Annette L. West

Incorporation of eicosapentaenoic and docosahexaenoic acids into lipid pools when given as supplements providing doses equivalent to typical intakes of oily fish

Background: Estimation of the intake of oily fish at a population level is difficult. The measurement of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in biological samples may provide a useful biomarker of intake. Objective: We identified the most appropriate biomarkers for the assessment of habitual oily fish intake and changes in intake by elucidating the dose- and time-dependent response of EPA and DHA incorporation into various biological samples that represent roles in fatty acid transport, function, and storage. Design: This was a double-blind, randomized, controlled intervention trial in 204 men and women that lasted 12 mo. EPA and DHA capsules were provided in a manner to reflect sporadic consumption of oily fish (ie, 1, 2, or 4 times/wk). EPA and DHA were assessed at 9 time points over 12 mo in 9 sample types (red blood cells, mononuclear cells, platelets, buccal cells, adipose tissue, plasma phosphatidylcholine, triglycerides, cholesteryl esters, and nonesterified fatty acids). Results: A dose response (P < 0.05) was observed for EPA and DHA in all pools except for red blood cell EPA (P = 0.057). EPA and DHA measures in plasma phosphatidylcholine and platelets were best for the discrimination between different intakes (P < 0.0001). The rate of incorporation varied between sample types, with the time to maximal incorporation ranging from days (plasma phosphatidylcholine) to months (mononuclear cells) to >12 mo (adipose tissue). Conclusions: Plasma phosphatidylcholine EPA plus DHA was identified as the most suitable biomarker of acute changes in EPA and DHA intake, and platelet and mononuclear cell EPA plus DHA were the most suitable biomarkers of habitual intake. This trial was registered at Current Controlled Trials (www.controlled-trials.com) as ISRCTN48398526.

Omega-3 Fatty Acid Supplementation Does Not Reduce Risk of Atrial Fibrillation After Coronary Artery Bypass Surgery A Randomized, Double-Blind, Placebo-Controlled Clinical Trial

Background—Omega-3 polyunsaturated fatty acids (n-3 PUFA) have been reported to reduce the risk of sudden cardiac death presumed to be due to fatal ventricular arrhythmias, but their effect on atrial arrhythmias is unclear. Methods and Results—Patients (n=108) undergoing coronary artery bypass graft surgery were randomly assigned to receive 2 g/d n-3 PUFA or placebo (olive oil) for at least 5 days before surgery (median, 16 days; range, 12 to 21 days). Phospholipid n-3 PUFA were measured in serum at study entry and at surgery and in right atrial appendage tissue at surgery. Echocardiography was used to assess left ventricular function and left atrial dimensions. Postoperative continuous ECG monitoring (Lifecard CF) for 5 days or until discharge, if earlier, was performed with a daily 12-lead ECG and clinical review if patients remained in the hospital beyond 5 days. Lifecard recordings were analyzed for episodes of atrial fibrillation (AF) ≥30 seconds (primary outcome). Clinical AF, AF burden (% time in AF), hospital stay, and intensive care/high dependency care stay were measured as secondary outcomes. One hundred three patients completed the study (51 in the placebo group and 52 in the n-3 PUFA group). There were no clinically relevant differences in baseline characteristics between groups. n-3 PUFA levels were higher in serum and right atrial tissue in the active treatment group. There was no significant difference between groups in the primary outcome of AF (95% confidence interval [CI], −6% to 30%, P=0.28) in any of the secondary outcomes or in AF-free survival. Conclusions—Omega-3 PUFA do not reduce the risk of AF after coronary artery bypass graft surgery. Clinical Trial Registration—www.ukcrn.org.uk. Identifier: 4437.

Fish oil N-3 polyunsaturated fatty acids selectively affect plasma cytokines and decrease illness in Thai schoolchildren: a randomized, double-blind, placebo-controlled intervention trial

OBJECTIVE To determine whether very long-chain n-3 polyunsaturated fatty acids (PUFAs) affect illness and selected plasma cytokines in schoolchildren. STUDY DESIGN Thai schoolchildren aged 9 to 12 years consumed milk containing placebo (soybean) oil (n = 86) or fish oil (n = 94) on 5 days per week for 6 months; the latter provided 200 mg eicosapentaenoic acid plus 1 g docosahexaenoic acid daily. Episodes and duration of illness were recorded, and plasma interleukin (IL)-2 receptor, IL-6, IL-10, and transforming growth factor (TGF)-beta1 concentrations and the fatty acid profile of plasma phosphatidylcholine determined. RESULTS After intervention, very long-chain n-3 PUFAs were higher in plasma phosphatidylcholine in the fish oil group than in the placebo group (P < .001). The fish oil group showed fewer episodes (P = .014) and shorter duration (P = .024) of illness (mainly upper respiratory tract) than the placebo group. Plasma IL-2 receptor, IL-10, and IL-6 were not affected by either treatment. Plasma TGF-beta1 increased in both groups, but the increase was smaller in the fish oil group, and at the end of supplementation TGF-beta1 concentration was lower in the fish oil group (P < .001). CONCLUSIONS Very long-chain n-3 PUFAs reduce illness, mainly infections, in healthy Thai schoolchildren.

Treating liver fat and serum triglyceride levels in NAFLD, effects of PNPLA3 and TM6SF2 genotypes: Results from the WELCOME trial

BACKGROUND & AIMS Genetic variation in both patatin-like phospholipase domain-containing protein-3 (PNPLA3) (I148M) and the transmembrane 6 superfamily member 2 protein (TM6SF2) (E167K) influences severity of liver disease, and serum triglyceride concentrations in non-alcoholic fatty liver disease (NAFLD), but whether either genotype influences the responses to treatments is uncertain. METHODS One hundred three patients with NAFLD were randomised to omega-3 fatty acids (DHA+EPA) or placebo for 15-18months in a double blind placebo controlled trial. Erythrocyte enrichment with DHA and EPA was measured by gas chromatography. PNPLA3 and TM6SF2 genotypes were measured by PCR technologies. Multivariable linear regression and analysis of covariance were undertaken to test the effect of genotypes on omega-3 fatty acid enrichment, end of study liver fat percentage and serum triglyceride concentrations. All models were adjusted for baseline measurements of each respective outcome. RESULTS Fifty-five men and 40 women (Genotypes PNPLA3 I148M, 148I/I=41, 148I/M=43, 148M/M=11; TM6SF2 E167K 167E/E=78, 167E/K+167K/K=17 participants) (mean ± SD age, 51 ± 11 years) completed the trial. Adjusting for baseline measurement, measured covariates and confounders, PNPLA3 148M/M variant was independently associated with percentage of DHA enrichment (B coefficient -1.02 (95% CI -1.97, -0.07), p=0.036) but not percentage of EPA enrichment (B coefficient -0.31 (95% CI -1.38, 0.75), p=0.56). This genotype was also independently associated with end of study liver fat percentage (B coefficient 9.5 (95% CI 2.53, 16.39), p=0.008), but not end of study triglyceride concentration (B coefficient -0.11 (95% CI -0.64, 0.42), p=0.68). CONCLUSIONS PNPLA3 148M/M variant influences the changes in liver fat and DHA tissue enrichment during the trial but not the change in serum triglyceride concentration.

The pattern of fatty acids displaced by EPA and DHA following 12 Months supplementation varies between blood cell and plasma fractions

Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are increased in plasma lipids and blood cell membranes in response to supplementation. Whilst arachidonic acid (AA) is correspondingly decreased, the effect on other fatty acids (FA) is less well described and there may be site-specific differences. In response to 12 months EPA + DHA supplementation in doses equivalent to 0–4 portions of oily fish/week (1 portion: 3.27 g EPA+DHA) multinomial regression analysis was used to identify important FA changes for plasma phosphatidylcholine (PC), cholesteryl ester (CE) and triglyceride (TAG) and for blood mononuclear cells (MNC), red blood cells (RBC) and platelets (PLAT). Dose-dependent increases in EPA + DHA were matched by decreases in several n-6 polyunsaturated fatty acids (PUFA) in PC, CE, RBC and PLAT, but were predominantly compensated for by oleic acid in TAG. Changes were observed for all FA classes in MNC. Consequently the n-6:n-3 PUFA ratio was reduced in a dose-dependent manner in all pools after 12 months (37%–64% of placebo in the four portions group). We conclude that the profile of the FA decreased in exchange for the increase in EPA + DHA following supplementation differs by FA pool with implications for understanding the impact of n-3 PUFA on blood lipid and blood cell biology.

Acute appearance of fatty acids in human plasma -- a comparative study between polar-lipid rich oil from the microalgae Nannochloropsis oculata and krill oil in healthy young males

BackgroundThe long-chain n-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have human health benefits. Alternatives to fish as sources of EPA and DHA are needed. Oil from the micro-algae Nannochloropsis oculata contains a significant amount of EPA conjugated to phospholipids and glycolipids and no DHA. Krill oil contains EPA and DHA conjugated to phospholipids. We compare the appearance of fatty acids in blood plasma of healthy humans after consuming a high fat meal followed by either algal oil or krill oil.MethodsTen healthy males aged 18-45 years consumed a standard high fat (55 g) breakfast followed by either algal oil (providing 1.5 g EPA and no DHA) or krill oil (providing 1.02 g EPA and 0.54 g DHA). All participants consumed both oils in random order and separated by 7 days. Blood samples were collected before the breakfast and at several time points up to 10 hours after taking the oils. Fatty acid concentrations (μg/ml) in plasma were determined by gas chromatography.ResultsFatty acids derived mainly from the breakfast appeared rapidly in plasma, peaking about 3 hours after consuming the breakfast, and in a pattern that reflected their content in the breakfast. There were time-dependent increases in the concentrations of both EPA and DHA with both algal oil (P < 0.001 for EPA; P = 0.027 for DHA) and krill oil (P < 0.001 for both EPA and DHA). The concentration of EPA was higher with algal oil than with krill oil at several time points. DHA concentration did not differ between oils at any time point. The maximum concentration of EPA was higher with algal oil (P = 0.010) and both the area under the concentration curve (AUC) and the incremental AUC for EPA were greater with algal oil (P = 0.020 and 0.006). There was no difference between oils in the AUC or the incremental AUC for DHA.ConclusionThis study in healthy young men given a single dose of oil indicates that the polar-lipid rich oil from the algae Nannochloropis oculata is a good source of EPA in humans.

Perioperative Immunonutrition in Well-Nourished Patients Undergoing Surgery for Head and Neck Cancer: Evaluation of Inflammatory and Immunologic Outcomes

Limited work is available on the benefits of nutritional support enriched with arginine and n-3 fatty acids in surgical patients with head and neck cancer, particularly if well-nourished. We conducted a pilot study in these patients to examine effects on inflammatory markers and clinical outcome. Patients scheduled for radical resection of the oral cavity were randomised to 5 day preoperative and 5 day postoperative Impact® (IMN, n = 4), or no preoperative supplementary nutrition and Isosource® postoperatively (STD, n = 4). Plasma fatty acids, C-reactive protein (CRP), tumour necrosis factor (TNF)-α, interleukin (IL)-6 and IL-10 were measured at baseline, day of surgery and on postoperative days (POD) 2, 4 and 10. Postoperative complications were recorded. The (eicosapentaenoic acid plus docosahexaenoic acid) to arachidonic acid ratio was significantly higher in IMN patients on POD 2, 4 and 10 (P < 0.01). While not statistically significant, CRP, TNF-α, and IL-6 concentrations were higher in the STD group on POD2 while IL-10 was lower. Median length of stay was 10 (range 10–43) days in the IMN group and 21.5 (7–24) days in the STD group. Five complications were seen in the STD group and two in the IMN group. The results support the need for a larger trial focusing on clinical outcome.

Compared with Daily, Weekly n–3 PUFA Intake Affects the Incorporation of Eicosapentaenoic Acid and Docosahexaenoic Acid into Platelets and Mononuclear Cells in Humans

Consumption of oily fish is sporadic, whereas controlled intervention studies of n–3 (ω-3) fatty acids usually provide capsules containing eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) as a daily dose. This methodologic study explored whether there are differences in the short-, medium-, and long-term incorporation of EPA and DHA into blood plasma and cells with the provision of identical amounts of EPA and DHA, equivalent to 2 oily fish servings per week (or 6.54 g/wk EPA and DHA), either intermittently (i.e., 1 portion twice per week) or continuously (i.e., divided into daily amounts). The study was part of a randomized, double-blind controlled intervention lasting 12 mo, with participants stratified by age and sex. There were 5 intervention groups, 2 of which are reported here: the 2 intermittent portions (2I) and 2 continuous portions (2C) groups. EPA and DHA were measured in plasma phosphatidylcholine, platelets, and blood mononuclear cells (MNCs) at 9 time points. Sixty-five participants completed the study (2I group, n = 30, mean age of 49.2 y; 2C group, n = 35, mean age of 50.6 y). The incorporation pattern over the 12-mo intervention was different between the 2 groups in all samples (P < 0.0001, time × treatment interaction). At the end of the 12-mo intervention, the 2C group had higher EPA, DHA, and EPA + DHA in platelets (all P < 0.01) and higher EPA and EPA + DHA in MNCs (both P < 0.05) compared with the 2I group. No significant differences were shown for plasma phosphatidylcholine EPA (P = 0.1), DHA (P = 0.15), EPA + DHA (P = 0.07), or MNC DHA (P = 0.06). In conclusion, the pattern of consumption does affect the incorporation of EPA and DHA into cells used as biomarkers of intake. The differences identified here need to be considered in the design of studies and when extrapolating results from continuous capsule-based intervention studies to dietary guidelines for oily fish consumption. This trial was registered at www.controlled-trials.com as ISRCTN48398526.

Impairment of lysophospholipid metabolism in obesity: altered plasma profile and desensitization to the modulatory properties of n–3 polyunsaturated fatty acids in a randomized controlled trial

BACKGROUND Plasma lysophospholipids have emerged as signaling molecules with important effects on inflammation, insulin resistance, and fatty liver disease, each of which is linked closely to obesity. Dietary n-3 (ω-3) polyunsaturated fatty acids (PUFAs) may be able to improve these conditions. OBJECTIVE The objective of this study was to assess the response of plasma lysophospholipids to obesity, n-3 PUFA consumption, and a high-fat meal challenge to better understand the role of lysophospholipid metabolism in the progression of obesity-related disorders. DESIGN We determined the concentrations of 8 lysophosphatidylcholines, 11 lysophosphatidylethanolamines, and 7 lysophosphatidylinositols in the plasma of 34 normal-weight and 38 obese subjects randomly assigned to consume corn oil (control) or n-3 PUFA-rich fish oil (3 g/d; n = 15-19/group) for 90 d. Blood samples were collected on the last day of the study under fasting conditions and 6 h after a high-fat meal (1135 kcal, 86 g fat) challenge. The profile of secreted lysophospholipids was studied in HepG2 cells under palmitate-induced steatosis. RESULTS Obese and normal-weight subjects had different profiles of plasma lysophospholipids. A multivariate combination of the 26 lysophospholipids could discriminate between normal-weight and obese subjects with an accuracy of 98%. The high-fat meal challenge altered the concentration of plasma lysophosphatidylcholines in an oil treatment-dependent manner in normal-weight but not obese subjects, suggesting that obesity impairs the sensitivity of lysophospholipid metabolism to n-3 PUFAs. Noncytotoxic steatosis in HepG2 cells affected the secretion pattern of lysophospholipids, partially resembling the changes observed in the plasma of obese subjects. CONCLUSIONS Obesity has a substantial impact on lysophospholipid metabolism, altering the plasma lysophospholipid profile and abolishing its sensitivity to dietary n-3 PUFAs. These effects could contribute to the onset or progression of alterations associated with obesity, such as inflammation, insulin resistance, and fatty liver disease. This trial was registered at www.controlled-trials.com as ISRCTN96712688.

Membrane fatty acids, oxidative burst and phagocytosis after enrichment of P388D1 monocyte/macrophages with essential 18-carbon fatty acids.

The fatty acid composition of cell membranes can be modified in cell culture. The role of different fatty acid families in modulating phagocytosis and oxidative burst is not clear and therefore the influence of 18-carbon polyunsaturated fatty acids (PUFA) on these processes was examined. The mouse monocyte/macrophage line P388D1 was cultured in medium supplemented with 2 or 20 µmol/l 18:2n–6 (linoleic acid; LA) or 18:3n–3 (α-linolenic acid; LNA) and fatty acid enrichment of the cells was tested after 8 days. The macrophages were activated with phorbol ester in order to promote oxidative burst and intracellular dihydrorhodamine oxidation was determined. To test phagocytosis capacity uptake of fluorescence-labeled Escherichia coli was determined. Activation of the transcription factor nuclear factor (NF)-kappaB was also determined. Cells grown in medium with 20 µmol/l LA contained 2- to 3-fold more n–6 PUFA including 4-fold more arachidonic acid. Cells grown in medium with 20 µmol/l LNA contained 4-fold more n–3 PUFA. Both LA and LNA enhanced phagocytosis and decreased oxidative burst, with little difference between the fatty acids. NF-kappaB activation at 1 h post-stimulation was not affected by adding LA or LNA to the culture medium. We conclude that the fatty acid composition of macrophages influences their ability to phagocytose and mount oxidative burst.