Vitamin C deficiency and risk of myocardial infarction: prospective population study of men from eastern Finland

Introduction

The oxidation of low density lipoprotein has been implicated in atherogenesis and in the progression of early atherosclerotic plaques.1 2 3 Experiments in rabbits have shown that atherogenesis can be inhibited by supplementation with antioxidants.1 4 5 6 7 Epidemiological follow up studies suggest that a high intake of vitamin E might be associated with a reduced risk of coronary events.8 9 Increased body stores of the transition metals iron and mercury, which catalyse lipid peroxidation, have been associated with excess risk of myocardial infarction.10 11 12

We found that a high titre of autoantibodies against oxidised low density lipoprotein was associated with accelerated progression of carotid atherosclerosis in a prospective nested case-control study.13 Consistently, associations have been reported between raised titres of antibodies against oxidised low density lipoprotein14 and reduced resistance to oxidation of low density lipoprotein15 and increased severity of atherosclerosis. Vitamin C is an important water soluble dietary antioxidant in humans. We previously reported an association between low plasma vitamin C concentrations and enhanced progression of atherosclerosis.16 Only a few studies have investigated the relation between the use of vitamin C supplements, dietary intake of vitamin C, or plasma vitamin C concentration and the risk of coronary disease, and their results are inconsistent.3 None of them has considered the role of vitamin C deficiency.

We investigated the association of vitamin C deficiency with the risk of myocardial infarction in middle aged men free of coronary disease from eastern Finland, a population with low average vitamin C concentrations and high mortality from coronary heart disease.

Subjects and methods

The Kuopio ischaemic heart disease risk factor study is a population study designed to investigate risk factors for cardiovascular diseases, atherosclerosis, and related outcomes in men from eastern Finland.18 The baseline examinations were carried out between March 1984 and December 1989. The study sample included 3235 eligible men aged 42, 48, 54, or 60 at the baseline examination; 2682 (82.9%) participated. Men with symptomatic coronary heart disease (n = 92) or ischaemia on maximal exercise testing (n = 359),19 or both (n = 585), were excluded from the current analyses. Of the remaining 1646 men, 1605 had data on plasma ascorbate concentration at baseline.

The examination protocol and measurements have been described.10 11 12 13 Plasma ascorbate concentrations were determined by a chromatographic method in deep frozen samples, which were stabilised in 5% metaphosphoric acid immediately after sampling.20 The coefficient of variation between 12 batches was 7.2%.20

Plasma ascorbate concentration was redetermined in a subsample of 401 men with hypercholesterolaemia in samples drawn 4-9 years after the baseline examination. Pearson's correlation between the baseline and the remeasurement values was 0.27. Plasma vitamin C concentration below <11.4 µmol/l (2.0 mg/l) was defined as being at high risk of developing clinical symptoms of hypovitaminosis C and is considered to be a limit for vitamin C deficiency.21 22 For these reasons we used this limit as the primary cut off value for plasma vitamin C concentration.

The main lipoprotein fractions were separated from fresh serum samples by ultracentrifugation and precipitation.23 The cholesterol and triglyceride contents of all lipoprotein fractions were measured enzymatically. Serum apolipoprotein B concentration was determined by an immunoturbidimetric method.12 Serum copper concentration was determined with an atomic-absorption spectrometric technique.24 Plasma fibrinogen concentration was determined on the basis of clotting of diluted plasma with excess thrombin.12 Hair mercury content was measured with a flow injection and amalgam system (FIAS-200) in a Perkin Elmer Zeeman 5000 Spectrometer (Norwalk, CT).12

The consumption of foods was assessed when blood was sampled by recording intake over four days with a questionnaire, which was checked at interview.25 The use of vitamin C tablets and vitamin C containing nutritional supplements was assessed in a self administered questionnaire.

The number of cigarettes, cigars, and pipefuls of tobacco currently smoked daily, the duration of regular smoking in years, and the history of myocardial infarction, angina, and other ischaemic heart disease were recorded using a self administered questionnaire, which was checked by an interviewer. A family history of coronary disease was defined as positive when the biological father, mother, sister, or brother of a subject had myocardial infarction, angina, or ischaemic heart disease.

The lifelong exposure to smoking (cigarette pack years) was estimated as the product of the number of years spent smoking and the number of tobacco products smoked daily at the time of examination. The consumption of alcohol in the previous 12 months was assessed with the Nordic alcohol consumption inventory, which contains 15 items.26 Socioeconomic status was measured with a summary index that combined measures of income, education, occupation, occupational prestige, standard of living, and housing conditions.27 Leisure time physical activity was assessed over the previous 12 months.28

Resting blood pressure was measured by one nurse with a random zero mercury sphygmomanometer.10 12 The mean of six systolic pressure values was used in these analyses. Respiratory gas exchange was measured breath by breath with the MGC 2001 system (Medical Graphics, Minneapolis, MI) during a symptom limited exercise test.19 Ischaemia in exercise electrocardiograms was coded manually by one cardiologist.10 12 Diabetes was defined as either previous diagnosis of diabetes mellitus or fasting blood glucose concentration >8.0 mmol/l.

Acute myocardial infarctions in the cohort were monitored by a registry that collected detailed diagnostic information of all such attacks in the study cohort in a prospective manner.29 30 Between March 1984 and December 1992 a fatal or non-fatal myocardial infarction was registered in 70 of the 1605 men at risk. The follow up period for individual subjects was up to 8.75 years, and the mean follow up time was about five years.

Risk factors for myocardial infarction were analysed using spss Cox proportional hazards models.31 32 All tests of significance were two sided. Relative hazards adjusted for risk factors (risks) were estimated as the antilogarithm of a coefficient for independent variables. Their confidence intervals were estimated with a macro based on the assumption of asymptotic normality of estimates.

Results

Plasma ascorbate concentration ranged from 1.7 µmol/l to 137 µmol/l, with a mean (SD) of 47.7 (23.3) µmol/l. There was a slow average increase in the mean plasma ascorbate concentration during the six years of baseline examinations (46.0, 43.2, 43.7, 49.4, 56.8, and 48.3 µmol/l; r = 0.12, P<0.0001 for linear trend). For this reason the year of examination was adjusted for in all statistical analyses.

Plasma ascorbate concentration decreased with increasing age (r = -0.17). The age specific plasma ascorbate mean concentrations were 55.7, 52.3, 44.3, and 46.6 µmol/l for those aged 42, 48, 54, and 60, respectively (P<0.0001 for linear trend). Of dietary factors, only the intake of fruits and berries (r = 0.276) and of vegetables (r = 0.255) had any notable adjusted correlations with plasma ascorbate concentration. Plasma ascorbate concentration also correlated with the dietary intake of carotenes (r = 0.159) and the sum of C14 to C16 saturated fatty acids (r = -0.096). Men with plasma vitamin C concentrations <11.4 µmol/l differed from men with higher plasma vitamin C concentrations in many respects (table 1). Men deficient in vitamin C were older, had smoked more in their life time, and were in lower socioeconomic groups; they also had lower maximal oxygen uptake, lower dietary carotene intake, a higher blood leucocyte count, lower dietary iron intake, less moderate to vigorous physical activity during leisure time, higher systolic blood pressure, higher alcohol intake, and higher coffee consumption.

Men who had vitamin C deficiency (plasma ascorbate <11.4 µmol/l) were at an increased risk of myocardial infarction in the subsequent years (table 2). The relative risk for this lowest category compared with the highest and after adjustment for age, season, and year of examination was 4.0 (95% confidence interval 1.7 to 9.4; P = 0.0012). There were no differences in the risk in the quarters of vitamin C concentration above this limit (table 2). Of 91 men with vitamin C deficiency, 12 (13.2%) had a myocardial infarction during the follow up, compared with 58 (3.8%) of the 1514 men with higher vitamin C concentrations (P = 0.00037, Fisher's exact test). The proportions of men with infarction in all categories from the lowest to the highest was 12 out of 91, 21 out of 382, 12 out of 383, 13 out of 376, and 12 out of 373 (P = 0.0013 for linear trend in proportions).

The strongest predictors of acute myocardial infarction after adjustment only for age (in years), year of examination (covariates for individual years), and season of the year (August to October v the rest of the year) in this study cohort were maximal oxygen uptake (inversely), the number of pack years smoked, blood leucocyte count, and low plasma ascorbate concentration (table 3). Serum apolipoprotein B, plasma fibrinogen, serum copper, and triglyceride concentrations, hair mercury content, blood haemoglobin concentration, systolic blood pressure, diabetes, moderate to vigorous physical activity during leisure time (inversely), serum high density lipoprotein-2 cholesterol concentration (inversely), family history of coronary disease, and low socioenonomic group were also significantly associated with the risk of myocardial infarction. Men who had vitamin C deficiency (plasma ascorbate concentration <11.4 µmol/l) had a 3.5-fold (1.8 to 6.7, P = 0.0002) risk of myocardial infarction after adjustment for age, season, and year of examination (table 3).

To estimate the independent impact of risk factors, all factors shown in table 3, as well as age, year of examination, season, and three dietary factors, were entered simultaneously in a Cox proportional hazards model. In this multivariate model only the number of cigarette pack years smoked, low plasma vitamin C concentration, family history of coronary heart disease, low maximal oxygen uptake, and high hair mercury content (in the order of strength) had significant associations with infarction risk (table 3). When all risk factors shown in table 3, as well as dietary intakes of tea, fibre, and saturated fats, were allowed for, men with plasma ascorbate concentrations <11.4 µmol/l had a relative risk of myocardial infarction of 2.5 (1.3 to 5.2, P = 0.0095) compared with those with higher plasma ascorbate concentrations.

Only 89 (5.5%) of our study subjects took supplements containing vitamin C. The exclusion of these men did not materially change the results. In fact, among the 1516 men who had not used vitamin C containing supplements during the previous week the relative risk for having a plasma ascorbate concentration <11.4 µmol/l was 3.7 (1.9 to 7.2, P = 0.0001) after adjustment for age, season, and year of examination. The use of vitamin C supplements had no significant association with the risk of myocardial infarction.

Discussion