Cristina Mosconi

Reduced plasma C-20 and C-22 polyunsaturated fatty acids in children with phenylketonuria during dietary intervention

The fatty acid composition of plasma and erythrocyte lipids was analyzed in 15 children with phenylketonuria (aged 3 to 12 years) during dietary treatment aimed to maintain plasma phenylalanine levels at less than 8 mg/dl (485 mumol/L), and compared with those of 12 matched control subjects. The diet of children with phenylketonuria provided less protein, with a very low proportion of animal proteins, less fat, but a higher proportion of linoleic acid as a percentage of calories, and a higher carbohydrate content versus that in the diet of control subjects. The children with phenylketonuria had higher plasma levels of oleic acid but lower levels of arachidonic (n-6) and n-3 fatty acids. Linoleic and eicosatrienoic (n-9) acid levels were the same in both groups. These changes in patients with phenylketonuria resemble those observed in vegetarians and may be due to the absence of preformed arachidonic acid and long-chain n-3 fatty acids in the phenylketonuric diet.

Prolonged retention of doubly labeled phosphatidylcholine in human plasma and erythrocytes after oral administration

The plasma kinetics of a preparation of dilinoleoyl phosphatidylcholine (DLPC) specifically labeled with3H in the choline moiety and with14C in the 2-fatty acid (FA) were evaluated in six healthy volunteers after oral administration. Retention of both isotopes in plasma exceeded expectations, with a half-life in the elimination phase of 172.2 h for3H and 69.7 h for14C. Up to 60 d after administration, there were still significant levels of radioactivity present in plasma. The relative stability of the [14C]FA label was demonstrated by the retention for more than 12 h of an isotope ratio close to that of the compound administered. The14C label of DLPC remained in position-2, as assessed by cleavage of plasma phospholipids with phospholipase A2. The [3H]choline label showed an early incorporation into high density lipoproteins and subsequently into low density lipoproteins (LDL); conversely, the14C radioactivity was rapidly incorporated into triacylglycerols that were mainly associated with very low density lipoproteins. Radioactivity measurements revealed that both isotopes remained the longest time in LDL. In red blood cell (RBC) lipids, [3H]choline radioactivity accumulated over time, with a plateau after 48 h, whereas FA radioactivity accumulated more rapidly and was followed by a progressive decay. Analysis of the isotope ratio in these cells suggested an early incorporation of lyso products followed by rapid transfer of FA from plasma. The RBC maintained considerable radioactivity for a prolonged time, thus acting as a possible reservoir for the DLPC administered. Our study showed that dilinoleoyl PC remained in plasma longer than predicted based on earlier studies, and that after absorption the FA label was found in position 2.

Diets rich in n-9, n-6 and n-3 fatty acids differentially affect the generation of inositol phosphates and of thromboxane by stimulated platelets, in the rabbit

We have studied the effects of semi-synthetic diets rich in either n-9 (olive oil, OO) or n-6 (corn oil, CO), or n-3 (fish oil, FO, as MaxEPA) fatty acids on the levels of major PUFA in platelet lipids, on the generation of inositol phosphates by [3H]inositol labelled platelets after stimulation with thrombin and of thromboxane B2 (TxB2) by platelet rich plasma (PRP) after stimulation with collagen. The predicted elevations of oleic (OA), linoleic (LA) and eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids were observed in platelet lipids of each animal group, but in the MaxEPA fed group accumulation of EPA was associated with depletion of linoleic acid (LA) rather than of arachidonic acid (AA). Basal levels of inositol-tris-phosphate (IP3) in platelets were lowest in the OO group and highest in the CO group, whereas the increment after thrombin stimulation (1 unit/ml NIH) was maximal in the OO group and minimal in the FO group. Instead, when generation of TxB2 by stimulated platelets was evaluated, no appreciable difference among the various groups could be detected, in accordance with the limited modifications of platelet AA content induced by the diets. The overall data indicate that dietary fatty acids modulate the pathway of inositol phosphate generation in rabbit platelets, independently of modifications of TxB2 production.

Phosphatidylinositol (PI) and PI-associated arachidonate are elevated in platelet total membranes of type IIa hypercholesterolemic subjects

The lipid composition (phospholipid distribution and fatty acid patterns of individual glycerophospholipids) and levels of lipid components (cholesterol, total and individual phospholipid classes, arachidonic acid) have been determined in total membranes of platelets from type IIa hypercholesterolemic (HC) and control (C) subjects. Levels of cholesterol and total phospholipid, relative to the protein content, were about 80% and 60% higher respectively in platelet total membranes from HC subjects. Small differences between the two groups of samples were observed for the phospholipid distribution and the fatty acid patterns. Concentrations of individual phospholipid classes, were on the average 60% higher in HC than in C platelet membranes, with an even greater difference for phosphatidylinositol (PI) and sphingomyelin. Levels of arachidonic acid, relative to the protein content, were also 60-80% higher in membranes from HC platelet with a more than 100% increase in PI. The higher levels of the eicosanoid precursor fatty acid in phospholipids and especially in PI, which is considered a donor pool for eicosanoid synthesis, may be a contributing factor for the greater thromboxane formation and enhanced aggregation, upon stimulation, of platelets from HC patients in comparison to platelets from control subjects.

Decrease of polyunsaturated fatty acids and elevation of the oleic/stearic acid ratio in plasma and red blood cell lipids of malnourished cancer patients

The fatty acids profiles of plasma and red blood cell lipids have been evaluated in 12 malnourished cancer patients in comparison with samples from eight healthy controls. In such patients, significantly lower levels of linoleic acid (LA) as percentage of total fatty acids were observed in plasma phospholipids (PL) and cholesterol esters (CE), and in red blood cells PL. The levels of arachidonic acid (AA) and the unsaturation index of the two lipid classes were also reduced in plasma CE but not in PL. In spite of the marked reduction of LA and, more generally, of total polyunsaturated fatty acids (PUFA), no elevation of eicosatrienoic acid (20:3 n-9) was observed, such acid being considered a typical index of essential fatty deficiency. Moreover, no modification of the parameters indicating impairment of the fatty acid desaturation activity was shown. In addition, the levels of palmitic and oleic acids were significantly higher in both plasma PL and CE and in red blood cells PL. The reported elevation of the oleic to stearic acid ratio in lipids of red blood cells from malnourished cancer patients, already observed by other authors, was confirmed in our study. This ratio was even more markedly elevated in plasma lipids of the patients. A very good correlation was found between the reduction of linoleic acid levels, especially in plasma CE, and weight loss, suggesting enhanced utilization of this fatty acid in association with extensive depletion of lipid stores, in this pathological state.

Increments of dietary linoleate raise liver arachidonate, but markedly reduce heart n−6 and n−3 fatty acids in the rat

Four diets containing 20% of energy (en%) as fat and with linoleic acid contents of 1.9, 3.1, 7.7 and 10.1 en%, respectively, were fed to one-month-old male rats for three months. The fatty acid profiles and the levels of the major n−6 and n−3 fatty acids in the lipids of plasma, liver, heart and kidney were measured. We found that with increasing concentrations of 18∶2n−6 in the diet, linoleic acid rose in plasma and in all organs, but long-chain n−6 and n−3 fatty acids responded differently. In liver, arachidonic acid increased and n−3 fatty acids were not significantly affected; in heart, both arachidonic and docosahexaenoic acids were progressively reduced; and in kidney, there was no change of n−6 and n−3. The results indicate that incremental changes in dietary, linoleate affect the levels of polyunsaturated fatty acids in liver and extrahepatic organs differently.

Vitamin E influences the effects of fish oil on fatty acids and eicosanoid production in plasma and circulating cells in the rat

An EPA enriched oil (MaxEPA, Seven Seas, U.K. containing 18% EPA and 12% DHA) alone or supplemented with 10 mg/ml/alpha tocopherol, was administered by gastric intubation at the dose of 3.2 ml/kg/day for a period of eight weeks to male rats fed a standard diet. An additional group of animals was treated with the same amount of olive oil. The administration of MaxEPA alone resulted, as expected, in accumulation of EPA and reduction of AA levels in plasma, platelet, red blood cell and PMNL phospholipids, when compared to values in the olive oil group. In addition, levels of linoleic acid were elevated, suggesting inhibition of the conversion of linoleic to arachidonic acid. Formation of i.r. TxB2 by stimulated PRP, of i.r. 6-keto-PGF1 alpha by perfused aortas, and of IR LTB4 and C4 by stimulated PMNL were reduced, but production of superoxide anion by PMNL was enhanced by MaxEPA treatment vs the olive oil treatment. Supplementation of MaxEPA with vitamin E caused a smaller reduction of 20:4 levels and a smaller increase of 20:5 levels in plasma and cell phospholipids and modified the effects of MaxEPA on eicosanoid and superoxide anion production, suggesting that lipid peroxidation may mediate some of the biological effects of omega 3 fatty acids.

Dietary n-9, n-6 and n-3 fatty acids modify linoleic acid more than arachidonic acid levels in plasma and platelet lipids and minimally affect platelet thromboxane formation in the rabbit

We have studied the effects of semisynthetic diets containing 5% by weight (12% of the energy) of either olive oil (70% oleic acid, OA) or corn oil (58% linoleic acid), or fish oil (Max EPA, containing about 30% eicosapentaenoic, EPA C 20:5 n-3, plus docosahexaenoic, DHA C 22:6 n-3, acids, and less than 2% linoleic acid), fed to male rabbits for a period of five weeks, on plasma and platelet fatty acids and platelet thromboxane formation. Aim of the study was to quantitate the absolute changes of n-6 and n-3 fatty acid levels in plasma and platelet lipid pools after dietary manipulations and to correlate the effects on eicosanoid-precursor fatty acids with those on platelet thromboxane formation. The major differences were found when comparing the group fed fish oil and depleted linoleic acid vs the other groups. The accumulation of n-3 fatty acids in various lipid classes was associated with modifications in the distribution of linoleic acid and arachidonic acid in different lipid pools. In platelets maximal incorporation of n-3 fatty acids occurred in phosphatidyl ethanolamine, which also participated in most of the total arachidonic acid reduction occurring in platelets, and linoleic acid, more than archidonic acid, was replaced by n-3 fatty acids in various phospholipids. The archidonic acid content of phosphatidyl choline was unaffected and that of phosphatidyl inositol only marginally reduced. Thromboxane formation by thrombin stimulated platelets did not differ among the three groups, and this may be related to the minimal changes of arachidonic acid in phosphatidyl choline and phosphatidyl inositol.

Vascular eicosanoids and platelet-aortic wall interactions in spontaneously hypertensive rats

We studied the aggregation of collagen and ADP-stimulated platelet-rich plasma (PRP) and the formation of thromboxane B2 (TxB2) by collagen-stimulated PRP in spontaneously hypertensive rats (SHR) and in Wistar-Kyoto control rats (WKY). In addition, we evaluated the inhibition of the aggregation of PRP following homologous or heterologous perfusions through isolated aortas, the release of 6-keto-prostaglandin (PG)F1 alpha from these arteries perfused with PRP, and the sensitivity of PRP to the antiaggregatory activity of the stable PGI2 analogue, iloprost, in both SHR and WKY. The lower activities (aggregation induced by ADP and collagen, collagen-stimulated TxB2 production) of SHR platelets, were not accompanied by morphological differences from WKY platelets. These changes were associated with a greater release of arterial 6-keto-PGF1 alpha, with greater platelet antiaggregatory activity of the arterial wall and with higher sensitivity of platelets to iloprost. The lower reactivity of platelets to aggregating agents, and the greater sensitivity to prostacyclin, associated with a greater production of arterial prostacyclin were the major changes observed in SHR animals. These alterations in the SHR vs. normotensive WKY may lead to an enhanced risk of hemorrhage in the hypertensive state.

N-6 and N-3 Fatty Acids in Plasma and Platelet Lipids, and Generation of Inositol Phosphates by Stimulated Platelets after Dietary Manipulations in the Rabbit

Dietary fatty acids modify the fatty acid composition of plasma and tissue lipids, and these changes appear, in turn, to modulate biochemical and functional parameters in various biological compartments. Modifications of the amounts and proportions of saturated and polyunsaturated fatty acids (PUFA) in the diet, for instance, influence the levels of plasma cholesterol and affect the aggregation of platelets (see Goodnight et al., 1982 for a review), possibly through modifications of the eicosanoid cascade (Galli et al.,1981). More specifically, the administration of polyunsaturated fatty acids of the n−3 series, such as eicosapentaenoic acid (EPA, 20:5 n−3) and docosahexaenoic acid (DHA, 22:6 n−3) results in quantitative and qualitative changes of eicosanoid production (Fischer and Weber, 1983), following the accumulation of this fatty acid in cell lipid pools (Siess et al.,1980). This effect reduces blood platelet-vessel wall interactions and the thrombotic potential.