the high dose group, from 3.86 to 4.56 mmol/liter and from 1.75 to 2.07 mmol/liter, respectively.
2.3 Plasma antioxidants.
2.3.1 Within-group changes.
Plasma concentrations of alpha-tocopherol, vitamin C and beta-carotene were 24.43 ± 7.06, 71.94 ± 36.34 and 0.29 ± 0.17 μmol/liter, respectively, at baseline and did not significantly change at 12 weeks in the placebo group (Fig. 1). Results were unchanged when plasma alpha-tocopherol was lipid standardized. Alpha-tocopherol levels increased twofold during the trial in the mid-dose group, from 28.63 ± 7.7 to 60.57 ± 17.4 μmol/liter (p < 0.01), and threefold in the high dose group, from 27.39 ± 10.10 to 88.72 ± 47.85 μmol/liter (p < 0.01). Vitamin C levels increased 1.5 fold in the mid-dose group, from 73.13 ± 36.34 to 107.31 ± 28.95 μmol/liter (p < 0.05), and nearly twofold in the high dose group, from 61.95 ± 22.14 to 118.67 ± 35.77 μmol/liter (p < 0.01). Beta-carotene levels also significantly increased with supplementation in the mid- and high dose groups, from 0.34 ± 2.01 to 1.99 ± 1.17 μmol/liter (p < 0.01) and from 0.29 ± 0.1 to 3.01 ± 1.5 μmol/liter (p < 0.05), nearly six- and tenfold, respectively.
2.3.2 Between-group changes.
A significant between-group difference at 12 weeks for plasma concentrations of alpha-tocopherol was observed (p = 0.0001, ANOVA), with significant pairwise comparisons between mid-dose versus placebo, high dose versus placebo and high versus mid-dose. Beta-carotene levels were significantly different between groups at 12 weeks (p = 0.0001, ANOVA), with significant pairwise comparisons between placebo versus mid-dose and placebo versus high dose. No statistically significant between-group differences were noted for vitamin C at 12 weeks (p = 0.057). However, the between-group differences for change from baseline to 12 weeks were significant for vitamin C (p = 0.0001), which was attributable to a significant pairwise comparison for high dose versus placebo. Both alpha-tocopherol and beta-carotene plasma levels showed significant between-group changes over time for high dose versus placebo and mid dose versus placebo (p < 0.05).
2.4 Lag phase.
2.4.1 Within-group changes.
Table 2also describes the change in lag phase after treatment at 6 and 12 weeks according to group assignment. In the placebo group, there is a significant increase in lag phase between baseline and 6 weeks, from 219.5 ± 69.4 to 258.2 ± 77.0 min (p < 0.01), with no significant increase at 12 weeks (256.8 ± 73.8 min) compared with that at baseline or 6 weeks. The mid-dose group experienced an increase in lag phase between baseline and 6 weeks, from 182.0 ± 100.6 to 267.1 ± 76.3 min (p < 0.01) and between 6 and 12 weeks, from 267.1 ± 76.3 to 311.8 ± 112.5 min (p < 0.01). The high dose group had a significant increase in lag phase at 12 weeks (391.1 ± 153.0 min) compared with that at 6 weeks (239.1 ± 97.3, p < 0.01) and baseline (190.1 ± 63.8 min, p < 0.01). There was no statistically significant increase in lag phase between baseline and 6 weeks.
2.4.2 Between-group changes.
Lag phase did not differ significantly between treatment groups or placebo at baseline or at 6 weeks. At 12 weeks there was a significant between-group difference in mean lag phase (p = 0.02, ANOVA). The change in lag phase from baseline to 6 weeks did not differ between groups, but the change from baseline to 12 weeks did differ (p = 0.04, ANOVA).
Fig. 2illustrates a significant dose response for mean percent change in lag phase between the treatment groups from baseline to 12 weeks (p = 0.004 for trend). The mean percent increase in lag phase from baseline to 12 weeks in the mid-dose group was 71% compared with a doubling of lag phase in the high dose group. The mean percent change in lag phase from baseline to 12 weeks was highly correlated with mean percent change in plasma alpha-tocopherol (r = 0.48, p = 0.002) and beta-carotene (r = 0.49, p = 0.002).
No significant within or between-group differences at baseline or at 12 week follow-up were noted for dietary intake of antioxidant nutrients, total calories, percent energy from fat, saturated fat, carbohydrate, protein or grams of alcohol/day.
2.5 Predictors of lag phase.
No statistically significant dichotomous predictors of lag phase were identified. Continuous predictors of lag phase are shown in Table 3. The only significant lipid predictor of lag phase was LDL-C (r = −0.39, p = 0.01). Significant inverse correlations existed for body mass index (r = −0.33, p < 0.05), heart rate (r = −0.34, p < 0.05) and percent body fat (r = −0.41, p < 0.01). Only heart rate retained significance when univariate predictors were examined in multiple regression models controlling for age, gender and LDL-C at 12 weeks and LDL-triglycerides. Heart rate was significantly associated with beta-blocker use (p = 0.02); however, heart rate still was a significant predictor of lag phase when beta-blocker use was controlled for in a multiple regression model (p = 0.02).
Supplementation with a combination of vitamin E (800 IU), vitamin C (1,000 mg) and beta-carotene (24 mg) was a significant predictor of lag phase in multiple regression models controlling for age and gender (p = 0.008, model R2= 0.21). When heart rate and beta-blocker use were added to the model, high dose group assignment still remained a significant predictor of lag phase (p = 0.02, model R2= 0.23).
The major finding of our studywas a significant reduction in the susceptibility of LDL-C to oxidation in patients with CVD supplemented daily for 12 weeks with a combination of 800 IU of vitamin E, 1,000 mg of vitamin C, and 24 mg of beta-carotene 24 daily compared with placebo. In addition, we observed a significant dose response for this high dose combination compared with a combination of 400 IU of vitamin E, 500 mg of vitamin C and 12 mg of beta-carotene and a placebo control.
This finding is consistent with data from a study by Jialal and Grundy who observed a reduction in the rate of LDL oxidation in 24 healthy men given 800 IU of dl-alpha-tocopherol in the form of soy bean capsules compared with placebo. In their study, indexes of oxidized LDL, time course curves of thiobarbituric acid-reacting substances (TBARS) activity or conjugated diene formation were significantly lower among treated subjects at 6 and 12 weeks than at baseline. We did not detect significant between-group differences until 12 weeks. This finding suggests that a longer exposure to antioxidant nutrients may be necessary to protect LDL against oxidation in patients with CVD compared with healthy subjects and may be important in the design of future studies of antioxidant supplementation in patients with CVD or those with cardiovascular risk factors because these groups may have significant biologic variability in response to antioxidant supplementation. An alternative explanation for the lack of a significant treatment effect at 6 weeks may be the significant increase in lag phase that was observed in our control group at 6 weeks.
We chose lag phase to evaluate the effect of antioxidant supplementation on LDL oxidation as opposed to measurement of the products of peroxidation or the maximal rate of oxidation. Lipophilic antioxidants, such as vitamin E and beta-carotene, are carried in LDL and may protect polyunsaturated fatty acids against oxidation by competing with them for free radicals. Once antioxidants are depleted from LDL, the propagating chain reaction of lipid peroxidation ensues, so an increase in the amount of antioxidants in LDL would be expected to delay the onset of this process. Change in lag phase is most likely the most appropriate measure of the effect of antioxidant vitamin supplementation on LDL oxidation because it reflects altered susceptibility of the LDL particle to oxidation.
In a more recent dose–response study by Jialal et al. , a minimal dose of alpha-tocopherol (400 IU) was necessary to prolong lag phase in 48 healthy men after 8 weeks of vitamin E supplementation ranging from 60 to 1,200 IU daily. Our mid-dose group, which included 400 IU of alpha-tocopherol supplementation, did not show a significant prolongation of lag phase compared with placebo at 12 weeks. This difference may reflect an increased dose of antioxidant vitamins required to alter LDL-C oxidizability in patients with CVD or may be due to a lack of power to detect a difference at that level of supplementation in our study.
Abbey et al. showed a significant increase in lag time after 3 months in 45 nonsmoking subjects supplemented with a daily dose of 18 mg of beta-carotene, 900 mg of vitamin C, 250 mg of d-alpha-tocoheryl succinate (equivalent to 50 mg of vitamin E) and 12 mg of elemental zinc. Plasma concentrations of beta-carotene, alpha-tocopherol and ascorbic acid increased fivefold: 55% and 27%, respectively. We observed similar or greater changes in plasma antioxidants in our mid-dose group but did not detect a significant increase in lag phase in that group compared with the placebo group. This result raises the possibility that patients with CAD may require more significant changes in plasma antioxidants to protect the LDL particle from susceptibility to oxidation relative to that in healthy subjects. However, this finding should be interpreted with caution because our power to detect a statistically significant increase in lag phase in the mid-dose group compared with the placebo group was low because of the small sample size. Substantial individual variability in the ability of antioxidants to protect LDL from oxidation may also exist in patients with CAD.
Our study was also consistent with a crossover study of eight healthy, mildly hyperlipidemic volunteers by Reaven et al. , who used in vitro assays of oxidation to show that long-term supplementation with large doses of vitamin E conferred increased protection to LDL. A linear increase in LDL alpha-tocopherol levels up to an intake of 800 IU/day in 20 healthy nonsmoking volunteers was demonstrated by Princen et al. . Simultaneously, the resistance of LDL to oxidation was increased in a dose-dependent manner and differed significantly from baseline even after ingestion of only 25 IU/day.
The finding of a significant increase in lag phase at 6 weeks in our placebo control group may have been due to chance. It is interesting that the trend was similar for all groups and raises the possibility that altered lifestyle habits, such as diet, in the initial phase of the study may have been associated with an improvement in lag phase in all groups, making it difficult to detect between-group differences at the 6 week time point. Beard et al. recently demonstrated an increase in lag phase in subjects participating in a 3-week residential program with access to high complex carbohydrate/low fat foods and formal exercise classes. The significant increase in plasma levels for all antioxidants at 12 weeks may have been able to overcome nonpharmacologic effects on lag phase at that point. Furthermore, we documented no significant within- or between-group differences in dietary variables at 12 weeks, so any alterations in dietary habits may have returned to baseline levels by the conclusion of the study.
An alternative explanation for the significant increase in lag phase at 6 weeks in the placebo group is the corresponding significant increase in HDL-C in the placebo group at 6 weeks. HDL-C is protective for LDL oxidation, and the increase in lag phase that was observed in the placebo group at 6 weeks may have been a reflection of the increase in HDL-C that occurred in the placebo group in that time frame. We were surprised by the finding that LDL-C and triglycerides were significantly increased from baseline in the high dose group. Tsai et al. has reported a significant reduction of serum thyroid hormone levels and an elevation of serum triglyceride levels in women receiving supplemental megadoses of vitamin E (600 IU). A recent review of the safety of vitamin E supplementation suggests inconsistent effects on serum lipid and lipoprotein levels. It is unclear whether the lipid alterations that we observed are adverse or simply reflect conformational changes in LDL, which we did not evaluate in the present study. The increase in LDL-C that was observed in the high dose group is not likely to explain the effect of treatment on lag phase. LDL-C was inversely related to lag phase, and therefore an increase in LDL-C would tend to attenuate the effect of treatment in the high dose group.
The prolongation in lag phase with high dose supplementation of antioxidant nutrients in our study is consistent with the results of the Cambridge Heart Antioxidant Study (CHAOS) . In that randomized, clinical trial of 2,002 patients with angiographically proven CVD, a nearly 50% reduction in the combined end point of CVD death and nonfatal events was demonstrated with supplementation of vitamin E (400 to 800 IU daily), attributable to a significant reduction in nonfatal myocardial infarction.
3.1 Study limitations.
Our study is limited because we used a combination of antioxidant vitamins, and therefore we cannot define the relative contribution of each individual component of the supplement we utilized to the inhibition of LDL oxidation. Because vitamin E is contained in the LDL particle at substantially higher concentrations than other antioxidant nutrients, we speculate that the alpha-tocopherol component would have a major impact on reducing the susceptibility of LDL to oxidation. Future studies should address the effects of vitamin E with and without other antioxidant supplements on LDL oxidation and other cardiovascular risk factors.
It is unlikely that the powerful effect of antioxidant supplementation on LDL oxidation that we observed can be explained by selection bias or confounding because this was a randomized clinical trial, and groups were well matched at baseline. Although beta-blocker use did differ between groups, there were no significant differences in baseline lag phase between groups, and drug usage did not change throughout the trial. If beta-blockers do confer a protective effect on LDL to oxidation, our results would be strengthened because the placebo control group had increased usage of beta-blockers. Our data suggest that it is heart rate and not beta-blocker use that is a significant multivariate predictor of lag phase. We questioned whether this finding may be explained by improved levels of fitness and attempted to evaluate exercise level as a predictor of lag phase. We were unable to detect significant effects of exercise on lag phase, which may be due to our small sample size and the small range of variation in exercise levels in our subjects. Future studies should address the role of exercise and fitness levels in predicting changes in the oxidizability of LDL.
We demonstrated a significant reduction in the susceptibility of LDL to oxidation, as evidenced by a doubling of mean lag phase, in patients with CAD supplemented with a combination of vitamin E (800 IU), vitamin C (1,000 mg) and beta-carotene (24 mg). These data lend support for a role of antioxidants in secondary prevention; however, the potential adverse effects of high dose vitamin supplementation deserves further investigation.
We are indebted to Terri Johnson, Elizabeth Maimon and Leslie Bluman for assistance with data collection. We are grateful to Jackie Toms for technical assistance in the preparation of the manuscript, and we sincerely appreciate the helpful comments and critical review of the manuscript by Kim Eagle, MD.
☆ This study was supported by a gift from Kay and Z. Harold Peplau.
- analysis of variance
- body mass index
- coronary artery disease
- cardiovascular disease
- high density lipoprotein
- high density lipoprotein cholesterol
- high performance liquid chromatography
- low density lipoprotein
- low density lipoprotein cholesterol
- total cholesterol
- Received December 1, 1996.
- Revision received April 14, 1997.
- Accepted April 24, 1997.
- The American College of Cardiology
- ↵(1996) Heart and Stroke Facts: 1996 Statistical Supplement (American Heart Association, Dallas (TX)).
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