Dietary (n-3) fatty acids alter plasma fatty acids and leukotriene B synthesis by stimulated neutrophils from healthy geriatric Beagles.

The study objective was to determine the effect of feeding food enriched in (n-3) fatty acids (FA) on plasma FA profiles and leukotriene B (LTB) synthesis by stimulated peripheral blood neutrophils from dogs. For 36 weeks, two groups of dogs (n = 5) were fed food that contained either a low ratio of (n-6)-(n-3) FA (1.31:1; fish oil-enriched food) or a high ratio of (n - 6)-(n-3) FA (40.6:1; corn oil-enriched food). Consumption of food enriched in fish oil resulted in higher plasma concentrations of eicosapentaenoic acid and docosahexaenoic acid and lower concentrations of arachidonic acid. Neutrophils from dogs fed fish oil-enriched food produced 7.6-fold more LTB(5)(P = 0.002), and the ratio of LTB(5)-LTB(4) concentrations was 8.3-fold higher (P < 0.001) compared with dogs fed corn oil-enriched food. Dietary FA can modulate leukotriene production by neutrophils in dogs, and suggests that foods enriched in (n-3) FA from fish oil may have value in the treatment of canine inflammatory diseases.

Supplementation of postmenopausal women with fish oil does not increase overall oxidation of LDL ex vivo compared to dietary oils rich in oleate and linoleate.

Although replacement of dietary saturated fat with monounsaturated and polyunsaturated fatty acids (MUFA and PUFA) has been advocated for the reduction of cardiovascular disease risk, diets high in PUFA could increase low density lipoprotein (LDL) susceptibility to oxidation, potentially contributing to the pathology of atherosclerosis. To investigate this possibility, 15 postmenopausal women in a blinded crossover trial consumed 15 g of sunflower oil (SU) providing 12.3 g/day of oleate, safflower oil (SA) providing 10.5 g/day of linoleate, and fish oil (FO) providing 2.0 g/day of eicosapentaenoate (EPA) and 1.4 g/day of docosahexaenoate (DHA). During CuSO(4)-mediated oxidation, LDL was depleted of alpha-tocopherol more rapidly after FO supplementation than after supplementation with SU (P = 0.0001) and SA (P = 0.05). In LDL phospholipid and cholesteryl ester fractions, loss of n-3 PUFA was greater and loss of n-6 PUFA less after FO supplementation than after SU and SA supplementation (P < 0.05 for all), but loss of total PUFA did not differ. The lag phase for phosphatidylcholine hydroperoxide (PCOOH) formation was shorter after FO supplementation than after supplementation with SU (P = 0.0001) and SA (P = 0.006), whereas the lag phase for cholesteryl linoleate hydroperoxide (CE18:2OOH) formation was shorter after FO supplementation than after SU (P = 0.03) but not SA. In contrast, maximal rates of PCOOH and CE18:2OOH formation were lower after FO supplementation than after SA (P = 0.02 and 0.0001, respectively) and maximal concentrations of PCOOH and CE18:2OOH were lower after FO supplementation than after SA (P = 0.03 and 0.0006, respectively). Taken together, our results suggest that FO supplementation does not increase the overall oxidation of LDL ex vivo, especially when compared with SA supplementation. Consequently, health benefits related to increased fish consumption may not be offset by increased LDL oxidative susceptibility.-- Higdon, J. V., S. H. Du, Y. S. Lee, T. Wu, and R. C. Wander. Supplementation of postmenopausal women with fish oil does not increase overall oxidation of LDL ex vivo compared to dietary oils rich in oleate and linoleate. J. Lipid Res. 2001. 42: 407--418.

Supplementation of postmenopausal women with fish oil rich in eicosapentaenoic acid and docosahexaenoic acid is not associated with greater in vivo lipid peroxidation compared with oils rich in oleate and linoleate as assessed by plasma malondialdehyde and F(2)-isoprostanes.

BACKGROUND: Although the replacement of dietary saturated fat with unsaturated fat has been advocated to reduce the risk of cardiovascular disease, diets high in polyunsaturated fatty acids (PUFAs) could increase lipid peroxidation, potentially contributing to the pathology of atherosclerosis. OBJECTIVE: The objective of this study was to examine indexes of in vivo lipid peroxidation, including free F(2)-isoprostanes, malondialdehyde (MDA), and thiobarbituric acid reacting substances (TBARS), in the plasma of postmenopausal women taking dietary oil supplements rich in oleate, linoleate, and both eicosapentaenoic acid and docosahexaenoic acid. DESIGN: Fifteen postmenopausal women took 15 g sunflower oil/d, providing 12.3 g oleate/d; safflower oil, providing 10.5 g linoleate/d; and fish oil, providing 2.0 g EPA/d and 1.4 g DHA/d in a 3-treatment crossover trial. RESULTS: Plasma free F(2)-isoprostane concentrations were lower after fish-oil supplementation than after sunflower-oil supplementation (P: = 0.003). When plasma free F(2)-isoprostane concentrations were normalized to plasma arachidonic acid concentrations, significant differences among the supplements were eliminated. Plasma MDA concentrations were lower after fish-oil supplementation than after sunflower-oil supplementation (P: = 0.04), whereas plasma TBARS were higher after fish-oil supplementation than after sunflower oil (P: = 0.003) and safflower oil (P: = 0.001) supplementation. When plasma MDA concentrations were normalized to plasma PUFA concentrations, significant differences were eliminated, but TBARS remained higher after fish-oil supplementation than after sunflower oil (P: = 0.01) and safflower-oil (P: = 0.0003) supplementation. CONCLUSIONS: With fish-oil supplementation, there was no evidence of increased lipid peroxidation when assessed by plasma F(2)-isoprostanes and MDA, although plasma TBARS was higher than with sunflower-oil and safflower-oil supplementation.

Oxidation of plasma proteins is not increased after supplementation with eicosapentaenoic and docosahexaenoic acids.

BACKGROUND: It is generally thought that as the intake of dietary polyunsaturated fatty acids increases, so should that of alpha-tocopherol, to protect the polyunsaturated fatty acids from increased in vivo peroxidation. However, there are little quantitative data about the concentration of alpha-tocopherol that is necessary when eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are consumed. OBJECTIVE: The purpose of this study was to measure changes produced in 2 indexes of lipid oxidation after supplementation with EPA and DHA from fish oil and 3 doses of RRR-alpha-tocopheryl acetate in postmenopausal women. DESIGN: Daily supplements of fish oil providing 2.5 g EPA and 1.8 g DHA and 0, 100, 200, or 400 mg alpha-tocopheryl acetate were given to 46 postmenopausal women in a 4-treatment, 4-period crossover design. RESULTS: The supplements increased plasma concentrations of EPA, DHA, and alpha-tocopherol. The fish-oil supplement increased the plasma concentration of thiobarbituric acid-reactive substances (TBARS) (P: = 0.0001) but not that of oxidatively modified protein, as indicated by the carbonyl content. The alpha-tocopheryl acetate and fish-oil supplements had no significant effect on plasma concentrations of TBARS or oxidized protein. CONCLUSIONS: Although these data show a small but statistically significant increase in oxidative stress on the basis of plasma TBARS concentrations after the consumption of EPA and DHA, the clinical relevance of this change is questionable. In addition, as supplements of alpha-tocopheryl acetate were added to the diet, neither the plasma TBARS concentration nor the protein oxidation changed. Consequently, the results of this study indicate that there is no basis for vitamin E supplementation after consumption of EPA and DHA.

Selenium supplementation of Chinese women with habitually low selenium intake increases plasma selenium, plasma glutathione peroxidase activity, and milk selenium, but not milk glutathione peroxidase activity.

Twenty-one pregnant women living in Xichang County, China, a selenium-deficient area, were divided into two groups and given either a placebo (n = 10) as yeast or selenium-enriched yeast tablets (n = 11) to provide 100 microg selenium per day. This supplementation was begun the last trimester of pregnancy and continued for 3 months after parturition. Plasma selenium levels and glutathione peroxidase (GPX) activity steadily declined in supplemented women, but a curvilinear response occurred in milk selenium and GPX activity in both supplemented and deficient women and in plasma selenium and GPX activity in deficient women. The milk selenium levels were higher in supplemented women but there were no differences in the milk GPX activity between the two groups of women. The plasma alpha-tocopherol concentrations declined after parturition in both groups but no differences were found between the two groups of women. Plasma thiobarbituric acid reactive substances declined in supplemented women but showed a curvilinear response in unsupplemented women, suggesting peroxidative stress in these women. GPX, selenium, and peroxidative responses in plasma and milk following parturition is advocated as a new method to assess selenium status of lactating women.

Higher total homocysteine concentrations and lower folate concentrations in premenopausal black women than in premenopausal white women.

BACKGROUND: Premenopausal black women have a greater rate of coronary artery disease (CAD) than do premenopausal white women. Plasma total homocysteine concentrations, a risk factor for CAD, have not been reported in premenopausal black women. OBJECTIVE: The purpose of this study was to compare plasma total homocysteine, folate, and vitamin B-12 concentrations in premenopausal black and white women. DESIGN: Eighty-nine black and 90 white, healthy, premenopausal women living in Portland, OR, were recruited. Dietary histories were obtained by using the Diet Habit Survey, a 40-item eating-behavior questionnaire. Plasma concentrations of total homocysteine, folate, and vitamin B-12 were measured. RESULTS: Black women had higher plasma total homocysteine (8.32 compared with 7.60 micromol/L;P = 0. 013), lower plasma folate (6.62 compared with 9.88 nmol/L;P < 0. 0001), and higher vitamin B-12 (355 compared with 283 pmol/L;P < 0. 001) concentrations than white women. White women had a greater rate of daily multivitamin supplement use (42.4% compared with 24.7%;P = 0.019) and ate more ready-to-eat cereal than did black women. After adjustment for multivitamin use and intake of ready-to-eat cereal, plasma total homocysteine concentrations did not differ significantly, but plasma folate remained significantly lower in the black women. None of the black women but 12.3% of the white women (P = 0.013) were homozygous for the cytosine to thymidine mutation at nucleotide 677 in the methylenetetrahydrofolate reductase gene. CONCLUSIONS: Black women had higher plasma total homocysteine and lower plasma folate concentrations than white women, largely because of lifestyle factors, which may contribute to the greater rate of CAD in premenopausal black than in white women.

Glutathione peroxidase activity modulates fatty acid profiles of plasma and breast milk in Chinese women.

Since little is known about the effect of selenium on the fatty acid profiles (FAP) of human breast milk, the purpose of this study was to measure the effect of habitual dietary selenium (Se) intake on this profile in plasma and breast milk. Subjects were lactating women from three locations in China where habitual selenium intakes are extremely low (Xichang), adequate (Beijing), or extremely high (Enshi). Plasma and milk samples were obtained within seven days of parturition (early samples) or within eighteen months postpartum (mature samples) and analyzed for selenium concentration, glutathione peroxidase (Gpx) activity and FAP. Plasma and milk selenium concentrations were significantly lower in the samples from women from Xichang and significantly higher in those from Enshi when compared to those from Beijing. Plasma Gpx activity, however, was higher in samples from Beijing than Xichang or Enshi. In contrast, the early breast milk samples had similar Gpx activity regardless of location. The mature samples, however, followed the same trend as plasma with the samples obtained from the women in Beijing having the highest activity. Of the unsaturated fatty acids examined, the concentration of linoleic acid, 18:2(n-6), in both plasma and milk was greater in the samples from Beijing when compared to those from Xichang or Enshi. Thus dietary selenium appears to influence the fatty acid composition in human breast milk, but influences Gpx activity only in mature milk samples.

Influence of long-chain polyunsaturated fatty acids on oxidation of low density lipoprotein.

Enrichment of low density lipoprotein (LDL) with long-chain fatty acids, such as eicosapentaenoic acid (EPA; 20:5 n-3) and docosahexaenoic acid (DHA; 22:6 n-3) found in fish oil, is thought to increase its oxidative susceptibility although such an increase has not been clearly demonstrated. The purpose of this study was to determine the composition and fatty acid concentration of LDL obtained from postmenopausal women given a supplement of fish oil and relate these values to its oxidative susceptibility. Fish oil supplementation significantly increased LDL concentration of EPA (P = 0.0001) and DHA (P = 0.0001) and decreased that of linoleic acid P = 0.006). The concentration of free cholesterol, cholesterol ester, phospholipids and protein was unchanged while triglyceride concentration increased 8% (P = 0.02). Cu2+-mediated oxidation resulted in a shorter lag time, slower oxidation rate and similar concentrations of conjugated dienes of EPA/DHA-enriched LDL than EPA/DHA-unenriched LDL. Stepwise multiple regression indicated that the primary predictor of oxidative susceptibility of LDL was linoleic acid, even after enrichment with EPA and DHA. The oxidation rate of EPA/DHA-unenriched LDL correlated with the cholesteryl ester concentration (P = 0.003) while that of EPA/DHA-enriched correlated with the concentration of phospholipids (P = 0.03). These data suggest that EPA/DHA-enriched LDL have decreased oxidative susceptibility and that surface lipids may mediate its rate of oxidation.

The ratio of dietary (n-6) to (n-3) fatty acids influences immune system function, eicosanoid metabolism, lipid peroxidation and vitamin E status in aged dogs.

We studied the effects of feeding experimental diets containing (n-6) to (n-3) fatty acid ratios of 31:1, 5.4:1, and 1.4:1 to 20 healthy female geriatric Beagles (9.5-11.5 y) for 8-12 wk on various indices of the immune response. Compared with the 31:1 diet, consumption of the 5.4:1 and 1.4:1 diets significantly increased (n-3) fatty acids in plasma (2.17 +/- 0.64, 9.05 +/- 0.64, 17.46 +/- 0.64 g/100 g fatty acids, respectively, P < 0.0001). Although supplementation with (n-3) fatty acids did not significantly alter the humoral immune response to keyhole limpet hemocyanin (KLH), it significantly suppressed the cell-mediated immune response based on results of a delayed-type hypersensitivity (DTH) skin test. The DTH response after intradermal injection of KLH at 24 h was significantly lower in the group consuming the 1.4:1 diet compared with the group consuming the 5.4:1 (P = 0.02) or the 31:1 diets (P = 0.04), and remained significantly suppressed at 48 h in the group fed 1.4:1 relative to the group fed 31:1. After consumption of the 1.4:1 diet, stimulated mononuclear cells produced 52% less prostaglandin E2 (PGE2) than those from dogs fed the 31:1 diet (224 +/- 74 and 451 +/- 71 pmol/L, respectively, P = 0.04). Plasma concentration of alpha-tocopherol was 20% lower in dogs fed the 1.4:1 diet compared with those fed the 31:1 diet (P = 0.04), and lipid peroxidation was greater in both plasma (P = 0.03) and urine (P = 0.002). These data suggest that although a ratio of dietary (n-6) to (n-3) fatty acids of 1.4:1 depresses the cell-mediated immune response and PGE2 production, it increases lipid peroxidation and lowers vitamin E concentration.

alpha-Tocopherol influences in vivo indices of lipid peroxidation in postmenopausal women given fish oil.

Although diets containing fish have been shown to be therapeutically valuable, the vitamin E requirement when large quantities of (n-3) fatty acids are consumed is not known. Additionally, as estrogens may function as an antioxidant, the requirement may be modified in postmenopausal women using hormone replacement therapy (HRT). Consequently, the purpose of this study was to measure the impact of graduated doses of RRR-alpha-tocopheryl acetate (TA) on in vivo indices of lipid peroxidation in postmenopausal women with and without hormone replacement therapy when given a supplement of fish oil. Forty-eight postmenopausal women, half receiving (+HRT) and half not receiving (-HRT) hormone replacement therapy, participated in a four-period, double-blind crossover trial. Each period lasted 5 wk followed by a 4-wk washout interval. During each period, the subjects consumed a 15-g supplement of fish oil and either 0, 100, 200, or 400 mg TA/d in a balanced, single square dosing order. Plasma levels of (n-3) fatty acids were significantly higher after fish oil supplementation; alpha-tocopherol concentration of plasma was significantly higher at each level of supplementation compared with the level without supplementation. Urinary excretion of thiobarbituric acid reactive substances (TBARS) and malondialdehyde, measured as the thiobarbituric-malondialdehyde adduct (TRA-MDA adduct), and the plasma concentration of the adduct were significantly greater after the fish oil supplement. Although urinary TBARS decreased linearly as the dose of TA increases (P < or = 0.05), urinary and plasma concentrations of TBA-MDA adduct did not. This study suggests that the evaluation of highly unsaturated fatty acids as oxidative stressors requires several measures of assessment.

Effects of interaction of RRR-alpha-tocopheryl acetate and fish oil on low-density-lipoprotein oxidation in postmenopausal women with and without hormone-replacement therapy.

We evaluated the effects of RRR-alpha-tocpheryl acetate (alpha-tocopheryl acetate) and hormone-replacement therapy (HRT) on the oxidative susceptibility of low-density lipoprotein (LDL) in postmenopausal women consuming a fish oil supplement. The independent effect of fish oil was also assessed. Forty-eight women, equally divided between women using and not using HRT, participated in a double-blind crossover trial. Each of the four periods lasted 5 wk and was followed by a 4-wk washout interval. During each period all subjects were given a 15-g supplement of fish oil and either 0 (placebo), 100, 200, or 400 mg alpha-tocopheryl acetate daily. LDL resistance to oxidative modification was assessed by calculating lag time, propagation rate, and maximum production of conjugated dienes. Supplementation with fish oil and placebo shortened lag time and slowed propagation rate in women both using and not using HRT. After subjects consumed fish oil, supplementation with alpha-tocopheryl acetate increased plasma and LDL alpha-tocopherol contents significantly and lengthened lag time (at even the lowest concentration) but had no significant effect on propagation rate or maximum production compared with values measured after consumption of fish oil alone. Women not using HRT had faster propagation rates and higher maximum production than women using HRT; after supplementation with fish oil and alpha-tocopheryl acetate these differences prevailed. Supplements as low as 100 mg alpha-tocopheryl acetate/d increase the resistance of LDL to oxidation when fish oil supplements are used. HRT and fish oil supplements may independently affect LDL oxidative susceptibility.

Effects of dietary oils and methyl ethyl ketone peroxide on in vivo lipid peroxidation and antioxidants in rat heart and liver.

Weanling male Sprague-Dawley rats were fed diets for four weeks which differed in their content of n-6 (corn oil; CO) and n-3 fatty acids (fish oil; FO), but were similar in their content of saturated and monounsaturated fatty acids and vitamin E. At the end of the four-week feeding period, each dietary group was subdivided into two groups. One group received a single placebo injection of alpha-tocopherol-stripped corn oil (TSCO); the other group received a single injection of the free radical generator; methyl ethyl ketone peroxide (MEKP), in TSCO. Twenty-four hours after injection, the effect of dietary oil and MEKP treatment on endogenous lipid peroxide (LPO) production (measured as methylene blue formed by the "Determiner LPO" assay), glutathione (GSH) and vitamin E content, and fatty acid composition of phosphatidylcholine and phosphatidylethanolamine in heart and liver from unfasted animals were measured. FO-fed rats had significantly heavier hearts and livers, increased levels of n-3 fatty acids in membrane phospholipids, and higher liver LPO levels than CO-fed rats. MEKP treatment resulted in significantly lower body weights and liver GSH levels. The data indicate that dietary n-3 fatty acids increase lipid peroxidation in liver somewhat more than in heart. The study also demonstrates that the effect of induced oxidative stress due to a single dose of MEKP on lipid peroxide formation and antioxidant status in tissues from unfasted animals was independent of the dietary oils.

The effect of porcine somatotropin supplementation in pigs on the lipid profile of subcutaneous and intermuscular adipose tissue and longissimus muscle.

The effect of porcine somatotropin (pST) on the lipid profiles of adipose tissue and muscle was investigated. Sixteen crossbred barrows were injected daily with either 3 mg of pST or a placebo. After slaughter, total lipid and fatty acid composition of raw subcutaneous (SC) adipose and intermuscular (IM) adipose tissue and longissimus muscle were determined. The SC adipose tissue from pST-treated pigs had a 7.5% decrease in total lipid content; specific fatty acids 16:0, 18:0, and 18:1(n-9)c decreased most. The IM fat from pST-treated pigs had lower levels of 16:0 and 20:0. There was no effect of pST treatment on the lipid profile of the longissimus muscle. The data suggest that pST treatment produces small but significant changes in the saturated fatty acid content of adipose tissue in pigs.

Effects of dietary selenium and fish oil (MaxEPA) on arachidonic acid metabolism and hemostatic function in rats.

This study investigated whether hemostatic function can be modified by both the consumption of fish oil and the level of dietary selenium. Male Sprague-Dawley rats were fed for 8 wk semipurified diets containing 7% corn oil (by wt) or 5.5% fish oil (MaxEPA) plus 1.5% corn oil with or without selenium supplementation. Consumption of the four diets caused no difference in weight gain, food intake or plasma malondialdehyde content. The selenium-supplemented rats had significantly higher levels of selenium and glutathione peroxidase activity in plasma. Fish oil feeding decreased ADP-induced platelet aggregation and increased bleeding time. The level of dietary selenium and type of oil interacted to influence the production of 6-keto-prostaglandin F1 alpha: more was produced when corn oil was fed in the selenium-deficient diets. These data suggest that the effect of dietary selenium on hemostatic function and the production of eicosanoids is minor.

Diet effects on membrane phospholipid fatty acids and mitochondrial function in BHE rats.

The effects of feeding a 5% corn oil or coconut oil diet on the composition of hepatic phospholipid fatty acids and on hepatic mitochondrial function were studied. Male BHE weanling rats were fed a 65% starch diet containing 5% corn or coconut oil. Rats were decapitated, and hepatic tissue was used for phospholipid fatty acid analysis and for the preparation of mitochondria. Mitochondrial ATPase activity, alpha-glycerophosphate and malate-aspartate shuttle activity, and succinate- or pyruvate-supported respiration were determined. Livers from rats fed the coconut oil diet had more saturated phospholipid fatty acids than those from rats fed the corn oil diet. ATPase activity and the activity of the malate-aspartate shuttle were not affected by diet. The activity of the alpha-glycerophosphate shuttle was greater in rats fed the coconut oil diet than in rats fed the corn oil diet. Succinate-supported state 3 respiration was not affected by diet, whereas succinate-supported state 4 respiration was higher in mitochondria from rats fed coconut oil than in rats fed corn oil. Evidence of uncoupling of pyruvate-supported respiration from ATP synthesis was observed in mitochondria from rats fed coconut oil but not in rats fed corn oil. These observations suggest that the inherent tendency of the BHE rat toward looser coupling of respiration to ATP synthesis is potentiated by the feeding of the highly saturated fat, hydrogenated coconut oil.

Effects of type of dietary fat and carbohydrate on gluconeogenesis in isolated hepatocytes from BHE rats.

The effect of type of dietary fat and carbohydrate on gluconeogenesis and ketogenesis by isolated hepatocytes was studied. BHE male weanling rats were fed one of six diets: 64% sucrose or cornstarch with 6% corn oil, 6% hydrogenated coconut oil, or a 1:2 mixture of the two oils. At 100 d of age the rats were anesthetized, and isolated hepatocytes were prepared. The cells were incubated with lactate, lactate and lysine, lactate and pyruvate, lactate and palmitate, lactate and linoleate, lactate and epinephrine or lactate and glucagon. The hepatocytes from the rats that had been fed hydrogenated coconut oil produced significantly more glucose than the rats fed either corn oil or a mixture of oils, regardless of the type of carbohydrate fed. Each of the additives in turn, except for epinephrine, stimulated glucose production above that obtained with lactate alone. However, when expressed as a percent increase above that from lactate there was no effect of fat type on the magnitude of this stimulation. We interpret these data to mean that, although the metabolic pathways function equally well in the hepatocytes isolated from rats fed hydrogenated coconut oil and rats fed corn oil, the flux through these pathways can be influenced by the type of dietary fat.