Whole-body utilization of n−3 PUFA in n−6 PUFA-deficient rats

We evaluated the utilization of α-linolenic acid (18∶3n−3) in growing rats consuming a diet deficient in n−6 PUFA. After 90 d, whole-body 18∶3n−3 accumulation was 55% lower, total n−3 PUFA accumulation was 21% lower, and 18∶3n−3 disappearance was 14% higher in n−6 PUFA-deficient rats. Part of the reduction of whole-body 18∶3n−3 in n−6 PUFA-deficient rats was due to the 25% increase in net conversion of 18∶3n−3 to long-chain n−3 PUFA. Despite adequate 18∶3n−3 intake, n−6 PUFA deficiency decreased the accumulation of 18∶3n−3 and total n−3 PUFA.     

Fatty acids,the immune response,and autoimmunity: A question of n−6 essentiality and the balance between n−6 and n−3

The essentiality of n−6 polyunsaturated fatty acids (PUFA) is described in relation to a thymus/thymocyte accretion of arachidonic acid (20∶4n−6, AA) in early development, and the high requirement of lymphoid and other cells of the immune system for AA and linoleic acid (18∶2n−6, LA) for membrane phospholipids. Low n−6 PUFA intakes enhance whereas high intakes decrease certain immune functions. Evidence from in vitro and in vivo studies for a role of AA metabolites in immune cell development and functions shows that they can limit or regulate cellular immune reactions and can induce deviation toward a T helper (Th)2-like immune response. In contrast to the effects of the oxidative metabolites of AA, the longer-chain n−6 PUFA produced by γ-linolenic acid (18∶3n−6, GLA) feeding decreases the Th2 cytokine and immunoglobulin (Ig)G1 antibody response. The n−6 PUFA, GLA, dihomo-γ-linolenic acid (20∶3n−6, DHLA) and AA, and certain oxidative metabolites of AA can also induce T-regulatory cell activity, e.g., transforming growth factor (IGF)-β-producing T cells; GLA feeding studies also demonstrate reduced proinflammatory interleukin (IL)-1 and tumor necrosis factor (TNF)-α production. Low intakes of long-chain n−3 fatty acids (fish oils) enhance certain immune functions, whereas high intakes are inhibitory on a wide range of functions, e.g., antigen presentation, adhesion molecule expression, Th1 and th2 responses, proinflammatory cytokine and eicosanoid production, and they induce lymphocyte apoptosis. Vitamin E has a demonstrable critical role in long-chain n−3 PUFA interactions with immune functions, often reversing the effects of fish oil. The effect of dietary fatty acids on animal autoimmune disease models depends on both the autoimmune model and the amount and type of fatty acids fed. Diets low in fat, essential fatty acid deficient (EFAD), or high in long-chain n−3 PUFA from fish oils increase survival and reduce disease severity in spontaneous autoantibody-mediated disease, whereas high-fat LA-rich diets increase disease severity. In experimentally induced T cell-mediated autoimmune disease, EFAD diets or diets supplemented with long-chain n−3 PUFA augment disease, whereas n−6 PUFA prevent or reduce the severity. In contrast, in both T cell- and antibody-mediated autoimmune disease, the desaturated/elongated metabolites of LA are protective. PUFA of both the n−6 and n−3 families are clinically useful in human autoimmune-inflammatory disorders, but the precise mechanisms by which these fatty acids exert their clinical effects are not well understood. Finally, the view that all n−6 PUFA are proinflammatory requires revision, in part, and their essential regulatory and developmental role in the immune system warrants appreciation.     

Influences of dietary n−3 long-chain PUFA on body concentrations of 20∶5n−3, 22∶5n−3, and 22∶6n−3 in the larvae of a marine teleost fish from Australian waters,the striped trumpeter (<Emphasis Type="Italic">Latris lineata</Emphasis>)

We determined the effect of dietary long-chain (≥C₂₀) PUFA (LC-PUFA), 20∶5n−3 and 22∶6n−3, on larval striped trumpeter (Latris lineata) biochemistry through early development and during live feeding with rotifers (Brachionus plicatilis). Rotifers were enriched using seven experimental emulsions formulated with increasing concentrations of n−3 LC-PUFA, mainly 20∶5n−3 and 22∶6n−3. Enriched rotifer n−3 LC-PUFA concentrations ranged from 10–30 mg/g dry matter. Enriched rotifers were fed to striped trumpeter larvae from 5 to 18 d post-hatch (dph) in a short-term experiment to minimize gross deficiency symptoms such as poor survival that could confound results. No relationships were observed between larval growth or survival with dietary n−3 LC-PUFA at 18 dph. The larval FA profiles generally reflected those of the rotifer diet, and significant positive regressions were observed between most dietary and larval FA at 10, 14, and 18 dph. The major exception observed was an inverse relationship between dietary and larval 22∶5n−3. The presence of 22∶5n−3 in elevated amounts when dietary 22∶5n−3. The presence of 22∶5n−3 in elevated amounts when dietary 22∶6n−3 was depressed suggests that elongation of 20∶5n−3 may be occurring in an attempt to raise body concentrations of 22∶6n−3. We hypothesize that accumulation of 22∶5n−3 might be an early indicator of 22∶6n−3 deficiency in larval fish that precedes a reduction in growth or survival. A possible role of 22∶5n−3 as a biochemical surrogate for 22∶6n−3 is discussed.     

Effects of dietary supplementation of rapeseed oil on metabolism of [1-<Superscript>14</Superscript>C]18∶1n−9, [1-<Superscript>14</Superscript>C]20∶3n−6, and [1-<Superscript>14</Superscript>C]20∶4n−3 in atlantic salmon heaptocytes

Atlantic salmon were fed fish meal-based diets supplemented with either 100% fish oil (FO) or 100% rapeseed oil (RO) from an initial weight of 85 g to a final average weight of 280 g. The effects of these diets on the capacity of Atlantic salmon hepatocytes to elogate, desaturate, and esterify [1-¹⁴C]18∶1n−9 and the immediate substrates for the Δ⁵ desaturase, [1-¹⁴C]20∶3 n−6 and [1-¹⁴C]20∶4n−3, were investigated. Radiolabeled 18∶1n−9 was mainly esterified into cellular TAG, whereas the more polyunsaturated FA, [1-¹⁴C]20∶3n−6 and [1-¹⁴C]20∶4n−3, were primarily esterified into cellular PL. More of the elongation product, [1-¹⁴C]20∶1n−9, was produced from 18∶1n−9 and more of the desaturation and elongation products, 22∶5n−6 and 22∶6n−3, were produced from [1-¹⁴C]20∶3n−6 and [1-¹⁴C]20∶4n−3, respectively, in RO hepatocytes than in FO hepatocytes. Further, we studied whether increased addition of [1-¹⁴C]18∶1n−9 to the hepatocyte culture media would affect the capacity of hepatocytes to oxidize 18∶1n−9 to acid-soluble products and CO₂. An increase in exogenous concentration of 18∶1n−9 from 7 to 100 μM resulted in a nearly twofold increase in the amount of 18∶1n−9 that was oxidized. The conversion of 20∶4n−3 and 20∶3n−6 to the longer-chain 22∶6n−3 and 22∶5n−6 was enhanced by RO feeding in Atlantic salmon hepatocytes. The increased capacity of RO hepatocytes to produce 22∶6n−3 was, however, not enought to achieve the levels found in FO hepatocytes. Our data further showed that there were no differences in the hepatocyte FA oxidation capacity and the lipid deposition of carcass and liver between the two groups.     

Paradoxical effect of n−3-containing vegetable oils on long-chain n−3 fatty acids in rat heart

Flaxseed, echium, and canola oils contain α-linolenic acid (18∶3n−3, ALA) in a range of concentrations. To examine their effect on elevating cardiac levels of long-chain n−3 FA, diets based on these n−3-containing vegetable oils were fed to rats for 4 wk. Sunflower oil, which contains little ALA, was a comparator. Despite canola oil having the lowest ALA content of the three n−3-containing vegetable oils, it was the most potent for elevating DHA (22∶6n−3) levels in rat hearts and plasma. However, the relative potencies of the dietary oils for elevation of EPA (20∶5n−3) in heart and plasma followed the same rank order as their ALA content, i.e., flaxseed>echium>canola>sunflower oil. This paradox may be explained by lower ALA intake leading to decreased competition for Δ6 desaturase activity between ALA and the 24∶5n−3 FA precursor to DHA formation.     

Effect of dietary n−6 and n−3 polyunsaturated fatty acids on peroxidizability of lipoproteins in steers

The susceptibility of major plasma lipoproteins to lipoperoxidation was studied in relation to the FA composition of their neutral and polar lipids in steers given PUFA-rich diets. Two trials used, respectively, 18 (“sunflower” experiment, S) or 24 (“linseed” experiment, L) crossbred Salers x Charolais steers. Each involved three dietary treatments over a 70-d period: a control diet (CS or CL diets) consisting of hay and concentrate, or the same diet supplemented with oilseeds (4% diet dry matter) fed either as seeds (SS or LS diets) or continuously infused into the duodenum (ISO or ILO diets). Compared with control diets, ISO and ILO treatments tended to decrease the resistance time of LDL and HDL classes to peroxidation, mainly owing to the enrichment of their polar and neutral lipids with PUFA. With diets SS and LS, sensitivity of major lipoprotein classes (LDL, light and heavy HDL) was not affected because ruminal hydrogenation of dietary PUFA decreased their incorporation into lipoparticles. ISO and ILO treatments induced a more important production of conjugated dienes and hydroperoxides generated by peroxidation in the three lipoprotein classes due to the higher amounts of PUFA esterified in lipids of the core and the hydrophilic envelope of particles. The production of malondialdehyde (MDA) increased in steers fed linseed supplements, indicating that MDA production did not occur with linoleic acid provided by sunflower oil supplements. Thus, plasma peroxidation of PUFA generates toxic products in steers fed diets supplemented with PUFA and can be deleterious for the health of the animal during long-term treatment.     

Dietary intakes and food sources of n−6 and n−3 PUFA in french adult men and women

The intake of individual n−6 and n−3 PUFA has been estimated in 4,884 adult subjects (2,099 men and 2,785 women), volunteers from the French SU.VI.MAX intervention trial. The food intakes of each subject were recorded in at least ten 24-h record questionnaires completed over a period of 2.5 yr, allowing the estimation of the daily intake of energy; total fat; and linoleic, α-linolenic, arachidonic, eicosapentaenoic (EPA), n−3 docosapentaenoic (DPA), and docosahexaenoic (DHA) acids. The mean total fat intake corresponded to 94.1 g/d (36.3% of total energy intake) in men and 73.4 g/d (38.1% of energy) in women. The intake of linoleic acid was 10.6 g/d in men and 8.1 g/d in women, representing 4.2% of energy intake; that of α-linolenic acid was 0.94 g/d in men and 0.74 g/d in women, representing 0.37% of energy intake, with a mean linoleic/α-linolenic acid ratio of 11.3. The mean intakes of long-chain PUFA were: arachidonic acid, 204 mg/d in men and 152 mg/d in women; EPA, 150 mg/d in men and 118 mg/d in women; DPA, 75 mg/d in men and 56 mg/d in women; DHA, 273 mg/d in men and 226 mg/d in women; long-chain n−3 PUFA, 497 mg/d in men and 400 mg/d in women. Ninety-five percent of the sample consumed less than 0.5% of energy as α-linolenic acid, which is well below the current French recommendation for adults (0.8% of energy). In contrast, the mean intakes of long-chain n−6 and n−3 PUFA appear fairly high and fit the current French recommendations (total long-chain PUFA: 500 mg/d in men and 400 mg/d in women; DHA: 120 mg/d in men and 100 mg/d in women). The intakes of α-linolenic acid, and to a lesser extent of linoleic acid, were highly correlated with that of lipids. Whereas the main source of linoleic acid was vegetable oils, all food types contributed to α-linolenic acid intake, the main ones being animal products (meat, poultry, and dairy products). The main source of EPA and DHA (and of total long-chain n−3 PUFA) was fish and seafood, but the major source of DPA was meat, poultry, and eggs. Fish and seafood consumption showed very large interindividual variations, the low consumers being at risk of insufficient n−3 PUFA intake.     

Distribution of 22∶6n−3 newly synthesized from 18∶3n−3 into glycerolipid classes from tissues of rainbow trout (<Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis>)

The distribution of D₅-22∶6n−3 following ingestion of a pulse of D₅-18∶3n−3 was measured quantitatively by GC-negative chemical ionization MS in lipid classes from liver, cecal mucosa, and brain from rainbow trout to further our understanding of the processes determining accretion and turnover of 22∶6n−3 in fish. The accretion of D₅-22∶6n−3 was expressed in two ways, as percent enrichment and as ng D₅-22∶6n−3/μg 22∶6n−3/mg D₅-18∶3n−3 eaten. In cecal mucosa at 2 d post-dose, PC was the most enriched lipid class followed by PE and then TAG. Enrichment fell in all lipid classes in cecal mucosa from 2 to 7 d post-dose of D₅-18∶3n−3. In liver, PC was also the most enriched lipid class at 2 d, but in this tissue all lipid classes were more enriched in D₅-22∶6n−3 by 7 d. When expressed in terms of the 22∶6n−3 content of the different lipid classes, TAG became relatively less important in cecal mucosa and more important in liver. Over a time course of 3 to 35 d, the percent enrichment of D₅-22∶6n−3 in liver peaked at 7 d in PC, PE, PS, and PI and fell rapidly in TAG from 3 d. PC from liver was the most enriched lipid class at 3 and 7 d, and thereafter PE was the most enriched lipid class. However, TAG had the highest specific activity at all times except 7 d. In brain, the enrichment of D₅-22∶6n−3 was very low in all lipid classes at 3 d and increased progressively to 35 d with PC and PE similarly enriched. TAG from brain had the highest specific activity at all times. This study is the first to present quantitative information on rates of accretion and depletion of newly synthesized 22∶6n−3 into the main lipid classes of fish tissues.     

Effect of 18∶1n−9, 20∶5n−3, and 22∶6n−3 on lipid accumulation and secretion by atlantic salmon hepatocytes

We have studied the effects of dietary FA on the accumulation and secretion of [³H]glycerolipids by salmon hepatocytes in culture. Atlantic salmon were fed diets supplemented with either 100% soybean oil (SO) or 100% fish oil (FO), and grew from an initial weight of 113±5 g to a final weight of 338 ±19 g. Hepatocytes were isolated from both dietary groups and incubated with [³H]glycerol in an FA-free medium; a medium supplemented with 0.75 mM of one of three FA—18∶1n−9, 20∶5n−3, or 22∶6n−3—or a medium supplemented with 0.75 mM of the sulfur-substituted FA analog tetradecylthioacetic acid (TTA), which cannot undergo β-oxidation. Incubations were allowed to proceed for 1,2,6, or 24 h. The rate of the secretion of radioactive glycerolipids with no FA added was 36% lower from hepatocytes isolated from fish fed the FO diet than it was from hepatocytes isolated from fish fed the SO diet. Hepatocytes incubated with 18∶1n−9 secreted more [³H]TAG than when incubated with no FA, whereas hepatocytes incubated with 20∶5n−3 or TTA secreted less labeled TAG than when incubated with no FA. This observation was independent of the feeding group. Hepatocytes incubated with 22∶6n−3 secreted the highest amounts of total [³H]glycerolipids compared with the other treatments, owing to increased secretion of phospholipids and mono- and diacylglycerols (MDG). In contrast, the same amounts of [³H]TAG were secreted from these cells as from cells incubated in an FA-free medium. The lipid-lowering effect of FO is thus independent of 22∶6n−3, showing that 20∶5n−3 is the FA that is responsible for the lipid-lowering effect. The ratio of TAG to MDG in lipids secreted from hepatocytes to which 20∶5n−3 or TTA had been added was lower than that in lipids secreted from hepatocytes incubated with 18∶1n−9 or 22∶6n−3, suggesting that the last step in TAG synthesis was inhibited. Morphometric measurements revealed that hepatocytes incubated with 20∶5n−3 accumulated significantly more cellular lipid than cells treated with 18∶1n−9, 22∶6n−3, TTA, or no treatment. The area occupied by mitochondria was also greater in these cells. The present study shows that dietary FO reduces TAG secretion from salmon hepatocytes and that 20∶5n−3 mediates this effect.     

Enzymatic fractionation and enrichment of n−9 PUFA

A single-cell oil from a Mortierella alpina mutant (TGM17 oil) contains n−9 PUFA: 14.3 wt% 6,9-octadecadienoic acid (18∶2n−9; n−9 LnA) and 17.1 wt% Mead acid (20∶3n−9; MA). Lipase screening indicated that Pseudomonas aeruginosa lipase acted strongly on n−9 LnA and weakly on MA, and Candida rugosa lipase acted weakly on the two PUFA. Hence, fractionation and enrichment of the two FA were conducted with the lipases. The first step was selective hydrolysis of IGM17 oil with P. aeruginosa lipase. The hydrolysis fractionated the oil into FFA containing 20.4 wt% n−9 LnA and 6.3 wt% MA, and acylglycerols containing 10.7 wt% n−9 LnA and 23.7 wt% MA. The FFA fraction was used for preparation of n−9 LnA-rich FFA. After removal of saturated FA, the FFA were esterified with lauryl alcohol (LauOH) using C. rugosa lipase. Two selective esterifications increased the n−9 LnA content to 54.0 wt% with 38.2% recovery of the initial content of TGM17 oil. The acylglycerol fraction obtained in the hydrolysis with P. aeruginosa lipase was used for preparation of MA-rich FFA. The acylglycerol fraction was hydrolyzed under alkaline conditions, and saturated FA were eliminated by urea adduct fractionation. Two selective esterifications of the FFA with LauOH increased the MA content to 60.2 wt% with 53.5% recovery. Thus, the two-step enzymatic process was effective for fractionation and enrichment of n−9 LnA and MA.     

Metabolism of n−3 and n−6 fatty acids in Atlantic salmon liver: Stimulation by essential fatty acid deficiency

Oxidation, esterification, desaturation, and elongation of [1-¹⁴C]18∶2n−6 and [1-¹⁴C]18∶3n−3 were studied using hepatocytes from Atlantic salmon (Salmo salar I.) maintained on diets deficient in n−3 and n−6 polyunsaturated fatty acids (PUFA) or supplemented with n−3 PUFA. For both dietary groups, radioactivity from 18∶3n−3 was incoporated into lipid fractions two to three times faster than from 18∶2n−6, and essential fatty acids (FFA) deficiency doubled the incorporation. Oxidation to CO₂ was very low and was independent of substrate or diet, whereas oxidation to acid-soluble products was stimulated by EFA deficiency. Products from 18∶2n−6 were mainly 18∶3n−6, 20∶3n−6, and 20∶4n−6, with minor amounts of 20∶2n−6 and 22∶5n−6. Products from 18∶3n−3 were mainly 18∶4n−3, 20∶5n−3, and 22∶6n−3, with small amounts of 20∶3n−3. The percentage of 22∶6n−3 in the polar lipid fraction of EFA-deficient hepatocytes was fourfold higher than in n−3 PUFA-supplemented cells. This correlated well with our other results obtained after abdominal injection of [1-¹⁴C]18∶3n−3 and [1-¹⁴C]18∶2n−6. In hepatocytes incubated with [4,5-³H]-22∶6n−3, 20∶5n−3 was the main product. This retrocon-version was increased by EFA deficiency, as was peroxisomal β-oxidation activity. This study shows that 18∶2n−6 and 18∶3n−3 can be elongated and desaturated in Atlantic salmon liver, and that this conversion and the activity of retroconversion of very long chain PUFA is markedly enhanced by FFA deficiency.     

Lipase-catalyzed incorporation of n−3 PUFA into palm oil

Two immobilized lipases, IM60 from Rhizomucor miehei and QLM from Alcaligenes sp., were used as biocatalysts for the modification of the FA composition of palm oil by incorporating n−3 PUFA. Acidolysis and interesterification reactions were conducted with hexane as organic solvent, and the products were analyzed by using GLC. After a 24-h incubation in hexane, there was an average incorporation of 20.8% EPA and 15.6% DHA into palm oil, respectively, while the percentages of palmitic and oleic acids in palm oil decreased by 28.8 and 11.8%, respectively. Higher EPA and DHA incorporation was obtained when EPAX (fish oil concentrate high in n−3 PUFA) was used in the ethyl ester form (interesterification reaction) than in the free acid form (acidolysis) in the presence of Lipozyme (IM60 lipase. Lipase QLM was found to discriminate against EPA, and it showed slightly better catalytic activity for DHA in the free acid form than in the ethyl ester form. Generally, as the mole ratio of the acyl donor to TAG increased, the percentage incorporation of EPA and DHA increased; however, reactions catalyzed by Lipozyme IM60 did not show increases in the incorporation beyond a TAG/EPAX mole ratio of 3. When limitations due to mass transfer were not a factor, an increase in the reactant amount also gave an increase in the percentage incorporation of the n−3 PUFA. Palm oil containing EPA and DHA was successfully produced and may be beneficial in certain food and nutritional applications.     

Induction of apoptosis and apoptotic mediators in balb/C splenic lymphocytes by dietary n−3 and n−6 fatty acids

The present study was designed to investigate the effect of diatery n−6 and n−3 polyunsaturated fatty acids (PUFA) on anti-CD3 and anti-Fas antibody-induced apoptosis and its mediators in mouse spleen cells. Nutritionally adequate semipurified diets containing either 5% w/w corn oil (n−6 PUFA) or fish oil (n−3 PUFA) were fed to weanling female Balb/C mice, and 24 wk later mice were sacrificed. In n−3 PUFA-fed mice, serum and splenocyte lipid peroxides were increased by 20 and 28.3% respectively, compared to n−6 PUFA-fed mice. Further, serum vitamin F levels were decreased by 50% in the n−3 PUFA-fed group, whereas higher anti0Fas- and anti-CD3-induced apoptosis (65 and 66%) and necrosis (17 and 25%), compared to the n−6 PUFA-fed group, were found when measured with Annexin V and propidium iodide staining, respectively. In addition, decreased Bcl-2 and increased Fas-ligand (Fas-L) also were observed in the n−3 PUFA-fed group compared to the n−6 PUFA-fed group. No difference in the ratio of splenocyte subsets nor their Fas expression was observed between the n−3 PUFA-fed and n−6 PUFA-fed groups, whereas decreased proliferation of splenocytes was found in n−3 PUFA-fed mice compared to n−6 PUFA-fed mice. In conclusion, our results indicate that dietary n−3 PUFA induces higher apoptosis by increasing the generation of lipid peroxides and elevating Fas-L expression along with decreasing Bcl-2 expression. A reduced proliferative response of immune cells also was observed in n−3 PUFA-fed mice.     

Speciation and Electronic Structure of La<Subscript>1−x</Subscript>Sr<Subscript>x</Subscript>CoO<Subscript>3−δ</Subscript> During Oxygen Electrolysis

Cobalt-containing perovskite oxides are promising electrocatalysts for the oxygen evolution reaction (OER) in alkaline electrolyzers. However, a lack of fundamental understanding of oxide surfaces impedes rational catalyst design for improved activity and stability. We couple electrochemical studies of epitaxial La_1?xSr_xCoO_3?δ films with in situ and operando ambient pressure X-ray photoelectron spectroscopy to investigate the surface stoichiometry, adsorbates, and electronic structure. In situ investigations spanning electrode compositions in a humid environment indicate that hydroxyl and carbonate affinity increase with Sr content, leading to an increase in binding energy of metal core levels and the valence band edge from the formation of a surface dipole. The maximum in hydroxylation at 40% Sr is commensurate with the highest OER activity, where activity scales with greater hole carrier concentration and mobility. Operando measurements of the 20% Sr-doped oxide in alkaline electrolyte indicate that the surface stoichiometry remains constant during OER, supporting the idea that the oxide electrocatalyst is stable and behaves as a metal, with the voltage drop confined to the electrolyte. Furthermore, hydroxyl and carbonate species are present on the electrode surface even under oxidizing conditions, and may impact the availability of active sites or the binding strength of adsorbed intermediates via adsorbate–adsorbate interactions. For covalent oxides with facile charge transfer kinetics, the accumulation of hydroxyl species with oxidative potentials suggests the rate of reaction could be limited by proton transfer kinetics. This operando insight will help guide modeling of self-consistent oxide electrocatalysts, and highlights the potential importance of carbonates in oxygen electrocatalysis.     

Modulation of respiratory syncytial virus-induced prostaglandin E<Subscript>2</Subscript> production by n−3 long-chain polyunsaturated fatty acids in human respiratory epithelium

Infection with respiratory syncytial virus (RSV) results in substantial infant morbidity and has been associated with the subsequent development of childhood asthma. Inflammatory mediators produced by both the epithelium and tissue leukocytes during RSV infection stimulate the release of chemotactic factors by the respiratory epithelium and the subsequent influx of inflammatory cells, predominantly neutrophils. We investigated the production of inflammatory mediators [prostaglandin E₂ (PGE₂), interleukin (IL)-1β, tumor necrosis factor α] and chemokines [IL-8, RANTES (regulation on activation, normal T cell expressed and secreted)] by alveolar epithelial cells in response to RSV infection. Infection of a human alveolar epithelial transformed cell line (A549 cells) with live RSV substantially increased production of PGE₂, IL-8, and RANTES. By altering cell membrane FA through incorporation of the long-chain PUFA (LCPUFA) arachidonic acid, EPA, and DHA, we were subsequently able to significantly modulate PGE₂ production by the infected epithelium. Because of the dynamic nature of the effects of PGE₂ on lung function, regulation of this prostaglandin during RSV infection by n−3 LCPUFA has the potential to significantly alter the disease process.     

Alteration of 20∶5n−3 and 22∶6n−3 fat contents and liver peroxisomal activities in fenofibrate-treated rainbow trout

Fish easily accumulate n−3 PUFA of exogenous origin, but the underlying mechanisms are not well established in the whole animal. This study was undertaken to investigate whether this feature was physiologically associated with mitochondrial and peroxisomal capacities that differentially affect FA oxidation. For this purpose, peroxisomal FA oxidation was increased by treating rainbow trout with fenofibrate, which strongly stimulates the peroxisome proliferator-activated receptor-α in rodents. Diets containing EPA and DHA, with or without fenofibrate added, were administered to male trout for 12 d. After treatment, neither liver hypertrophy nor accumulation of fat was apparent within the liver and muscle cells. However, fenofibrate treatment decreased the contents of EPA and DHA in the liver, white muscle, and intraperitoneal fat tissue, which represented (per whole body) at least 280 mg less than in controls. Carnitine-dependent palmitate oxidation rates, expressed per gram of liver, were slightly increased by fenofibrate when measured from tissue homogenates and were unchanged when calculated from isolated mitochondria, relative to control fish. The treatment altered neither carnitine palmitoyltransferase I activity rates, expressed per gram of liver, nor the sensitivity of the enzyme to malonyl-CoA inhibition, but did increase the malonyl-CoA content (+45%). Meanwhile, fenofibrate increased (by about 30%) the peroxisome-related activities, i.e, catalase, carnitine-independent palmitate oxidation, acyl-CoA oxidase, and the peroxisomal FA-oxidizing system, relative to the control group. The data strongly suggest that the induction of peroxisomal activities, some of which being able to oxidize very long chain FA, was responsible for the lower contents of EPA and DHA in the body lipids of fenofibrate-treated trout.     

Intramuscular injection of antigens and adjuvant preferentially decreases 18∶2n−6 and 18∶3n−3 in pig neck muscle

Linoleic (18∶2n−6) and α-linolenic acids (18∶3n−3) have many important physiological functions including immunomodulation. We tested how immunization influences the metabolism of 18∶2n−6 and 18∶3n−3 in the neck muscle of pigs. At 35 d old, pigs received either an intramuscular neck injection containing hen egg white lysozyme (HEWL), killed Mycobacterium tuberculosis, and Freund’s complete adjuvant (immunized) or PBS (control). At 49 d old, immunized pigs received a booster injection of HFWI and Freund’s incomplete adjuvant, and the control pigs received PBS into the neck. At 56 d old, all pigs received an intradermal injection of Mycobacterium bovis into the hind leg to induce a delayed-type hypersensitivity (DTH) reaction. At 57 d old, immunized pigs had a twofold increase in serum haptoglobin, a 10-fold increase in antibodies to HEWL, and the skinfold at the DTH reaction site was 10 times thicker than the controls. Both 18∶2n−6 and 18∶3n−3 (% composition) were approximately 25% lower in muscle IG, 40% lower in FFA, 50% lower in phospholipids, but not different in cholesteryl esters of the neck muscle of immunized pigs. The antigens in this model induce an increased response in the innate (haptoglobin), humoral (antibodies), and cellular (DTH) immune systems as well as a preferential decrease of 18∶2n−6 and 18∶3n−3 in the inflamed neck muscle. It appears that 18∶2n−6 and 18∶3n−3 are preferentially metabolized (possibly β-oxidized) in response to antigens.     

Effects of diets enriched in n−6 or n−3 fatty acids on cholesterol metabolism in older rats chronically fed a cholesterol-enriched diet

Hypocholesterolemic effects in older animals after long-term feeding are unknown. Therefore, aged rats (24 wk of age) fed a conventional diet were shifted to diets containing 10% perilla oil [PEO; oleic acid+linoleic acid+α-linolenic acid; n−6/n−3, 0.3; polyunsaturated fatty acid/saturated fatty acid (P/S), 9.6], borage oil [oleic acid+linoleic acid+α-linolenic acid; n−6/n−3, 15.1; P/S, 5.3], evening primrose oil (FPO; linoleic acid+γ-linolenic acid; P/S, 10.5), mixed oil (MIO; oleic acid+linoleic acid+γ-linolenic acid+α-linolenic acid; n−6/n−3, 1.7; P/S, 6.7), or palm oil (PLO; palmitic acid+oleic acid+linoleic acid; n−6/n−3, 25.3; P/S, 0.2) with 0.5% cholesterol for 15 wk in this experiment. There were no significant differences in the food intake and body weight gain among the groups. The liver weight in the PEO (n−6/n−3, 0.3) group was significantly higher than those of other groups in aged rats. The serum total cholesterol and very low density lipoprotein (VLDL) +intermediate density lipoprotein (IDL)+low density lipoprotein (LDL)-cholesterol concentrations of the PLO (25.3) group were consistently higher than those in the other groups. The serum high density lipoprotein cholesterol concentrations of the PEO (0.3) and EPO groups were significantly lower than in the other groups at the end of the 15-wk feeding period. The liver cholesterol concentration of the PLO (25.3) group was significantly higher than those of other groups. There were no significant differences in the hepatic LDL receptor mRNA level among the groups. Hepatic apolipoprotein (apo) B mRNA levels were not affected by the experimental conditions. The fecal neutral steroid excretion of the PLO (25.3) group tended to be low compared to the other groups. The results of this study demonstrate that both n\t-6 fatty acid and n\t-3 fatty acids such as \gg-linolenic acid and \ga-linolenic acid inhibit the increase of serum total cholesterol and VLDL+IDL+LDL-cholesterol concentrations of aged rats in the presence of excess cholesterol in the diet compared with dietary saturated fatty acid.     

Differential effect of N-ethyl maleimide on Δ6-desaturase activity in human fetal liver toward fatty acids of the n−6 and n−3 series

The effect of N-ethyl-maleimide (NEM) on Δ5-and Δ6-desaturase activities and the incorporation of substrates and products into different microsomal lipid classes and phospholipid (PL) subclasses were studied in human fetal liver microsomes, obtained after legally approved therapeutic abortion. Desaturase activities were measured by a radiochemical method using reversed-phase high-performance liquid chromatography (HPLC). After nonphospholipid (NPL) and PL separation on silica cartridges, the radioactivity in different lipids of the NPL group was assessed by two-dimensional thin-layer chromatography, and their fatty acid (FA) composition by gas-liquid chromatography. The PL subclasses were separated, and the distribution of radioactivity between products and substrates was determined in PL subclasses. NEM inhibited the Δ5- and Δ6-desaturase activities in the n−6 series of FA but not the Δ6-desaturase activity in the n−3 series, which suggests the existence of two distinct Δ6-desaturases, one for the n−6 series and another for the n−3 series. Whether NEM was present or absent, most of the radioactivity was recovered in the free FA form (about 80%). The desaturation products, obtained in the presence or absence of NEM, were preferentially incorporated into PL, suggesting a channeling of the newly synthesized FA toward microsomal PL. The comparison of the distribution of substrates and products incorporated into the different PL classes showed that most of the labeled FA were incorporated into phosphatidylcholine and to a lesser degree into phosphatidylethanolamine.