Effect of dietary EPA and DHA on murine blood and liver fatty acid profile and liver oxylipin pattern depending on high and low dietary n6-PUFA.

The intake of long-chain n3-polyunsaturated fatty acids (PUFA), which are associated with beneficial health effects, is low in the Western diet, while the portion of dietary n6-PUFA and hence the n6/n3-PUFA ratio is high. Strategies to improve the n3-PUFA status are n3-PUFA supplementation and/or lowering n6-PUFA intake. In the present study, mice were fed with two different sunflower oil-based control diets rich in linoleic (n6-high) or oleic acid (n6-low), either with low n3-PUFA content (∼0.02%) as control or with ∼0.6% eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA). The n6-low diet had only little or no effect on levels of arachidonic acid (ARA) and its free oxylipins in liver tissue. Supplementation with EPA or DHA lowered ARA levels with an effect size of n6-high < n6-low. Blood cell %EPA + DHA reached >8% and >11% in n6-high and n6-low groups, respectively. Elevation of EPA levels and EPA derived oxylipins was most pronounced in n6-low groups in liver tissue, while levels of DHA and DHA derived oxylipins were generally unaffected by the background diet. While the n6-low diet alone had no effect on blood and liver tissue ARA levels or n3-PUFA status, a supplementation of EPA or DHA was more effective in combination with an n6-low diet. Thus, supplementation of long-chain n3-PUFA combined with a reduction of dietary n6-PUFA is the most effective way to improve the endogenous n3-PUFA status.

Long-term stimulation of toll-like receptor-2 and -4 upregulates 5-LO and 15-LO-2 expression thereby inducing a lipid mediator shift in human monocyte-derived macrophages.

Macrophage polarization switches during the course of inflammation along with the lipid mediators released. We investigated the lipid mediator formation in human monocyte-derived macrophages during in vitro differentiation and pathogen stimulation. For this, peripheral blood monocytes were differentiated into M1 (CSF-2/IFNγ) or M2 (CSF-1/IL-4) macrophages followed by stimulation with the toll-like receptor (TLR) ligands zymosan (TLR-2), Poly(I:C) (TLR-3) or bacterial lipopolysaccharides (TLR-4) mimicking fungal, viral and bacterial infection, respectively. Expression of enzymes involved in lipid mediator formation such as 5- and 15-lipoxygenases (LO), the 5-LO activating protein and cyclooxygenase-2 (COX-2) was monitored on mRNA and protein level and lipid mediator formation was assessed. In addition, cytokine release was measured. In vitro differentiation of human peripheral blood monocytes to M1 and M2 macrophages considerably attenuated 5-LO activity. Furthermore, while TLR-2 and -4 stimulation of M1 macrophages primarily triggered pro-inflammatory cytokines and lipid mediators, persistent stimulation (16 h) of human M2 macrophages induced a coordinated upregulation of 5- and 15-LO-2 expression. This was accompanied by a marked increase in IL-10 and monohydroxylated 15-LO products in the conditioned media of the cells. After additional stimulation with Ca2+ ionophore combined with supplementation of arachidonic, eicosapentaenoic and docosahexaenoic acid these cells also released small amounts of SPM such as lipoxins and resolvins. From this we conclude that activation of TLR-2 or -4 triggers the biosynthesis of pro-inflammatory 5-LO and COX-2 derived lipid mediators in human monocyte-derived M1 macrophages while persistent stimulation of M2 macrophages induces a shift towards pro-resolving 15-LO derived oxylipins.

Single-Dose SDA-Rich Echium Oil Increases Plasma EPA, DPAn3, and DHA Concentrations.

The omega-3 (n3) polyunsaturated fatty acids (PUFA) eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are associated with health benefits. The primary dietary source of EPA and DHA is seafood. Alpha-linoleic acid (ALA) has not been shown to be a good source for EPA and DHA; however, stearidonic acid (SDA)-which is naturally contained in echium oil (EO)-may be a more promising alternative. This study was aimed at investigating the short-term n3 PUFA metabolism after the ingestion of a single dose of EO. Healthy young male subjects (n = 12) ingested a single dose of 26 g of EO after overnight fasting. Plasma fatty acid concentrations and relative amounts were determined at baseline and 2, 4, 6, 8, 24, 48, and 72 h after the ingestion of EO. During the whole examination period, the participants received standardized nutrition. Plasma ALA and SDA concentrations increased rapidly after the single dose of EO. Additionally, EPA and DPAn3 concentrations both increased significantly by 47% after 72 h compared to baseline; DHA concentrations also significantly increased by 21% after 72 h. To conclude, EO increases plasma ALA, SDA, EPA, DPAn3, and DHA concentrations and may be an alternative source for these n3 PUFAs.

Activation of Lipid Mediator Formation Due to Lipoprotein Apheresis.

Lipoprotein apheresis reliably reduces low-density lipoprotein (LDL) cholesterol in patients with atherosclerotic disease and therapy-refractory hypercholesterolemia or elevated lipoprotein (a) (Lp(a)). Besides lowering lipoproteins and triglycerides, apheresis also decreases levels of essential omega-6 and omega-3 polyunsaturated fatty acids (n-6 and n-3 PUFAs) in blood plasma. In contrast, heparin-induced extracorporeal low-density lipoprotein precipitation (HELP) lipid apheresis might increase the formation of potentially pro-inflammatory and pro-thrombotic lipid mediators derived from n-6 and n-3 PUFAs. The study presented here analyzed lipid mediator profiles in the plasma of patients with hyperlipidemia treated by one of three different apheresis methods, either HELP, direct absorption (DA), or membrane filtration (MDF), in a direct pre- and post-apheresis comparison. Using gas chromatography and liquid chromatography tandem mass spectrometry (LC-MS/MS) we were able to analyze fatty acid composition and the formation of lipid mediators called oxylipins. Our data illustrate-particularly in HELP-treated patients-significant decreases of essential omega-6 and omega-3 polyunsaturated fatty acids in blood plasma but significant increases of PUFA-derived lipoxygenase-, as well as cyclooxygenase- and cytochrome P450-derived lipid mediators. Given that n-3 PUFAs in particular are presumed to be cardioprotective and n-3 PUFA-derived lipid mediators might limit inflammatory reactions, these data indicate that n-3 PUFA supplementation in the context of lipid apheresis treatment might have additional benefits through apheresis-triggered protective n-3 PUFA-derived lipid mediators.

Effects of a low and a high dietary LA/ALA ratio on long-chain PUFA concentrations in red blood cells.

There is a debate about the optimal dietary ratio of the parent n6 fatty acid linoleic acid (LA) and n3 fatty acid alpha-linolenic acid (ALA) to promote an efficient conversion of ALA to EPA and DHA, which have implications for human health. The aim of the present study was to compare the effects of a low-LA/high-ALA (loLA/hiALA) diet with a high-LA/low-ALA (hiLA/loALA) diet on fatty acid concentrations in red blood cells (RBCs). Fifteen omnivore healthy men (mean age 26.1 ± 4.5 years) with a low initial EPA/DHA status (sum (∑) EPA + DHA% of total fatty acids in RBC at baseline: 4.03 ± 0.17) received both diets for two weeks with a nine-week wash-out phase in between. Fatty acid intake of the subjects was tightly controlled. Concentrations [µg mL-1] and relative amounts [% of total fatty acids] of fatty acids in RBCs were analyzed at baseline (day 0), day 7 and 14 by means of GC-FID. The dietary LA/ALA ratios were 0.56 ± 0.27 : 1 and 25.6 ± 2.41 : 1 and led to significantly different changes of ALA, LA, EPA and ∑EPA + DHA concentrations in RBCs. In the course of the loLA/hiALA diet ALA and EPA concentrations and relative amounts of ∑EPA + DHA increased, whereas LA concentrations decreased. The DHA concentration was unaffected. The hiLA/loALA diet led to slightly decreased EPA concentrations, while all other fatty acid concentrations remained constant. Compared to our previous study, where we simply increased the ALA intake, our results show that ALA supplementation combined with a reduced LA intake (loLA/hiALA diet) more efficiently enhanced EPA blood concentrations. The absence of changes in the PUFA pattern in consequence of a LA/ALA ratio of 25.6 ± 2.41 : 1 suggests that the high LA/ALA ratio of the Western diet already leads to a saturation and a further increase of the ratio does not affect the PUFA pattern.

A diet rich in omega-3 fatty acids enhances expression of soluble epoxide hydrolase in murine brain.

Several studies suggest that intake of omega-3 polyunsaturated fatty acids (n3-PUFA) beneficially influences cognitive function. However, effects on the adult brain are not clear. Little is known about the impact of dietary intervention on the fatty acid profile in adult brain, the modulation in the expression of enzymes involved in fatty acid biosynthesis and metabolism as well as changes in resulting oxylipins. These questions were addressed in the present study in two independent n3-PUFA feeding experiments in mice. Supplementation of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA, 1% each in the diet) for 30days to adult NMRI and C57BL/6 mice led to a distinct shift in the brain PUFA pattern. While n3-PUFAs EPA, n3 docosapentaenoic acid and DHA were elevated, many n6-PUFAs were significantly decreased (except, e.g. C20:3 n6 which was increased). This shift in PUFAs was accompanied by immense differences in concentrations of oxidative metabolites derived from enzymatic conversion of PUFAs, esp. arachidonic acid whose products were uniformly decreased, and a modulation in the activity and expression pattern of delta-5 and delta-6 desaturases. In both mouse strains a remarkable increase in the soluble epoxide hydrolase (sEH) activity (decreased epoxy-FA concentrations and epoxy-FA to dihydroxy-FA-ratios) as well as sEH expression was observed. Taking the high biological activity of epoxy-FA, e.g. on blood flow and nociceptive signaling into account, this finding might be of relevance for the effects of n3-PUFAs in neurodegenerative diseases. On any account, our study suggests a new distinct regulation of brain PUFA and oxylipin pattern by supplementation of n3-PUFAs to adult rodents.

Lipid Class Specific Quantitative Analysis of n-3 Polyunsaturated Fatty Acids in Food Supplements.

Supplementation products containing n-3 PUFA from marine sources serve a large market. Although the amount of eicosapentaenoic acid and docosahexaenoic acid in the products is provided by the manufacturer, no or little information is available on their lipid pattern. Therefore, we quantitatively analyzed the fatty acid pattern in the lipid fractions triglycerides, phospholipids, ethyl esters, and free fatty acids in supplementation products by means of solid phase extraction and gas chromatography. Twelve products from the European and U.S. markets containing fish, krill, algal, or plant oil were analyzed. Total n-3 PUFA content ranged from 68 g/100 g fat (fish oil) to 42 g/100 g fat (algal oil) to 17 g/100 g fat (krill oil). On the basis of the n-3 PUFA containing lipid class, the supplements can be separated dominantly in ethyl ester, re-esterified triglyceride, triglyceride, and phospholipid containing products. Algae-based products contained natural triglycerides, krill oils a complex mixture of phospholipids, triglycerides, and free fatty acids, and fish oil products either ethyl esters, re-esterified triglycerides, or triglycerides. Even products of the same class and source showed distinct differences in their lipid pattern. A specification of the lipid composition of n-3 PUFA products would allow distinguishing the different (qualities of) supplements.

Effects of docosahexaenoic acid supplementation on PUFA levels in red blood cells and plasma.

INTRODUCTION: Polyunsaturated fatty acids (PUFA) are metabolized in a complex network of elongation, desaturation and beta oxidation. MATERIAL AND METHODS: The short (1 and 3 wk), and long term (6 and 12 wk) effect of 1076mg/d docosahexaenoic acid (DHA, free of eicosapentaenoic acid (EPA)) on (absolute) PUFA concentrations in plasma and red blood cells (RBC) of 12 healthy men (mean age 25.1±1.5 years) was investigated. RESULTS: RBC DHA concentrations significantly (p<0.001) increased from 28±1.6µg/mL to 38±2.0µg/mL (wk 1), 52±3.3µg/mL (wk 3), 68±2.6µg/mL (wk 6), and 79±3.5µg/mL (wk 12). Arachidonic acid (AA) concentrations declined in response to DHA treatment, while the effect was more pronounced in plasma (wk 0: 183±9.9µg/mL, wk 12: 139±8.0µg/mL, -24%, p<0.001) compared to RBC (wk 0: 130±3.7µg/mL, wk 12: 108±4.0µg/mL, -16%, p=0.001). Furthermore, an increase of EPA concentrations in plasma (wk 0: 15±1.5µg/mL, wk 1:19±1.6µg/mL, wk 3: 27±2.3µg/mL, wk 6: 23±1.2µg/mL, wk 12: 25±1.7µg/mL, p<0.001) and RBC (wk 0: 4.7±0.33µg/mL, wk 1: 6.7±1.3µg/mL, wk 3: 8.0±0.66µg/mL, wk 6: 6.9±0.44µg/mL, wk 12: 6.7±0.45µg/mL, n.s.) was observed suggesting a retroconversion of DHA to EPA. CONCLUSION: Based on PUFA concentrations we showed that DHA supplementation results in increased EPA levels, whereas it is not known if this impacts the formation of EPA-derived lipid mediators. Furthermore, shifts in the entire PUFA pattern after supplementation of EPA or DHA should be taken into account when discussing differential physiological effects of EPA and DHA.