document: cardiovascular disease is the leading cause of mortality in many economically developed nations accounting for about 30% of all deaths   and its incidence is still increasing . 
   ongoing research aims to investigate and prevent the early development of cardiovascular risk factors such as atherosclerosis , hypertension , dyslipidemia , chronic inflammation , and insulin resistance . 
   the beneficial effects of n-3 polyunsaturated fatty acids ( n-3 pufas ) were proved in several observational and experimental studies . 
   the lipid lowering action of n3-pufas was detected at the beginning , so these nutrients were used for the treatment of dyslipidemic disorders . 
   their anti - inflammatory , antithrombotic , antiatherosclerotic , and antiarrhythmogenic effects were observed later . 
   low - grade chronic inflammation is now recognized as a prominent process in the development of atherosclerosis and coronary heart disease . 
   the induction of inflammation may well provide a link between hyperlipidemia and atherogenesis [ 2 , 3 ] . 
   atherosclerosis is now considered a  systemic disease  featured by low - grade arterial inflammatory lesions that can develop through the disease progression .   in physiological conditions , endothelial cells 
   synthesize and release adequate amounts of nitric oxide ( no ) and prostaglandins ( such as pge2 and pge3 ) and maintain a downstream balance between pro- and anti - inflammatory molecules . 
   however , in the presence of atherosclerosis this balance disrupts leading towards an increase production of proinflammatory cytokines as interleukins 1 , 2 , and 6 ( il-1 , 2 , and 6 ) and tumor necrosis factor  ( tnf- ) , with further progression of the disease . 
   these pro - inflammatory cytokines can induce oxidative stress by enhancing the production of reactive oxygen species ( ros ) by monocytes , macrophages , and leukocytes . 
   pufas and their eicosanoid derivatives may play a significant role modulating the inflammatory response ( figure 1 ) . 
   unsaturated fatty acids are referred to as pufas when two or more double bounds are present . 
   there are two pufas families , omega-3 ( n-3 ) and omega-6 ( n-6 ) fatty acids . 
   they differ in location of the last double bond relative to the terminal methyl end of the molecule . 
   the human body can produce almost all fatty acids , except linoleic acid ( la , c18:2 n-6 , precursor to the n-6 series of fatty acids ) and -linoleic acid ( ala , c18:3n-3 , precursor to the n-3 series of fatty acids ) . 
   these two pufas are named  essential fatty acids  because the body can not synthesize them . 
   endogenous conversion ( elongation and desaturation ) of the initial c18 pufa precursors results in the synthesis of longer - chain counterparts such as eicosapentaenoic acid ( epa ) and docosahexaenoic acid ( dha ) in the n-3 family and dihomo--linoleic acid ( dgla ) and arachidonic acid ( aa ) in the n-6 family . in humans 
   the biochemical pathways converting ala to epa and epa to dha are limited : 0.28% of ala is converted to epa ( generally more in women ) and 04% of ala to dha [ 812 ] . 
   results from pilot studies suggest that ala conversion is also limited to function as a surrogate for fish consumption . 
   thus , tissue and circulating epa and dha levels are primarily related to their dietary intake . 
   fish is the major food source of long - chain n-3 pufas , including epa , dha , and docosapentaenoic acid ( dpa ) , while ala is a plant n-3 fatty acid mainly found in seeds , nuts , and their oils . 
   thus , plant sources of n-3 fatty acids can not currently be considered as a replacement for seafood - derived n-3 pufas . 
   this suggests that n-3 fatty acids derived from different sources might have their own specific effects on cardiovascular risk markers . 
   linoleic acid is thought to decrease the conversion of ala into epa and dha by competing for the n-6-desaturase enzyme . 
   previous studies showed that genetic variations in this enzyme may be related to cardiovascular disease . furthermore , because the two fatty acid pathways are mutually exclusive ( i.e. , n-3 fatty acid can not become n-6 fatty acid and vice versa ) , a balanced intake of ala and la as precursors or of their longer - chain products epa , dha , and aa is required . 
   there is a vast amount of epidemiologic evidence of a cardioprotective effect of fish oil - derived epa and dha and it was confirmed in randomized controlled trials [ 1720 ] . 
   several intervention studies , such as the  gruppo italiano per lo studio della sopravvivenza nell'infarto miocardico  ( gissi)-prevenzione trial and the cardiovascular health study , have shown that an increased intake of eicosapentaenoic acid ( epa , c20:5n-3 ) and docosahexaenoic acid ( dha , c22:6n-3 ) lowers the risk of coronary heart disease ( chd ) [ 21 , 22 ] . 
   the first gissi - prevenzione trial , a randomized open label study in 11324 italian patients with recent myocardial infarction , demonstrated that patients had a 15% lower combined risk of mortality , nonfatal myocardial infarction , and stroke upon supplementation for 3.5 years with 850  mgday-1 of lc n-3 pufa . 
   the relative risk of cardiovascular mortality was also decreased by 30% and that of sudden death by 45% . 
   the gissi - prevenzione trial demonstrated that a significant protective effect could be obtained with doses much lower than those previously considered necessary for significant beneficial effects . 
   the second gissi - hf study , a randomized double - blind placebo - controlled trial in 6975 italian patients with chronic heart failure , revealed a moderate decrease in both all - cause mortality admissions to hospital for cardiovascular disease upon supplementation for an average of 3.9 years with 1  g of lc n-3 pufa daily . 
   again , the beneficial effects were seen in a population already treated with recommended therapies . 
   the  japan eicosapentaenoic acid ( epa ) lipid intervention study ,  ( jelis ) trial , was performed on 18645 japanese men and women with hypercholesterolaemia treated with statins . 
   supplementation with 1.8  g epa daily decreased major coronary events by 19% over 4.6 years . 
   this may be evident in japanese population where a consistent reduction of chd is observed probably because of a high background seafood intake [ 25 , 26 ] . in summary , a modest intake of lc 
   however , higher doses and longer duration of intervention , as reported for the jelis study , have the potential to protect from non - fatal chd events . 
   presently , there is evidence to suggest that the two lc n3-pufas  epa ( eicosapentaenoic acid , 20:5n3 ) and dha ( docosahexaenoic acid , 22:6n3)exert pleiotropism ( akin to those effects which have been reported for certain cardiovascular drugs ) by modulating a range of diverse target mechanisms involved in cardiovascular disease development . [ 2730 ] . 
   since the appearance of purified forms of dha in the market in the 1990s , researchers have started to investigate the differential effects of epa and dha on cardiovascular health . however , the number of human studies is still limited in this field and the independent effects of epa and dha on various cardiovascular outcomes are yet to be firmly established . 
   furthermore , the incorporation of epa and dha into the cell membrane influences its organisation , fluidity , and permeability , as well as the activity of transmembrane proteins , including receptors , enzymes , and ion channels . 
   both epa and dha modulate k , na , and ca channel activities in myocardial cells , regulating myocyte electrical excitability and contractility [ 3234 ] . 
   these effects are concentration dependent and appear to be mediated by the action of epa and dha on membrane fluidity , although other mechanisms , such as a direct binding of n-3 lcp to the channel , may be involved . 
   furthermore , animal studies support the increasing evidence that dha rather than epa is preferentially incorporated into the myocardial cell membrane . 
   collectively , these findings help to explain the antiarrhythmic and heart rate ( hr)-lowering effects observed by dha but not by epa in humans . in addition , incorporation of dha into the membrane of cardiomyocytes influences the beta adrenergic system to a greater extent than epa , that is , potentially another important mechanism involved in the hypotensive and anti - arrhythmic action of dha . 
   furthermore , dha incorporation into the membrane of endothelial cells stimulates atp release from the endothelium , increasing vasodilation by stimulating nitric oxide ( no ) release . 
   the induction of no release , together with the decrease in noradrenaline levels , may also account for the bp - lowering effect of dha . a recent meta - analysis including thirty randomized controlled trials showed that prolonged fish oil intake may reduce hr , especially in populations with a high baseline hr . in dyslipidemic males and postmenopausal women 
   , this decrease appears to be mediated by dha rather than epa [ 37 , 41 , 42 ] . 
   in contrast , no significant effect of either epa or dha on hr was observed in healthy males for similar dosage and treatment duration . 
   hr variability ( hrv ) is a strong predictor of cvd , including sudden cardiac death , arrhythmic chd , and atrial fibrillation . 
   fish oils have shown anti - arrhythmic properties in animal studies , and several clinical and epidemiological studies have reported an association between hrv and n-3 lcp blood cell levels and/or fish oil intake [ 4446 ] . 
   short - term trials ( 612 week ) in healthy individuals showed no or low effect of a daily intake of 1.23.6  g of ala on blood lipids , ldl - oxidation , lipoprotein ( a ) , and apolipoproteins a - i and b [ 8 , 47 ] . 
   long - term treatment with high - dose ala ( 40  g / daily ) showed a beneficial effect by reduction of body weight and blood ldl / total cholesterol ratio [ 47 , 48 ] . 
   previous evidence favored recommendation for modest dietary consumption of ala ( 2 - 3  g daily ) in the primary and secondary prevention of chd . 
   the recently published case - control study by campos et al .   shows a strong inverse associations between ala status - intake and nonfatal mi . 
   recent data support the assumption that ala may protect against atherosclerosis [ 50 , 51 ] . 
   however , data from two recent epidemiologic studies suggest that high tissue ala is related to an increased rather than decreased risk of fatal cardiovascular events   and sudden death . 
   also , n-3 pufas may interfere with cvd with other physiological effects : the reduction of triglyceride ( tg ) synthesis is well recognized . moreover 
   , the reduction of hepatic vldl synthesis may contribute to this protective effect reducing the availability of fatty acids for triglyceride synthesis due to the decreased 
   de novo  lipogenesis ( dnl ) , increasing the beta - oxidation of fatty acids , reducing the delivery of nonesterified fatty acids to the liver , reducing the activity of tg synthesis by hepatocytes , and finally increasing the hepatic synthesis of phospholipids [ 5459 ] .   in both experimental models and human studies , 
   a different effect was reported in some studies between epa and dha : both epa and dha produced a significant reduction of tgs and ldl - cholesterol , raising effects of fish oil [ 61 , 62 ] . 
   recently , buckley and coworkers   showed that a four - week supplementation with dha significantly reduced tg levels in normolipidemic human subjects by 22% , while epa decreased tg levels by 15% without reaching statistical significance . in another four - week interventional study , 
   both epa and dha reduced postprandial tg without affecting fasting tg levels in healthy human subjects . 
   however , epa and dha seemed to reduce triglyceridaemia to the same extent when given for a long - enough period [ 60 , 61 , 6772 ] . 
   studies in humans show that an average intake of n-3 pufas 3 - 4  g / daily decreases serum triacylglycerol concentration by 2530% in a dose - dependent manner . 
   the same intake does not affect total cholesterol but increases ldl cholesterol by 510% and hdl cholesterol by 13% . 
   the increase in ldl cholesterol is mainly due to a rise of the larger and the potentially less atherogenic ldl particles . 
   hypertension ( ah ) and dyslipidemia ( dlp ) are the most important and frequent risk factors for cvd in the general population , with a prevalence of 5080% according to different authors [ 7579 ] . 
   on the other hand almost half of the patients with increased total cholesterol ( tc ) have systo - diastolic hypertension [ 8083 ] . 
   modifications of erythrocyte - membrane fatty acids ( fa ) composition are an early indicator of the development of ah and lipid disorders . 
   thus , modifications of erythrocyte - membrane fa are fairly subtle indicators of lipid metabolism pathology which manifest themselves much earlier than changes in plasma lipoproteins . 
   modifications of fatty acids composition of the cell membrane lipid matrix play an important role in the ah pathogenesis . 
   changes of the fatty acids ( fa ) composition with decrease of essential pufas may result in increase of membrane microviscosity , activation of pro - inflammatory eicosanoids synthesis and increased sensitivity of the smooth muscle cells in artery walls to the influence of vasoconstrictors . 
   that is the reason of the great interest in the study of the spectrum of erythrocyte lipids in hypertensive dyslipidemic patients . 
   deficiency of n-3 pufa is a distinctive feature of the modification of erythrocyte - membrane fa in hypertensive dyslipidemic patients . 
   deficiency of endogenous n-3 pufas may lead to changes in physicochemical properties of cell membranes , activation of the synthesis of proinflammatory and vasoconstrictive eicosanoids , and finally induction of a systemic inflammatory syndrome . 
   thus , structural and functional changes in erythrocyte cell membranes develop in hypertension as a part of the common systemic disorder of lipid metabolism . 
   the fa disorganization of the cell membrane may cause the development of hyperlipidemia and the progression of hypertensive disease . 
   thus , an important role of fa in the pathogenesis of cardiovascular disease seems clear . 
   a large part of the cardioprotective actions of lc n-3-pufas is likely to be mediated by the vascular endothelium [ 28 , 8688 ] . 
   this thin monolayer of cells plays a central role in cardiovascular homeostasis and function via the production of a range of potent autocrine and paracrine biochemical mediators that control the tone of vascular smooth muscle cells [ 31 , 89 , 90 ] . 
   vascular endothelium is also the site for the inception , progression , and clinical manifestations of atherosclerosis [ 9193 ] . 
   multifactorial endothelial dysfunction induced by dlp , toxins , cigarette - smoking , and so on may be the early event in the development of atherosclerosis . 
   the endothelium becomes  proadhesive  and induces an increased adhesion of circulating monocytes that subsequently infiltrate the arterial intima . at this level endothelial cells or macrophages 
   release reactive oxygen species that oxidize circulating ldl into oxidized - ldl and create the  lipid streak  . in the evolution from the lipid streak to the atherosclerotic plaque , 
   numerous cytokines that cause infiltration of smooth muscle layer by leukocytes and fibroblasts and promote platelet adhesion are involved in turn   ( figures 2 and 3 ) . during the migration over the vessel wall 
   , monocytes are exposed to various proinflammatory stimuli that contribute to their differentiation into macrophages . 
   other macrophages induce a local inflammatory compartment within the vassel wall , that is , continuously powered by cytokines , activated t - helper cells and scavanger receptor activators . 
   the inhibition of this self - maintaining process may reduce atherogenesis as demonstrated in mutant mice deficient for macrophage colony stimulating factor ( m - csf ) or macrophage migration inhibitory factor ( mif ) [ 95 , 96 ] . in more advanced lesions , cell debris , apoptotic cells , and free cholesterol particles 
   accumulate in the lipid - rich necrotic core of the plaque , are sometimes separated from the blood only by a thin fibrous cap . 
   the number of inflammatory cells within the lesion is strongly associated with the risk of rupture of the so - called  vulnerable plaque .  at last , the process may propagate intraluminal thrombosis , thereby distal ischemia of heart or brain tissue [ 97 , 98 ] . through their positive antithrombotic , lipid - lowering , proendothelial functional activities , n-3 
   in fact supplementation of n-3 pufas improves flow - mediated arterial dilatation by an increased endothelial synthesis of nitric oxide [ 86 , 99111 ] . 
   several , although not all , clinical trials have also found that consumption of n-3 pufas lowers circulating markers of endothelial dysfunction , such as e - selectin , vascular cell adhesion molecule-1 , and intracellular adhesion molecule-1 [ 74 , 112 , 113 ] . 
   thus , normalization of endothelial function could partly mediate the protective effects of n-3 pufa against cvd . a recent randomized clinical trial established the role of pufas on plaque stability . 
   patients with carotid artery atherosclerosis were randomized to take placebo , fish oil ( n-3 pufas ) , or seed oil ( n-6 pufas ) . 
   subjects treated with n-3 pufas showed higher epa and dha concentration , reduction of monocytes and macrophages , and a thicker fibrous cap compared to controls and group treated with n-6 pufas , all changes that can enhance the stability of atherosclerotic plaques . 
   by contrast , increased assumption of n-6 pufas did not affect carotid plaque fatty acid composition or stability . 
   plaques stability may explain the reductions in nonfatal and fatal cardiovascular events , associated with increased n-3 pufas intake . 
   the triglyceride - lowering , antiatherogenic , antithrombotic , and anti - inflammatory effects of n-3 pufas were clearly demonstrated in human studies . 
   conversely , their anti - arrhytmogenic effect was showed only in vitro and in animal experiments , as confirmation in humans has been limited by the absence of reliable physiological measures or biomarkers to quantify antiarrhythmic potential . 
   inflammation is a physiological response to tissue trauma or infection , but leukocytes , which are the effector cells of the inflammatory process , have powerful capabilities of tissue remodelling . 
   thus , the passage of leukocytes from the bloodstream into inflamed tissue is tightly regulated to ensure their precise localization . 
   recruitment of circulating neutrophils into the tissue stroma occurs during the early phases of inflammation . in this process 
   , peptide agonists of the chemokine family are assumed to provide a chemotactic stimulus capable of supporting the migration of neutrophils across vascular endothelial cells , through the vessel wall and out into the tissue stroma . 
   although an initial chemokine stimulus is essential for the recruitment of flowing neutrophils by endothelial cells , stimulated by the inflammatory cytokine ( tnf- ) , transit across the endothelial monolayer is regulated by additional stimuli . a new step in the neutrophil recruitment process that relies upon a lipid - mediated signal to regulate the migration of neutrophils across endothelial cells 
   this signal is supplied by the metabolism of arachidonic acid into the eicosanoid prostaglandin - d2 ( pgd2 ) by cyclooxygenase ( cox ) enzymes . 
   this step in the neutrophil recruitment process was revealed when the epa was used as an alternative substrate for cox enzymes , leading to the generation of prostaglandin - d3 ( pgd3 ) [ 117120 ] . 
   this alternative eicosanoid inhibited the migration of neutrophils across endothelial cells competing for the pgd2 receptor [ 121 , 122 ] . 
   pgd2 signalling is subordinate to the chemokine - mediated activation of neutrophils , but without the sequential delivery of this signal , neutrophils fail to penetrate the endothelial cell monolayer . the ability of dietary epa to inhibit this process reveals an unsuspected level of regulation in the migration of inflammatory leukocytes . 
   this may contribute to clarify the interactions between pufas and the inflammatory system and direct research on novel therapeutic agents that target the inflammatory system with greater affinity and/or specificity than dietary supplements of n-3-pufas . 
   the inflammation healing mechanisms are of main interest , and in recent years , new endogenous anti - inflammatory and proresolving lipid mediators generated from pufas ( lipoxins , resolvins , protectin , and maresin ) were uncovered . 
   lipid mediator metabolomics of self - resolving inflammatory exudates recently highlighted a new family of potent anti - inflammatory and proresolving mediators . 
   serhan and coworkers identified families of novel bioactive mediators derived from arachidonic acid ( aa - derived lipoxins ) , eicosapentaenoic acid ( epa - derived e - series resolvins ) , and docosahexaenoic acid ( dha - derived d - series resolvins , protectin , and maresin ) [ 124135 ] . 
   these lipid mediators promote resolution through enhanced clearance of apoptotic pmns , chemokines , cytokines , and microbial products by macrophages [ 136 , 137 ] . 
   n-3 pufas may act through several mechanisms , for instance , by preventing conversion of the n-6 pufa arachidonic acid to pro - inflammatory eicosanoids and by their conversion to potent anti - inflammatory mediators such as resolvins [ 124126 , 138 ] . 
   a possible biological mechanism underlying the beneficial effects of lc n-3 pufas on inflammation and endothelial function is that they may compete with n-6 fatty acids for prostaglandin and leukotriene synthesis at the cyclooxygenase and lipoxygenase levels . 
   lc n-3 pufas from fish or fish oil modulate prostaglandin metabolism by increasing the active vasodilator and inhibitor of platelet aggregation prostaglandin e3 , the weak platelet aggregator and vasoconstrictor thromboxane a3 , and the weak inducer of inflammation leukotriene b5 . otherwise , the production of thromboxane a2 , a potent platelet aggregator and vasoconstrictor , and leukotriene b4 , an inducer of inflammation and a powerful inducer of leukocyte chemotaxis and adherence is reduced . another suggested mechanism is that lc n-3 pufas may bind reactive oxygen species because of their multiple double bonds and lead to a decreased production of hydrogen peroxide . 
   hydrogen peroxide is a critical activator of the nuclear factor-b system of transcription factors that controls the coordinated expression of adhesion molecules and of leukocyte - specific chemoattractants upon cytokine stimulation . 
   he and coworkers   found independent inverse associations of lc n-3 pufas and nonfried fish with il-6 , crp and mmp3 . 
   the relevance of these biomarkers of inflammatory and endothelial activation in the atherogenic process is well recognized . 
   previous investigations suggest that both crp and il-6 , two systemic inflammatory markers , are independent predictors of cvd and may play an important role in atherogenesis . 
   in addition , mmp3 is suggested as an independent prognostic factor in stable coronary artery disease . 
   soluble intercellular adhesion molecule-1 ( sicam-1 ) is thought to be a key factor in the adherence of monocytes to the endothelium and subsequent transmigration into the intima ( figure 2 ) . 
   the role of sicam-1 in the pathogenesis of inflammation and atherosclerosis has been confirmed in experimental models . 
   previous investigations also indicate that sicam-1 is an independent predictor of cvd apart of other traditional risk factors . 
   .   observed that fried fish but not non - fried fish consumption was significantly inversely related to sicam-1 . 
   this finding is not expected since the frying process may reduce the content of lc n-3 pufas and produce trans - fatty acids . 
   one possible explanation is that fried fish consumption may be a marker of relatively unhealthy lifestyle ; those who had high fried fish consumption were more likely to suffer from dyslipidemia and were likely to be under treatment with medications ( e.g. , statins ) , which may lower sicam-1 . however , the inverse association between fried fish intake and sicam-1 level remained significant when patients taking cholesterol - lowering medications were excluded . 
   lc n-3 pufas are believed to affect inflammatory processes mainly through two pathways : endothelial activation and changes in eicosanoid production , or a combination of the two . 
   the first , predicated on the observation that n-3 pufas regulate the transcription of endothelial cell inflammatory genes by downregulating the activity of the nuclear factor - kb , predicts changes in the levels of adhesion receptor and chemokine expression after supplementation with epa [ 149151 ] . 
   the second hypothesis proposes that upon endothelial cell activation , epa may compete with the n-6-pufa arachidonic acid ( aa ; 20:4n-6 ) for cyclooxygenase enzymes ( cox1 and cox2 ) after both fatty acids are released from membrane phospholipids by endogenous phospholipases . 
   a large amount of investigations suggest , the cardioprotective effects of lc n-3 pufas epa and dha intake in human subjects , because of their lipid lowering , hypotensive , anti - arrhythmic , and anti - thrombotic properties . 
   moreover , studies performed in the last twenty years showed heterogeneous effects of different n-3 pufas on various cardiovascular outcomes , which may be of paramount relevance in primary and secondary prevention of cardiovascular disease . 
   recent in vitro investigations as well as clinical studies also demonstrated that lc - n3-pufas significantly interact with inflammation - related mechanisms , such as endothelial activation , modification of eicosanoid metabolism , and resolution of the inflammatory process . 
   low - grade chronic inflammation is present in several diseases and is characterized by abnormal circulating levels of pro- and anti - inflammatory cytokines . 
   n-3 pufas may modulate inflammation , that is , suggested by the reduction of plasma inflammatory cytokines ( tnf- , il-6 ) and inflammatory markers as high sensitive c reactive protein , observed after the intake of epa and dha . 
   low - grade chronic inflammation plays a key role in the induction and progression of atherosclerosis and consequently of cardiovascular disease . 
   taking into consideration the pleiotropic nature of their actions , we suggest that dietary intake of lc n-3 pufas may lead to improvements in cardio - metabolic health parameters of their antioxidant , anti - inflammatory , and antiarrhytmic actions .
summary: phospholipids play an essential role in cell membrane structure and function . 
   the length and number of double bonds of fatty acids in membrane phospholipids are main determinants of fluidity , transport systems , activity of membrane - bound enzymes , and susceptibility to lipid peroxidation . 
   the fatty acid profile of serum lipids , especially the phospholipids , reflects the fatty acid composition of cell membranes . 
   moreover , long - chain n-3 polyunsatured fatty acids decrease very - low - density lipoprotein assembly and secretion reducing triacylglycerol production . n-6 and n-3 polyunsatured fatty acids are the precursors of signalling molecules , termed 
   eicosanoids ,  which play an important role in the regulation of inflammation . 
   eicosanoids derived from n-6 polyunsatured fatty acids have proinflammatory actions , while eicosanoids derived from n-3 polyunsatured fatty acids have anti - inflammatory ones . 
   previous studies showed that inflammation contributes to both the onset and progression of atherosclerosis : actually , atherosclerosis is predominantly a chronic low - grade inflammatory disease of the vessel wall . 
   several studies suggested the relationship between long - chain n-3 polyunsaturated fatty acids and inflammation , showing that fatty acids may decrease endothelial activation and affect eicosanoid metabolism .