Studies on the metabolic fate of n-3 polyunsaturated fatty acids

Several different processes involved in the metabolic fate of docosahexaenoic acid (DHA, C22:6n-3) and its precursor in the biosynthesis route, C24:6n-3, were studied. In cultured skin fibroblasts, the oxidation rate of [1-14C] 24:6n-3 was 2.7 times higher than for [1-14C]22:6n-3, whereas [1-14C]22:6n-3 was incorporated 7 times faster into different lipid classes than was [1-14C]24:6n-3. When determining the peroxisomal acyl-CoA oxidase activity, similar specific activities for C22:6(n-3)-CoA and C24:6(n-3)-CoA were found in mouse kidney peroxisomes. Thioesterase activity was measured for both substrates in mouse kidney peroxisomes as well as mitochondria, and C22:6(n-3)-CoA was hydrolyzed 1.7 times faster than C24:6(n-3)-CoA. These results imply that the preferred metabolic fate of C24:6(n-3)-CoA, after its synthesis in the endoplasmic reticulum (ER), is to move to the peroxisome, where it is beta-oxidized, producing C22:6(n-3)-CoA. This DHA-CoA then preferentially moves back, probably as free fatty acid, to the ER, where it is incorporated into membrane lipids.     

The metabolic role of n-3 polyunsaturated fatty acids: relationship to human disease.

1. An increasing body of evidence suggests that certain types of fat have beneficial effects on human health. 2. Fish oils in particular have been shown to exert positive effects on atherosclerosis, heart disease and carcinogenesis. 3. These positive effects are thought to be mediated through eicosanoids derived from polyunsaturated fatty acids of the n-3 family which are present in large quantities in fish oils.     

Protection against in vivo liver ischemia-reperfusion injury by n-3 long-chain polyunsaturated fatty acids in the rat

Abstract

N-3 polyunsaturated fatty acids (n-3 PUFA) affect inflammatory processes. This study evaluated the effects of dietary supplementation with fish oil on hepatic ischemia-reperfusion (IR) injury in the rat. Parameters of liver injury (serum transaminases and histology) and oxidative stress (serum 8-isoprostanes and hepatic GSH and GSSG), were correlated with NF-κB DNA binding and FA composition and inflammatory cytokine release. N-3 PUFA supplementation significantly increased liver n-3 PUFA content and decreased n-6/n-3 PUFA ratios. IR significantly modified liver histology and enhanced serum transaminases, 8-isoprotanes and inflammatory cytokines, with net reduction in liver GSH levels and net increment in those of GSSG. Early increase (3 h) and late reduction (20 h) in NF-κB activity was induced. All IR-induced changes were normalized by n-3 PUFA supplementation. In conclusion, prevention of liver IR-injury was achieved by n-3 PUFA supplementation, with suppression of oxidative stress and recovery of pro-inflammatory cytokine homeostasis and NF-κB functionality lost during IR.     

Intravenous infusion of n-3 polyunsaturated fatty acids.

Dietary supplementation with n-3 polyunsaturated fatty acids (PUFA) is regarded as beneficial for the prevention and treatment of atherosclerosis and thrombosis and chronic inflammatory diseases like rheumatoid arthritis and psoriasis. It may be possible to treat some acute diseases like acute myocardial infarction or acute rejection of grafted organs if it is possible to make n-3 PUFA take effect quickly (in hours instead of days). Three sets of experiments were done. In Experiment 1, emulsion of trieicosapentaenoyl-glycerol (EPA-TG) and tridocosahexaenoyl-glycerol was infused through rabbit ear veins, and the leukotriene B4/B5 production from polymorphonuclear leukocytes was measured at different time points by high-performance liquid chromatography. In Experiment 2, delayed type hypersensitivity (DTH) of mice was measured with sheep red blood cells as an antigen. Pure n-3 PUFA emulsions or a control solution were infused through tail veins just before the second challenge of the antigen. DTH was measured 24 hr after the second challenge. In Experiment 3, human natural killer cell activity was measured using K562 target cells before and after the infusion of pure EPA-TG emulsion to an antecubital vein. Leukotriene B4 production by rabbit polymorphonuclear leukocytes was depressed by 40% by EPA-TG infusion. DTH was suppressed almost completely by n-3 PUFA infusion. Natural killer cell activity was suppressed almost completely by EPA-TG infusion in 8 hr. DTH, natural killer cell activity, and leukotriene B4 production are probably related to acute rejection of grafted organs.(ABSTRACT TRUNCATED AT 250 WORDS)     

Beneficial effects of the n-3 long-chain polyunsaturated fatty acids in surgical patients: updating the evidence

The use of n-3 polyunsaturated fatty acids in surgical patients has risen by the fact that this may attenuate systemic and acute inflammatory responses secondary to surgical trauma through modulation of inflammatory mediators and cell membrane interactions. Moreover, the inclusion of n-3 fatty acids in clinical trials as part of the therapy in patients, who expect to undergo a surgical stress, suggests benefits on clinical progress. Therefore, the objective of this article is to review data from n-3 polyunsaturated fatty acid effects on biochemical parameters and on reduced length of hospitalization, number of infections, and mortality as main clinical outcomes in human surgical patients.     

n-3 polyunsaturated fatty acids and the cardiovascular system

n-3 Polyunsaturated fatty acids, mainly those contained in fish oils, are candidates for inclusion in secondary prevention programmes for coronary heart disease, based on the results of recent randomized trials in humans. Marine n-3 polyunsaturated fatty acids retard coronary atherosclerosis and appear to prevent fatal arrhythmias; and they decrease mortality subsequent to myocardial infarction.     

The antidepressant role of dietary long-chain polyunsaturated n-3 fatty acids in two phases in the developing brain

In this work we investigated the effect from fish oil (FO) supplementation, rich in n-3 fatty acids, on an antidepressant effect on adult rats in Phase A (supplementation during pregnancy and lactation) and phase B (supplementation during post-weaning until adulthood). During Phase A, female rats, used as matrix to obtain male rats, were divided in three groups: FO (daily supplemented), CF (coconut fat daily supplemented) and control (not supplemented). Our results showed that adult rats whose mothers were supplemented with FO during Phase A and rats supplemented during phase B demonstrated a significantly decreased immobility time when compared to control and CF groups. There was no difference in neither motor activity nor anxiety behavior in the three groups excluding false positive results. Our results suggest that n-3 fatty acids supplementation during Phases A and B had a beneficial effect on preventing the development of depression-like behavior in adult rats.     

Effect of n-3 and n-6 polyunsaturated fatty acids on hare reproductive performances

The study was carried out on 42 breeder couples (42 males and 42 females) of European brown hare (Lepus europaeus), divided into three groups fed three different experimental diets (14 couples/treatment). Two diets were supplemented with n-3 and n-6 polyunsaturated fatty acids (PUFAs; 2% of linseed oil and soybean oil, respectively) and were compared with a control diet supplemented with a monounsaturated fatty acids (2% of olive oil). During the experimental period (from 15 April to 30 September), the following parameters were recorded: days from the beginning of trial to the first parturition, parturition interval, number of parturitions, number of leverets born (alive and dead), dead during suckling, the total number of leverets weaned and feed intake per cage (of males, females and leverets until weaning). Feed intake was not influenced by treatments. In hares fed n-3 and n-6 diets, the days from the beginning of the trial to the first parturition and the parturition interval were similar and were lower compared with control group (63.1 v. 70.6 days, and 37.8 v. 40.9 days, respectively; P < 0.05). Hares from n-6 group had a higher (P < 0.05) number of parturitions per cage during the experimental period than the n-3 and control group that showed a similar value (3.00 v. 2.36, respectively). The total number of leverets born per cage and parturition in n-6 and n-3 groups increased with respect to those fed control diet (P < 0.05). The leverets' mortality rate at birth was higher in n-6 than in n-3 and control group (3.50 v. 2.17, respectively; P < 0.05). In control group, leverets' mortality rate during suckling was lower with respect to n-3 (P < 0.05) and n-6 (P < 0.05), showing the highest value for the latter (P < 0.05). In spite of this higher mortality, the number of leverets weaned per cage and parturition was higher (P < 0.05) in n-6 compared with n-3 group, being the latter higher than the control group (3.12, 2.79 and 2.43, respectively). Our results show that the dietary PUFAs, particularly n-6 supplementation, have a positive influence on the reproductive performances of the European brown hare.     

The effects of n-3 polyunsaturated fatty acids by gavage on some metabolic enzymes of rat liver

In this experimental study, the effect of fish n-3 fatty acids was studied on the some important enzymes of carbohydrate metabolism, hexokinase (HK), glucose-6-phosphate dehydrogenase (G6PD), 6-phosphogluconate dehydrogenase (6PGD), lactate dehydrogenase (LDH), and malate dehydrogenase (MDH) in rat liver. Wistar albino rats of experimental group (n= 9) were supplemented fish omega-3 fatty acids (n-3 PUFA) as 0.4 g/kg bw. by gavage for 30 days in addition to their normal diet. Isotonic solution was given to the control group (n= 8) by the same way. At 30th day, the rats were killed by decapitation under ether anesthesia, autopsied and liver was removed. Spectrophotometric methods were used to determine the activities of above-mentioned enzymes in the liver. The n-3 PUFA caused increases in the activities of HK, G6PD, LDH, and MDH in comparison with control. These increases were statistically significant (P < 0.01) except 6PGD activity. As a result, n-3 PUFA may regulate the metabolic function of liver effectively by increasing HK, G6PD, 6PGD, LDH, and MDH enzyme activities of rat liver when added in enough amounts to the regular diet.     

NGF blocks polyunsaturated fatty acids biosynthesis in n-3 fatty acid-supplemented PC12 cells

Regulation of polyunsaturated fatty acid (PUFA) biosynthesis in proliferating and NGF-differentiated PC12 pheochromocytoma cells deficient in n-3 docosahexaenoic acid (DHA 22:6n-3) was studied. A dose- and time-dependent increase in eicosapentaenoic acid (EPA, 20:5n-3), docosapentaenoic acid (DPA, 22:5n-3) and DHA in phosphatidylethanolamine (PtdEtn) and phosphatidylserine (PtdSer) glycerophospholipids (GPL) via the elongation/desaturation pathway following alpha-linolenic acid (ALA, 18:3n-3) supplements was observed. That was accompanied by a marked reduction of eicosatrienoic acid (Mead acid 20:3n-9), an index of PUFA deficiency. EPA supplements were equally effective converted to 22:5n-3 and 22:6n-3. On the other hand, supplements of linoleic acid (LNA, 18:2n-6) were not effectively converted into higher n-6 PUFA intermediates nor did they impair elongation/desaturation of ALA. Co-supplements of DHA along with ALA did not interfere with 20:5n-3 biosynthesis but reduced further elongation to 22-hydrocarbon PUFA intermediates. A marked decrease in the newly synthesized 22:5n-3 and 22:6n-3 following ALA or EPA supplements was observed after nerve growth factor (NGF)-induced differentiation. NGF also inhibited the last step in 22:5n-6 formation from LNA. These results emphasize the importance of overcoming n-3 PUFA deficiency and raise the possibility that growth factor regulation of the last step in PUFA biosynthesis may constitute an important feature of neuronal phenotype acquisition.     

n-6 and n-3 polyunsaturated fatty acids differentially modulate oncogenic Ras activation in colonocytes

Ras proteins are critical regulators of cell function, including growth, differentiation, and apoptosis, with membrane localization of the protein being a prerequisite for malignant transformation. We have recently demonstrated that feeding fish oil, compared with corn oil, decreases colonic Ras membrane localization and reduces tumor formation in rats injected with a colon carcinogen. Because the biological activity of Ras is regulated by posttranslational lipid attachment and its interaction with stimulatory lipids, we investigated whether docosahexaenoic acid (DHA), found in fish oil, compared with linoleic acid (LA), found in corn oil, alters Ras posttranslational processing, activation, and effector protein function in young adult mouse colon cells overexpressing H-ras (YAMC-ras). We show here that the major n-3 polyunsaturated fatty acid (PUFA) constituent of fish oil, DHA, compared with LA (an n-6 PUFA), reduces Ras localization to the plasma membrane without affecting posttranslational lipidation and lowers GTP binding and downstream p42/44(ERK)-dependent signaling. In view of the central role of oncogenic Ras in the development of colon cancer, the finding that n-3 and n-6 PUFA differentially modulate Ras activation may partly explain why dietary fish oil protects against colon cancer development.     

Folate and long-chain polyunsaturated fatty acids in psychiatric disease

Schizophrenia, autism and depression do not inherit by Mendel's law, and the search for a genetic basis seems unsuccessful. Schizophrenia and autism relate to low birth weight and pregnancy complications, which are associated with developmental adaptations by "programming". Epigenetics might constitute the basis of programming and depend on folate status and one-carbon metabolism in general. Early folate status of patients with schizophrenia might be compromised as suggested by (i) coinciding incidences of schizophrenia and neural tube defects (NTDs) in the Dutch hunger winter, (ii) coinciding seasonal fluctuations in birth of patients with schizophrenia and NTDs, (iii) higher schizophrenia incidence in immigrants and (iv) higher incidence in methylene tetrahydrofolate reductase 677C-->T homozygotes. Recent studies in schizophrenia and autism point at epigenetic silencing of critical genes or chromosomal loci. The long-chain polyunsaturated fatty acids (LCPUFA), arachidonic acid (AA, from meat) and docosahexaenoic acid (fish) are components of brain phospholipids and modulators of signal transduction and gene expression. Patients with schizophrenia and, possibly, autism exhibit abnormal phospholipid metabolism that might cause local AA depletion and impaired eicosanoid-mediated signal transduction. National fish intakes relate inversely with major and postpartum depressions. Five out of six randomized controlled trials with eicosapentaenoic acid (fish) have shown positive effects in schizophrenia, and 4 of 6 were favorable in depression and bipolar disorders. We conclude that folate and LCPUFA might be important in both the etiology and severity of at least some psychiatric diseases.     

Comparative utilization of n-3 polyunsaturated fatty acids by cultured human Y-79 retinoblastoma cells

The Y-79 retinoblastoma cell, a cultured human line derived from the retina, was utilized as a model for investigating the metabolism of n-3 polyunsaturated fatty acids in neural tissue. When cultures were incubated with 5 microM linolenic (18:3), eicosapentaenoic (20:5) or docosahexaenoic (22:6) acids, a low concentration probably representative of physiologic levels, the amount incorporated was 20:5 congruent to 18.3 greater than 22:6. Regardless of which fatty acid was provided, 65-75% of the total uptake accumulated in phosphatidylethanolamine and ethanolamine plasmalogen, suggesting that these phospholipids play an important role in n-3 polyunsaturated fatty acid metabolism. A small amount of 22:6 was converted to 20:5, which was recovered in phosphatidylinositol and phosphatidylserine. Therefore, one metabolic function of 22:6 may be to serve as an intracellular storage pool for the formation of 20:5 through retroconversion. When any of the n-3 polyunsaturates was available, the main fatty acid that accumulated in the cell phospholipids was 22:6. The extent to which 22:6 accumulated, however, depended on the particular n-3 polyunsaturated fatty acid that was available. This suggests that the 22:6 content of a neural cell, and any cellular function dependent on 22:6 content, may be regulated by changes in the type of n-3 polyunsaturate available to the nervous system.     

Differential induction of electrophile-responsive element-regulated genes by n-3 and n-6 polyunsaturated fatty acids

In this study the n-3 polyunsaturated fatty acids (PUFAs) eicosapentaenoic acid and docosahexaenoic acid appear to be effective inducers of electrophile-responsive element (EpRE) regulated genes, whereas the n-6 PUFA arachidonic acid is not. These n-3 PUFAs need to be oxidized to induce EpRE-regulated gene expression, as the antioxidant vitamin E can partially inhibit the PUFA induced dose-dependent effect. Results were obtained using a reporter gene assay, real-time RT-PCR and enzyme activity assays. The induction of EpRE-regulated phase II genes by n-3 PUFAs may be a major pathway by which n-3 PUFAs, in contrast to n-6 PUFAs, are chemopreventive and anticarcinogenic.     

Feeding long-chain n-3 polyunsaturated fatty acids during gestation increases intestinal glucose absorption potentially via the acute activation of AMPK

The current study utilized Ussing chambers to examine the impact of supplementing maternal gestation and/or lactation diets with n-3 polyunsaturated fatty acids (PUFA) provided via a protected fish oil (PFO) product on intestinal fatty acid profiles and ex vivo glucose uptake in the jejunum of weanling piglets. Jejunum tissues were enriched with n-3 PUFA as a result of feeding the sows the PFO during gestation and/or lactation (P<.05). Glucose uptake improved by twofold (P<.042) in intestinal preparations obtained from the offspring of sows fed PFO during gestation or throughout gestation/lactation versus lactation alone. This was also reflected in the jejunum protein expressions of glucose transporter 2 (GLUT2) and sodium-dependent glucose transporter 1 (SGLT1). Furthermore, adding docosahexaenoic acid (DHA) or an AMP-activated protein kinase (AMPK) agonist to the chamber buffer improved glucose uptake (P<.05) in intestinal preparations obtained from the offspring fed the control diet, devoid of the PFO product and containing minimal concentrations of n-3 PUFA. Collectively, these data indicate two important points. First, long-term exposure to n-3 PUFA via the maternal gestation diet effectively enhances glucose uptake in the weanling piglet, and the underlying mechanism may be associated with changes in the intestinal fatty acid profile. Secondly, there is an apparent direct and acute effect of DHA that is achieved within a time frame that precludes substantial changes in the intestinal fatty acid profile. Additionally, both mechanisms may involve activation of AMPK. Thus, n-3 PUFA delivered in utero and postnatally via the maternal diet may help the offspring adapt quickly to rapidly changing diets early in life and allow optimal nutrient uptake.     

The effects of n-3 and n-6 polyunsaturated fatty acids on plasma lipids and fatty acids of treated phenylketonuric children

Dietary-treated phenylketonuric patients (PKUs) display low levels of long-chain polyunsaturated fatty acids (PUFA) in plasma lipids. In a 6-month clinical trial we observed a decrease of triglycerides and an increase of n-3 long-chain PUFA in plasma of PKUs supplemented with fish oil, while no major differences in respect to the baseline values were found in a group supplemented with blackcurrant oil. A more complete source of long-chain PUFA of both the n-6 and n-3 series should be investigated for dietary supplementation of PKU patients.     

Differential modulation of Toll-like receptors by fatty acids: preferential inhibition by n-3 polyunsaturated fatty acids

Human subjects consuming fish oil showed a significant suppression of cyclooxygenase-2 (COX-2) expression in blood monocytes when stimulated in vitro with lipopolysaccharide (LPS), an agonist for Toll-like receptor 4 (TLR4). Results with a murine monocytic cell line (RAW 264.7) stably transfected with COX-2 promoter reporter gene also demonstrated that LPS-induced COX-2 expression was preferentially inhibited by docosahexaenoic acid (DHA, C22:6n-3) and eicosapentaenoic acid (EPA, C20:5n-3), the major n-3 polyunsaturated fatty acids (PUFAs) present in fish oil. Additionally, DHA and EPA significantly suppressed COX-2 expression induced by a synthetic lipopeptide, a TLR2 agonist. These results correlated with the preferential suppression of LPS- or lipopeptide-induced NF kappa B activation by DHA and EPA. The target of inhibition by DHA is TLR itself or its associated molecules, but not downstream signaling components. In contrast, COX-2 expression by TLR2 or TRL4 agonist was potentiated by lauric acid, a saturated fatty acid. These results demonstrate that inhibition of COX-2 expression by n-3 PUFAs is mediated through the modulation of TLR-mediated signaling pathways. Thus, the beneficial or detrimental effects of different types of dietary fatty acids on the risk of the development of many chronic inflammatory diseases may be in part mediated through the modulation of TLRs.