EAL

Cardiovascular Disease and Micronutrients

Citation:

Study Design:

Class:

- Click here for explanation of classification scheme.

Quality Rating:

Research Purpose:

To test the effect of vitamin E and C supplements, in moderate doses, on the progression of carotid atherosclerosis in men and post-menopausal women with hypercholesterolemia.

Inclusion Criteria:

Hypercholesterolemia (serum cholesterol at least 193mg per dL)

Exclusion Criteria:

Regular intake of antioxidants, acetosalicylate or other drugs with antioxidant properties
Severe obesity (BMI above 32kg/m²)
Type 1 diabetes
Uncontrolled hypertension (sitting diastolic blood pressure above 105mmHg)
Any condition limiting mobility
Severe disease shortening life expectancy
Pre-menopausal women
Women taking oral estrogen therapy.

Description of Study Protocol:

Recruitment: Volunteers were screened over the phone and eligible persons were invited to participate
Design: Randomized clinical trial
Blinding used: IMT measurement technician was blinded to participant supplementation. First three years of study were double-blinded. Second three years were open study.

Intervention

First three years of study:

91mg per day alpha-tocopherol (corresponding to 100mg per day alpha-tocopheryl acetate and 136 IU vitamin E), twice daily
250mg slow-release ascorbic acid, twice daily
Both 91mg per day alpha-tocopherol and 250mg slow-release ascorbic acid in a single tablet, twice daily
Placebo, twice daily.

Study Years Three through Six:

All supplemented subjects receive supplement combination (91mg per day alpha-tocopherol and 250mg slow-release ascorbic acid in a single tablet, twice daily
Placebo subjects receive no supplementation.

Statistical Analysis

Intention-to-treat included 440 participants for whom ultrasound examination was available at six years
Subjects randomized in strata and a priori power calculations based on analysis in men. Statistical analysis also done in men and women separately.
Non-parametric Mann-Whitney test to examine differences between treatment groups
General linear modeling of SPSS 10.0 for Windows for comparisons taking into account covariates
Two-sided P-values
Linear regression slope of the mean CCA-IMT over all points of follow-up time was used as the primary outcome measure
- Mean CCA-IMT from the right and left side was averaged, then slope computed across time-specific means
- Simple difference between the final mean IMT at 72 months and baseline mean IMT calculated for confirmation.

Data Collection Summary:

Timing of Measurements

Common carotid artery intima-media thickness was measured at baseline and follow-up visits at six, 12, 18, 24, 30, 36 and 72 months
Ascorbic acid measured by high-pressure liquid chromatography
Alpha-tocopherol measured by reversed phase high-pressure liquid chromatography
Plasma F₂-isoprostane concentrations measured by gas chromatography-mass spectrometry
Concomitant diseases and medications recorded at study Years Zero, One, Three and Six.

Dependent Variables

Common carotid artery intima-media thickness measured by high-resolution ultrasonography: Severity of atherosclerosis classified using four-categorical scale separately for right and left common carotid artery.

Independent Variables

91mg per day alpha-tocopherol, twice a day
250mg slow release ascorbic acid, twice a day
Both 91mg per day alpha-tocopherol and 250mg slow-release ascorbic acid, twice a day (the only independent variable tested in Years Three to Six)
Placebo (unsupplemented in Years Three to Six).

Control Variables

Gender
Baseline mean CCA-IMT
Classification of severity of right and left CCA
Baseline vitamin C concentration
Three indicator variables for summer baseline examination months.

Description of Actual Data Sample:

Initial N: 520 (256 males, 264 post-menopausal females)
Attrition (final N): 440 (212 men, 228 women) completed the study and six-year ultrasound re-examination. Of the 390 subjects randomized to supplementation, 335 continued study after three years and 256 (76.4%) adhered to supplements for six years.
Age: Middle-aged (45 to 69 years)
Ethnicity: Danish, Finnish and Swedish
Other relevant demographics: None given
Anthropometrics: Not given except BMI above 32kg/m² excluded
Location: Denmark and Finland.

Summary of Results:

Mean Annual Increase of the Mean Common Carotid Artery Intima-Media Thickness (CCA-IMT)	Supplemented Subjects (vitamins C+E)	Non-supplemented Subjects	Statistical Significance
Linear slope across study period	0.0118±0.0136mm	0.0156±0.0182mm	P=0.007
Simple difference between 72-month and baseline	0.0103±0.014mm	0.0134±0.016mm	P=0.007

Other Findings

CCA-IMT progression

Small reduction in mean CCA-IMT during first year of study, after which CCA-IMT started to progress linearly. Lines began to diverge after first year.
Among those who took their supplements (N=256), the CCA-IMT linear slope was 0.0111mm per year (29% treatment effect, P=0.004)
Among those who took their supplements (N=256), the CCA-IMT simple difference between 72-month and baseline assessments was 0.0095±0.012mm per year
In men, difference between treatment groups in slope (P=0.008) and difference (P=0.002) was statistically significant. In women, neither was statistically significant.
- In both men and women combined, there was a 26% treatment effect in slope (95% CI, 5 to 46; P=0.014) and a 30% treatment effect in those who took their supplements (95% CI, 10 to 51; P=0.003)
- In all men, treatment effect in slope was 33% (95% CI, 4 to 62; P=0.024) and in all women, 14% (95% CI, -11 to 44, not significant)
  - Among compliant men, treatment effect was 39% (95% CI, 9 to 69, P=0.019).
  - Among compliant women, treatment effect was 17% (95% CI, -11 to 44, P=0.235).
- IMT mean simple difference in men and women combined was 0.0137mm in the non-supplemented group and 0.0102mm in the supplemented group, a 25% treatment effect (95% CI, 2 to 49; P=0.034)
- In men, the average annual increase of CCA-IMT was 0.0162mm in the non-supplemented group and 0.0103mm in the supplemented group, a 37% treatment effect (95% CI, 4 to 69; P=0.028). Difference was not significant in women.
Treatment effect was greater in participants who already had plaques in CCAs at baseline (54% in those who had at least one plaque obstructing greater than 20% arterial lumen)
Treatment effect also larger in those with low baseline vitamin C plasma levels
Six-year change in mean CCA-IMT was inversely correlated with change in plasma alpha-tocopherol concentration in men, but not women.

36-month change in F₂-Isoprostane concentrations

In men, F₂-Isoprostane concentration was significantly reduced in vitamin E group, but was increased in placebo and vitamin C groups: Treatment effect of vitamin E (mean -6.2ng per L; 95% CI, -2.2 to -10.2) was statistically significant (P=0.003, N=100)
In women, F₂-Isoprostanes decreased more in placebo group than in either vitamin E or C groups.

36-month change in HDL-cholesterol

Mean HDL-cholesterol increased more significantly at three years in men who received vitamin C supplement than placebo (P=0.025)
Vitamin E had no effect on HDL-cholesterol in men
Neither vitamin E nor C had any effect on HDL in women.

Author Conclusion:

This trial showed that long-term vitamin E and slow-release vitamin C supplementation combined, in reasonable doses, can slow the progression of common carotid atherosclerosis, especially in men.

Funding Source:

University/Hospital:	Academy of Finland
In-Kind support reported by Industry:	Yes

Reviewer Comments:

Quality Criteria Checklist: Primary Research
Relevance Questions
	1.	Would implementing the studied intervention or procedure (if found successful) result in improved outcomes for the patients/clients/population group? (Not Applicable for some epidemiological studies)	Yes
	2.	Did the authors study an outcome (dependent variable) or topic that the patients/clients/population group would care about?	Yes
	3.	Is the focus of the intervention or procedure (independent variable) or topic of study a common issue of concern to dieteticspractice?	Yes
	4.	Is the intervention or procedure feasible? (NA for some epidemiological studies)	Yes

Validity Questions
1.	Was the research question clearly stated?		Yes
	1.1.	Was (were) the specific intervention(s) or procedure(s) [independent variable(s)] identified?	Yes
	1.2.	Was (were) the outcome(s) [dependent variable(s)] clearly indicated?	Yes
	1.3.	Were the target population and setting specified?	Yes
2.	Was the selection of study subjects/patients free from bias?		Yes
	2.1.	Were inclusion/exclusion criteria specified (e.g., risk, point in disease progression, diagnostic or prognosis criteria), and with sufficient detail and without omitting criteria critical to the study?	Yes
	2.2.	Were criteria applied equally to all study groups?	Yes
	2.3.	Were health, demographics, and other characteristics of subjects described?	Yes
	2.4.	Were the subjects/patients a representative sample of the relevant population?	Yes
3.	Were study groups comparable?		Yes
	3.1.	Was the method of assigning subjects/patients to groups described and unbiased? (Method of randomization identified if RCT)	Yes
	3.2.	Were distribution of disease status, prognostic factors, and other factors (e.g., demographics) similar across study groups at baseline?	Yes
	3.3.	Were concurrent controls or comparisons used? (Concurrent preferred over historical control or comparison groups.)	Yes
	3.4.	If cohort study or cross-sectional study, were groups comparable on important confounding factors and/or were preexisting differences accounted for by using appropriate adjustments in statistical analysis?	N/A
	3.5.	If case control study, were potential confounding factors comparable for cases and controls? (If case series or trial with subjects serving as own control, this criterion is not applicable.)	N/A
	3.6.	If diagnostic test, was there an independent blind comparison with an appropriate reference standard (e.g., "gold standard")?	N/A
4.	Was method of handling withdrawals described?		No
	4.1.	Were follow-up methods described and the same for all groups?	Yes
	4.2.	Was the number, characteristics of withdrawals (i.e., dropouts, lost to follow up, attrition rate) and/or response rate (cross-sectional studies) described for each group? (Follow up goal for a strong study is 80%.)	Yes
	4.3.	Were all enrolled subjects/patients (in the original sample) accounted for?	No
	4.4.	Were reasons for withdrawals similar across groups?	Yes
	4.5.	If diagnostic test, was decision to perform reference test not dependent on results of test under study?	N/A
5.	Was blinding used to prevent introduction of bias?		No
	5.1.	In intervention study, were subjects, clinicians/practitioners, and investigators blinded to treatment group, as appropriate?	No
	5.2.	Were data collectors blinded for outcomes assessment? (If outcome is measured using an objective test, such as a lab value, this criterion is assumed to be met.)	Yes
	5.3.	In cohort study or cross-sectional study, were measurements of outcomes and risk factors blinded?	N/A
	5.4.	In case control study, was case definition explicit and case ascertainment not influenced by exposure status?	N/A
	5.5.	In diagnostic study, were test results blinded to patient history and other test results?	N/A
6.	Were intervention/therapeutic regimens/exposure factor or procedure and any comparison(s) described in detail? Were interveningfactors described?		Yes
	6.1.	In RCT or other intervention trial, were protocols described for all regimens studied?	Yes
	6.2.	In observational study, were interventions, study settings, and clinicians/provider described?	N/A
	6.3.	Was the intensity and duration of the intervention or exposure factor sufficient to produce a meaningful effect?	Yes
	6.4.	Was the amount of exposure and, if relevant, subject/patient compliance measured?	Yes
	6.5.	Were co-interventions (e.g., ancillary treatments, other therapies) described?	No
	6.6.	Were extra or unplanned treatments described?	No
	6.7.	Was the information for 6.4, 6.5, and 6.6 assessed the same way for all groups?	Yes
	6.8.	In diagnostic study, were details of test administration and replication sufficient?	N/A
7.	Were outcomes clearly defined and the measurements valid and reliable?		Yes
	7.1.	Were primary and secondary endpoints described and relevant to the question?	Yes
	7.2.	Were nutrition measures appropriate to question and outcomes of concern?	Yes
	7.3.	Was the period of follow-up long enough for important outcome(s) to occur?	Yes
	7.4.	Were the observations and measurements based on standard, valid, and reliable data collection instruments/tests/procedures?	Yes
	7.5.	Was the measurement of effect at an appropriate level of precision?	Yes
	7.6.	Were other factors accounted for (measured) that could affect outcomes?	Yes
	7.7.	Were the measurements conducted consistently across groups?	Yes
8.	Was the statistical analysis appropriate for the study design and type of outcome indicators?		Yes
	8.1.	Were statistical analyses adequately described and the results reported appropriately?	Yes
	8.2.	Were correct statistical tests used and assumptions of test not violated?	Yes
	8.3.	Were statistics reported with levels of significance and/or confidence intervals?	Yes
	8.4.	Was "intent to treat" analysis of outcomes done (and as appropriate, was there an analysis of outcomes for those maximally exposed or a dose-response analysis)?	Yes
	8.5.	Were adequate adjustments made for effects of confounding factors that might have affected the outcomes (e.g., multivariate analyses)?	Yes
	8.6.	Was clinical significance as well as statistical significance reported?	No
	8.7.	If negative findings, was a power calculation reported to address type 2 error?	N/A
9.	Are conclusions supported by results with biases and limitations taken into consideration?		Yes
	9.1.	Is there a discussion of findings?	Yes
	9.2.	Are biases and study limitations identified and discussed?	Yes
10.	Is bias due to study's funding or sponsorship unlikely?		Yes
	10.1.	Were sources of funding and investigators' affiliations described?	Yes
	10.2.	Was the study free from apparent conflict of interest?	Yes