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Journal of Nutrition in Gerontology and
Geriatrics
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The Potential of Flavanol and
Procyanidin Intake to Influence Age-
Related Vascular Disease
Roberta R. Holt PhD a , Christian Heiss MD b , Malte Kelm MD b & Carl
L. Keen PhD c
a Department of Nutrition, University of California, Davis, California,
USA
b Division of Cardiology, Pulmonology, and Vascular Medicine,
University Duesseldorf, Medical Faculty, Duesseldorf, Germany
c Department of Nutrition and Internal Medicine, University of
California, Davis, California, USA
Version of record first published: 13 Aug 2012
To cite this article: Roberta R. Holt PhD, Christian Heiss MD, Malte Kelm MD & Carl L. Keen PhD
(2012): The Potential of Flavanol and Procyanidin Intake to Influence Age-Related Vascular Disease,
Journal of Nutrition in Gerontology and Geriatrics, 31:3, 290-323
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The Potential of Flavanol and Procyanidin
Intake to Influence Age-Related
Vascular Disease
ROBERTA R. HOLT, PhD
Department of Nutrition, University of California, Davis, California, USA
CHRISTIAN HEISS, MD, and MALTE KELM, MD
Division of Cardiology, Pulmonology, and Vascular Medicine, University Duesseldorf,
Medical Faculty, Duesseldorf, Germany
CARL L. KEEN, PhD
Department of Nutrition and Internal Medicine, University of California,
Davis, California, USA
Advancing age is an independent major risk factor for cardio-
vascular disease (CVD). Age-associated impairments in the control
of inflammation, excessive oxidative stress, and reduced cellular
repair can all contribute to the development and progression of
CVD. Current recommendations for both the primary and second-
ary prevention of CVD promote lifestyle modifications that include
the adoption of healthy dietary patterns, such as the consumption of
diets rich in plant foods, as these have been associated with a lower
lifetime risk for the development of CVD. The potential for a diet rich
in plant foods to be cardiovascular protective is also supported by
prospective studies that suggest the intake of foods providing high
amounts of certain phytochemicals, in particular flavanols and
procyanidins, reduce the risk for CVD. These observations are
further supported by a number of dietary intervention trials that
show improvements in vascular function and reduced platelet
reactivity following the consumption of high flavanol foods. In the
current article we review a selection of these studies, and comment
on some of the potential mechanisms that have been postulated to
underlie the health effects of flavanol and procyanidin-rich foods.
Address correspondence to Carl L. Keen, Department of Nutrition and Internal Medicine,
University of California, One Shields Avenue, Davis, CA 95616, USA. E-mail: clkeen@ucdavis.edu
Journal of Nutrition in Gerontology and Geriatrics, 31:290–323, 2012
Copyright #Taylor & Francis Group, LLC
ISSN: 2155-1197 print=2155-1200 online
DOI: 10.1080/21551197.2012.702541
290
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KEYWORDS age, cardiovascular, epicatechin, flavanol,
procyanidin
INTRODUCTION
Cardiovascular disease (CVD) is the leading cause of morbidity and mortality
in the developed world. While advances in the diagnosis and standard of care
for CVD have been significant over the past decade (1), it is predicted that the
health burden of CVD will continue to grow given our aging populations (2).
The pathogenesis of CVD is multifactorial in nature, being viewed in part as a
chronic inflammatory disease (3) that is promoted by platelets and a dysregu-
lation of peripheral blood and vascular cells, including leukocytes, smooth
muscle, and endothelial cells (4–7). Cardiovascular risk factors, such as dysli-
pidemia, hypertension, and smoking, can promote the inflammatory process.
Apart from these traditional risk factors, advancing age is an independent
major risk factor for CVD. Age-associated impairments in the control of
inflammation, excessive oxidative stress, and reduced cellular repair can all
contribute to the development and progression of CVD.
Current recommendations for both the primary and secondary preven-
tion of CVD promote lifestyle modifications including increases in the level
of one’s physical activity, smoking cessation, and the adoption of dietary pat-
terns aimed at the maintenance of a healthy body weight, blood pressure,
blood glucose levels, and appropriate blood lipid and lipoprotein profiles.
While weight loss alone can often improve many of the above parameters,
there is compelling evidence that the consumption of diets rich in plant foods,
such as fruits, vegetables, cocoa, tea, wine, and nuts, can be associated with a
lower lifetime risk for the development of CVD (8–13). The potential for a diet
rich in plant foods to be cardioprotective over the entire lifecycle is supported
by findings in the Cardiovascular Risk in Young Finns Study showing that
high fruit and vegetable intakes in childhood are positively associated with
good vascular function in adulthood (14). A plethora of epidemiological
studies show inverse associations between the intake of plant food–rich diets
and CVD risk, yet the mechanisms by which these diets mediate their ben-
eficial health effects are a subject of vigorous debate (15), although they are
certainly multifactorial in nature. For example, numerous fruits and vegeta-
bles are low in calories and fat, high in fiber, and often have a favorable
sodium=potassium ratio. Similarly, in addition to well-recognized essential
vitamins and minerals, such as vitamins A, C, K, folate and magnesium, plant
foods can contain a diverse array of bioactive phytochemicals that can influ-
ence the integrity and functioning of the vascular system in a positive manner.
One specific class of phytochemicals, the flavonoids, has generated con-
siderable interest as numerous epidemiological studies suggest an inverse
association between the intake of flavonoid-rich foods and the risk of CVD
Flavanol and Procyanidin’s Influence on Vascular Disease 291
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(16–18). Flavonoids can be found in high concentrations in diverse fruits,
vegetables, nuts, herbs, spices, and seeds (19). Flavonoids share a common
three-ring structure further defined by the presence or not of a ketone group
FIGURE 1 Flavonoid subclasses and associated foods. Flavonoids are divided into 6
subclasses (1A, B), which can be further subdivided (not pictured). For example, flavanols
can be further subdivided into flavans, flavan-3-ols (1B), flavan-4-ols, and flavan-3,4-diols.
The oligomers of flavanol monomeric units are known as the proanthocyanidins. The
proanthocyanidin subclasses and their monomeric subunits (in parentheses) are listed (1B)
as well as a sampling of foods contained in each flavonoid class (21, 25).
292 R. R. Holt et al.
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in the C ring, a double bond between carbons 2 and 3, or a hydroxyl group at
carbon 3 (19). There are six basic subclasses of flavonoids (Figure 1A):
flavones, anthocyanins, flavanones, flavonols, isoflavones, and the flavanols,
including the flavanol oligomers, the proanthocyanidins that are further sub-
divided into 16 species (Figure 1B), including the procyanidins, oligomers of
the flavan-3-ols catechin and epicatechin, and the prodelphinidins, oligomers
of the gallocatechins (20, 21). (See also the article by Corcoran, McKay, and
Blumberg in this same issue, wherein basic flavonoid chemistry is described
in greater detail.) The interpretation of data from epidemiological as well as
dietary intervention studies on the influence of flavonoids on cardiovascular
health can be complicated by the fact that often seemingly subtle differences
in chemical structures that exist across the flavonoid subclasses can translate
into marked differences in their absorption, metabolism, and bioactivity (22,
23). Illustrative of this, it has been reported that healthy human subjects show
pronounced differences in their ability to absorb the four catechin stereoi-
somers (23, 24) and there is evidence the isomers can have different biological
activities. This becomes important given the recognition that there are sub-
stantial variations in flavonoid profiles among specific plant foods; while
any specific food can include a number of the flavonoid classes, typically
one class predominates. Examples of foods for specific flavonoid classes
are depicted in Figure 1 (25). Further complicating the interpretation of the
vast majority of diet studies is the fact that the amounts of certain flavonoids
within a food can vary greatly depending on varietal, agricultural practices,
storage and processing, and cooking techniques. To work toward a goal of
making dietary recommendations for flavonoids and cardiovascular health,
it is important to assess the results from controlled dietary intervention trials,
at different life stages, using physiologically significant outcomes, and
well-characterized and standardized foods with appropriate controls (26,
27). Particular diseases of concern for the elderly include the CVDs such as
coronary artery disease (CAD), hypertension, and type II diabetes, as well
as, cancer, osteoporosis, and neurological dysfunction. To date, dietary inter-
vention trials suggest that increasing flavanols and procyanidins in the diet
will improve outcomes for a number of the previously mentioned age-related
diseases=disorders. In the following discussion we will focus our attention on
a review of epidemiological evidence and findings from dietary intervention
trials that have evaluated the effects of this subclass of flavonoids on cardio-
vascular health.
FLAVANOLS IN THE DIET AND CARDIOVASCULAR RISK
Examples of potentially rich dietary sources of flavanols and proanthocyani-
dins (oligomers of flavanols) include cocoa products (920–1220 mg=100 g),
raw beans (350–550 mg=200 g), apricots (100–250 mg=200 g), green tea
Flavanol and Procyanidin’s Influence on Vascular Disease 293
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(100–800 mg=L), black tea (60–500 mg=L), blackberries (130 mg=100 g), and
red wine (80–300 mg=100 mL) (28). To date, accurate estimates of typical
dietary intakes of flavanols have been largely hindered by the often poor pre-
cision of dietary intake records and food frequency questionnaires as well as
by the limited information available in many of the early food compositional
databases with respect to the flavonoid content of specific foods. Illustrative
of this, an early key paper that suggested the intake of high flavonoid diets
was associated with a reduced risk for vascular disease (The Zutphen Elderly
Study) was based on a limited analysis of 40 foods, comprising of two sub-
classes of flavonoids (flavonols and flavones) (29). Results presented in this
paper suggested that typical flavonoid intakes in a healthy adult population
were less than 50 mg=day. Recent studies in both the United States and Europe
that have utilized more extensive flavonoid nutrient databases available from
sources such as the U.S. Department of Agriculture (USDA) (30) and the
Phenol Explorer (25) indicate that daily flavanol and proanthocyanidin
intakes on the order of 40–214 mg=day and 235–455 mg=day, respectively,
may be more typical of the general population (16, 31, 32).
Within the past few years there has been increasing support for the idea
that it is important to distinguish between the dietary intake of flavanol mono-
mers versus the intake of long chain flavanol oligomers, This need is driven by
the fact that while the absorption of the monomers is quite high, only small
amounts of the oligomers are absorbed (33). While at one point it was gener-
ally accepted, based on in vitro experiments, that the oligomers were largely
degraded in the intestinal tract into monomers that were then absorbed,
recent data suggest this is not the case (34). In addition to this, there is
accumulating evidence that the flavanol monomers and their oligomers can
have different biological effects with respect to the gastrointestinal microbiota
and mucosal immunity (35–39). Reflecting the previously mentioned
advances in our understanding of the different nutritional impacts of flavanol
monomers versus their oligomers, some recent food composition tables
provide data for the separate categories.
As commented on previously, differences in agricultural practices and
food processing and cooking can have marked effects on the flavanol content
of certain foods. For example, with cacao, ()-epicatechin is the primary
flavonoid (40); however, flavanol and procyanidin content among cacao
varieties can be highly variable (41), with fermentation and drying practices
adding to the variability of the final epicatechin content (41–43). Similar issues
occur with tea and wine (44). In addition, epimerization of the flavanols can
occur during cocoa alkalization and heating processes (45, 46). While it has
been argued that epimerization of flavanols may occur in the intestinal tract,
recent feeding studies with the four pure stereoisomers suggest this does not
occur to any significant extent (23, 24). The importance of this is underscored
by the observation that the absorption of ()-epicatechin is much higher than
that of ()-catechin. Importantly, the determination whether epimerization has
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occurred within a food product requires advanced analytic techniques such as
chiral chromatography (45). Standard high performance liquid chromato-
graphy protocols do not detect whether epimerization has taken place, thus
there can be a substantial under- or overestimating of the total ()-epicatechin
and (þ)-catechin content of many foods that are listed in current databases. A
recently published Association of Analytical Communities (AOAC) method
should help facilitate routine laboratory analysis of flavanol stereoisomers (47).
Evidence in support of the concept that a chronic high intake of dietary
flavanols and procyanidins is beneficial with respect to vascular health is pro-
vided by data from the indigenous peoples of the San Blas islands, the Kuna
Indians, a population that has been reported to be characterized by very low
incidences of age-related hypertension and mortality from cardiovascular dis-
ease (48, 49). It has been reported that CVD mortality of the Kuna who live on
the San Blas islands is considerably lower that of the other Pan-American
populations (9 vs. 83 age-adjusted deaths per 100,000). The determinants of
this effect seem to be predominantly environmental rather than genetic, given
that this protection is lost on migration of Kuna Indians to Panama City (48). It
has been reported that the typical Kuna diet includes 3 to 4 cups a day of a
cocoa beverage (8–10 ounces per cup) made from locally grown cacao that
is particularly high in flavanols and procyanidins (50). If one uses an estimate
of 30 g of cocoa powder per cup (51, 52), 3 to 4 cups of cocoa beverage repre-
sents an intake of 1764–2352 mg=day of flavanols and procyanidins (50, 53).
Supporting the idea that the low incidence of hypertension of the islanders is
due in part to their high-flavanol diets is the observation that when Kuna
move to the mainland, they typically develop age-related hypertension (49,
54), which cannot be explained from an increase in added salt in the diet
as this intake, along with added sugars, is reduced compared to the islander
diet. However, those that migrate to the mainland have been reported to
adopt diets that are much lower in cocoa intake as well as in fruit and fish con-
sumption (49). Mechanistically, the intake of flavanol and procyanidin-rich
cocoa has been shown to increase circulating metabolites of the vasodilator
nitric oxide (NO) (see later) (27). That the high cocoa consumption of the
Kuna living on the islands contributes to the low incidence of hypertension
observed in the island dwellers is further supported by the observation that
urinary nitrite and nitrate excretion, potential markers of NO, are higher in
Kuna living on the islands than those living on the mainland (52).
A number of large prospective cohort studies provide additional
evidence for a primary protective effect of flavanols and procyanidins. In
the Zutphen Elderly Study conducted in Holland, elderly men were followed
for over 15 years. In the initial five-year follow-up period, a significant inverse
association between CAD mortality and the intake of specific flavonoid-rich
foods was detected (29). As stated, these initial findings were based on an
analysis of just 40 foods primarily in the flavonoid subclasses of flavonols
and flavones (29) with the flavonol quercetin representing 63%of the total
Flavanol and Procyanidin’s Influence on Vascular Disease 295
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flavonol intake. Tea consumption, which is high in flavanols, was inversely
associated with CAD mortality with an age, diet, and cardiovascular risk factor
adjusted risk ratio (RR) of 0.45 at the highest levels of intake. That specific
flavanols in the tea, such as catechin, may contribute to these epidemiological
findings was suggested in a follow-up analysis of the data (55). Recent studies
that have incorporated the flavanol and proanthocyanidin databases from the
USDA and Phenol explorer into the dietary analyses have helped to refine esti-
mates of the intakes of dietary flavanols and proanthocyanidins that may provide
positive vascular benefits. Median proanthocyanidin intakes starting at 175 mg=
day, but not flavanol intake, were inversely associated with CAD mortality in the
34,489 postmenopausal women of the Iowa Women’s Health Study (IWHS) (18).
It is important to note that within any analysis, the definition of what constitutes a
flavanol- or proanthocyanidin-rich food will influence the results. In the IWHS,
apples, red wine, and green and black teas were defined as flavanol-rich,
whereas apples, chocolate, and seeded grapes were classified as
proanthocyanidin-rich (18). This approach is consistent with respect to how
others have analyzed the health effects of these foods; however, given that some
of these foods can contain substantial amounts of both flavanols and proantho-
cyanidins, their contributions to both nutrient pools should be considered. This
strategy was employed in the Cancer Prevention Study II Nutrition Cohort of
38,180 men and 60,289 women (17), which defined flavanol-rich foods as apples,
black tea, blueberries, chocolate, and red wine and kept these same foods as
proanthocyanidin-rich but added mixed nuts, peanuts, strawberries, and
walnuts. As a result, although an 18%lower risk of CVD mortality in the highest
quintiles of total flavonoid intake was observed compared to the lowest quintiles
of intake (17), possible sex differences were detected for specific flavonoid sub-
classes. A multivariate-adjusted median flavanol intake of at least 11.8 mg=day
was associated with reduced CVD mortality in women but not in men. For the
proanthocyanidins, an intake of 132 mg=day was associated with protection in
womencomparedto379mg=day in men (17). Apart from CVD mortality,
flavanol and proanthocyanidin intakes arealsoassociatedwithreductionsin
select CVD risk factors. In the Nurses’ Health Study I and II and the Health
Professional Follow-up Study a pooled RR of 0.92 for incident hypertension
was observed with anthocyanin intake, but not for flavanols or flavanol oligo-
mers, which included proanthocyanidins (16). However, an association between
reducedhypertensionincidenceandthehighest and lowest quintiles of catechin
and epicatechin intake was observed for those 60 years of age and younger. The
Kupio Ischaemic Heart Disease Risk Factor Study first demonstrated a relation-
ship between carotid artery intima media thickness (CA-IMT) and the risk for
future coronary heart events; with RR increasing 15%per 0.1 mm increase in
CA-IMT (56). In the nutrient analysis of this cross sectional study flavanol
consumption was inversely associatedwithCA-IMTthickness(57).
Evidence that specific flavanol- and proanthocyanidin-rich foods, such as
cocoa and chocolate, may drive some epidemiological findings is provided
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from several recent prospective studies. Buijsse and colleagues (58) reported an
inverse relationship between chocolate consumptionand CVD risk (myocardial
infarction (MI), stroke, 8 years follow-up) in the Potsdam Arm of the European
Prospective Investigation into Cancer and Nutrition. In this large cohort
(n ¼19,357) of both sexes, the authors observed that in the lowest quartile of
chocolate consumption (1.7 g=day) 106 MI and strokes occurred, whereas 61
events occurred (relative risk (RR) 0.61) in the quartile with the highest
chocolate consumption (7.5 g=day). In the latter group, both systolic and dias-
tolic blood pressures were significantly lower (1 mm Hg) than the referent
low-chocolate quartile. Baseline blood pressure explained 10%–12%of the risk
reduction. Counterintuitive to the idea that a high vegetable intake is beneficial,
the subgroup with the lowest risk was the group with the lowest vegetable
intake; however, this group also had the highest chocolate intake (58). Similar
protection has been observed with cocoa products, including chocolate, in eld-
erly men of the Zutphen Study (12). In this study, a 3.7 mmHg reduction of sys-
tolic and 2.1 mmHg reduction of diastolic blood pressures were observed in
individuals with the highest tertile of cocoa intake (>2.3 g=day) compared to
the lowest tertile of intake (<0.36 g=day), with a 47%and 50%reduction,
respectively, in all-cause and cardiovascular mortality (12). While this dataset
is suggestive for both the primary and secondary prevention of CVD, the use
of food frequency questionnaires or dietary recalls may severely limit the pre-
cision of the estimates of intake for specific flavanols and proanthocyanidins. In
addition, with foods such as cocoa and chocolate, the content of bioactive fla-
vanols and proanthocyanidins within any individual product varies markedly,
depending on food manufacturing processes and recipe formulations.
Finally, these studies are limited with respect to proving cause and effect
relationships, or in giving mechanistic insights as the observed associations
may be due to strong confounders. An important potential confounder in this
context is reporting bias. For example, individuals may under report certain
foods, such as chocolate, that are perceived to be unhealthy. Assuming, for
the sake of argument, that the flavanols provided through the consumption
of cocoa and chocolate containing products are indeed contributing to vascu-
lar health, a potential consequence of underreporting is that a given food pro-
duct, such as chocolate, may appear to have stronger potential cardiovascular
benefits, with respect to the amount of the food that is needed to provide
these effects, than what really is needed. Illustrative of this potential problem,
in the IWHS the consumption of chocolate at a seemingly low level of once a
week was associated with marked reductions in the risk for vascular disease
(18). If this is not considered when designing dietary intervention trials, a false
negative may result if the intake level chosen is too low. Overall, while the
data from epidemiological studies can be directional, randomized controlled
dietary intervention studies using well-characterized food products and
suitable controls are needed to causally link high intakes of flavanol and
proanthocyanidin-rich foods with prevention of CVD.
Flavanol and Procyanidin’s Influence on Vascular Disease 297
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INFLUENCE OF FLAVANOLS ON VASCULAR HEALTH
Vascular homeostasis is maintained through multiple complex interactions
between the endothelium and the underlying smooth muscle cells and con-
nective tissues of the vessel wall. A number of mediators produced from
the endothelium are known to regulate vascular tone, including vasodilators
such as NO, prostacyclin, and endothelium-derived hyperpolarizing factor,
and vasoconstrictors such as endothelin-1. At the same time these mediators,
especially NO, can have marked effects on vascular smooth muscle cell remo-
deling, platelet activation, and inflammation. It is known that cardiovascular
risk factors can modulate vascular homeostasis through the perturbation of
endothelium-derived mediators, as well as through the remodeling of the vas-
culature. Importantly, there is substantial crosstalk between the endothelium
and the vascular smooth muscle that can be affected by known cardiovascular
risk factors as well as lifestyle choices such as diet.
Advancing age is an independent major risk factor for CVD. Substantial
changes in systemic hemodynamics occur with advancing age because
aortic and large central artery stiffness occurs in addition to traditional cardio-
vascular risk factors (59). This increase in arterial stiffness is associated with
elevated pulse pressures (i.e., difference between systolic and diastolic press-
ure) and pulse wave velocity that lead to an increase in mechanical forces on
the microvasculature and increase in central blood pressure. Coupled with
age-related increases in endothelial dysfunction of the peripheral arteries,
the increased mechanical force of the pulse wave promotes structure and
function abnormalities of the microvasculature, particularly in sensitive
organs, for example, the kidney and brain (59, 60), and explains, in part, a
number of pathological states, such as renal dysfunction, hypertension,
stroke, and cognitive disorders often observed with age.
To study the effects of flavanol and procyanidin intake on cardiovascular
health, a number of noninvasive surrogate outcome measures that are affec-
ted by cardiovascular risk factors, including the vascular aging process,
have been employed. These surrogate endpoints are thought to reflect key
pathophysiologically relevant entities implied in CVD development and
progression. These endpoints include vascular function, blood pressure,
blood lipids, glucose tolerance, platelet reactivity, inflammatory markers,
and circulating progenitor cells. Several measures of vascular function have
been employed in dietary intervention studies, each of which measure
distinctly different physiological parameters. Flow-mediated dilation (FMD)
utilizes ultrasound to assess the diameter of large conduit arteries, such as
the brachial artery, as a measure of endothelium-dependent vasodilation after
reactive hyperemia induced shear stress (61). Peripheral arterial tonometry
(PAT) measures digital blood volume changes from reactive hyperemia-
induced shear stress and is considered a measure of microvascular reactivity
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(62). Finally, pulse wave velocity (PWV) is an assessment of arterial stiffness
(60). Although these vascular measures have their distinct differences, studies
have shown either an association or similar responses between FMD and PAT
(63–66), and FMD and PAT to arterial stiffness (67). In addition, a number of
studies show an association of FMD, PAT, or PWV response to cardiovascular
risk factors (62, 65, 66, 68–73). It is important to note that all of these techni-
ques have their limitations as well as the need for standardization of proce-
dures in order to properly interpret results across studies (74). For example,
it is known that baseline brachial artery diameter influences the shear stress
that is developed from reactive hyperemia, and ultimately, the amount of
dilation of the brachial artery. Therefore, in order to gauge potential mechan-
isms of action, it is important that for FMD studies brachial artery diameter and
sheer stress are reported. Moreover, manual single frame inspection of vessel
diameters can lead to significant operator error. To increase precision, the use
of edge detection and wall tracking software is currently recommended (74).
While it is appreciated that randomized controlled clinical intervention
studies are the gold standard for generating the type of evidence that is
needed to causally link the intake of specific dietary factors, such as flavanols,
with a reduced risk for diseases such as CVD (27), the complex matrix of foods
makes the proper design of these studies difficult. Key to success is the use of
food products that are well characterized for micro- and macronutrient, and
phytochemical content. Not only does this help define the level of bioactive
needed for physiological effect but importantly it lends to the design of con-
trols that are compositionally matched for macronutrient content and other
potential bioactive compounds. This is critical for flavanol-rich foods such
as cocoa, tea, and red wine, which can be high in bioactive components such
as theobromine, caffeine, and alcohol, respectively. While this may be more
readily achieved with beverages such as cocoa and tea, determining the
proper control for whole foods, such as fruits and nuts, can be difficult, and
make blinding prohibitive, unless the food is substantially changed from its
natural state, such as through the use of extracts or freeze-dried powders.
Influence of Flavanol Intake on Vascular Function
Several dietary intervention trials have tested the effects of flavanol-rich foods
on vascular function including cocoa, chocolate, red wine, and tea. Investiga-
tions on the effects of red wine intake on vascular reactivity have typically
been complicated by the lack of standardization and characterization of the
wines, and the vascular effects of alcohol itself. For example, it is apparent
in studies using alcoholic beverages, such as red wine, vodka, or ethanol
alone, that a significant increase in baseline brachial artery diameter (prior
to reactive hyperemia) occurred, resulting in a reduced FMD. This is in con-
trast to the intake of dealcoholized red wine, which does not induce changes
Flavanol and Procyanidin’s Influence on Vascular Disease 299
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in baseline brachial artery response, and does increase FMD (75, 76). That the
phytochemicals in red wine affect vascular reactivity is further suggested by
improvements in FMD observed after acute (hours) or short-term (weeks)
intake of dealcoholized red wine by smokers (77) and CAD patients.
Significant increases in FMD have been consistently shown both after
acute or short-term intakes of either black or green tea (78–82). In healthy
individuals, a beverage made from brewing 6 g of green tea increased FMD
by 3.7%30 minutes after consumption. The control drink containing 125 mg
of caffeine in water did not significantly change FMD; however, the sublingual
nitroglycerin control performed 120 minutes after tea consumption increased
brachial artery dilation, indicating improvements in both endothelial-
dependent and independent relaxation after tea intake (78). An increase in
both endothelial-dependent and independent relaxation response was also
observed after short-term intake of black tea in individuals with mild hyperch-
olesterolemia (81). In CAD patients, both acute (2 hour) and short-term (4
weeks) consumption of black tea (450 mL and 900 mL, respectively) improved
FMD by about 3.5%versus baseline and water controls (80). Interestingly,
dietary flavonoid intake, and not traditional risk factors, was an independent
predictor of baseline FMD (80). In addition, studies have shown a dose-
dependent response of tea flavanol intake, with FMD response increasing
from baseline with 100–400 mg per day of total flavanols for 1 week, and with
800 mg providing further enhancements of the FMD response (83).
Impressively, in a recent survey of 25 studies, all but two (84, 85) reported
significantly improved vascular function after acute or short-term cocoa or choc-
olate intake in healthy individuals (52, 86–97), those who smoke (98–100), are
diabetic (101), with CAD (102–104) or heart failure (105), and in the elderly
(106, 107). The reported improvements in FMD ranged from 1.4%to 6%at
1–2 hours after a single intake of cocoa or chocolate (52, 87, 93, 98, 100, 102,
107). Similar to tea, the effects observed were dose dependent and are associa-
ted with plasma levels of flavanol metabolites (107). Specifically, the plasma
concentrations of both epicatechin and its metabolite epicatechin-7-O-glucuro-
nide have been reported to predict the magnitude of the FMD response (52). In
addition, acute intakes of flavanol-rich cocoa or chocolate have been reported to
enhancePATresponseby68%(52), significantly reduce measures of arterial
stiffness (87), and increase coronary artery diameter after a cold pressor test
by 4.3%in heart transplant patients (103). The magnitude of the acute response
was similar to that of short-term intake. Short-term intake of flavanol- and
procyanidin-rich cocoa for 8 days to 3 months (ranging from 450 mg to
960 mg per day) have been reported to increase endothelium-dependent FMD
responses, with significant improvements in vascular reactivity from baseline
exhibited after three days of intake that were sustained throughout the trial
(99, 101). Either a decrease or no change in endothelium-independent dilation
was reported in the few studies that have measured this response post intake
(87, 101, 105).
300 R. R. Holt et al.
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Currently there are a limited number of studies that have examined the
vascular effects of flavanol intake in older populations (>60 years; Table 1).
In this population, cocoa intake increases the response as measured both by
FMD and pulsatile digital blood volume; however, potentially different
mechanisms are suggested between healthy younger adults and the elderly,
as high-flavanol cocoa intake produced a more robust blood volume
response in the elderly that was only partially attenuated with nitric oxide
synthase (NOS) inhibition (106). Recently, Monahan and colleagues (107),
reported dose-dependent increases in FMD response at ranges of epicatechin
intakes lower than previously found in other populations (6.3–96 mg); how-
ever, it is important to note that theobromine levels in this study were not
controlled. As theobromine can potentially act as a vasodilator (108), further
studies will need to be undertaken to determine whether the response to fla-
vanol intake is different in healthy young versus elderly populations, or if
there is a synergistic effect between epicatechin and theobromine. With that
said, a large portion of the vascular studies to date are double-blind con-
trolled studies using a low-flavanol cocoa, suggesting that the positive vascu-
lar effects are primarily due to the intake of these specific flavonoids from the
cocoa. Consistent with this thinking, a single dose of 1–2 mg per kg of epica-
techin significantly increased both FMD and PAT as soon as 1 hour after
intake, providing direct evidence that dietary intake of flavanols can improve
vascular reactivity on an acute basis (52).
Complementary to these studies on vascular reactivity, short-term choc-
olate or cocoa consumption has been shown to decrease systolic (SBP) and
diastolic blood pressure (DBP). For the most part, a single dose of cocoa or
chocolate providing less than 100 mg of epicatechin has shown no effect on
blood pressure (86, 87, 103, 109); however, chocolate or sugar-free cocoa
containing 800 mg of total flavanols and procyanidins and 10–22 mg of epica-
techin significantly reduced blood pressure compared to their respective con-
trols 2 hours after intake (93). It is known that impaired vascular function is
associated with exaggerated exercise-induced increases in blood pressure.
A reduced exercise induced blood pressure response in overweight and
obese individuals was observed 2 hours after the intake of cocoa containing
700 mg of flavanols and procyanidins (95). Otherwise, most studies observing
positive blood pressure effects utilize a cocoa and chocolate intake period of
2–18 weeks (92, 94, 96, 104, 110–112). These studies have reported changes in
either the office setting or with ambulatory (24 hour) monitoring ranging from
2–12 mmHg for SBP and 2–9 mmHg for DBP. It is important to note the dis-
tinct differences in study design and products tested. Three studies were
double-blind controlled, using a corresponding control cocoa that was low
in flavanols and procyanidins but equal in theobromine content (92, 104,
110). In these studies the effective range of epicatechin was 36–200 mg per
day. In the remaining studies, open-control designs were used, providing a
control, typically white chocolate, which was similar in macronutrient content
Flavanol and Procyanidin’s Influence on Vascular Disease 301
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TABLE 1 Randomized, Controlled Trials in Older Subject Populations (>60 Years Old)
Intervention Control
Total flavanol
and procyanidins=
epicatechin=
catechin (mg)
content in
intervention
Theobromine
controlled?
Study
design
Subject
population
Mean
age
(Years)
N
(%Male) Results
Studies With Intervention Product Containing <50 mg Epicatechin
Taubert, D
2007
111
Dark
chocolate
White
chocolate
(control)
30=5.1=1.7 No Short-term intake
(18 weeks);
Open-label
parallel-arm
Hypertensive
(stage 1) or
prehypertensive
adults
63 44 (45) #SBP, DBP, and
"S-nitrosoglu-
tathione after
12 and 18
weeks of
intake
Monagas, M
2009
187
Cocoa þmilk Milk 425.7=46.08=10.41 No Short-term intake
(2 weeks);
Open-label
crossover
Diabetics or
3þcardiovascu-
rdiovascular risk
factors
69.7 42 (45) No change in BP
Desch, S
2010
110
Dark
chocolate
6 g or 25 g per
day
None 6 g: ND=5=ND
25 g: ND=21=ND
No Short-term intake
(3 months);
Single blind,
parallel-arm
Hypertensive
(stage 1) or
prehypertensive
adults; and has
CAD, PAD or
diabetes
66 71 (70) #24 h Mean
ambulatory BP
and SBP after 3
months of
either 6 g or
25 g of
chocolate per
day
#24 h DBP after 3
months of 25 g
of chocolate
per day
Mellor, DD
2010
188
High flavanol
chocolate
Low flavanol
chocolate
<2mg
epicatechin
ND=17=ND Unknown Short-term intake
(8 weeks);
Double blind,
crossover
Diabetic patients 68 12 (58) No change BP
and CRP
302
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Studies With Intervention Product Containing >50 mg Epicatechin
Farouque,
HMO
2006
83
High flavanol
chocolate
and cocoa
Low flavanol
chocolate
and cocoa
High flavanol
products:
444=107=ND
Low flavanol
products:
19.6=4.7=ND
Yes Acute and
short-term
intake
(6 weeks);
Double blind,
parallel-arm
CAD patients 61 40 (75) No change in
FMD,
acetylcholine
stimulated
forearm blood
flow, or arterial
compliance
No change in BP
Balzer, J
2008
100
HFC LFC
17 mg
epicatechin
4 mg catechin
371=78.9=19.7
963=203=50.8
Yes Acute intake
double blind,
parallel-arm
Diabetic patients 64.7 10 (80) Dose dependent
"FMD
Balzer, J
2008
100
HFC LFC (control)
17 mg
epicatechin
4 mg catechin
963=203=50.8 Yes Acute and
short-term
intake (30
days);
Double blind,
parallel-arm
Diabetic patients 64 41 (29) "FMD 2 h after
intake at 1, 8
and 30 days of
intake
"Baseline FMD 8
and 30 days
after HFC
intake
No change in
MAP or CRP
Monahan, KD
2011
106
HFC
Dose
response
LFC
0mg
epicatechin
0 mg catechin
2 g: 420=6.3=3.0
5 g: 420=17.7=8.1
13 g: 840=45=21.6
26 g: 1470=96=48
No Acute intake
double blind,
crossover with
or without
exercise
Healthy adults 63 23 (39) "FMD (5, 13,
26 g).
"SBP (2 and 26 g)
"DBP (0, 2, 13,
26 g)
"MAP (2, 13,
26 g)
Note. Only randomized placebo-controlled studies using a food product characterized for at least epicatechin content are presented. Studies organized by those
providing less or greater than 50 mg of epicatechin in the active treatment.
Not determined, ND; High Flavanol Cocoa, HFC; Low Flavanol Cocoa, LFC; Flow-mediated Dilation, FMD; Nitric Oxide, NO; L-N
G
-monomethyl-arginine (L-NMMA)
used for inhibition of NO synthase; Blood Pressure, BP; Systolic BP, SBP; Diastolic BP, DBP; Mean Arterial Pressure, MAP; Heart Rate, HR; C-reactive Protein, CRP;
Hours, h; Coronary Artery Disease, CAD; Peripheral Arterial Disease, PAD.
303
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to the test chocolate except for the amount of polyphenols, and included
theobromine. For the most part these studies did not measure the epicatechin
content of the test product, but one study provided 111 mg of epicatechin and
two studies as low as 5 mg per day (96, 111, 112). The chocolates providing
5 mg of epicatechin a day significantly lowered both SBP and DBP after 3
months (111, 112). This is in contrast to multiple studies that have provided
similar levels of epicatechin in a low flavanol and procyanidin cocoa (with
theobromine levels also controlled), where marked effects on blood pressure
or FMD have not been observed (85, 104, 111).
Table 1 presents the flavanol and procyanidin content, experimental
design, and food product control of studies in older healthy adults or those
with CVD. The studies are divided by epicatechin content that is greater or
lesser than 50 mg as recent meta-analyses have described an association of
epicatechin intake to blood pressure (113, 114), with greater effects observed
at intake levels greater than 50 mg (113), and a predicted reduction of SBP of
4.1 mmHg and 2.0 mmHg for DBP with epicatechin intakes of 25 mg using a
nonlinear regression model of randomized controlled trials (114). It is also
important to note that changes in blood pressure at lower epicatechin intakes
may be due in part to the use of an open-label study design rather than
through a vasoactive effect of the flavanols or theobromine, as evidenced
from a recent study in older adults (mean age 62 years) with pre- or stage
1 hypertension that were given cocoa with either natural or triple the amount
of theobromine for three weeks. After three weeks, no change in peripheral
blood pressure was observed (taken 2 hours after cocoa intake), while
24-hour ambulatory blood pressure increased (115), suggestive that theobro-
mine does not play a role in the vascular response observed with cocoa or
chocolate intake. Until studies are performed using control chocolates that
are suitable for blinding, and solely deficient in flavanols and procyanidins,
the putative difference in blood pressure response between cocoa and choc-
olate will remain unresolved.
The potential for flavanol- and procyanidin-rich foods to influence
blood pressure and improve vascular function is significant as epidemiologi-
cal evidence suggests that mid-life hypertension as well as other cardiovascu-
lar risk factors is associated with impaired cognitive function, Alzheimer
disease, and dementia (116, 117). In addition, measures of vascular health
including CA-IMT and arterial stiffness have also been associated with cogni-
tive impairment independent of other cardiovascular risk factors (116). That
certain diets, such as those rich in fruits and vegetables, can reduce the risk
for cognitive disorders is suggested by epidemiological evidence, but, to
date, the epidemiological findings need to be confirmed with dietary inter-
vention trials (116). Short-term intake (1–2 weeks) of 900 mg of flavanols
and procyanidins has been reported to increase cerebral blood flow and
blood flow velocity of the middle cerebral artery in healthy elderly subjects
(106, 118). Whether this increase in blood flow translates to improved
304 R. R. Holt et al.
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neurological function is still a matter of debate. A recent large prospective
study observed a negative association between catechin and proanthocyani-
din intake in midlife on executive function 13 years later (119). In dietary
intervention trials, a single study in the elderly showed no cognitive improve-
ment with 6 weeks of approximately 750 mg of flavanol and procyanidin
intake (120), while 30 days of chocolate containing 250 or 500 mg of flava-
nols and procyanidins measured significant differences in the activation of
steady state evoked potentials that suggest an improvement in spatial work-
ing memory in middle-age adults (121). Moreover, three studies have
reported both acute and short-term improvements in cognitive measures in
healthy young adults (122–124).
Flavanol Intake and Platelet Reactivity
Platelet hyper-reactivity is associated with cardiovascular risk factors and
reduced survival in those with a previous myocardial infarct (125, 126).
Aspirin and additional antiplatelet strategies have been employed for both pri-
mary and secondary prevention of acute events of occlusive arterial disease
(127–130). It has been shown that a number of foods can reduce platelet reac-
tivity including berries (131), grapes (132–134), and cocoa products (135).
Curiously, in a study initially designed to examine the genetic components
of varied aspirin responses GeneSTAR (Genetic Study of Aspirin Responsive-
ness) a reduction in platelet reactivity, measured as prolonged epinephrine
and collagen (EPI-Col) PFA-100 closure time and reduced urinary thrombox-
ane excretion was observed in individuals who declared chocolate consump-
tion in a dietary recall for the 24-hours prior to testing (136). Dietary
intervention trials confirm the potential for flavanol-rich cocoa and chocolate
consumption to produce positive effects on platelet reactivity using a number
of different outcome measures. A reduction in both adenosine diphosphate
(ADP)- and EPI-Col PFA-100 closure times was observed 2 and 6 hours after
intake of 897 mg of flavanols and procyanidins from cocoa. In addition,
reduced expression of platelet activation markers (glycoprotein IIb=IIIA and
P-Selectin) after stimulation with both epinephrine and ADP has been
reported. Murphy and collaborators extended these initial observations by
looking at 2 weeks of 280 mg of flavanols and procyanidin supplementation
(137). Twenty-eight days of flavanol and procyanidin intake decreased both
ADP- and collagen-induced platelet whole blood platelet aggregation and
secondary ATP release after collagen activation. P-Selectin expression was
also reduced (137). Chocolate consumption has been reported to reduce pla-
telet reactivity. In healthy subjects, chocolate consumption was observed to
reduce collagen-induced platelet aggregation (138), and shear-stress–initiated
platelet adhesion was attenuated 2 hours after chocolate consumption in both
healthy subjects and heart transplant patients (103, 139). Activated platelets
can shed membrane particles of less than a micrometer in diameter termed
Flavanol and Procyanidin’s Influence on Vascular Disease 305
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microparticles. Microparticles are present in healthy individuals but are
known to be increased in a number of chronic diseases including CVD
(140). A reduction in ADP-stimulated microparticle formation was observed
6 hours after cocoa intake (51). Currently, the body of data from acute and
short-term dietary intervention trials looking at the effects of flavanol- and
procyanidin-rich foods on platelet function suggests that the most consistent
positive effects occur after the consumption of cocoa and chocolate. This is in
agreement with a recent critical review of 25 controlled intervention trials that
considered the potential benefits of numerous dietary polyphenols (141).
POTENTIAL MECHANISMS
NO Availability
In general, the dietary intervention trials discussed support the epidemiologi-
cal observations that diets rich in flavanols and procyanidins reduce an indivi-
dual’s risk for CVD; how these nutrients might work at a mechanistic level is an
area of intense investigation. Evidence strongly suggests that the consumption
of flavanol-rich foods can facilitate NO- and endothelium-dependent relax-
ation of arteries. The infusion of the NOS inhibitor L-N
G
-monomethyl-arginine
(L-NMMA) into healthy adults and young adult smokers prior to flavanol- and
procyanidin-rich cocoa consumption inhibited the FMD response in the bra-
chial artery (52, 98). Caution in interpretation of these findings is warranted
as these results do not exclude other mechanisms that may indirectly enhance
FMD (74, 142). With that said, if baseline brachial artery diameters and shear
stress do not change between treatments and measurements, one could
reasonably deduce that the observed improvements in FMD response are in
part NO-mediated. Additional evidence from dietary intervention studies con-
firms a significant contribution of NO to the vascular response after flavanol
and procyanidin intake. As soon as an hour after the intake of flavanol- and
procyanidin-rich foods, the plasma levels of NO metabolites are significantly
increased (52, 98, 104, 143), and these increases correlate to plasma levels
of epicatechin metabolites and FMD response (52, 98, 143). The mechanism
behind these observations has been described as an enhancement of the cir-
culating NO pool (144). For the elderly, that flavanol intake increases NO avail-
ability would be beneficial, as it has been shown that older individuals exhibit
decreased endothelial function compared to younger adults (98, 145, 146) as
well as reduced NO availability during exercise (107, 146). While FMD
responses are improved in older subjects after cocoa intake (107), the exact
mechanism(s) have not yet been fully explored and studies in healthy younger
adults may not translate to older individuals, as shown recently by Wray and
colleagues, where FMD responses were reduced in the young, but improved
in older subjects, after antioxidant supplementation (145). NO is produced via
the L-arginine and NOS pathway, and with a half-life of approximately 1–2
306 R. R. Holt et al.
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milliseconds, will directly act on its biologic targets in close proximity to its
production or metabolized into nitrite, nitrate, S-nitrosothiols, and other spe-
cies that can have biological activities similar to NO itself (147). Therefore,
the circulating NO pool may include nitrate and nitrite, and a number of
‘‘nitrosylated’’ species of albumin, glutathione, and hemoglobin, all which
are considered as NO storage forms that can be utilized as needed (144). Using
these techniques it was shown that exercise-induced increases in plasma
nitrite are lower in older subjects compared to younger adults (146).
Interpretation of this data set can be complicated by the observation that
basal levels of plasma nitrite and urinary excretion of nitrate are increased 2
hours after the intake of 200 mg of pure epicatechin or quercetin (143). While
these data could suggest an increase in basal NOS activity, or improvements in
NO availability through attenuation of reactive oxygen species (ROS) produc-
ing systems, it is known that dietary nitrate is reduced to nitrite by oral and
gastrointestinal microflora, with nitrite subsequently reduced to NO in the sto-
mach potentially enhanced by dietary procyanidins (147–151). Importantly,
substantial amounts of dietary nitrate are continually secreted from the circu-
lation back into the saliva, and can persist in the plasma and enterosalivary
circulation with a half-life of 5–8 hours (147, 148), corresponding to peak
urinary excretion that itself does not return to basal excretion levels until 24
hours after a high-nitrate meal (152). Although often dismissed as an inter-
fering artifact of the diet (153), dietary nitrate and its interaction with flavo-
noids may represent an additional vasculoprotective mechanism of diets
rich in fruits and vegetables (148).
That circulating nitrite is increased after flavanol intake is particularly
important to consider as NO is continually formed from and recycled back
to nitrite and nitrate (147). Blood flow is regulated by the balance between
tissue oxygen demand and supply, in that as oxygen demand increases
and supply decreases, blood flow is increased. It has been hypothesized that
heme-group oxygen saturation, specifically on hemoglobin and myoglobin,
are key sensors for this response (147, 154). Moreover, as hemoglobin
becomes desaturated, such as during oxygen tissue exchange in the arteriole
and capillary beds, deoxyhemoglobin can act as a nitrite reductase to pro-
duce NO and increase blood flow in a NOS independent manner (155).
NO can subsequently be reduced back to nitrite. This process can be impor-
tant particularly during intense exercise and possibly during procedures such
as FMD and PAT, which use reactive hyperemia as a vascular stimulus. To
date the limited number of studies that have examined shear stress responses
after flavanol intake report no change in shear stress. Importantly, vascular
measures that do not employ reactive hyperemia as a vasodilatory stimulus,
such as PWV and pulsatile blood volume, are also increased after flavanol
intake (87, 90, 106), as these measures are also influenced by NO (156,
157). The data currently suggest that hypoxic vasodilation has a limited role
in the mechanism of improved FMD or PAT response after flavanol intake
Flavanol and Procyanidin’s Influence on Vascular Disease 307
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(104, 107); however, studies that are designed to specifically eliminate this
possibility are needed.
Circulating Angiogenic Cells
Recent data suggest that flavanol- and procyanidin-rich diets improve specific
aspects of endothelial repair. Repair of endothelial lesions is facilitated by
mobilization from the bone marrow of circulating angiogenic cells (CACs, pre-
viously termed endothelial progenitor cells) (158). Enumeration of these cells
is commonly performed via the positive identification of early hematopoietic
lineage markers CD34 and CD133, and the vascular endothelial growth factor
(VEGF) receptor-2 marker KDR by flow cytometry (159). A reduced number of
CD34
þ
=KDR
þ
cells is associated with reduced vascular function and increased
risk of cardiovascular events and mortality(160, 161). Importantly, the number
and functional capacity of these cells was inversely associated with age and
endothelial function (160, 162–165, 166). Two to four weeks of tea and cocoa
intake in smokers and CAD patients, respectively, increased both FMD
response and circulating CD34
þ
=KDR
þ
cells, with cocoa intake also increasing
circulating CD133
þ
=KDR
þ
cells (82, 104). Both studies looked at the functional
characteristics of CAC cells, produced from specific plating conditions that
measure endothelial marker expression and the proliferating characteristics
of adherent cells after seven days of culture (159). Only tea consumption by
smokers showed an increase in cell number (82); whereas cocoa consumption
in CAD patients produced functional cells but did not exhibit an increased pro-
liferating or chemotactic capability compared to CAC cultured from patients
after low flavanol and procyanidin intake (104). Nevertheless, both beverages
increased CAC, which are known to migrate from the bone marrow after
receiving pro-angiogenic signals from ischemic areas, such as stromal
cell-derived factor-1aand VEGF. Importantly, NO and NOS activityplay crucial
roles in the mobilization and function of these cells. It has been reported that
CD34
þ
=KDR
þ
and CD133
þ
=KDR
þ
cells are increased with nitrate treatment
(167). CAC and endothelial cell migration depends on NOS activity and che-
mokinesis is enhanced by NO donors, which act synergistically with VEGF
in assays of chemotaxis (168). Furthermore, reduced eNOS activity and chemo-
tactic ability was observed in CACs from CAD patients compared to healthy
controls (168). Whether the increase in CAC after flavanol and procyanidin
intake is due to changes in angiogenic signaling or possibly an increase in
NO availability or NOS activity is untested as the a potential in vivo flavanol
effect on CAC activity may likely be lost after seven days cell culture (104).
Platelet Reactivity
An increase in NO availability after flavanol and procyanidin intake may also
reduce platelet reactivity. Production of NO, nitroso species, and PGI
2
from
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endothelium-reduced platelet activation and adhesion through the elevation
of intracellular cAMP and cGMP that maintain intracellular Ca
2þ
low (169–
171). In addition to increasing NO, within hours of consuming flavanol- and
procyanidin-rich chocolate plasma PGI
2
was significantly increased (172,
173). Apart from the endothelium, platelets produce NO, which can act to
reduce platelet recruitment to the growing thrombus. As there is a known
association between reduced platelet NO production and increased risk for
acute coronary syndrome mortality (174), incorporation of foods into the diet
that improve platelet NO availability could potentially reduce cardiovascular
mortality. The consumption of grape juice for 14 days increased platelet NO
and reduced platelet superoxide ðO
2Þproduction in healthy volunteers
(134). Platelet activation is characterized by a large release of ROS including
O
2and hydrogen peroxide (H
2
O
2
) and is thought to be dependent on a
balance between NO, platelet redox state, and oxidative stress (174).
Specific platelet agonists such as collagen, thrombin, and arachidonic
acid (AA) activate platelet NADPH oxidase (NOX) to produce O
2, which sub-
sequently enhances platelet aggregation response to platelet ADP release
from dense granules (174–177). Catechin and quercetin have been reported
to synergistically reduce platelet aggregation and adhesion, in vitro, via a
reduction in NOX induced O
2production, and increased NO production
(178, 179). This finding was not observed with either flavonoid alone. In
addition, ()-epicatechin alone increased NO and reduced platelet ROS
and NOX activation in platelets from smokers, in vitro (100). ADP-Col closure
times were also prolonged in samples with NOX inhibitors or NO donors
(178), whether this is a potential mechanism for the reported prolonged
ADP-Col closure times after flavanol- and procyanidin-rich intake (51, 137,
173, 180–182) is currently unknown, as well as the significance of any of
the previously mentioned in vitro mechanisms to the ex vivo results of dietary
intervention trials. Furthermore, these studies were performed with PRP, thus,
it is unknown if these results are reproducible in whole blood aggregations
studies, which will provide some oxidant defense and NO from RBC (183).
Moreover, measures of ROS release, GPIIb=IIIa expression, and aggregation=
closure time are end events of activation. Little is known as to whether flava-
nol intake is preventing ROS production, increasing NO, or simply preventing
activation through platelet receptor interactions. As described, chocolate con-
sumption increased PFA-closure times in the GeneSTAR study. Not only were
closure times modified, but also urinary thromboxane was reduced (136). A
decrease in plasma thromboxane was also observed after black tea and wine
intake (184, 185). Aspirin inhibits cyclooxygenase (COX)-1 production of
thromboxane from AA (186). A variety of platelet agonists activate this lipid
signaling pathway, including collagen, to produce thromboxane that can sub-
sequently act on additional platelets to promote thrombus formation. A num-
ber of flavonoids, including catechin, reduced thromboxane production from
Flavanol and Procyanidin’s Influence on Vascular Disease 309
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collagen-stimulated platelets (i.e., receptor mediated), but not thromboxane
production from AA (i.e., nonreceptor mediated) activated platelets. Further-
more, catechin has been reported to attenuate thromboxane receptor ligand
binding, although at a very high concentration (160 mM) in washed platelets,
suggestive that flavanols may affect platelet reactivity, in part, through antag-
onism of the thromboxane receptor (187).
FUTURE DIRECTIONS AND TAKE AWAY POINTS
Data from short-term dietary interventional trials demonstrate the potential of
flavanol- and procyanidin-containing foods to modulate a number of surro-
gate markers of vascular homeostasis in healthy individuals as well as in those
with age-related diseases such as diabetes and CVD. In addition, the intake of
foods that are high in flavanols and procyanidins may be protective with
respect to cognitive decline later in life. While the current body of research
in the area of dietary flavanols and health is promising, data concerning the
effects of flavanol and procyanidin intake on accepted clinical endpoints of
cardiovascular health, specifically long-term intake data on cardiovascular
events, clinical symptoms, and death (27) are still lacking. The fact that high
intakes of antioxidant supplements over the long-term were found to produce
significant adverse events (188) emphasizes the need for long-term intake stu-
dies to define the safety of prolonged intake of high amounts flavanol- and
procyanidin-rich foods. Information gathered to date suggests that dietary
intakes of flavanols and procyanidins up to 3.5 g a day is protective against
CVD; however, whether this is a safe range of intake for a sustained period
of time in all individuals, and at different life stages, such as those who are
pregnant or elderly, merits additional investigation (27). While the obser-
vation that supplements of ()-epicatechin enhances vascular reactivity pro-
vides direct evidence that dietary flavanols are bioactive, additional research is
needed to further define the specific mechanisms of actions of this important
class of nutrients.
TAKE AWAY POINTS
.Flavanol- and procyanidin-containing foods likely modulate surrogate
markers of vascular homeostasis in healthy individuals and may be ben-
eficial in counteracting age-related diabetes, CVD, and cognitive decline.
.While evidence to date suggests that dietary intakes up to 3.5 g a day can be
protective against CVD, long-term studies are needed to define the safety of
prolonged intake of high amounts of flavanol- and procyanidin-rich foods,
including at different life stages, such as in pregnancy and in older adults.
.Additional research is needed to define the specific mechanisms of action
of ()-epicatechin, a bioactive flavonoid thought to enhance vascular
reactivity.
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