A risk factor

long-chain
for
polyunsaturated cardiovascular
fatty acids disease
and
health benefits
Translated from «Oméga-3 et bénéfice santé», published by the Centre d'Etude et d'Information
sur les Vitamines, Roche Vitamines France, Neuilly-sur-Seine.
I. Introduction

In the 1970s researchers struck by the low incidence of coronary

artery disease among Greenland Inuits, began to study the
nutritional properties of omega-3 polyunsaturated fatty acids
(PUFAs): Inuits consume large amounts of oily fish and marine
mammals with a particularly high omega-3 long-chain (LC) fatty
acid content. The observation that Greenland emigrants developed
the same incidence of cardiovascular disease as the host populations
soon eliminated any notion of genetic protection. The researchers
therefore associated the weak incidence of coronary heart disease
among the Inuits with their diet.

Many studies have since confirmed the cardiovascular benefits of

omega-3 LC fatty acids. They have also shown that these acids have
many biological functions, in particular an important role in the
development of the brain. They may also be involved in the preven-
tion of certain cancers.

This document reviews the most significant clinical and metabolic

findings related to omega-3 LC PUFAs, as described in the literature.

3
 II. Omega-3 and omega-6
polyunsaturated fatty acids
1. Definition
PUFAs are natural constituents of animal and vegetable fats. They
are made up of a carbon chain with a methyl group at one terminal
and an acid group at the other. These fatty acids are said to be
polyunsaturated because they contain several double bonds.

■
H3C
Omega-3 LC PUFAs
Eicosapentaenoic acid (EPA)
Docosahexaenoic acid (DHA) COOH
Stearic acid (saturated)

H3C 3 5
1

2 4 6 COOH
Linoleic acid (n-6 PUFA)

H3C 1 3
COOH
-linolenic acid (ALA, n-3 PUFA)

H3C 1 3
COOH
eicosapentaenoic acid (EPA, n-3 PUFA)

2
COOH

H3C 1 3

docosahexaenoic acid (DHA, n-3 PUFA)

Figure 1: PUFA structures
4
■ PUFAs are classified by the position of the first double bond coun-
Since the body cannot ting from the methyl terminal. Thus the first double bond of n-3
synthesize linoleic or PUFAs, also known as omega-3 or 3 PUFAs, is three carbon atoms
-linolenic acid, these are from the methyl terminal; in n-6 PUFAs (omega-6 or 6 PUFAs) it is
termed ‘essential’ fatty six carbon atoms from the methyl terminal.
acids.
Omega-3 Omega-6

-linolenic Linoleic
acid acid
(ALA, C18:3) (C18:2)

Prostaglandins -linolenic
■ acid
The enzyme reactions C18:4 (GLA, C18:3)
responsible for LC fatty acid Thromboxanes

formation fail to satisfy the

body’s requirements. C20:4
These fatty acids must Dihomo--
therefore also be supplied linolenic acid
in the diet to avoid the risk (DHLA, C20:3)
of deficiency. Eicosapentaenoic
Leucotrienes

acid
(EPA, C20:5)
Arachidonic
Prostaglandins acid
(AA, C20:4)

C22:5 Thromboxanes

C22:4
Docosahexaenoic
acid
C22:5
(DHA, C22:6)
Figure 2: PUFA metabolism
5
 Two PUFAs, linoleic acid (omega-6 PUFA) and a-linolenic acid
(omega-3 PUFA), are termed essential in that they cannot be
synthesized by the body (whether animal nor human) and have to
be supplied by the diet.
The body stores these fatty acids, uses them as an energy
source, or transforms them, using elongase and desaturase
enzymes common to both PUFA families, into elongated
PUFAs with additional points of desaturation.
However, these reactions are slow and limited, and may fail to
satisfy the body’s LC PUFA requirements.
The diet needs to supply appropriate amounts not only of
essential fatty acids but also of their LC derivatives.

2. Sources
Omega-6 PUFAs are found mainly in vegetable products. Linoleic
acid is found in soybean, corn, nut, sunflower and olive oils, -lino-
lenic acid (GLA) in borage, evening primrose and blackcurrant seeds,
and omega-6 LC PUFAs such as arachidonic acid mainly in animal
products.
Omega-3 PUFAs are found in vegetable products (rapeseed, soybean
and nut oils) and animal sources (mainly fish and fish oils). The
richest sources of Omega-3 LC PUFAs are fish, particularly oily fish
(mackerel, herring and salmon) while smaller amounts are found in
meat (Table 1).
■ Source (EPA + DHA) g/100 g
Omega-3 LC PUFAs are
found mainly in oily fish. Mackerel 2.5
Salmon 1.8
Herring 1.6
Tuna 1.6
Beef 0.25
Table 1: Lamb (leg) 0.5
Omega-3 LC PUFA content of common
animal products1 Pork 0.7
6

■ The cell membrane is a barrier characterized by highly selective
PUFAs in the body:
– energy source
– cell membrane
constituents
– eicosanoid precursors
■ The omega-3 and omega-6 PUFA families compete for the enzymes
The omega-3/omega-6 ratio
is critical: the recommended
ratio is between 1/4 and
1/10, the ideal being 1/5.
3. Role and metabolism of polyunsaturated fatty acids
permeability. It is involved in the process of energy transformation; it
also controls the flow of information between cells, and contains
receptors sensitive to external stimuli.
The cell membrane consists of a double phospholipid layer
incorporating both omega-3 and omega-6 PUFAs (EPA and
DHA, and arachidonic acid, respectively).
The particular cis configuration of PUFA double bonds endows these
molecules with fluid-like physical properties.
LC PUFAs account for the flexibility of biological membranes.
They also act as messengers. 20-carbon PUFAs, e.g. dihomo--lino-
lenic acid (C20:3 omega-6), arachidonic acid (C20:4 omega-6) and
eicosapentaenoic acid (C20:5 omega-3), are the precursors of the
prostaglandin and leukotriene eicosanoids (Figure 2) which behave
as local hormones by acting on cell regulation.
Via the eicosanoids, PUFAs are involved in inflammation, the
modulation of synaptic transmission, the regulation of blood
flow and ionic transport etc.
by which they are metabolized; the eicosanoids which are synthe-
sized as a result can have antagonistic actions depending on their
PUFA precursor. It is therefore critical to maintain a proper omega-
3/omega-6 balance. The recommended ratio is between 1/4 and
1/10, the ideal being 1/5 (§ VII – Recommendations).
Altering this balance can have functional repercussions and
may predispose to the development of diseases, e.g. cardio-
vascular diseases, immune disorders, and inflammatory
conditions. For the same reason, dietary intakes in line with
the optimal omega-3/omega-6 ratio appear to help in the
prevention of such diseases.
7
 III. Omega-3 polyunsaturated fatty acids
and cardiovascular diseases

1. Epidemiology
Omega-3 PUFA research was originally inspired by the very low
incidence of cardiovascular diseases among the Greenland Inuits
(10–30% lower than in Denmark) 2,3. The Inuit diet is based on oily
fish and marine mammals containing high omega-3 LC PUFA
concentrations. The average Inuit consumes 10 g of omega-3 PUFA
for every 3000 kcal, therefore having high plasma levels of DHA
and EPA and low levels of arachidonic acid. This results in a lower
platelet reactivity and a longer bleeding time. The same metabolic
profile has been observed in other populations with a high fish
consumption (e.g. Japan) 4,5.

Several epidemiological studies have found an association between

regular fish consumption and a decrease in cardiovascular morta-
lity 6–11. The US Physicians’ Health Study showed that consuming a
weekly portion of fish decreased prospective cardiovascular and
overall mortality by 30% and 45%, respectively10. The Seattle case
control study from 1988 to 1994 in subjects with no cardiovascular
history showed that cardiac arrest rates were halved in persons
consuming a weekly portion of oily fish. This protective effect was
associated with a change in the lipid composition of the red cell
■ membranes11.
Regular (twice-weekly) Above five portions a week, fish consumption provides no additional
consumption of fish with a cardiovascular benefit12–14.
high omega-3 LC PUFA
content decreases cardio- Secondary intervention studies in groups of survivors of a first
vascular incident severity cardiovascular incident receiving dietary fish or fish oil supplemen-
and lowers mortality by tation vs placebo have given similar results15–17. The Diet and
30%. Reinfarction Trial (DART) in 2033 men showed a 29% lower morta-
8
lity risk in consumers of 300 g fish per week for 2 years vs non
consumers15. However no significant influence was observed on the
risk of a new cardiovascular incident.

The GISSI study, which included patients hospitalized for myocardial

infarction no more than 3 months before the start of the study,
confirmed these findings16. After 3.5 years, the cardiovascular
mortality rate in the group supplemented with omega-3 PUFA 1
g/day (comprising EPA and DHA 850–880 mg/day), was 30% lower
than in the placebo group. Omega-3 PUFA supplementation also
decreased the sudden death rate by 45%. Lower rates of non fatal
myocardial infarctions and strokes were also observed.

Epidemiological studies have shown that omega-3 fish oils have

protective cardiovascular effects which have been confirmed in
secondary prevention studies. The GISSI study, which is the most
recent, has clearly shown that omega-3 LC PUFA consumption
decreases cardiovascular mortality.

In summary, all these studies suggest that omega-3 LC PUFAs

and their metabolic products help to prevent some of the risk
factors involved in cardiovascular diseases18,19.

2. Omega-3 long-chain polyunsaturated fatty acids

and cardiovascular risk factors

a) Coagulation and platelet aggregation

Under physiological conditions, the coronary circulation adapts its
blood flow to myocardial oxygen demand. At rest and during
exercise, a balance is therefore struck between oxygen supply and
demand. In coronary heart diseases, this balance is upset and the
myocardium no longer receives the oxygen it requires. The main
9
 consequence of coronary heart disease is myocardial ischemia, i.e. a
decrease and sometimes a complete interruption in the blood supply
to an area (or areas) of myocardium. If this situation persists, the
myocardial cells suffocate and die; the result is myocardial necrosis
or infarction.
The leading cause of coronary heart disease is atherosclerosis20, the
main feature of which is the atheromatous plaque, a deposit of
lipids and cell debris in the intima layer of the artery which partially
blocks the lumen. It takes several years to form and may eventually
fissure, triggering the development of a blood clot which adheres to
the plaque and leads to myocardial infarction. This outcome, acti-
vated by irreversible platelet aggregation, is precipitated by slowing
of the circulation over the plaque.

Any substance which modulates platelet function has a major

influence on the severity of myocardial infarction.

■ Omega-3 LC PUFAs act on platelet aggregation mainly via their role

As eicosanoid precursors, in eicosanoid synthesis. Eicosanoids are produced by all cells from
omega-3 LC PUFAs enhance the 20-carbon PUFAs contained in their membrane phospholipids.
blood fluidity. Eicosanoids vary in structure and bioactivity according to their PUFA
precursor (arachidonic acid: omega-6 family; and EPA: omega-3
family). The thromboxanes produced by platelets and prostaglandins
produced by vascular endothelium are involved in regulating hemo-
stasis and vasomotility. Thromboxane A2 (TXA2), whose precursor is
arachidonic acid, is a vasoconstrictor which stimulates platelet
aggregation. It promotes hemostasis and its pathological equivalent,
thrombosis. Conversely, prostaglandin I2 (PGI2), which is derived
from arachidonic acid, is an antihemostatic and antithrombotic
(Figures 2 & 3).
A high omega-3 PUFA diet promotes the formation of 3-series
prostanoids21. Prostaglandin I3 (PGI3) is similar in activity to prosta-
glandin I2 (PGI2)but thromboxane A3 (TXA3) is a less effective hemo-
static than thromboxane A2 TXA2.
10
 A high omega-3 PUFA diet has antihemostatic and antithrom-
botic effects by altering the balance between the various
eicosanoids22.

■ ENDOTHELIAL
PLATELETS
Omega-3 LC PUFAs decrease CELLS

platelet aggregation and

promote vasodilatation.
They have antihemostatic
and antithrombotic activity.

Arachidonic Arachidonic
EPA EPA
acid acid

TXA2 TXA3 PGI2 PGI3

platelet little anti-aggregation anti-aggregation

aggregation biological activity vasodilatation

Figure 3: Simplified pathway of

prostanoid production from PUFAs22

EPA and DHA modulate hemostatis and vasomotility by

affecting the expression of enzymes involved in eicosanoid
synthesis23,24.

Although the mechanisms differ, the action of omega-3 PUFAs on

platelet function could be compared to that of aspirin. Aspirin irre-
versibly blocks platelet cyclooxygenase and hence thromboxane A2
and Thromboxane A3 production but causes virtually no inhibition of
prostaglandin I2 production. The resulting hemostatic and anti-
thrombotic properties account for the use of aspirin in the primary
and secondary prevention of cardiovascular diseases25,26.
11
 Several studies have shown that omega-3 PUFAs have antiaggregant
activity and lengthen the bleeding time4,27–33. EPA achieves its
maximum antiaggregant effect at a dose of 6 g/day 34.

However, at doses between 1 to 3.5 g DHA and EPA daily,

the antithrombotic and antihemostatic activities of omega-3
PUFA remain within physiological limits, with no increase in
hemorrhagic risk 28,35.

b) Omega-3 long-chain polyunsaturated fatty acids and blood

pressure
Hypertension is one of the major cardiovascular risk factor,
accounting for 34% of deaths in developed countries and increasing
the risk of heart failure 5-fold. It causes hardening and loss of
elasticity in the artery wall so that arteries are no longer able to
absorb the systolic pressure waves. Hypertension also promotes
atherosclerosis by causing microtrauma that allow LDL to
penetrate the artery wall. In addition, it is also responsible for the
left ventricular hypertrophy often associated with coronary heart
diseases.
■
Omega-3 LC PUFAs Supplementation with omega-3 LC PUFAs at relatively high
lower blood pressure in doses (3–9 g) markedly lowers systolic and diastolic blood
hypertensive patients. pressure in moderately hypertensive subjects 36–40.

The effects in normotensive subjects are more disputed 41-45. An

intervention study of non fish-consuming patients with moderate
hypertension (diastolic: 85–110 mm Hg; systolic: < 180 mm Hg)
showed that supplementation with 6 g/day fish oil containing 85%
EPA and DHA (i.e. approximately 5 g) significantly lowered systolic
and diastolic blood pressure. Fish oil supplementation did not lower
blood pressure in subjects consuming fish at least 3 times a week
or in those whose plasma omega-3 phospholipid levels exceeded
175.1 mg/l46. Véricel et al. reported a hypotensive effect on systolic
12

■ these “remnants” from the circulation involves the formation of
The postprandial response
occurs approximately 60
minutes after ingestion and
lasts 6–8 hours. It is modu-
lated by the amount and
composition of ingested
lipids.
but not diastolic pressure at much lower omega-3 PUFA doses (DHA:
150 mg; EPA: 30 mg)32.
When combined with the beta-blocker propranolol, omega-3
PUFAs have a synergistic hypotensive effect42. They also lower the
triglyceride levels which tend to be raised by the use of antihyper-
tensive drugs.
The hypotensive effect of omega-3 PUFAs correlates with the
plasma levels of omega-3 LC PUFA phospholipids, suggesting
that this effect is due not only to the effect of these PUFAs
on membrane fluidity but also on their ability to influence
the balance of the prostanoids controlling constriction and
dilatation of the artery wall.
c) Omega-3 long-chain polyunsaturated fatty acids and
plasma lipids
Dietary triglycerides are hydrolysed by salivary and pancreatic
enzymes to monoglycerides and fatty acids. They are then resynthe-
sized by enterocytes as triglycerides which form chylomicrons when
combined with proteins. Chylomicrons transport triglycerides in the
blood to the liver and other tissues (Figure 4).
Chylomicron catabolism involves lipases located mainly in the
vascular endothelium of tissues with a high fatty acid requirement
(heart muscle, adipose tissue, mammary gland etc.). This results in
the formation of triglyceride-depleted ‘remnants’. Clearance of
triglyceride-rich high density lipoproteins (HDL-cholesterol) and very
low density lipoproteins (VLDL). VLDL, which are synthesized in the
liver from remnants, carbohydrates and proteins, are transformed by
hydrolysis into low density lipoproteins (LDL-cholesterol) which are
highly atherogenic.
The postprandial response refers to the appearance of chylomicrons
in the circulation. It occurs approximately 60 minutes after ingestion
13
Triglyceride metabolism Lipase
1
1. Hydrolysis of dietary lipids by
salivary and pancreatic enzymes into
monoglycerides and fatty acids.
2
2. Formation of chylomicrons by GENERAL CIRCULATION
entherocytes. They transport
triglycerides to the liver and to
other tissues. GENERAL
CIRCULATION
3. Catabolism of chylomicrons by
lipases into remnants and fatty Chylomicrons
acids. Fatty acids are distributed 3
Adipocytes
among tissues like muscles, brain, Free
fatty
and adipose tissue acids

Lipase
4. Clearance of the Remnants from the
Remnants
blood by liver receptors.
Formation of triglyceride rich VLDL
lipoproteins from remnants, alcohol, Brain
carbohydrates and proteins in the
liver.
Remnant receptor
HDL
5. Hydrolysis of VLDL into LDL.
Triglycerides are split off. 4
LIVER

6. LDL particles are cleared by

peripheral tissues through a specific
LDL receptor. Free
LDL receptor fatty
Modified (oxidized) LDL particles acids
are preferably taken up by
macrophages, they are particularly Muscles
VLDL
atherogenic.
Lipase
5
5
Modified LDL

LDL 6
Scavenger receptor

Peripheral cell
LDL receptor

Macrophage
of the arterial
wall

Figure 4: Triglyceride metabolism

14

■ Several epidemiological studies have shown that small
High fasting or postprandial
triglyceride levels are
associated with increased
cardiovascular risk.
and lasts 6–8 hours. The response is modulated by the amount and
composition of ingested dietary lipids. The amplitude of the post-
prandial response is accentuated by advancing age and a sedentary
lifestyle. It is higher in men than in women.
A normal and effective postprandial response rapidly
eliminates the chylomicrons and promotes the formation
of HDL-cholesterol 47, 48.
The fasting triglyceride level can be viewed as a marker of the post-
prandial triglyceric response. In the fasting state, a high plasma
triglyceride level is a sign of abnormal postprandial metabolism and
is generally associated with other cardiovascular risk factors.
High postprandial triglyceride concentrations lead to the formation
of remnants and small, very dense particles of LDL-cholesterol which
are particularly atherogenic. They also tend to lower the levels of
HDL-cholesterol, which is thought to be less atherogenic as it helps
to clear cholesterol from the circulation 49–51.
particles of LDL-cholesterol, low HDL-cholesterol levels and
high plasma triglyceride concentrations are associated with
a high mortality from coronary heart diseases 52–54.
Some investigators have argued that high postprandial triglyceride
levels are a specific marker of cardiovascular risk 55–62. However, this
risk correlates closely with all other, already recognized risk factors.
All nutritional factors that can help to control high plasma trigly-
ceride levels have therefore a potential role in cardiovascular disease
prevention.
15
 Many studies have shown that omega-3 LC PUFAs lower high
plasma triglyceride levels63. They do so by lowering hepatic
triglyceride synthesis and by decreasing the release of trigly-
ceride-rich VLDL into the blood 64–69.

■ EPA and DHA act via transcription factors (small proteins that bind
Omega-3 LC PUFAs lower to the regulatory domains of a gene). Binding of an omega-3 LC
plasma triglyceride levels. PUFA to a transcription factor modifies the structure of the factor
and its ability to activate or inhibit the target gene (Figure 5). To
date, two transcription factors are known to interact with LC PUFAs:
peroxisome proliferator activated receptor (PPAR) and sterol regula-
tory element binding protein (SREBP)70,71. PPAR appears to be
involved in the activation of fatty acid oxidation65 and SREBP in the
inhibition of triglyceride synthesis pathways.

Omega-3 LC PUFAs in conjunction with transcription factors target

the genes governing cell triglyceride production and those activating
■ oxidation of excess fatty acids in the liver72-74. As a result of
Omega-3 LC PUFAs inhibition of fatty acid synthesis and an increase in fatty acid cata-
decrease endogenous VLDL bolism, the amount of substrate available for triglyceride production
production. decreases, as does the entry of triglycerides into the blood.
Omega-3 LC PUFAs also control postprandial blood lipid levels by
decreasing endogenous VLDL production75-78.

Regular consumption of oily fish, like omega-3 LC PUFA

supplementation, lowers postprandial triglyceride levels
in both normal subjects30,33,45 and those with hyperlipi-
demia77, 79–82. The totality of these effects has been observed
at omega-3 PUFA daily doses from 1 to 7 g.

■ Recently several studies have been performed with low doses of

Low doses of EPA (0.17 g) omega-3 PUFAs. Daily supplementation with DHA 0.11 g and EPA
and DHA (0.11 g) lower 0.17 g lowered triglyceride levels by 22% in postmenopausal
triglyceride levels. women with normal triglyceride levels 63. Sorensen et al.83 also
16
 Omega-3 fatty acid

Blood circulation

cytosol

nucleus

inactivated SREBP

inactivated PPAR

activated
activated
PPAR
SREBP

transcription
or repression

SRE Gene PPRE Gene

showed a significant decrease in triglyceride levels after supplemen-

tation with omega-3 PUFAs 0.91 g daily. Marckmann et al.84 found
a 16–18% decrease in postprandial triglycerides in the group
supplemented with omega-3 LC PUFAs vs a decrease of only
10–13% in the control group supplemented with sunflower oil.
Very recently Visioli et al. showed that consuming 500 ml daily of
omega-3 LC PUFA-enriched milk (EPA: 240 mg/l; DHA: 360 mg/l)
17
 lowered triglyceride levels by 19% and increased HDL-cholesterol
levels by 19%85. Although the study population was small, these
results confirm those already achieved with higher doses of omega-
3 LC PUFAs and suggest that milk enhances omega-3 PUFA bioavai-
lability. EPA and DHA are identical in their ability to lower triglyce-
rides86.

Hypertriglyceridemia is now recognized as a cardiovascular

risk factor. Both DHA and EPA lower raised triglyceride levels
and DHA is particularly effective in the postprandial response.
Omega-3 LC PUFAs are thus of documented nutritional
benefit in the prevention of cardiovascular risk factors.

d) Omega-3 long-chain polyunsaturated fatty acids and

arrhythmias
Arrhythmias are characterized by irregular electrical activity of heart
muscle. They are potentially fatal and often result in sudden death.
An epidemiological study showed that the arachidonic/docosa-
■
hexaenoic acid (AA/DHA) ratio in the heart cell membranes of
Death rates from cardiac
subjects dying from a sudden cardiac cause was consistently higher
arrest are lower in fish
than in age-matched controls87. Siscovick et al.11 found lower
consumers whose cardiac
cardiac arrest rates in fish consumers whose cell membranes had a
cell membranes have high
high omega-3 LC PUFA content. This suggests a relationship
DHA levels.
between the PUFA composition of the myocyte membrane and
death from cardiac arrest.

High levels of omega-3 LC PUFAs –DHA in particular –

■ enhance cardiomyocyte protection in certain disease states.
Omega-3 LC PUFA
supplementation of Sellmayer et al.88 showed a significant decrease in ventricular
2.4 g/day decreases arrhythmias in a small population consuming 2.4 g EPA and DHA.
ventricular arrhythmias. This study was confirmed by Christensen et al.89,90 who also
18

■ average of 30%.
The severity of post-infarct
arrhythmias correlates with
the omega-3 LC PUFA
content of the cardiomyo-
cyte membranes.
■
An antiarrhythmic effect is
observed when DHA
accounts for 20% of
membrane lipids.
showed a significant decrease in ventricular arrhythmias after
supplementation with omega-3 LC PUFA 4.3 g daily. Omega-3 LC
PUFAs have a moderate antiarrhythmic effect compared to drugs
such as flecainide or encainide91. However, whereas these drugs
impair survival after myocardial infarction, omega-3 PUFA supple-
mentation increases survival after a cardiovascular accident by an
Omega-3 PUFAs seem to be involved in the control of such
key cardiac parameters as rhythm and energy production92–94.
✔ Effects on ischemia and reperfusion
Ischemia and reperfusion, as occur most acutely in infarction, are
situations which generate arrhythmias. It has been shown in the rat,
monkey and dog that the severity of such arrhythmias can be corre-
lated with the type of fatty acids which the animal has previously
ingested. Susceptibility to arrhythmias is markedly decreased in
animals previously fed a fish oil diet95-98.
The results show effective antiarrhythmic activity when DHA
accounts for approximately 20% of the fatty acids in the
cardiomyocyte membranes.
This level of omega-3 PUFA composition can only be achieved by
supplementation with DHA itself (not its precursors).
✔ Effect on adrenoceptors
Attempts to identify the precise mechanism of the cytoprotective
effect led to work on the adrenoceptors which are recognized
factors in arrhythmia. Adrenoceptors are a group of membrane
proteins whose function in the heart is to transmit the neuroendo-
crine message of the catecholamines (adrenaline and its derivatives)
with respect to rhythm and force of contraction.
19

■ the presence of DHA has proved determinant, causing a decrease in
Membrane PUFA content
modulates cardiac
adrenoceptor response.
■
DHA has the same basic
activity as a -blocker.
■
In ischemia, cell membranes
with a high omega-3 LC
PUFA content have a
decreased oxygen
requirement.
It has been known for many years that the cardiac adrenoceptor
response depends on the membrane PUFA content. In this regard,
the production of cyclic AMP, the intracellular mediator99.
Ischemia is associated with a decrease in cardiomyocyte
-adrenergic response, which is more marked in membranes
with a high DHA content100.
In other words, the activity of DHA is similar in principle to that of
-blockers, a group of key cardiology drugs used to decrease the
cardiac effects of catecholamines. The effect of omega-3 PUFAs on
cardiac adrenergic function may help to account for their antiar-
rhythmic and cytoprotective activity.
✔ Effect on energy production
Another key element in myocardial cytoprotection is related to
energy production. At all moments the myocardium is producing a
large amount of energy in the form of ATP. Production from oxygen
and energy substrates (mainly fatty acids and glucose) takes place in
the inner mitochondrial membrane. Studies in isolated rat hearts
have shown that the hearts of animals fed an omega-3 LC PUFA
diet produce ATP at a lower oxygen cost. Their mitochondrial
membranes also use energy more effectively92.
This moderate reduction in the oxygen requirement is of little
consequence in normal circumstances. But in ischemia, due to
the oxygen deprivation, it is a key element in myocardial
protection.
Post-ischemic recovery of mitochondrial function is also markedly
enhanced in animals receiving fish oil.
20

■
Omega-3 LC PUFAs by their
involment in the eicosanoid
balance are also involved in
the cardiac rhythm and
contractions.
This as yet unexplained metabolic effect of omega-3 LC PUFAs
on energy production could be an important factor in myocar-
dial cytoprotection since it is based on the principle of
reducing the oxygen cost of cardiac contraction.
✔ Effect on cardiac prostaglandins
The influence of arachidonic acid metabolites on cardiac rhythm and
contraction has been increasingly elucidated. Hypoxia activates the
synthesis of prostaglandin E2 while reoxygenation activates the
synthesis of prostaglandin I2. Synthesis of these prostaglandins is
decreased in normal cardiomyocytes enriched with omega-3 LC
PUFA. In hypoxia, however, enrichment of the membranes with
those fatty acids may have little effect on the increased production
of prostaglandin E2, but in reoxygenation they inhibit the synthesis
of prostaglandin I2101.
✔ The ion channel question
The use of isolated cardiomyocytes as an experimental model of
ischemia and reperfusion has shown that electrical activity decreases
more rapidly during hypoxia in cells with a high omega-3 LC PUFA
content98. Recovery after reperfusion is also more rapid and effec-
tive. Yet in isolated cardiomyocytes and under normal conditions,
membrane electrical activity (resulting from the actions of the
various ion channels) is not significantly altered by the membrane
omega-3 LC PUFA content.
Recent studies suggest that omega-3 LC PUFA activity is
specific to certain ion channels, possibly through an ‘acute’
effect by free fatty acids rather than a ‘chronic’ effect by
membrane-bound fatty acids102.
21
 Such channel effects could influence arrhythmias but the mechanism
remains unelucidated. It may involve a direct effect on the channels
concerned or a general effect on membrane characteristics, fluidity
in particular.

■ 3. Conclusion
Omega-3 LC PUFAs
moderate cardiovascular Prevention of such a multifactorial entity as cardiovascular disease
risk factors such as means taking all the risk factors involved into account. A number of
elevated triglyceride drug treatments are currently available which slow the course or
levels, arrhythmias and reduce the incidence of cardiovascular diseases by acting on these
hypertension. risk factors (blood pressure, cholesterol, triglycerides etc). But as the
age of the population increases, the long-term efficacy of these
treatments is largely unknown and there is a high risk of side
effects.

Many of the risk factors involved in cardiovascular diseases

have a dietary link.

A properly balanced diet could therefore help to delay the first

symptoms and hence the age at which drugs first need to be taken.
In this regard, there have been many clinical and experimental
studies with omega-3 LC PUFAs. Technological progress, particularly
in molecular biology, has provided a number of biochemical and
metabolic explanations for the clinical and epidemiological data. The
most striking concern the role of omega-3 LC PUFAs in the control
of postprandial triglyceride levels, but exhaustive investigations can
also be found in the literature on the effects of these PUFAs on
blood pressure and cardiac arrhythmias.

22
Dietary correction of the omega-3/omega-6 ratio in healthy
subjects and omega-3 LC PUFA supplementation at nutritional
doses combined with tailored treatments in high-risk subjects
constitute solidly documented strategies in cardiovascular
disease prevention.

23
 IV. Omega-3 long-chain polyunsaturated
fatty acids and inflammation
Inflammation, the body’s non specific response to trauma and
chemical or microbial aggression, is classically associated with rubor,
tumor, calor and dolor (redness, swelling, heat and pain). These
symptoms result from the release of mediators derived primarily
from leukocyte activation. They include two eicosanoid mediators,
prostaglandin E2 and leukotriene B4, whose precursor in both cases
is arachidonic acid.

Omega-6 Omega-3

inhibition
excitation Arachidonic acid Eicosapentaenoic acid

PROSTAGLANDIN E2 LEUKOTRIENE B4

Stimulus

n local
inflammation

n fever
n blood
coagulation

n septic shock
Macrophages or neutrophils T and B lymphocytes
Figure 6: Omega-3 PUFAs and the
inflammatory response (Calder103)
24
 These mediators are produced via reactions involving cyclooxygenase
and 5-lipoxidase, respectively (Figure 2).

EPA and DHA are competitive inhibitors of arachidonic acid

oxygenation by cyclooxygenase. In addition, EPA is a
substrate of cyclooxygenase, but it promotes the formation
of prostaglandin E3 and leukotriene B5 which are less active
proinflammatory agents than the corresponding derivatives
of omega-6 precursors (Figure 6).

Omega-3 LC PUFA ingestion also lowers the arachidonic content of

the cell membranes and hence its availability for eicosanoid
synthesis104,105.

Other mediators such as cytokines, interleukins and tumor necrosis

factor (TNF) have a proinflammatory cellular action. EPA and DHA
inhibit the production of cytokines IL-1 and TNF- via a still
unelucidated mechanism103,106.

■ Thus a diet high in omega-3 PUFA modulates the inflammatory

The omega-3/omega-6 ratio response by inhibiting arachidonic acid metabolism and promoting
modulates the inflammatory the production of much less inflammatory mediators.
response.
Omega-3 LC PUFAs are similar in action to certain anti-
inflammatory drugs which inhibit mediator production.

Under normal circumstances, the inflammatory response has a

protective and beneficial function. But when it becomes chronic or
exaggerated, it can give rise to diseases such as rheumatoid arthritis,
Crohn’s disease and asthma. Omega-3 LC PUFAs appear to relieve
the symptoms of these inflammatory diseases.

25
 1. Omega-3 long-chain polyunsaturated fatty acids
and rheumatoid arthritis
■ Epidemiological evidence supports the hypothesis that consumption
Regular fish consumption of oily fish prevents the development of rheumatoid arthritis : preva-
correlates with a lower lence is decreased among Inuits eating large amounts of fish and
incidence of rheumatoid marine mammals with a high omega-3 LC PUFA content107; rheu-
arthritis. matoid arthritis also takes less severe forms in Faroe Islanders, who
have a high-fish diet108; lastly, a case-control study showed that the
consumption of baked or grilled fish is associated with a decreased
risk of rheumatoid arthritis109 (particularly rheumatoid factor-positive
disease, which is considered to have a poorer outcome)109.

These results suggest that omega-3 LC PUFA consumption may help

to prevent rheumatoid arthritis. They support the hypothesis of
dietary factor involvement in the development or clinical expression
of the disease.

Clinical studies using omega-3 LC PUFAs in the treatment of

rheumatoid arthritis, mostly conducted vs placebo, differ from each
other in treatment duration, dose, coadministration of non steroidal
antiinflammatory drugs (NSAIDs), and placebo used.

Most observed significant clinical improvement vs baseline in

the active treatment arm (3–6 g/day), and in some cases vs
the placebo arm 110–116.

The clinical parameters included the number of painful joints,

number of swollen joints, morning stiffness, and global impression
(patients and/or investigators). The clinical improvement observed
was not generally associated with a significant change in the labora-
tory indicators of disease activity (erythrocyte sedimentation rate and
C-reactive protein). The results indicated that clinical effect was not
generally observed until 8 weeks, or more commonly 12 weeks.
26
 Curtis et al. have recently argued that omega-3 LC PUFAs also
influence the expression and activity of catabolic factors involved in
the breakdown of joint cartilage117.

A metaanalysis has confirmed that omega-3 LC PUFAs have a

dose-dependent effect on the clinical parameters of rheumatoid
arthritis118. They also appear to decrease NSAID use in rheumatoid
patients119,120.

2. Omega-3 long-chain polyunsaturated fatty acids

and Crohn’s disease

Crohn’s disease is a chronic inflammatory bowel disease. Despite

much research over many years, its cause remains unknown. The
sole undisputed risk factors are smoking and a first-degree family
history. Infection (measles virus) and diet (an excess of rapid sugars
and saturated fats) have been accused but the charge remains
■ unproven. In Japan, the incidence of Crohn’s disease is steadily
A high saturated-fat diet increasing. This correlates with an increased proportion of animal
may predispose to Crohn’s fat in the diet and a decreased consumption of omega-3 LC PUFA,
disease. suggesting that the latter may be protective in this regard121.

Several studies have shown benefit from omega-3 LC PUFA

supplementation in Crohn’s disease and inflammatory bowel
disease 122 –126.

An Italian study showed that a high omega-3 PUFA preparation (EPA

1.8 g/day and DHA 0.9 g/day) helped to prevent relapse in Crohn’s
disease127. Other studies have found more disappointing results:
comparing omega-3 LC PUFA supplementation vs a low carbo-
hydrate diet and vs a control group, Lorenz-Meyer et al. found no
benefit in favor of dietary intervention128. The discrepancy between
27
these studies could be due to patient selection, the doses of
omega-3 supplementation but also to the type of placebo129. Some
placebos, e.g. olive oil, have shown benefit in several inflammatory
bowel conditions. Also, Belluzzi et al.127 used enteric-coated
capsules that resist gastric juice acidity for approximately 30
minutes, releasing the omega-3 LC PUFAs in the jejunum rather
than the stomach, thus enhancing absorption128. This could account
for the efficacy of omega-3 LC PUFA supplementation in Crohn’s
disease observed in this study. Other studies are currently running in
Europe.

3. Conclusion

Research into the involvement of omega-3 LC PUFAs in inflamma-

tory disease is relatively recent, accounting for the few clinical data
that are available.

Nevertheless, eicosanoid involvement in inflammatory and

immune mechanisms suggests that the omega-3 precursors of
E3 prostaglandins and B5 leukotrienes may play a beneficial
role in inflammatory disease.

As recent studies show, this role appears confined to the disease

symptoms, with omega-3 LC PUFAs enabling patients to decrease
their doses of NSAIDs and hence minimize the related side effects.

28
 V. Omega-3 long-chain polyunsaturated
fatty acids and cancers
The molecular and biological properties associated with the interac-
tions between omega-3 LC PUFAs and other nutrients (omega-6,
antioxidants), give a strong indication of the role of these fatty
acids in cancer prevention and cancer treatment130,131.

Correlations have been drawn between the low incidence of breast

cancer in Inuit and Japanese women and their high consumption of
fish and omega-3 fatty acids132-134. The incidence tends to increase
as the diet changes to include a greater proportion of meat and
saturated fats132,135. Many animal studies have shown a protective
effect by EPA and DHA in mammary cancer136, colon cancer137-139
and liver cancer140, in addition to an antimetastatic effect141-144.
However, the overall results of the epidemiologic case-control
studies are mixed. Most show no convincingly significant effects of
omega-3 LC PUFAs in cancer prevention145.

■ Some investigators argue that it is the omega-3/omega-6

There is a correlation ratio which modulates the preventive effects146 –149. In the
between the incidence of case-control study by Simonsen et al., the omega-3/omega-6
some cancers and an ratio was inversely proportional to the cancer incidence147.
imbalance in the
omega-3/omega-6 ratio. Others have concentrated on the highly cytotoxic lipoperoxides
derived from fatty acid oxidation : omega-3 LC PUFAs may inhibit
tumor growth by inducing tumor cell apoptosis via these lipoper-
oxides150.

Noding et al. observed that the sensitivity of tumor cells to DHA and
its oxidation products, depends on their antioxidant status151.
The presence of vitamin E blunts antitumor activity and apoptosis
induction by omega-3 LC PUFAs152. Timmer-Bosscha et al. found
29
 that DHA potentiated the toxicity of an anticancer agent (cis-
diamine-dichloroplatinum II) in malignant human embryonic cell
lines153.

■ In a prospective study, Bougnoux et al. observed an enhanced

The breast tumor adipose response to chemotherapy in breast cancer patients with high
tissue DHA level is a adipose tissue DHA levels.
predictor of
chemosensitivity. They also showed that the DHA level was a predictor of chemosen-
sitivity154. DHA enhances the tumor cell response to prooxidants155.
The tumor cell toxicity of omega-3 LC PUFAs thus seems due to a
decrease in the efficacy of antioxidant defence mechanisms during
malignant transformation.

This accounts for the specificity of omega-3 LC PUFA action

against tumors vs normal cells and offers promising prospects
in terms of combination chemotherapy regimens and dietary
prevention.

30

■ after adipose tissue. LC PUFA levels are particularly high in the retina
Omega-3 LC PUFAs are
heavily involved in cerebral
and visual function.
■ and release neurotransmitters, the medium must contain LC
Dietary LC fatty acids are
essential to neuronal
functions.
VI. Omega-3 long-chain polyunsaturated
fatty acids and brain function
Nervous tissue has the second highest concentration of fatty acids
and cerebral cortex. DHA can account for up to 50% of phospho-
lipid fatty acids in these tissues, suggesting that it is heavily involved
in neuronal and visual functions156 –158. Vision would in fact be
impossible without the presence of DHA.
For ethical reasons it is obviously very difficult to investigate
the effect of fatty acids on brain functions in humans. Know-
ledge in this field is therefore based essentially on data from
animal models.
The role of fatty acids in nervous tissue is primarily structural. As a
component of the architecture, they are involved in the function of
the neuronal membrane. A change in the essential fatty acid
composition of the synaptic membranes can affect neuronal func-
tions in terms not only of membrane receptors, ion channels and
enzymes159-165, but also of the transmission of intra- and intercel-
lular signals generated by second messengers derived from LC fatty
acid precursors166-169.
For cultured nerve cells to differentiate, multiply, and take up
PUFAs with docosahexaenoic acid (DHA, omega-3) and
arachidonic acid (AA, omega-6)170.
These fatty acids are supplied in the diet either directly as LC fatty
acids or as short-chain precursors which are elongated and desatu-
rated in the liver then transported to the brain cells171,172. Dietary
supplementation with large amounts of fish oil alters the brain fatty
acid profile and causes a decrease in omega-6 LC fatty acids
combined with an increase in DHA173,174.
31
 1. Omega-3 long-chain polyunsatured fatty acids and
cognitive functions
■ In rats a diet low in omega-3 causes behavioral disturbances and
Psychiatric symptoms are markedly impairs learning but has little effect on motility175 –177. In
often associated with low humans, low plasma omega-3 concentrations have been observed in
omega-3 blood levels. association with some psychiatric symptoms178 –180. Hibbeln et al.
suggested that omega-3 deficiency increased the risk of clinical
depression and behavioral disturbance; they reported an inverse
correlation between fish consumption and the prevalence of
depression178. Edwards et al. observed that erythrocyte membrane
omega-3 levels were lower in depressed patients than in controls;
the severity of depression was inversely proportional to these levels
and to dietary omega-3 intake181,182. Maes et al. also found omega-
3 fatty acid deficiency in depressed patients and suggested that
impaired omega-3 fatty acid metabolism was involved in clinical
depression183.

In a recent study, Kalmijn et al. observed an inverse correlation

between fish consumption and the development of dementia,
including Alzheimer’s disease, and explained it on the basis of the
antithrombotic and antiatherogenic properties of omega-3
PUFAs184. The Dutch Zutphen study had previously shown an
association between atherosclerosis and an increased risk of
Alzheimer’s disease185.

There remain limited data on omega-3 PUFA involvement in

psychiatric disease. Overall, the available studies show that
patients are deficient in these fatty acids. However, it has yet
to be shown whether this deficiency increases the risk of such
disease, or whether such disease, by influencing dietary
behavior, increases the risk of deficiency.

32
 2. Omega-3 long-chain polyunsaturated fatty acids
and brain development
■ Omega-3 LC PUFAs play a now well-documented role in the brain
Omega-3 LC PUFAs are development of the foetus and child. However, a consensus has yet
essential for neonatal brain to be reached on the recommended dietary allowance (RDA)186.
development. Premature infants require particular attention as they have no
reserve adipose tissue. These reserves are not generally built up until
the third trimester. Premature infants are therefore closely depen-
dent on dietary intake.

Differences in LC fatty acid status were first observed by comparing

breast-fed infants with parenterally fed infants and infants fed
formulas containing an imbalance of precursor PUFAs. An
imbalanced dietary PUFA intake in premature infants halves the
DHA/arachidonic acid ratio compared to term infants of equivalent
corrected age. It is also associated with impaired visual function158.
Similar effects have been reported in term infants fed a low
-linolenic acid formula for 4 months186,187. However, it should be
noted that the consequences of omega-3 LC fatty acid deficiency
are less dramatic than those due to a deficiency of the -linolenic
acid precursor.

The main studies of formula supplementation with DHA- and

EPA-rich fish oil in premature infants have shown that, after
6 weeks, retinal electrophysiology parameters in infants
receiving the supplemented formula are similar to those in
breast-fed infants188,189.

In term infants, despite an adequate intake of essential precursor

fatty acids (linoleic acid and -linolenic acid) at recommended levels,
omega-3 and omega-6 LC fatty acid levels in red cell phosphati-
dylethanolamine remain low in infants formula-fed for 4–6 months
compared to breast-fed controls190.
33
Breast-fed infants and formula-fed infants supplemented
with LC PUFAs (DHA and arachidonic acid) perform better at
problem solving and language learning than infants receiving
non supplemented formula191,192.

These findings suggest that omega-3 LC PUFA supplementation is

important in the third trimester and the first months of life for a
child’s subsequent brain development, and in particular for its
learning potential and behavior. Other studies are still needed to
determine the optimal daily dose of these fatty acids in neonates
and children. Nevertheless, we already know enough to be able to
recommend simultaneous supplementation with linoleic acid and
-linolenic acid, together with DHA and arachidonic acid in the
correct omega-3/omega-6 ratio.

34
VII. Recommendations
Our current diet encourages an excessive intake of saturated fats
without meeting our dietary requirements for PUFAs, in particular
omega-3 PUFAs. Yet it is now accepted that a limited intake of
saturated fatty acids and compliance with a correct omega-3/
omega-6 PUFA ratio have documented health benefits, in particular
in cardiovascular prevention.

The omega-6/omega-3 ratio should be between 4/1 and 10/1,

the ideal being 5/1.

Omega-3 and omega-6 fatty acids share metabolic pathways and

thus compete with one another, in particular for eicosanoid
synthesis. Depending on their fatty acid precursor, the eicosanoid
derivatives differ in their biological potential, with correspondingly
different impacts on health.

Various bodies have issued PUFA guidelines (Table 2). In particular,

the Food and Drug Administration (FDA) and American Heart
Association (AHA) attribute the cardiovascular benefits of PUFAs to
EPA and DHA. However, they consider that in contrast to the highly
positive findings in secondary intervention studies, the scientific
observations in the general population, although significant, require
further investigations. Nevertheless they recommend a regular
omega-3 LC PUFA intake of 0.9–1 g/day.

35
 Source Omega-6/Omega-3 Omega-3 EPA + DHA
ratio recommendations
Nordic Nutrition Committee, None 0.5%* (1-2 g/day)
1989193
NATO Workshop, None 0.27%*
1989194 (0.8 g/day)
Scientific Review Committee, 5:1 - 6:1.5 0.5%* (1-2 g/day)
Canada,1990195
British Nutrition Task Force, 6:1 0.5%* (1.1 g/day)
1992196
Scientific Committee for Food, 4.5:1 - 6:1.5 0.5%* (1-2 g/day)
EU, 1993197
FAO/WHO Expert Committee, 5:1 - 10:1
1994198
Committee on Medical Aspects None 0.2%* 0.1-0.2 g/day
of Food Policy, 1991, 1994199,200
National Nutrition council None 0.5%* (1-2 g/day)
(Norway), 1996201
NIH Workshop, 1%* (2.22 g ALA) 0.3%* (0.65 g/day)
1999202
ANC 2000, 5:1 2-2.5 g/day 0.12 g/day (DHA)
France203
FDA, 2000204 1 g/day
AHA, 2000205 0.9 g/day
The Japanese Society of Nutrition 4 :1
and Food Science, RDA for the
Japanese, 6th revision 2000
*% calorie intake.

Table 2: PUFA recommended dietary allowances

36
VIII. Conclusion
The great number of studies on omega-3 LC PUFAs reflect the
interest aroused by this fatty acid family in the scientific community.
The studies have shown the importance of omega-3 LC PUFAs in
the diet, even though these micronutrients are not considered
essential as they are synthesized in low amounts by the body. They
are critical to brain development and function. They are therefore
particularly important in pregnant and breastfeeding women and in
infants.

Clinical data and secondary intervention studies have provided

evidence for the prophylactic potential of omega-3 LC PUFAs in
various conditions, such as cardiovascular diseases, chronic
inflammatory states, and some cancers. The clinical data have a
biochemical and molecular explanation. On the one hand, omega-3
LC PUFAs are the precursors of the eicosanoids involved in immune
and inflammatory reactions; on the other, they act on the expression
of target genes encoding proteins involved in fatty acid metabolism
– an action which partly accounts for their role in the prevention of
cardiovascular diseases and some cancers.

Our occidental type of diet provides a surplus of saturated fats and

omega-6 fatty acids without meeting our requirements for omega-3
fatty acids, and for omega-3 LC PUFAs in particular. The omega-3/
omega-6 ratio must therefore be corrected, and the overall lipid
intake decreased. The eventual result of such a change in dietary
habits should be an impact on the incidence of cardiovascular,
inflammatory and malignant diseases.

In the particular case of cardiovascular diseases, an increase in

omega-3 LC fatty acid intake can be expected to moderate the main
risk factors and hence delay the requirement for drugs whose long-
term effects are not always fully understood. Once such drugs
37
become necessary, synergistic dietary therapy with omega-3 LC
PUFA supplementation should help to lower the drug doses and
hence minimize side effects.

The growing interest in omega-3 LC PUFAs appears wholly justified.

They have a particular place in new nutritional strategies for cardio-
vascular risk reduction.

38
■ References
1. Givens D.I. et al., Sources of n-3 polyunsaturated fatty acids
additional to fish oil for livestock diets. A review. in Nutrition
Abstracts and Reviews: Series B. Livestock feed and feeding,
2000, 1-19.
2. Kromann N. et al., Epidemiological studies in the Upernavik
district, Greenland. Acta Med. Scand., 1980, 208: 401-406.
3. Bjerregaard P. et al., Mortality from ischaemic heart disease and
cerebrovascular disease in Greenland. Int. J. Epidemiol., 1988,
17: 514-519.
4. Hirai A. et al., Effect of oral administration of highly purified
eicosapentaenoic acid and docosahexaenoic acid on platelet
function and serum lipid in hyperlipidemic patients. Adv. Prosta-
glandin Thromboxane Leukot. Res., 1989, 19: 627-630.
5. Kagawa Y. et al., Eicosapolyenoic acids of serum lipids of
Japanese islanders with low incidence of cardiovascular diseases.
J. Nutr. Sci. Vitaminol., 1982, 28: 441-453.
6. Kromhout D. et al., The inverse relation between fish consump-
tion and 20 years mortality from coronary heart disease. N. Engl.
J. Med., 1985, 312: 1205-1209.
7. Daviglus M. et al., Benefit of a favorable cardiovascular risk-
factor profile in middle age with respect to Medicare costs. N.
Engl. J. Med., 1998, 339: 1122-1129.
8. Dolecek T. A. et al., Dietary polyunsaturated fatty acids and
mortality in the multiple risk factor intervention trial (MRFIT). in
Health effects of w-3 polyunsaturated fatty acids in seafoods,
1991, 205-216.
9. Rodriguez B.L. et al., Fish intake may limit the increase in risk of
coronary heart disease morbidity and mortality among heavy
smokers: The Honolulu Heart Program. Circulation, 1996, 94:
952-956.
10. Albert C. et al., Fish consumption and risk of sudden cardiac
death. JAMA, 1998, 7: 23-28.
39
11. Siscovick D.S. et al., Dietary intake and cell membrane levels of
long-chain n-3 polyunsaturated fatty acids and the risk of
primary cardiac arrest. JAMA, 1995, 274: 1363-1367.
12. Ascherio A. et al., Dietary intake of marine n-3 fatty acids, fish
intake, and the risk of coronary disease among men. N. Engl. J.
Med., 1995, 332: 997-982.
13. Curb J.D. et al., Fish consumption and mortality from coronary
heart disease. N. Engl. J. Med., 1985, 313: 821.
14. Vollset N., Fish consumption and mortality from coronary heart
disease. N. Engl. J. Med., 1985, 313: 820-821.
15. Burr M.L. et al., Effect of changes in fat, fish, and fibre intakes
on death and myocardial reinfarction: diet and reinfarction trial
(DART). Lancet, 1989, 2: 757-761.
16. Trial G.P., Dietary supplementation with n-3 polyunsaturated
fatty acids and vitamin E after myocardial infarction: results of
the GISSI-Prevenzione trial. Lancet, 1999, 354: 447-455.
17. von Schaky C. et al., The effect of dietary w-3 fatty acids on
coronary atherosclerosis. A randomized double-blind, placebo-
controlled trial. Ann. Intern. Med., 1999, 130: 554-562.
18. Schmidt E.B. et al., Omega-3 fatty acids. Current status in
cardiovascular medecine. Drugs, 1994, 47: 405-424.
19. O'Keefe J. et al., From Inuit to implementation: omega-3 fatty
acids come of age. Mayo Clin. Proc., 2000, 75: 607-614.
20. Lusis A.J., Atherosclerosis. Nature, 2000, 407: 233-241.
21. Leaf A. et al., Cardiovascular effects of n-3 fatty acids. N. Engl.
J. Med., 1988, 318: 549-557.
22. Gerster H., The use of n-3 PUFAs (fish oil) in enteral nutrition.
Int. J. Vitam. Nutr. Res., 1995, 65: 3-20.
23. Lemaitre D. et al., Effects of fatty acids on human platelet
glutathione peroxidase: possible role of oxidative stress.
Biochem. Pharmacol., 1997, 53: 479-486.
24. Krämer H.J. et al., Fish oil, fatty acids and human platelets: Dose-
dependent decrease in dienoic and increase in trienoic thrombo-
xane generation. Biochem. Pharmacol., 1996, 52: 1211-1217.
40
25. Eritsland J. et al., Long-term effects of n-3 polyunsaturated fatty
acids on haemostatic variables and bleeding episodes in patients
with coronary artery disease. Blood Coagul. Fibrinolysis, 1995, 6:
17-22.
26. Steering Committee of the Physicians' Health Study Research
Group, Final report on the aspirin component of the ongoing
Physicians' Health Study. N. Engl. J. Med., 1989, 321: 129-135.
27. Dyerberg J. et al., Haemostatic function and platelet polyun-
saturated fatty acids in Eskimos. Lancet, 1979, 2:433-435.
28. von Schacky C. et al., Long-term effects of dietary marine w-3
fatty acids upon plasma and cellular lipids, platelet function and
eicosanoids formation in human. J. Clin. Invest., 1985, 76:
1626-1631.
29. Freese R. et al., a-Linolenic acid and marine long-chain n-3 fatty
acids differ slightly in their effects on hemostatic factors in
healthy subjects. Am. J. Clin. Nutr., 1997, 66: 591-598.
30. Brown A.J. et al., Fish and fish oil intake: effect on haematolo-
gical variables related to cardiovascular disease. Thromb. Res.,
1991, 64: 169-178.
31. Levine P. et al., Dietary supplementation with omega-3 fatty
acids prolongs platelet survival in hyperlipidemic patients with
atherosclerosis. Arch. Intern. Med., 1989, 149: 1113-1116.
32. Véricel V. et al., The influence of low intake of n-3 fatty acids on
platelets in elderly people. Atherosclerosis, 1999, 147: 187-192.
33. Agren J. et al., Fish diet, fish oil and docosahexaenoic acid rich
oil lower fasting and postprandial plasma lipid levels. Eur. J. Clin.
Nutr., 1996, 50: 765-771.
34. Li X. et al., Dose response of dietary fish oil supplementations
on platelet adhesion. Arterioscler. Thromb., 1990, 11: 39-46.
35. Gerster H., n-3 Fish oil polyunsaturated fatty acids and bleeding.
J. Nutr. Environ. Med., 1995, 5: 281-296.
36. Knapp H., Omega-3 fatty acids, endogenous prostaglandins and
blood pressure regulation in humans. Nutr. Rev., 1989, 47: 301-
313.
41
37. Morris M.C. et al., Does fish oil lower blood pressure? A meta-
analysis of controlled trials. Circulation, 1992, 88: 523-533.
38. Appel L.J. et al., Does supplementation of diet with fish oil
reduce blood pressure? Arch. Intern. Med., 1993, 153: 1429-
1438.
39. Knapp H.R., Polyunsaturates, endogenous eicosanoids, and
cardiovascular disease. J. Am. Coll. Nutr., 1990, 9: 344-351.
40. Gerster H., Fish oil (n-3 long-chain PUFAs) and blood pressure.
Nutrition, 1993, 17: 15-24.
41. Singer P. et al., Lipid and blood presure-lowering effect of
mackerel diet in man. Atherosclerosis, 1983, 49: 99-108.
42. Singer P. et al., Fish oil amplifies the effect of propranolol in mild
essential hypertension. Hypertension, 1990, 16: 682-691.
43. Rogers S. et al., Effects of fish oil supplement on serum lipids,
blood pressure, bleeding time, haemostatic and rheological
variables. A double blind randomised controlled trial in healthy
volunteers. Atherosclerosis, 1987, 63: 137-143.
44. Haglund O. et al., Effect of a new fluid fish oil concentrate,
eskimo-3, on triglyceride, cholesterol, fibrinogen and blood
pressure. J. Int. Med., 1990, 227: 347-353.
45. Blonk M. et al., Dose-response effects of fish-oil supplementa-
tion in healthy volunteers. Am. J. Clin. Nutr., 1990, 52: 120-127.
46. Bonaa K. et al., Effect of eicosapentaenoic and docosahexaenoic
acids on blood pressure in hypertension. A population-based
intervention trial from the Tromso study. N. Engl. J. Med., 1990,
322: 795-801.
47. De Bruin T. et al., Postprandial decrease in HDL cholesterol
and HDL apo A-I in normal subjects in relation to triglyceride
metabolism. Am. J. Physiol., 1991, 260: E492-E498.
48. Williams C., Dietary interventions affecting chylomicron and
chylomicron remnant clearance. Atherosclerosis, 1998, 141:
S87-S92.
49. Griffin B. et al., Influence of dietary fatty acids on the athero-
genic lipoprotein phenotype. Nutr. Res. Rev., 1995, 8: 1-26.
42
50. Roche H. et al., Postprandial triacylglycerolaemia: nutritional
implications. Prog. Lipid Res., 1995, 34: 249-266.
51. Sethi S. et al., Postprandial lipoprotein metabolism. Nutr. Res.
Rev., 1993, 6: 161-183.
52. Criqui M. et al., Plasma triglyceride level and mortality from
coronary heart disease. N. Engl. J. Med., 1993, 328: 1220-1225.
53. Manninen V. et al., Joint effects of serum triglyceride and LDL
cholesterol and HDL cholesterol concentrations on coronary
heart disease risk in the Helsinki Heart Study. Implications for
treatment. Circulation, 1991, 85: 37-45.
54. Gardner C. et al., Association of small low-density lipoprotein
particles with the incidence of coronary artery disease in men
and women. JAMA, 1996, 18: 875-881.
55. Stampfer M.J. et al., A prospective study of triglyceride level,
low-density lipoprotein particle diameter and risk of myocardial
infarction. JAMA, 1996, 276: 882-888.
56. Austin M. et al., Epidemiology of triglycerides, small dense low-
density lipoprotein and lipoprotein as risk factors for coronary
heart disease. Lipid Disorders, 1994, 78: 99-115.
57. Miller M. et al., Normal triglyceride levels and coronary artery
disease events: The Baltimore coronary observational long-term
study. J. Am. Coll. Cardiol., 1998, 31: 1252-1257.
58. NIH Consensus Conference, Triglyceride, high-density lipopro-
tein, and coronary heart disease. NIH Consensus Development
Panel on Triglyceride, High-Density Lipoprotein and Coronary
Heart Disease. JAMA, 1993, 269: 505-510.
59. Roche H. et al., Postprandial coagulation factor VII activity: The
effect of monounsaturated fatty acids. Br. J. Nutr., 1997, 77:
537-549.
60. Clifton P. et al., Effect of dietary cholesterol on postprandial lipo-
proteins in three phenotypic groups. Am. J. Clin. Nutr., 1996,
64: 361-367.
61. Hokanson J. et al., Plasma trigyceride level is a factor for cardio-
vascular disease independent of high-density lipoprotein cholest-
43
 erol level: a meta-analysis of population-based prospective
studies. J. Cardiovasc. Risk, 1996, 3: 213-219.
62. Miller M., Is hypertriglyceridaemia an independent risk factor for
coronary heart disease? Eur. Heart J., 1998, 19: H18-H22.
63. Weber P. et al., Triglyceride-lowering effect of omega-3 LC-poly-
unsaturated fatty acids. A review. Nutr. Metab. Cardiovasc. Dis.,
2000, 10: 28-37.
64. Nenseter M. et al., Effect of dietary supplementation with n-3
polyunsaturated fatty acids on physical properties and metabo-
lism of low density lipoprotein in humans. Atheroscler. Thromb.,
1991, 12: 369-379.
65. Keller H. et al., Fatty acids and retinoids control lipid metabolism
through activation of peroxisome proliferator-activated receptor-
retinoid X receptor heterodimers. Proc. Natl. Acad. Sci. U S A,
1993, 90: 2160-2164.
66. Gottlicher M. et al., Fatty acids activate a chimera of the clofibric
acid-activated receptor and the glucocorticoid receptor. Proc.
Natl. Acad. Sci. U S A, 1992, 89: 4653-4657.
67. Bordin P. et al., Effects of fish oil supplementation on apolipo-
protein B100 production and lipoprotein metabolism in normoli-
pidaemic males. Eur. J. Clin. Nutr., 1998, 58: 104-109.
68. Haglund O. et al., Effects of fish oil alone and combined with
long chain (n-6) fatty acids on some coronary risk factors in male
subjects. J. Nutr. Biochem., 1998, 9: 629-635.
69. Lu G. et al., Omega-3 fatty acids alter lipoprotein subfraction
distributions and the in vitro conversion of very low density lipo-
proteins to low density lipoproteins. J. Nutr. Biochem., 1999, 10:
151-158.
70. Clarke S.D. et al., Peroxisome proliferator-activated receptors: a
family of lipid-activated transcription factors. Am. J. Clin. Nutr.,
1999, 70: 566-571.
71. Clarke S. et al., Polyunsaturated fatty acid regulation of hepatic
gene transcription. J. Nutr., 1996, 126: 1105S-1109S.

44
72. Forman B.M. et al., Hypolipidemic drugs, polyunsaturated fatty
acids, and eicosanoids are ligands for peroxisome proliferator-
activated receptors alpha and delta. Proc. Natl. Acad. Sci. U S A,
1997, 94: 4312-4317.
73. Kliewer S.A. et al., Fatty acids and eicosanoids regulate gene
expression through direct interactions with peroxisome prolife-
rator-activated receptors alpha and gamma. Proc. Natl. Acad.
Sci. U S A, 1997, 94: 4318-4323.
74. Jump D. et al., Dietary polyunsaturated fatty acid regulation of
gene transcription. Prog. Lipid Rev., 1996, 35: 227-241.
75. Williams C., Postprandial lipid metabolism: effects of dietary
fatty acids. Proc. Nutr. Soc., 1997, 56: 679-692.
76. Roche H. et al., Long-chain n-3 polyunsaturated fatty acids and
triacylglycerol metabolism in the postprandial state. Lipids, 1999,
34: S259-S265.
77. Zampellas A. et al., Polyunsaturated fatty acids of the n-6 and
n-3 series: effects on postprandial lipid and apolipoprotein levels
in healthy men. Eur. J. Clin. Nutr., 1994, 48: 842-848.
78. Harris W.S. et al., Effects of low saturated fat, low cholesterol
fish oil supplement in hypertriglyceridemic patients. A placebo-
controlled trial. Ann. Intern. Med., 1988, 109: 465-470.
79. Harris W. et al., Fish oils in hypertriglyceriemia: a dose-response
study. Am. J. Clin. Nutr., 1990, 51: 399-406.
80. Radack K. et al., n-3 Fatty acid effects on lipids, lipoproteins,
and apolipoproteins at very low doses: results of a randomized
controlled trial in hypertriglyceridemic subjects. Am. J. Clin.
Nutr., 1990, 51: 599-605.
81. Beil F. et al., Dietary fish oil lowers lipoprotein in primary hyper-
triglyceridemia. Atherosclerosis, 1991, 90: 95-97.
82. Sirtori C. et al., One-year treatment with ethyl esters of omega-
3. Atherosclerosis, 1998, 137: 419-427.
83. Sorensen N. et al., Effect of fish-oil-enriched margarine on plasma
lipids, low-density- lipoprotein particle composition, size, and
susceptibility to oxidation. Am. J. Clin. Nutr., 1998, 68: 235-241.
45
84. Marckmann P. et al., Dietary fish oil (4g daily) and cardiovascular
risk markers in healthy men. Atheroscler. Thromb. Vasc. Biol.,
1997, 17: 3384-3391.
85. Visioli F. et al., Very low intakes of n-3 fatty acids incorporated
into bovine milk reduce plasma triacylglycerol and increase HDL-
cholesterol concentrations in healthy subjects. Pharmacol. Res.,
2000, 41: 571-576.
86. Hansen J.B. et al., Effects of highly purified eicosapentaenoic
acid and docosahexaenoic acid on fatty acid absorption, incor-
poration into serum phospholipids and postprandial triglyceri-
demia. Lipids, 1998, 33: 131-138.
87. Gudbjarnason S., Dynamics of n-3 and n-6 fatty acids in phos-
pholipids of heart muscle. J. Int. Med., 1989, 225: 117-128.
88. Sellmayer A.M.D. et al., Effects of dietary fish oil on ventricular
premature complexes. Am. J. Cardiol., 1995, 76: 974-977.
89. Christensen J. et al., n-3 Fatty acids and ventricular extrasystoles
in patients with ventricular tachyarrhythmias. Nutr. Res., 1995,
15: 1-8.
90. Christensen J.H. et al., Effect of fish oil on heart rate variability
in survivors of myocardial infarction: a double blind randomised
controlled trial. BMJ, 1996, 312: 677-678.
91. The Cardiac Arrhythmia Pilot Study (CAPS) investigators., Effect
of encainide, flecainide, imipramine and moricizine on ventri-
cular arrhythmias during the year after acute myocardial infarc-
tion: The CAPS. Am. J. Cardiol., 1988, 61: 501-509.
92. Grynberg A. et al., Acides gras omega-3 et prévention cardio-
vasculaire. Cah. Nutr. Diét., 1997, 32: 107-114.
93. Kang J. et al., Antiarrhythmic effects of polyunsaturated fatty
acids. Recent studies. Circulation, 1996, 94: 1174-1780.
94. Nair S.S. et al., Prevention of cardiac arrhythmia by dietary (n-3)
polyunsaturated fatty acids and their mechanism of action.
J. Nutr., 1997, 127: 383-393.
95. Kang J.X. et al., Prevention of fatal cardiac arrhythmias by poly-
unsaturated fatty acids. Am. J. Clin. Nutr., 2000, 71: 202-207.
46
 96. Siebert B.D. et al., Cardiac arrhythmia in rats in response to
dietary n-3 fatty acids from red meat, fish oil and canola oil.
Nutr. Res., 1993, 13: 1407-1418.
97. Pepe S. et al., Dietary fish oil confers direct antiarrhythmic
properties on the myocardium of rats. J. Nutr., 1996, 126: 34-
42.
98. Durot I. et al., Influence of phospholipid long chain polyun-
saturated fatty acid composition on neonatal rat cardiomyocyte
function in physical conditions and during glucose-free
hypoxia-reoxygenation. Mol. Cell. Biochem., 1997, 175: 253-
262.
99. Grynberg A. et al., Membrane docosahexaenoic acid vs. eico-
sapentaenoic acid and the beating function of the cardiomyo-
cyte and its regulation through the adrenergic receptors. Lipids,
1996, 31: S205-S210.
100. Delerive P. et al., Hypoxia-reoxygenation and polyunsaturated
fatty acids modulate adrenergic functions in cultured cardio-
myocytes. J. Mol. Cell. Cardiol., 1999, 31: 377-386.
101. Oudot F. et al., Eicosanoid synthesis in cardiomyocytes:
influence of hypoxia, reoxygenation and polyunsaturated fatty
acids. Am. J. Physiol., 1995, 268: H308-H315.
102. Leaf A. et al., Experimental studies on antiarrhythmic and anti-
seizure effects of polyunsaturated fatty acids in excitable
tissues. J. Nutr. Biochem., 1999, 10: 440-448.
103. Calder P.C., n-3 Polyunsaturated fatty acids and cytokine
production in health and disease. Ann. Nutr. Metab., 1997, 41:
203-234.
104. Endres S. et al., The effect of dietary supplementation with n-3
polyunsaturated fatty acids on the synthesis of interleukin-1
and tumor necrosis factor by mononuclear cells. N. Engl. J.
Med., 1989, 320: 265-271.
105. Denzlinger C. et al., Modulation of the endogenous leuko-
triene production by fish oil and vitamin E. J. Lipid Mediat. Cell
Signal., 1995, 11: 119-132.
47
106. James M. et al., Dietary polyunsaturated fatty acids and inflam-
matory mediator production. Am. J. Clin. Nutr., 2000, 71:
343S-348S.
107. Horrobin D.F., Low prevalences of coronary heart disease (CHD),
psoriasis, asthma and rheumatoid arthritis in Eskimos: are they
caused by high dietary intake of eicosapentaenoic acid (EPA), a
genic variation of essential fatty acid (EFA) metabolism or a
combination of both? Med. Hypotheses, 1987, 22: 421-428.
108. Recht L. et al., Hand handicap and rheumatoid arthritis in a
fish-eating society (the Faroe Islands). J. Int. Med., 1990, 227:
49-55.
109. Shapiro J. et al., Diet and rheumatoid arthritis in women: a
possible protective effect of fish consumption. Epidemiology,
1996, 7: 256-263.
110. Kremer J.M., n-3 Fatty acid supplements in rheumatoid
arthritis. Am. J. Clin. Nutr., 2000, 71: 349-351.
111. van der Temple et al., Effects of fish oil supplementation in
rheumatoid arthritis. Ann. Rheum. Dis., 1990, 49: 76-80.
112. Sköldstam L. et al., Effect of six months of fish oil supplemen-
tation in stable rheumatoid arthritis. A double-blind, controlled
study. Scand. J. Rheumatol., 1992, 21: 178-185.
113. Lau C. et al., Effects of fish oil supplementation on non stero-
idal anti-inflammatory drug requirement in patients with mild
rheumatoid arthritis - A double-blind placebo controlled study.
Br. J. Rheumatol., 1993, 32: 982-989.
114. Kremer J., Effects of modulation of inflammatory and immune
parameters in patients with rheumatic and inflammatory
disease receiving dietary supplementation of n-3 and n-6 fatty
acids. Lipids, 1996, 31: S243-S247.
115. Volker D. et al., Dietary n-3 fatty acid supplementation in rheu-
matoid arthritis-mechanisms, clinical outcomes, controversies,
and future directions. J. Clin. Biochem. Nutr., 1996, 20: 83-97.
116. Ariza-Ariza R. et al., Omega-3 fatty acids in rheumatoid
arthritis: an overview. Sem. Arthritis Rheum., 1998, 27: 366-370.
48
117. Curtis C.L. et al., n-3 Fatty Acids specifically modulate catabolic
factors involved in articular cartilage degradation. J. Biol.
Chem., 2000, 275: 721-724.
118. Fortin P. et al., Validation of a meta-analysis: The effects of fish
oil in rheumatoid arthritis. J. Clin. Epidemiol., 1995, 48: 1379-
1390.
119. Kremer J.M. et al., Effects of high-dose fish oil on rheumatoid
arthritis after stopping nonsteroidal antiinflammatory drugs:
clinical and immune correlates. Arthritis Rheum., 1995, 38:
1107-1114.
120. Kjeldsen-Kragh J. et al., Dietary omega-3 fatty acid supplemen-
tation and naproxen treatment in patients with rheumatoid
arthritis. J. Rheumatol., 1992, 19: 1531-1536.
121. Shoda R. et al., Epidemiologic analysis of Crohn disease in
Japan: increased dietary intake of n-6 polyunsaturated fatty
acids and animal protein relates to the increased incidence of
Crohn disease in Japan. Am. J. Clin. Nutr., 1996, 63: 741-745.
122. Salomon P. et al., Treatment of ulcerative colitis with fish oil n-3
fatty acid: an open trial. J. Clin. Gastroenterol., 1990, 12: 157-161.
123. Aslan A. et al., Fish oil fatty acid supplementation in active
ulcerative colitis: a double blind, placebo-controlled, crossover
study. Am. J. Gastroenterol., 1992, 87: 432-437.
124. Loeschke K. et al., n-3 Fatty Acids only delay early relapse of
ulcerative colitis in remission. Dig. Dis. Sci., 1996, 41: 2087-
2094.
125. Stenson W. et al., Dietary supplementation with fish oil in ulce-
rative colitis. Ann. Intern. Med., 1992, 116: 609-614.
126. Grimminger F. et al., Influence of intravenous n-3 lipid supple-
mentation on fatty acid profiles and lipid mediator generation
in a patient with severe ulcerative colitis. Eur. J. Clin. Invest.,
1993, 23: 706-715.
127. Belluzzi A. et al., Effect of an enteric-coated fish-oil preparation
on relapses in Crohn's disease. N. Engl. J. Med., 1557, 334:
1557-1560.
49
128. Lorenz-Meyer H. et al., Omega-3 fatty acids and low carbohy-
drate diet for maintenance of remission in Crohn disease. A
randomized multicenter trial. Scand. J. Gastroenterol., 1996,
31: 778-785.
129. Belluzzi A. et al., Polyunsaturated fatty acids and inflammatory
bowel disease. Am. J. Clin. Nutr., 2000, 71: 339-342.
130. Bougnoux P., n-3 Polyunsaturated fatty acids and cancer. Curr.
Opin. Clin. Nutr. Metab. Care, 1999, 2: 121-126.
131. Rose D., Dietary fatty acids and cancer. Am. J. Clin. Nutr.,
1997, 66: 998-1003.
132. Ziegler R. et al., Migration patterns and breast cancer risk in
Asian-American women. J. Nat. Cancer Inst., 1993, 85: 1819-
1827.
133. Hirayama T., Epidemiology of breast cancer with special refe-
rence to the role of diet. Prev. Med., 1978, 7: 173-195.
134. Lanier A. et al., Cancer incidence in Alaska natives. Int. J.
Cancer, 1976, 18: 409-412.
135. Landymore R.W. et al., Effects of low-dose marine oils on
intimal hyperplasia in autologous vein grafts. J. Thorac. Cardio-
vasc. Surg., 1996, 98: 788-791.
136. Noguchi M. et al., Chemoprevention of DMBA-induced
mammary carcinogenesis in rats by low dose EPA and DHA. Br.
J. Cancer, 1997, 75: 348-353.
137. Kim D.Y. et al., Stimulatory effects of high-fat diets on colon
cell proliferation depend on the type of dietary fat and site of
the colon. Nutr. Cancer, 1998, 30: 118-123.
138. Paulsen J.E. et al., A fish oil-derived concentrate enriched in
eicosapentaenoic and docosahexaenoic acid as ethyl esters
inhibits the formation and growth of aberrant crypt foci in rat
colon. Pharmacol. Toxicol., 1998, 82: 28-33.
139. Calder P.C. et al., Dietary fish oil suppresses human colon
tumour growth in athymic mice. Clin. Sci., 1998, 94: 303-311.
140. Calviello G. et al., Dietary supplementation with eicosapentae-
noic and docosahexaenoic acid inhibits growth of Morris hepa-
50
 tocarcinoma 3924A in rats: Effects on proliferation and
apoptosis. Int. J. Cancer, 1998, 75: 699-705.
141. Sensaki H. et al., Dietary effects of fatty acids on growth and
metastasis of KPL-1 human breast cancer cells in vivo and in
vitro. Anticancer Res., 1998, 18: 1621-1628.
142. Connolly J.M. et al., Effects of dietary menhaden oil, soy and a
cyclooxygenase inhibitor on human breast cancer cell growth
and metastasis in nude mice. Nutr. Cancer, 1997, 29: 48-54.
143. Hubbard N. et al., Alteration of murine mammary tumorige-
nesis by dietary enrichment with n-3 fatty acids in fish oil.
Cancer Lett., 1998, 124: 1-7.
144. Rose D. et al., Omega-3 fatty acids as cancer chemopreventive
agents. Pharmacol. Ther., 1999, 83: 217-244.
145. Willett W., Specific fatty acids and risks of breast and prostate
cancer: dietary intake. Am. J. Clin. Nutr., 1997, 66: 1557-1563.
146. Sasaki S. et al., An ecological study of the relationship between
dietary fat intake and breast cancer mortality. Prev. Med.,
1993, 22: 187-202.
147. Simonsen N. et al., Adipose tissue omega-3 and omega-6 fatty
acid content and breast cancer in the EURAMIC study. Am. J.
Epidemiol., 1998, 147: 342-352.
148. Simopoulos A., Omega-3 fatty acids in health and disease and
in growth and development. Am. J. Clin. Nutr., 1991, 54: 438-
463.
149. Sasaki T. et al., Effects of dietary n-3 to n-6 polyunsaturated
fatty acid ratio on mammary carcinogenesis in rats. Nutrition
Cancer, 1998, 30: 137-143.
150. Hawkins R.A. et al., Apoptotic death of pancreatic cancer cells
induced by polyunsaturated fatty acids varies with double bond
number and involves an oxidative mechanism. J. Pathol., 1998,
185: 61-70.
151. Noding R. et al., Effects of polyunsaturated fatty acids and their
n-6 hydroperoxides on growth of five malignant cell lines and
the significance of culture media. Lipids, 1998, 33: 285-293.
51
152. Lhuillery C. et al., Suppresion of the promoter effect of polyun-
saturated fatty acids by the absence of dietary vitamin E in
experimental mammary carcinoma. Cancer Letters, 1997, 114:
233-234.
153. Timmer-Bosscha H. et al., Differential effects of all-trans-reti-
noic acid, docosahexaenoic acid, and hexadecylphosphocholine
on platin-induced cytotoxicity and apoptosis in a cisplatin-sensi-
tive and resistant human embryonal carcinoma cell line. Cancer
Chemother. Pharmacol., 1998, 41: 469-476.
154. Bougnoux G., E. et al., Cytotoxic drugs efficacy correlates with
adipose tissue docosahexaenoic acid level in locally advanced
breast carcinoma. Br. J. Cancer, 1999, 79: 1765-1769.
155. Germain E. et al., Enhancement of doxorubicin cytotoxicity by
polyunsaturated fatty acids in the human breast tumor cell line
MDA-MB-231: Relationship to lipid peroxidation. Int. J. Cancer,
1998, 75: 578-583.
156. Bazan N. et al., Metabolism of arachidonic and docosahexae-
noic acids in the retina. Prog. Lipid Res., 1986, 25: 595-606.
157. Clandinin M.T. et al., Intrauterine fatty acid accretion rates in
human brain: implications for fatty acid requirements. Early
Human Dev., 1980, 4/2: 121-129.
158. Martinez M., Tissue levels of polyunsaturated fatty acids during
early human development. J. Pediatr., 1992, 4: S129-S138.
159. Lundbaek J. et al., Lysophospholipids modulate channel func-
tion by altering the mechanical properties of lipid bilayers. J.
Gen. Physiol., 1994, 104: 645-673.
160. Salem N. et al., The nervous system has an absolute molecular
species requirement for proper function. Mol. Membr. Biol.,
1995, 12: 131-134.
161. Fong T.M. et al., Correlation between acetylcholine receptor
function and structural properties of membranes. Biochemistry,
1986, 25: 830-840.
162. Malnoë A. et al., Effect of in vivo modulation of membrane
docosahexaenoic acid levels on the dopamine-dependent
52
 adenylate cyclase activity in the rat retina. J. Neurochem.,
1990, 55: 1480-1485.
163. Witt M.R. et al., Characterization of the influence of unsatu-
rated free fatty acids on brain GABA/benzodiazepine receptor
binding in vitro. J. Neurochem., 1994, 62: 1432-1439.
164. Miller B. et al., Potentiation of NMDA receptor currents by
arachidonic acid. Nature, 1992, 355: 722-725.
165. Delion S. et al., [alpha]-Linolenic acid dietary deficiency alters
age-related changes of dopaminergic and serotoninergic
neurotransmission in the rat frontal cortex. J. Neurochem.,
1996, 66: 1582-1591.
166. Hudson C. et al., CNS signal transduction in the pathophysio-
logy and pharmacotherapy of affective disorders and schizoph-
renia. Synapse, 1993, 13: 278-293.
167. Graber R. et al., Fatty acids and cell signal transduction. J. Lipid
Mediat. Cell Signal., 1994, 9: 91-116.
168. de la Presa Owens S. et al., Diverse, region-specific effects of
addition of arachidonic and docosahexanoic acids to formula
with low or adequate linoleic and alpha-linolenic acids on
piglet brain monoaminergic neurotransmitters. Pediatr. Res.,
2000, 48: 125-30.
169. McGahon B.M. et al., Age-related changes in synaptic func-
tion: analysis of the effect of dietary supplementation with w-3
fatty acids. Neuroscience, 1999, 94: 305-314.
170. Loudes C. et al., Release of immunoreactive TRH in serum-free
cultures of mouse hypothalamic cells. Develop. Brain Res.,
1983, 9: 231-234.
171. Bourre J.M. et al., Effect of polyunsaturated fatty acids on fetal
mouse brain cells in culture in a chemically defined medium.
J. Neurochem., 1983, 41: 1234--1242.
172. Sprecher H. et al., Reevaluation of the pathways for the biosyn-
thesis of polyunsaturated fatty acids. J. Lipid Res., 1995, 36:
2471-2477.

53
173. Bourre J.M. et al., Effect of increasing amounts of dietary fish
oil on brain and liver fatty composition. Biochim. Biophys. Acta,
1990, 1043: 149-152.
174. Philbrick D.J. et al., Ingestion of fish oil or a derived n-3 fatty
acid concentrate containing eicosapentaenoic acid (EPA) affects
fatty acid compositions of individual phospholipids of rat brain,
sciatic nerve and retina. J. Nutr., 1987, 117: 1663-1670.
175. Wainwright P., Nutrition and behaviour: the role of n-3 fatty
acids in cognitive function. Br. J. Nutr., 2000, 83: 337-339.
176. Zimmer L. et al., Chronic n-3 polyunsaturated fatty acid defi-
ciency alters dopamine vesicle density in the rat frontal cortex.
Neurosci. Lett., 2000, 284: 25-28.
177. Innis S., Essential fatty acids in growth and development. Prog.
Lipid Res., 1991, 30: 39-103.
178. Hibbeln J. et al., Do plasma polyunsaturates predict hostility
and depression? in Nutrition and fitness: Metabolic and beha-
vioral aspects in health and disease, 1997, Karger: New York,
175-186.
179. Fenton W. et al., Essential fatty acids, lipid membrane abnor-
malities and the diagnosis and treatment of schizophrenia. Biol.
Psychiatry, 1999, 47: 8-21.
180. Stoll A. et al., Omega-3 fatty acids in bipolar disorder. Arch.
Gen. Psychiatry, 1999, 56: 407-412.
181. Edwards R. et al., Omega-3 polyunsaturated fatty acid levels in
the diet and in red blood cell membranes of depressed
patients. J. Affect. Disord., 1998, 48: 149-155.
182. Peet M. et al., Depletion of omega-3 fatty acid levels in red
blood cell membranes of depressive patients. Biol. Psychiatry,
1998, 43: 315-319.
183. Maes M. et al., Lowered w3 polyunsaturated fatty acids in
serum phospholipids and cholesteryl esters of depressed
patients. Psychiatry Res., 1999, 85: 275-291.

54
184. Kalmijn S. et al., Polyunsaturated fatty acids, antioxidant and
cognitive function in very old men. Am. J. Epidemiol., 1997,
145: 33-41.
185. Feskens E.J. et al., A longitudinal study on diet, risk factors and
cardiovascular disease in an aging cohort: the Zutphen Study.
Neth. J. Cardiol., 1993, 6: 200-204.
186. Uauy R. et al., Role of essential fatty acids in the function of
the developing nervous system. Lipids, 1996, 31: S167-S176.
187. Birch E. et al., Dietary essential fatty acid supply and visual
acuity development. Invest. Ophthalmol. Vis. Sci., 1992, 33:
3242.
188. Decsi T. et al., Essential fatty acids in full term infants fed
breast milk or formula. Arch. Dis. Child., 1995, 72: F23-F28.
189. Uauy R. et al., Safety and efficacy of omega-3 fatty acids in the
nutrition of very low birth weight infants: soy oil and marine oil
supplementation of formula. J. Pediatr., 1994, 124: 612-620.
190. Kohn G. et al., Diet and the essential fatty acid status of term
infants. Acta Paediatr. Suppl., 1994, 402: 69-74.
191. Willatts P. et al., Effect of long chain polyunsaturated fatty
acids in infant formula on problem solving at 10 months of
age. Lancet, 1998, 352: 688-691.
192. Agostini C. et al., Neurodevelopmental quotient of healthy
term infants at 4 months and feeding practice: the role of
long-chain polyunsaturated fatty acids. Pediatr. Res., 1995, 38:
262-266.
193. PNUN, Nordishe Naringrekomendationer Andra Uplagen, The
National Food Agency, Sweden, 1989.
194. Simopoulos A. Summary of the NATO advanced research
workshop on dietary omega-3 and omega-6 fatty acids : biolo-
gical effects and nutritional essentiality. The Journal of Nutri-
tion, 1989, 119: 521-528.
195. Scientific Review committee, Nutrition recommendations
Ottawa: Minister of National Health and Welfare, Canada,
1990.
55
196. British Nutrition Foundation, Unsaturated fatty acids: Nutritional
and physiological significance. Task and Force Report. London,
Chapman and Hall, 1992.
197. Reports of the Scientific Committee for Food, Nutrient and
energy intakes for the European community, 1993, 52-60.
198. FAO/WHO, Fats and oils in human nutrition (Report of a joint
expert consultation), Fat and Nutrition paper 57, Rome: FAO,
1994.
199. Department of Health, Report on Health and Social Subjects,
N°41. Dietary reference values for food energy and nutrients
for the UK. Report of the Committee on Medical Aspects of
Food Policy, London, HMSO, 1991.
200. Department of Health, Report on Health and Social Subjects,
N°46. Nutritional aspect of cardiovascular disease. Report of
the Committee on Medical Aspects of Food Policy, London,
HMSO, 1994.
201. Nordic Nutrition Recommendation, Scandinavian J. Nutrition,
1996, 40: 161-165
202. Simopoulos A. et coll., Essentiality of and recommendated
dietary intakes for omega-6 and omega-3 fatty acids. Ann.
Nutr. Metab., 1999, 43: 127-130.
203. AFSSA-CNERNA-CNRS, Apports nutritionnels conseillés pour la
population française. eds. TEC&DOC, 2000.
204. Food and drug Administration, Letter regarding dietary supple-
ment health claim for omega-3 fatty acids and coronary heart
disease. http://vm.cfsan.fda.gov/~dms/ds-ltr11.html, 2000.
205. AHA dietary Guidelines, revision 2000: A statement for health
care professionals from the nutrition committee of the
American Heart Association. Circulation, 2000, 102:2296-
2311.

56
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