Fatty acids represent the main part of which food triglycerides are made and are molecules with important structural, energy and metabolic functions.
The fatty acids found in nature are distinguished, based on the absence or presence of double bonds, in saturated and unsaturated, respectively. The unsaturated fatty acids, in turn, differentiate into mono-unsaturated or poly-unsaturated in relation to the number of double bonds present in their molecule. The increase in the number of double bonds, or the passage from saturated fatty acids to progressively more “unsaturated” molecules, constitutes a structural element of primary functional importance: the presence in series of double bonds gives the carbon chain the progressive increase of the points of twist, just at the double bond. At a three-dimensional level this involves the growing tendency of polyunsaturated fatty acids (PUFAs) to assume a less rigid conformation which, on a morpho-functional level, corresponds to an increasing fluidity of the membranes within which the PUFA are incorporated. The main n-3 PUFA, contained almost exclusively in fish, are eicosapentaenoic acid (EPA, C20: 5 n-3) and docosahexaenoic acid (DHA, C22: 6 n-3), the last of which is most represented in the body human. The main n-6 PUFAs, mainly contained in vegetable oils and in meat, are represented by linolenic acid (GLA, C18: 3 n-6) and arachidonic acid (AA, C20: 4 n-6).
Both n-6 and n-3 PUFA series are incorporated into biological membranes, in the form of phospholipid and glycolipid components. At this level they compete for the same enzymatic system, as their metabolism is completely separate and their interconversion is not possible. The metabolic pathways of the linoleic and α-linolenic series use, in fact, the same enzymes for elongation and desaturation reactions which from linoleic acid and α-linolenic acid lead, respectively, to the formation of arachidonic acid (PUFAn-6) and eicosapentaenoic acid (PUFA n-3). EPA can be further extended and desaturated to docosahexaenoic acid (DHA), which in the human body is the most represented n-3 PUFA; the latter constitutes an EPA reserve and can therefore exercise its cellular effects both directly and indirectly through conversion into EPA.
In recent years various experimental and clinical studies have been conducted, which have demonstrated the numerous favorable effects of n-3 PUFAs. The first identified biological effect was the hypotrigliceridemizing one, whereby these compounds were initially used for the treatment of dyslipidemia. Subsequently, the effects of anti-aggregation, anti-thromothrombosis and, recently, the anti-arrhythmogenic effect (1-5) were highlighted.
ANTITROMBOTIC AND EMOREOLOGICAL EFFECT
After appropriate stimulation, the PUFAs are released from cell membranes and, due to the cyclooxygenase and lipooxygenase enzymes, give rise to active metabolites, called eicosanoids (prostaglandins, thromboxanes, leukotrienes and other biologically active compounds). From the arachidonic acid derive the A2 thromboxane series (TXA2), the 4 series leukotrienes (LTB4, LTC4, etc.) and the series 2 prostaglandins (PGI2, PGD2, etc.) formed respectively in platelets, leukocytes and endothelial cells , while the EPA is metabolized, at these sites, to thromboxane A3 (TXA3), leukotrienes of series 5 (LTB5, LTC5, etc.) and prostaglandins of series 3 (PGI3, PGD3, etc.). While the vasodilating and anti-aggregating effects of the prostaglandins of series 2 and 3 are considered equivalent, the TXA2 and the leukotrienes of the series 4, derived from arachidonic acid, have a pro-aggregating / vasoconstrictor and pro-inflammatory effect respectively higher than the analogous eicosanoids derived from the EPA (6). The higher intake of n-3 PUFA in the diet leads to a shift in the synthesis of eicosanoids by substitution at the level of arachidonic acid cell membranes with EPA, as a substrate of the same enzyme system. This may partly explain the favorable biological effects of n-3 PUFAs compared to n-6 PUFAs, although probably other mechanisms are operative in determining a beneficial antithrombotic effect of n-3 PUFAs, such as: 1. strengthening of fibrinolysis and reduction of fibrinogen; 2. reduction of the activity of lipoprotein (a), a powerful prothrombotic factor as well as proaterogenic. The mechanisms listed above account for the positive effects of n-3 polyunsaturated fatty acids on platelet aggregation, with a related increase in bleeding time. It has also been shown that a greater incorporation of n-3 PUFA into the red blood cell membrane is able to increase its fluidity and therefore its mechanical and osmotic deformability, favoring its circulating at the level of the microcirculation.
EFFECTS ON THE LIPID STRIP
The effects of n-3 PUFA fatty acids on triglycerides have been the most studied: an average reduction of my triglyceride of around 30% is confirmed and the hypoglyceridemic effect appears to be dose-dependent (5-6). The underlying mechanism appears to be a decreased hepatic synthesis of fatty acids, triglycerides, VLDL, as well as an increased catabolism of VLDL to LDL, since omega-3 rich VLDLs would be
more susceptible to the action of lipoproteinlipases. Recently an action on PPARs nuclear receptors (PerixosomeProliferatorActivatedReceptors) has also been hypothesized (7). The effects of n-3 PUFAs on other lipids are much discussed. They modestly increase HDL levels, thanks to the reduction of free fatty acids in the plasma, which would cause a reduced transfer, mediated by CholesterolEsterTransferProtein, of cholesterol esters from HDL to LDL and VLDL (8). The effect of omega-3 fatty acids on the plasma concentrations of LDL is much less certain: a modest increase in this lipoprotein fraction has been described, due to a greater size of the individual particles, rather than to their numerical increase. This is therefore an antiatherogenic effect of n-3 PUFAs taking into account that only small and dense LDLs are associated with a greater atherogenic risk. Finally, some recent studies have shown that n-3 PUFAs reduce post-prandial lipemia, a factor that seems to have an independent role in the development of atherosclerosis: this effect seems to be related to a greater activity of lipoproteinlipase on lipoproteins, on chylomicrons and maybe even VLDL.
EFFECTS ON GLUCID METABOLISM
Taking into account the central role of insulin resistance in the development of diabetes, some studies wanted to verify whether n-3 PUFAs could influence the action of insulin at the level of the corresponding receptors. In the rat it was shown that an increased concentration of omega-3 improves membrane fluidity and therefore the interaction between insulin and its receptor. All this has not yet been demonstrated in humans, where in literature they are reported conflicting data, although from the latest meta-analyzes it would appear that the administration of PUFA n 3 up to 3 g / day is safe and has a neutral effect on both glycated hemoglobin and fasting blood glucose.
EFFECTS ON ARTERIAL PRESSURE AND ON THE VASCULAR TONE
Omega-3 fatty acids reduce blood pressure both in healthy subjects and in patients with arterial hypertension (9) through different mechanisms: suppression of prostanoids with vasoconstrictor action; increased production and release of nitroxide; reduction of plasma norepinephrine concentration; regulation of calcium accumulation in the cell; increased fluidity of plasma membranes.
As previously stated, leukotrienes from the 5 series derive from omega-3 fatty acids, which, compared to those of the 4 series (derived from arachidonic acid), have a much more modest vasoconstrictor and pro-inflammatory action, about ten times lower. This can have important consequences both on atherosclerosis and on other inflammatory pathologies (rheumatoid arthritis, Crohn’s disease, psoriasis, atopic dermatitis). The role played by inflammation in the genesis of atherosclerotic plaque and in the conditions of instability and rupture of the same is now universally accepted. It is therefore likely that omega-3 fatty acids, through the attenuation of inflammatory processes, can play a central role in the prevention of both the atherosclerotic process and the acute events related to plaque rupture.
Several studies have amply demonstrated how endothelial dysfunction, induced by various factors such as toxins, shear stress, cigarette smoke, dyslipidemia, and the initial event in the development of atherosclerosis: the endothelium becomes “pro-adhesive”, inducing a increased adhesion of the monocytic circulars, which subsequently infiltrate the interior of the arterial wall. At this level they form the lipid strip, the first morphologically detectable event in atherosclerosis, recalling LDL oxidized by free radicals released by the endothelium itself or by macrophages. There are several factors that mediate the interaction of endothelium-leukocytes, such as chemokines (N-formyl peptides, complement components, leukotrienes B4, PAF), selectins, adhesion proteins (ICAM1, ICAM2, ICAM3, VCAM-1) which they recognize as ligands, some integrins expressed on the leukocyte membrane. In the evolution from stria lipidica to atherosclerotica plaque several cytokines are implicated, which cause the infiltration of leukocytes, smooth muscle cells and fibroblasts and promote platelet adhesion. Unstable plates are those particularly rich in lipids and coated with a thin fibrous cap, which are at risk of fissuring. They represent the last evolutionary stage of the atheroma that leads to unstable angina, infarction and all the complications related to it, first of all the onset of fatal arrhythmias. The n-3 PUFAs are positively inserted within the pathogenetic mechanisms of atherosclerosis thanks to their lipid-lowering, antithrombotic, hemoreological and endothelial activation effects. Numerous studies have demonstrated the effects of n-3 PUFA on endothelial activation such as a reduced production of pro-inflammatory cytokines (IL-1, IL6, TNF α), of PDGF (potent pyogen and chemokine for smooth muscle cells), as well as a reduced expression of tissue factor of monocytes and endothelial adhesion molecules (10) , and an increased release of nitroxide by the endothelium. In vitro experiments conducted on endothelial cell cultures have shown that the addition of DHA a few hours or days before stimulation with cytokines (IL-1, IL-1 α, TNF α, IL-4 and LPS) significantly inhibits the endothelial activation in its various phases, including the expression of adhesion molecules, such as VCAM-1, E-selectin and, to a lesser extent, ICAM-1 (11). This effect is related to the incorporation of n-3 PUFAs in cell membranes, while it is inversely proportional to the n-6 PUFA content. The DHA concentrates more in the phosphatidyl-ethanolamine pool, which is notoriously more represented in the inner part of the membrane plasma and therefore in a strategic position to alter intracellular transduction pathways. Finally, n-3 PUFAs inhibit the NF-kB nuclear transcription factor that controls the coordinated expression of different adhesion and chemotactic molecules, specific for leukocytes. A recent randomized trial demonstrated the role of polyunsaturated n-3 fatty acids on plaque stability. In this study, 188 patients with carotid atherosclerosis conditioning a critical stenosis, pending thromboendarterectomy intervention, were enrolled; these patients were randomized to placebo, fish oil (PUFA n-3) and seed oil (PUFA n-6). Patients who received n-3 PUFAs had a high concentration of EPA and DHA at the level of atherosclerotic plaque, a reduction in the infiltration of monocytes and macrophages, a fibrous cap more often with respect to controls, and to patients treated with PUFA n-6. All these factors certainly contribute to plaque stability and could therefore justify the reduction of fatal and non-fatal cardiovascular events observed in primary and secondary clinical trials with n-3 PUFA.
The polyunsaturated fatty acids n-3, born as drugs to be used in the control of dyslipidemias, have unpredictably demonstrated an antiarrhythmic efficacy, representing a new pharmacological option in the treatment of ventricular arrhythmias, and therefore of Sudden Cardiac Death (MCI), in affected patients from post-infarct heart disease. It is known that MCI represents a frequent occurrence, being able to begin as a first manifestation of acute ischemia in 25% of cases or as a late complication of heart attack or ischemic heart disease in 75% of cases. The mechanisms hypothesized in inducing a greater electrical membrane stability by PUFA n-3 would be multiple. The production of less harmful eicosanoids (TXA3 and LTB5) determines a slight vasoconstrictor and inflammatory tissue response and therefore, a reduction in infarcts and a lower production of superoxide radicals, favoring the electric instability of the infarct areas. The n-3 PUFAs are able to modulate the sympathetic-vagal balance in favor of the latter, when there is a sympathetic overactivity. In fact, studies conducted by analyzing heart rate variability (HRV), used as a surrogate end-point for arrhythmic and MCI events, have documented how the administration of n-3 PUFAs determines an increase in RR variability both in patients and in patients with high arrhythmic risk (patients infarcted with left ventricular dysfunction, patients with chronic renal failure on dialysis, diabetic patients), both in healthy subjects and how this is correlated with concentrations of EPA and DHA in cell membranes. The increase in n-3 PUFA content in platelet membranes is related to an increase in heart rate variability, expressed as SDNN (Standard Deviation of Normal to Normal Intervals). Finally, n-3 PUFAs modulate the conductance of membrane ion channels, through the modification of the physical state and, therefore, of the fluidity characteristics of the lipid bilayer. The n-3 PUFAs act at the level of the Na ++ channels causing a shift of the threshold for the opening of the channels towards a more positive potential value, so that only a stimulation greater than 40-50% can induce the potential of action. At the level of the Ca ++ channels, the n-3 PUFAs would determine an inhibition of the voltage-dependent L-type currents, reducing the cytosolic concentration of this ion, which is arrhythmogenic when excessive (ischemia, decompensation, digitalis intoxication) and, above all, decreasing the fluctuations in the concentration of cytosolic Ca ++ before contraction, responsible for the appearance of post-potentials. A mechanism more recently hypothesized about the modulating effect exerted by n-3 PUFAs at the level of ion channels, would be represented by an alteration of the tension that membrane phospholipids exert on the channel itself, with consequent conformational change and alteration of ionic conductance ( 12). These effects on membrane ion channels have been tested in a series of experimental works conducted mainly on isolated myocytes treated with arrhythmogenic substances (for example toxic levels of calcium or ouabainine). Similarly, studies conducted in vivo on experimental animals (dogs with previous extended anterior infarction induced by ligation of the anterior interventricular) showed a protective effect of n-3 PUFAs against ischemic-induced fatal ventricular arrhythmias.
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