Dietary flavonoid intake and cardiovascular risk: a population-based cohort study

The results of this population-based cohort study suggest that higher dietary intakes
of flavonoids may be associated with a reduced CV risk score and a 40–50% lower risk
of non-fatal CV events.

This is intriguing since in our cohort the consumption of some flavonoid-rich foods
inversely associated with CV risk such as cocoa, soybean and tea 13], 17], 28], 29], is infrequent, being fruits and red wine the main sources of flavonoids. Epidemiological
studies have suggested that a Mediterranean diet reduces the CV risk 42] and a high concentration of flavonoids has been found in fruits, vegetables, red
wine and other elements of the Mediterranean diet. However, there is inconsistent
evidence on the role of flavonoids derived from these foods and CV risk, since previous
studies reported either a decreased CV incidence and mortality with increased intake
of apples, pears, and red wine 8], 12], 16], 24], 25], or no significant effect 6], 10], 21], 26], 27].

We have found both a lower CV risk score at baseline and at follow-up in the higher
tertile of flavonoid intake. Intriguingly, the increase in the score from enrolment
to the end of follow-up was higher in those individuals. Accordingly, the consumption
of flavonoid-rich food has been associated with lower systolic blood pressure 15], 43], 44], lower total cholesterol 44], higher HDL cholesterol values 44]–46].

Benefits of flavonoids on blood pressure, lipid values, insulin resistance, and flow-mediated
dilatation seem to derive above all from soy, cocoa and tea, as suggested by systematic
reviews 47], 48]. However, more recently, flavonoids from fruits and vegetables have been reported
to reduce the risk of diabetes mellitus and to improve microvascular reactivity and
inflammatory status 49]–51]. Accordingly, although a small number of incident CV events occurred in our cohort,
the risk of non-fatal CV events was significantly lower in individuals with the higher
intake of total and all subclasses of flavonoids, but flavones and isoflavones, which
were consumed at negligible concentrations in our cohort. Therefore, the dietary intakes
of flavonoids seems relevant for healthy CV outcomes at relatively low concentrations,
since most inverse associations with CV risk score and non-fatal CV events appeared
with intermediate or low intakes of specific subclasses, suggesting that even small
amounts may be beneficial. However, a threshold of intake is probably needed, under
which these compounds are unlike to be active.

Flavonoids can inhibit or induce a large variety of enzyme systems, involved in pathways
regulating platelet aggregation, inflammatory and immune responses 1], 52], 53]. Furthermore, by their antioxidant properties, flavonoids may protect tissues against
oxygen free radicals and lipid peroxidation, thus contributing to the prevention of
atherosclerosis, chronic inflammation and cancer 1], 52], 53]. Because of their antioxidant and chelating properties, flavonoids may inactivate
reactive oxygen species (ROS) and counteract the oxidation of circulating LDL particles
52]–54]. Other anti-atherogenic actions proposed for these compounds are: reduction of the
activity of enzymes increasing ROS production; inhibition of HMG-CoA reductase, cholesteryl
ester transfer protein (CEPT), angiotensin-converting enzyme, signal transducers and
activators of transcription (STAT), and glucose transporters; synthesis of nitric
oxide; inhibition of platelet activation and function; anti-angiogenetic effects;
improvement in endothelial function, vascular fragility, cellular permeability 54]–56]. The anti-inflammatory properties of flavonoids may be due to the inhibition of NF-?B
activation and adhesion molecule expression; suppression of the activity and secretion
of inflammatory cells; reduction of the concentrations of CRP and cytokines 57], 58].

The associations with fatal events were controversial in our cohort. No significant
association was found with CV mortality, probably because the number of fatal CV events
was low. Otherwise, many flavonoid subclasses, such as flavan-3-ols, anthocyanidins
and flavanones were inversely associated with all-cause mortality. Previous studies
have reported a reduced total and/or CV mortality with proanthocyanids 19], flavan-3-ols, 11], 19], 25], anthocyanidins 16], 19], flavonols 13], 19], flavanones 16], 18], flavones 16], 19], and isoflavones 17], 44]–47]. On the other hand, other authors reported no protective effects of total or specific
subclasses of flavonoids on mortality 7], 12], 21], 22], 27].

These highly divergent results among studies might be due to differences in nutritional,
socio-cultural and ethnic characteristics.

The median intakes of flavonoids are highly variable among studies, and values ranging
from 50 to 450 mg have been reported in European studies 54]. In particular, the following median intakes have been described for Mediterranean
countries: 92 mg/day in Greece 59] and 332.4 mg/day in Spain 60]. On the other hand, in non-Mediterranean countries, the median consumption of flavonoids
was much lower, varying from 203 mg/day in US population 19] to 88 mg/day in Sweden, and 13 mg/day in Finland 61]. Our values were between these extreme intakes, in line with other Italian data 62], 63]. The high consumption of red wine and fruits, such as apples and citrus fruits, in
our Italian cohort justify the higher intake of total flavonoids and proanthocyanidins
with respect to other non-Mediterranean cohorts 19], 61]. On the other hand, the low consumption of tea, justified the lower intakes of flavon-3-ols
(in particular epigallocatechin 3-gallate, epicatechin 3-gallate and epigallocatechin)
with respect to UK and Ireland 61], and the negligible use of soy explain why the intake of isoflavones and flavones
was much lower in our cohort when compared with Asian studies 17].

In most studies, the higher consumption of flavonoids was associated with an overall
healthy dietary and metabolic pattern, in line with our results 8], 10]–12], 16]–21], 25], 26], 49]. Our cohort indeed included individuals with a low level of education, differently
from previous studies performed in samples where most participants had at least a
high school education 13], 16], 17], 19], 26].

Finally, many compounds tend to be present in the same foods: for example, in our
cohort, individuals with lower intakes of flavonoids, ate less fiber and antioxidant
vitamins and more saturated fats. It is therefore difficult to ascertain the independent
effect of dietary components because of multicollinearity. However, our associations
remained significant after adjusting for these dietary factors, thus suggesting that
a higher flavonoid intake might not merely be an indicator of a healthier lifestyle.

Limitations

The EPIC questionnaire was not originally designed to measure flavonoid intake, but
it has been extensively used and validated for this purpose 60], 64], 65].

The flavonoid intake might have been underestimated because of the limitations of
the food composition databases. It should be noted that the presence of particular
flavonoids in vegetables and fruits depends on the crop variety, location and type
of cultivation. The adaptability of the USDA database to the Italian diet is questionable.
The absorption and microbial transformation in the gut of specific subclasses of flavonoids
vary considerably, therefore the different flavonoid bioavailability could have an
impact on the associations between the assumption of these compounds and chronic diseases.
In general, flavonoid subclasses are present simultaneously in foods and establishing
which of the compound is responsible for the potential biological effect is difficult.
We relied on dietary intake from the questionnaire administered at one point in time;
thus misclassification of dietary exposure might have occurred if individuals have
changed their diets during the follow-up. Furthermore, measurement error in collecting
self-reported dietary intake is inevitable and our observational study was prone to
the possibility of unmeasured confounding.

However, the recent versions of the USDA database includes more cooked foods 2], because in culinary preparations important losses in flavonoid content occur, and
is the most complete and used database in the estimation of flavonoid intake. Moreover,
we have referred also to a European database, and the USDA has been already used for
the Italian population 62]–64]. We have used a validated instrument and, both at baseline and at follow-up, the
associations between flavonoid intakes and the CV risk score were consistent. Measurement
errors and misclassification was likely to be random and would have attenuated the
association found. We have took care to adjust for many potential confounders. Finally,
we have studied a large population-based cohort from a localized region, with a high
level of participation, which could have limited the number of potential confounders.