FA Supplementation Research Articles

216 EFFECTS OF DIFFERENT POLYUNSATURATED FATTY ACID SUPPLEMENTATION DURING THE POSTPARTUM PERIODS OF EARLY LACTATING DAIRY COWS ON INSULIN RESISTANCE AND SOMATOTROPIC AXIS

During periods of negative energy balance, GH cannot stimulate hepatic IGF-1; the condition is termed GH resistance (Donaghy and Baxter, 1996). Studies show that insulin may be a key metabolic signal that regulates the coupling of the somatotropic axis (Butler et al., 2003). The objectives were to test the hypothesis that α-linoleic acid (n-3), or linoleic acid (n-6) could promote insulin resistance in dairy cows. For this goal, Holstein cows were assigned randomly to be fed either 1-soybean whole roast (S, n = 4), or 2-linseed extruded (L, n = 4), or 3-palm oil use a source of saturated fatty acid (C, n = 4), respectively, from calving to 60 DIM. Three diets were formulated to have equal concentrations of DM, ME, and CP, but to differ in ratio of n-3/n-6 PUFA (4.2, 1.2, and 3.2 for S, L, and C treatments, respectively). Supplementation of FA was at 1.5% of dietary dry matter. There was no difference between groups (mean ± SEM) in parity (3.0 ± 1.90) or BCS at calving day (3.2 ± 0.07). At Day 60 postpartum, liver tissues (~100 mg) were collected by needle biopsy as described by Rhoads et al. (2008) with minor modifications. Liver tissue was washed with sterile PBS, placed in a screw-cap micro centrifuge tube, snap-frozen in liquid nitrogen, and stored at –80°C until RNA extraction. We applied real-time PCR for measurement of mRNA abundance. Real-time PCR was executed in triplicate. Reaction mixtures for real-time PCR included 1.5 µL of cDNA as template, 7.5 µL of SYBRR Green PCR Master Mix (ABI, USA), 1 µL of forward primer, 1 µL of reverse primer, and 4 µL of double distilled water. The efficiencies for GHR1A, insulin receptor (IR), IGF-I, and IGFBP-2 were 1.22, 0.99, 1.02, and 1 with an R2 of 0.99 or above for the regression. We used the 2–ΔΔCt method in this model. Rotor Gene analysis software (version 6.0) was used to analyse all the results from the PCR assays. Data were analysed using the MIXED procedure of SAS (2000, SAS Institute Inc., Cary, NC, USA), in a completely randomised design. Results showed that supplementation with a high level of flaxseed on treatment L for 2 months increased levels of IR mRNA compared with S and C treatments (P < 0.05). The levels of IR mRNA were not different between S and C treatments (P = 0.202). There was also a strong tendency towards a 2.2-fold increase (P = 0.06) in GHR1A gene expression in cows on the flaxseed diet compared with the other diets, also there were no significant difference between S and C treatments. The IGF-I and IGFBP mRNA were not different between the treatments (P > 0.05). In conclusion, n-3 fatty acids promoted insulin resistance via increasing insulin receptor signalling that could improve milk production and specifically fertility in lactating dairy cows.

178 20-HETE CONTRIBUTES TO THE BLOOD PRESSURE REDUCTION FOLLOWING OMEGA-3 FATTY ACID SUPPLEMENTATION IN PATIENTS WITH CHRONIC KIDNEY DISEASE

Background and Objectives: Omega-3 fatty acids (n-3FA), specifically eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), are protective against cardiovascular disease. n-3FA have multiple biological effects including benefits on blood pressure (BP). This study aimed to determine whether the reduction in 24-h ambulatory BP following supplementation with n-3 FA in patients with chronic kidney disease (1), was related to changes in the cytochrome P450 metabolite 20-HETE, a regulator of vascular tone and BP, and implicated in the pathogenesis of hypertension. Design and Methods: In a double-blind, placebo-controlled intervention, patients were randomized to either n-3FA (4 g), coenzyme Q10 (200 mg), both supplements, or control (4 g), daily for 8 weeks. All measures including 24-h ambulatory BP, plasma 20-HETE and F2-isoprostanes, and platelet fatty acids were recorded at baseline and after 8 weeks. Results: n-3FA, but not coenzyme Q10, reduced 24-h ambulatory BP by -3.3±0.7/-2.9 ± 0.5 mmHg (both P<0.0001). n-3 FA significantly reduced plasma 20-HETE by 28% (1217 ± 72 to 876 ± 69 pmol/L). Plasma 20-HETE levels were lower in females than males (P = 0.004) and significantly related to platelet EPA (b = -108.0, P = 0.013) and arachidonic acid (b = 20.7, P = 0.008), and plasma F2-isoprostanes (b = 0.7, P<0.0001). The fall in BP was positively associated with the reduction in plasma 20-HETE in regression analyses that adjusted for baseline BP (24-h SBP, P = 0.018 and 24-h DBP, P = 0.028). Conclusion: Reduced 20-HETE may play an important role in mediating the fall in BP following n-3 FA supplementation in patients with chronic kidney disease. Reference: Mori TA, et al. J Hypertension 2009;27:1863.

Ω3 fatty acids may reduce hyperlipidemia in pediatric renal transplant recipients

Life expectancy after pediatric renal transplantation remains lower than that of the normal population largely due to cardiovascular morbidity and mortality. Hyperlipidemia is a potentially modifiable risk factor for cardiovascular morbidity. Retrospective chart review of all available pediatric renal transplant patients (26) in a single center with assessment of anthropometry, renal function, steroid, calcineurin or mTOR inhibitor exposure and Ω3 FA supplementation. Eighteen transplant recipients without Ω3 FA supplementation served as control. Nutrition and supplement surveys were conducted with standardized questionnaires. Fasting cholesterol values were compared using the latest value prior to start of Ω3 FA and at last follow-up. Eight patients (five receiving mTOR inhibitor) started Ω3 FA supplementation at a mean dose of 29.2 ± 12 mg of EPA/kg and 16.1 ± 7.4 mg DHA/kg body weight. Median duration of treatment was 2.5 yr (range 0.8-5.9 yr) and their total fasting cholesterol at last follow-up dropped significantly from 5.08 ± 0.97 (control group 3.77 ± 0.81, p = 0.0084) to 4.17 ± 0.54 mm (p = 0.0158). High-density lipoprotein cholesterol increased not significantly from 1.74 ± 0.49 to 2.02 ± 0.93 mm. No patient had increased bleeding. Supplementation of omega-3 FAs may reduce hyperlipidaemia after pediatric renal transplantation.

Ethanol and acetaldehyde mediate folic acid and human papillomavirus-induced proliferation of oral squamous cell carcinoma cells in vitro

Background Although great scientific emphasis has been placed upon HPVas the primary cause of cervical cancers and its involvement in carcinogenic progression of other cancers, less attention has been focused on the secondary factors that are associated with progression from subclinical HPV infection to invasive carcinoma. Among the secondary factors that limit virus production and carcinogenic progression is CpG methylation of the HPV genome. Several studies now confirm that CpG site-specific methylation of HPV DNA, mediated in part by folate availability, is sufficient to suppress neoplastic progression. In contrast, demethylation or hypomethylation of HPV-DNA sequences is required for transformation, revealing the importance of preferential DNA methylation at CpG sites in the HPV long control region (LCR) between L1 and E6 HPV genes, in addition to the tumor suppressor sites in p53 exons 248 and 273. Because HPV has the potential to initiate oncogenesis, and also to modulate oral cancer growth and folate plays a central role in mediating the availability of methyl groups for CpG-specific DNA methylation (modulating both p53 and HPV mRNA expression) – an investigation of these inter-connected and inter-related mechanisms in oral cancers must be undertaken. In addition, ethanol and acetaldehyde may also play critical roles in determining folate availability, and are primary risk factors for the development of oral cancers, which makes the evaluation of these interconnected metabolic pathways critically important. Materials and methods Using a comprehensive series of integrated in vitro assays, including proliferation, viability and mRNA analysis using RT-PCR, distinct effects of ethanol and acetaldehyde administration were observed in the oral cancer cell lines, CAL27, SCC15 and SCC25. In addition, the growth modulating effects of HPV infection and FA supplementation were also examined.

Effects of DHA-rich n-3 fatty acid supplementation on gene expression in blood mononuclear leukocytes: the OmegAD study.

BackgroundDietary fish oil, rich in n-3 fatty acids (n-3 FAs), e.g. docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), regulate inflammatory reactions by various mechanisms, e.g. gene activation. However, the effects of long-term treatment with DHA and EPA in humans, using genome wide techniques, are poorly described. Hence, our aim was to determine the effects of 6 mo of dietary supplementation with an n-3 FA preparation rich in DHA on global gene expression in peripheral blood mononuclear cells.Methods and FindingsIn the present study, blood samples were obtained from a subgroup of 16 patients originating from the randomized double-blind, placebo-controlled OmegAD study, where 174 Alzheimer disease (AD) patients received daily either 1.7 g of DHA and 0.6 g EPA or placebo for 6 months. In blood samples obtained from 11 patients receiving n-3 FA and five placebo, expressions of approximately 8000 genes were assessed by gene array. Significant changes were confirmed by real-time PCR. At 6 months, the n-3 FAs group displayed significant rises of DHA and EPA plasma concentrations, as well as up- and down-regulation of nine and ten genes, respectively, was noticed. Many of these genes are involved in inflammation regulation and neurodegeneration, e.g. CD63, MAN2A1, CASP4, LOC399491, NAIP, and SORL1 and in ubiqutination processes, e.g. ANAPC5 and UBE2V1. Down-regulations of ANAPC5 and RHOB correlated to increases of plasma DHA and EPA levels.ConclusionsWe suggest that 6 months of dietary n-3 FA supplementation affected expression of genes that might influence inflammatory processes and could be of significance for AD.Trial RegistrationClinicalTrials.gov NCT00211159

Abstract 5432: The effect of folic acid supplementation on chemosensitivity of colon cancer cells to 5-fluorouracil in a xenograft model

Abstract Background: As a mediator of one-carbon transfer reactions, folate is an essential cofactor in nucleotide biosynthesis. Interruption of intracellular folate metabolism and the consequent inhibition of tumor growth has been the basis for chemotherapy using antifolates and 5-fluorouracil (5FU). A growing body of evidence suggests that folic acid (FA; the synthetic form) may promote the progression of established colorectal (pre)neoplastic lesions. Folate blood levels in North America have dramatically increased over the past decade owing to FA fortification and widespread supplement use. Furthermore, over 50% of newly diagnosed colorectal cancer patients use vitamin supplements containing FA in addition to chemotherapeutic agents whose mechanisms of action are based on interruption of folate metabolism. This study therefore investigated whether FA supplementation can affect chemosensitivity of colon cancer cells to 5FU, the cornerstone of colorectal cancer treatment, in vivo. Methods: Male athymic nude mice were placed on the control diet containing 2 mg FA/kg diet and were inoculated with human HCT116 colon cancer cells. When xenografts were established, the animals were randomized to receive a diet containing 2, 8, or 25 mg FA/kg diet for eight weeks. Within each diet group, the animals were further randomized to receive saline vehicle or 5FU (20 mg/kg) + leucovorin (1 mg/kg) for five days. The primary endpoints were the rate of xenograft growth and the final volume. At necropsy, blood was collected and xenografts were harvested and evidence of metastasis was determined. Results: Plasma folate concentrations of animals on the 2 mg FA diet were significantly lower than those of animals fed the 8 and 25 mg FA diets (p&amp;lt;0.05). Xenograft folate concentrations significantly reflected the supplemental levels of FA in a dose-responsive manner (p=0.002). The effect of dietary levels of FA and 5FU alone were not significant, p=0.907 and p=0.193, respectively, but the interaction effect was significant (p&amp;lt;0.007). 5FU was beneficial only to animals fed the 2 mg FA diet (p&amp;lt;0.0001). In contrast, animals fed the 8 mg FA diet fared worse on 5FU (p=0.048) whereas animals fed the 25 mg FA diet demonstrated no effect of 5FU (p=0.31). All animals exhibited macroscopically evident lymph node involvement, but no distant metastases, irrespective of dietary levels of FA. Conclusion: Our data suggests that FA supplementation may adversely affect chemosensitivity to 5FU chemotherapy, although a dose-responsive relationship was not evident. Results of this study pose a legitimate public health concern given the drastically increased intake of FA from supplements, particularly among recently diagnosed colorectal cancer patients and from mandatory fortification. Further studies are warranted to confirm our findings and to elucidate the underlying mechanisms. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5432. doi:1538-7445.AM2012-5432

Inflammation and Insulin Resistance

Current evidence indicates that chronic activation of the proinflammatory state may be an underlying mechanism for insulin resistance. Together, these findings suggest that increased macrophage activation can be an important contributor to the pathogenesis of cellular insulin resistance. To assess this concept, we have used genetic deletion of key molecules controlling the inflammatory pathway, in mouse KO studies. Macrophage specific deletion of IKKβ or JNK1 disables the cytokine stimulated NFκB or JNK1/AP2 proinflammatory pathways leading to decreased tissue inflammatory markers, enhanced insulin sensitivity, and protection from the effects of high fat diet to cause insulin resistance. The enhanced insulin sensitivity and protection against insulin resistance were manifested in liver, muscle, and adipose tissue, indicating that inhibition of the inflammatory pathway within macrophages leads to a global state of heightened insulin sensitivity. Taking another approach to this issue, omega‐3 fatty acids (ω‐3 FAs), DHA and EPA, exert anti‐inflammatory effects, but the mechanisms are poorly understood. We found show that the G protein‐coupled receptor 120 (GPR120) functions as an ω‐3 FA receptor/sensor. Stimulation of GPR120 with ω‐3 FAs or a chemical agonist causes broad anti‐inflammatory effects in monocytic RAW 264.7 cells and in primary intraperitoneal macrophages. All of these effects are abrogated by GPR120 knockdown. Since chronic macrophage‐mediated tissue inflammation is a key mechanism for insulin resistance in obesity, we fed obese WT and GPR120 knockout mice a high fat diet with or without ω‐3 FA supplementation. The ω‐3 FA treatment inhibited inflammation and enhanced systemic insulin sensitivity in WT mice, but was without effect in GPR120 knockout mice. Thus, GPR120 is a functional ω‐3 FA receptor/sensor and mediates potent insulin sensitizing and anti‐diabetic effects in vivo by repressing macrophage‐induced tissue inflammation.

Omega-3 Fatty Acids and Vitamin D in Cardiology

Dietary modification and supplementation play an increasingly important role in the conservative treatment of cardiovascular disease. Current interest has focused on n-3 polyunsaturated fatty acids (PUFA) and vitamin D. Clinical trial results on this subject are contradictory in many aspects. Several studies indicate that n-3 PUFA consumption improves vascular and cardiac hemodynamics, triglycerides, and possibly endothelial function, autonomic control, inflammation, thrombosis, and arrhythmia. Experimental studies show effects on membrane structure and associated functions, ion channel properties, genetic regulation, and production of anti-inflammatory mediators. Clinical trials evaluating a possible reduction in cardiovascular disease by n-3 PUFA have shown different results. Supplementation of vitamin D is common regarding prevention and treatment of osteoporosis. But vitamin D also seems to have several effects on the cardiovascular system. Vitamin D deficiency appears to be related to an increase in parathyroid hormone levels and can predispose to essential hypertension and left ventricular hypertrophy, increased insulin resistance, and eventually to atherosclerosis and adverse cardiovascular events. Randomized prospective clinical trials are needed to determine whether vitamin D and omega-3 FA supplementation therapy should be recommended as a routine therapy for primary or secondary prevention of cardiovascular disease.

Maternal omega-3 fatty acid supplementation protects against lipopolysaccharide-induced white matter injury in the neonatal rat brain

Objectives: Periventricular leukomalacia (PVL) is the predominant form of brain injury in premature infants, and no specific treatment currently exists for this condition. We have evaluated whether maternal omega-3 fatty acid (ω3 FA) treatment reduces endotoxin-induced PVL in the developing rat brain. Methods: Wistar rats with dated pregnancies were fed a standard diet or a diet enriched in ω3 FA (70% docosahexaenoic acid + 30% eicosapentaenoic acid mixture) during gestation. Intraperitoneal injection of lipopolysaccharide (LPS) was administered consecutively on the 18th and 19th embryonic days to establish the endotoxin-induced PVL rat model. The animals were divided into four groups: (i) control, (ii) PVL, (iii) PVL+low-dose ω3 FA and (iv) PVL+high-dose ω3 FA. At day P7, apoptosis and hypomyelination in periventricular white matter were evaluated by immunohistochemical assessments. Results: High-dose maternal ω3 FA treatment reduced brain weight loss. Maternal ω3 FA treatment given either in low or high doses greatly decreased caspase-3 immunoreactivity and increased myelin basic protein immunoreactivity, indicating a decrease in apoptosis and hypomyelination. Conclusion: Considering that no specific treatment is available for PVL, maternal ω3 FA supplementation may provide a nutritional strategy to limit periventricular white matter damage caused by infections during pregnancy.

Supplementation effect(s) of organic acids and/or lipid to plant protein-based diets on juvenile yellowtail, Seriola quinqueradiata Temminck et Schlegel 1845, growth and, nitrogen and phosphorus excretion

A feeding experiment was conducted to investigate the effect of organic acids and/or lipid supplementation on growth, utilization and environmental loading of nitrogen (N) and phosphorus (P) in juvenile yellowtail fed fishmeal (FM) and plant protein (PP) diets. Six diets as FM (FM-based), FM+P (FM with inorganic P), FM+L (FM with lipid), PP+CA (PP with citric acid), PP+L+CA (PP with lipid and citric acid) and PP+L+FA (PP with formic acid) were formulated. Yellowtails were fed each of the diets in duplicate groups; once a day, 6 days a week to near satiation at water temperature 19.0–25.0 °C for 16 weeks. Fishmeal with inorganic P gave the best growth while PP+L+FA the lowest. However, growth increased in PP+CA and PP+L+CA. Addition of lipid significantly increased N and P retention resulting in significant reduction in N and P excretion. Citric acid and FA supplementation to PP diets also increased retention of P; hence, its excretion was lowered. Thus, CA, FA and lipid in juvenile yellowtail diets can help to partially replace FM with PP sources and reduce inorganic P use to minimize environmental loading from aquafeeds.

Endothelial microparticle formation in moderate concentrations of homocysteine and methionine in vitro.

Microparticles (MPs) are small membrane vesicles released by stimulated or apoptotic cells, including the endothelium. Hyperhomocysteinemia (HHcy) is a blood disorder characterized by an increase in the plasma concentrations of total homocysteine (Hcy). The plasma Hcy level is determined by environmental factors (dietary habits, i.e. the intake of folic acid, FA) and genetic factors (N 5,N 10-methylenetetrahydro-folate reductase, MTHFR, polymorphism 677C>T). To evaluate whether moderate Hcy concentrations induce endothelial MP formation, the role of FA supplementation and the influence of MTHFR polymorphism were analysed. Human umbilical vein endothelial cells (HUVEC) were treated in vitro with 50 μM of Hcy and methionine (Met). The MP number and apoptotic phenotype were analyzed using flow cytometry. Increasing doses of FA (5, 15 and 50 μM) were used to reduce the HHcy effect. The MTHFR 677C>T polymorphism was determined. HUVEC stimulated by Hcy produced significantly more MPs than HUVEC under the control conditions: 3,551 ± 620 vs 2,270 ± 657 kMP (p = 0.02). Supplementation with FA at concentrations of 5, 15 and 50 μM reduced the MP count in the cell culture supernatant to 345 ± 332, 873 ± 329, and 688 ± 453 kMP, respectively (p = 0.03). MTHFR 677C>T heterozygosity was associated with a significant increase in MP formation after stimulation with Hcy compared to the control conditions: 3,617 ± 152 vs 1,518 ± 343 kMP (p = 0.02). Furthermore, the MTHFR genotype altered MP formation after Met loading. On average, 24% of the entire MP population was apoptotic (annexin V-positive). Endothelial function impairment due to HHcy is related to MP shedding, which may involve platelets and other blood and vascular cells. MP shedding is a physiological response to moderate HHcy.

Knowledge on and intake of Folic acid supplementation in pregnant women in Denmark

Results 82% had knowledge of FA supplementation and 89% had a FA supplementation. 51% followed national recommendations. We found statistically significant correlation between higher educational level and knowledge about FA supplementation, actual supplementation of FA and FA supplementation in correlation with national recommendations. No statistical relations were found between number of pregnancies or age and FA related parameters. Family, friends, general practitioner (GP) and the internet were main information sources.

2,4-Dinitrochlorobenzene으로 유도된 아토피 피부염 동물모델에서 비파엽 및 삼백초 추출발효물의 항아토피 활성

본 연구는 in vitro 실험결과 아토피 피부염의 억제 효능이 기대되는 비파엽 추출물(BI군), 삼백초 추출물(SA군), 비파엽 추출물과 삼백초 추출 혼합물(FB군) 그리고 삼백초와 비파엽 추출 혼합 발효물(FA군)을 경구 투여하여 DNCB 처리한 아토피 실험동물을 대상으로 아토피 질환 개선에 대한 유효성을 평가하였다. 혈청 중의 IgE 수치를 측정한 결과, DNCB군에서 227.21±60.04 ng/mL를 나타내었으며, FA군에서는 154.78±78.99 ng/mL로 IgE 생성량이 감소되었다. 혈청내의 히스타민 농도는 DNCB군에서 1.90±0.04 ng/mL였으며, FA, FB, BI, SA군에서 각각 1.47±0.20 ng/mL, 1.41±0.07 ng/mL, 1.61±0.05 ng/mL, 1.08±0.07 ng/mL로 DNCB군과 비교 시 유의적인 감소를 나타내었다. 세라마이드 총 함량을 측정한 결과, FA군에서 1,350±357 μg/g으로 DNCB와 비교 시 세라마이드 함량이 유의적으로 증가되었다. IFN-γ의 경우, DNCB군에서 2.5±0.5 ng/mL로 분석되었으며, FA, FB, BI 및 SA군에서 각각 7.1±1.3 ng/mL, 7.7±0.2 ng/mL, 11.5±0.6 ng/mL 및 6.9±0.2 ng/mL로 DNCB 군과 비교 시 모두 유의적인 증가를 나타내었다. 특히 비파엽 및 삼백초 추출물 혼합 발효물은 등 부위 피부의 육안 증상을 관찰한 결과, 가피형성과 발적 감소가 가장 우수하여 아토피 피부염 질환 개선에 도움을 줄 수 있을 것으로 기대된다.

GPR120 Is an Omega-3 Fatty Acid Receptor Mediating Potent Anti-inflammatory and Insulin-Sensitizing Effects

Omega-3 fatty acids (omega-3 FAs), DHA and EPA, exert anti-inflammatory effects, but the mechanisms are poorly understood. Here, we show that the G protein-coupled receptor 120 (GPR120) functions as an omega-3 FA receptor/sensor. Stimulation of GPR120 with omega-3 FAs or a chemical agonist causes broad anti-inflammatory effects in monocytic RAW 264.7 cells and in primary intraperitoneal macrophages. All of these effects are abrogated by GPR120 knockdown. Since chronic macrophage-mediated tissue inflammation is a key mechanism for insulin resistance in obesity, we fed obese WT and GPR120 knockout mice a high-fat diet with or without omega-3 FA supplementation. The omega-3 FA treatment inhibited inflammation and enhanced systemic insulin sensitivity in WT mice, but was without effect in GPR120 knockout mice. In conclusion, GPR120 is a functional omega-3 FA receptor/sensor and mediates potent insulin sensitizing and antidiabetic effects in vivo by repressing macrophage-induced tissue inflammation.

Meta‐analysis of cancer incidence in folic acid supplementation trials

It has been suggested by several research groups that folate is procarcinogenic. Data in the 1940s of folate supplementation indicated that it enhanced the proliferation of cancer cells. Folate is a unique vitamin because its coenzymes are required for de novo adenine, guanine, and thymine nucleotide biosynthesis; therefore, it is essential for DNA synthesis and repair, and RNA synthesis, and structurally related folate analogs, such as methotrexate, are used as anticancer agents. Using a meta‐analysis weighted for the duration of folic acid (FA, pteroylglutamic acid) supplementation, we analyzed the cancer incidence of 5 published large prospective FA supplementation trials that lasted 1 year or more involving men and women. These articles were selected from over 1,100 identified by a PubMed search that was done in June 2009, and skin cancer was excluded by the original investigators or by us. Our data suggest that cancer incidence was higher in the FA supplemented groups than the non‐FA supplemented groups (relative risk = 1.26 [95% confidence interval: 1.05–1.51]). FA supplementation trials should be performed with careful monitoring of cancer incidence by investigators, and funding agencies should provide long‐term support. In the US, solid monitoring systems to detect side effects, such as increased cancer risk, should have been established before the food FA fortification mandate.

Effects of n–3 fatty acids on macro- and microvascular function in subjects with type 2 diabetes mellitus

Recent evidence supports the protective effects of n-3 (omega-3) fatty acids (n-3 FAs), such as eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on vascular function. We investigated the effects of EPA and DHA on postprandial vascular function in subjects with type 2 diabetes mellitus. In a double-blind, placebo-controlled, randomized, crossover manner, 34 subjects with type 2 diabetes mellitus received daily either 2 g purified EPA/DHA (termed n-3 FAs) or olive oil (placebo) for 6 wk. At the end of this period, we measured macrovascular (brachial ultrasound of flow-mediated dilatation; FMD) and microvascular [laser-Doppler measurements of reactive hyperemia (RH) of the hand] function at fasting and 2, 4, and 6 h after a high-fat meal (600 kcal, 21 g protein, 41 g carbohydrates, 40 g fat). Fasting vascular function remained unchanged after n-3 FAs and placebo. Postprandial FMD decreased from fasting after placebo, with a maximum decrease (38%) at 4 h-an effect that was significantly reduced (P = 0.03 for time x treatment interaction) by n-3 FA supplementation (maximum decrease in FMD was at 4 h: 13%). RH remained unchanged after placebo, whereas it improved significantly (P = 0.04 for time x treatment interaction) after n-3 FA supplementation (maximum increase was at 2 h: 27%). In subjects with type 2 diabetes mellitus, 6 wk of supplementation with n-3 FAs reduced the postprandial decrease in macrovascular function relative to placebo. Moreover, n-3 FA supplementation improved postprandial microvascular function. These observations suggest a protective vascular effect of n-3 FAs.

Mechanistic insights into folic acid-dependent vascular protection: Dihydrofolate reductase (DHFR)-mediated reduction in oxidant stress in endothelial cells and angiotensin II-infused mice: A novel HPLC-based fluorescent assay for DHFR activity

Folate supplementation improves endothelial function in patients with hyperhomocysteinemia. Mechanistic insights into potential benefits of folate on vascular function in general population however, remain mysterious. Expression of dihydrofolate reductase (DHFR) was markedly increased by folic acid (FA, 50 μmol/L, 24 h) treatment in endothelial cells. Tetrahydrofolate (THF) is formed after incubation of purified DHFR or cellular extracts with 50 μmol/L of substrate dihydrofolic acid. THF could then be detected and quantified by high performance liquid chromatography (HPLC) with a fluorescent detector (295/365 nm). Using this novel and sensitive assay, we found that DHFR activity was significantly increased by FA. Furthermore, FA improved redox status of Ang II treated cells by increasing H 4B and NO bioavailability while decreasing superoxide (O 2 − ) production. It however failed to restore NO levels in DHFR siRNA-transfected or methotrexate pre-treated cells, implicating a specific and intermediate role of DHFR. In mice orally administrated with FA (15 mg/kg/day, 16 days), endothelial upregulation of DHFR expression and activity occurred in correspondence to improved NO and H 4B bioavailability, and this was highly effective in reducing Ang II infusion (0.7 mg/kg/day, 14 days)-stimulated aortic O 2 − production. 5′-methyltetrahydrofolate (5′-MTHF) levels, GTPCH1 expression and activity remained unchanged in response to FA or Ang II treatment in vitro and in vivo. FA supplementation improves endothelial NO bioavailability via upregulation of DHFR expression and activity, and protects endothelial cells from Ang II-provoked oxidant stress both in vitro and in vivo. These observations likely represent a novel mechanism (intermediate role of DHFR) whereby FA induces vascular protection.

Folate rescues lithium-, homocysteine- and Wnt3A-induced vertebrate cardiac anomalies.

Elevated plasma homocysteine (HCy), which results from folate (folic acid, FA) deficiency, and the mood-stabilizing drug lithium (Li) are both linked to the induction of human congenital heart and neural tube defects. We demonstrated previously that acute administration of Li to pregnant mice on embryonic day (E)6.75 induced cardiac valve defects by potentiating Wnt-beta-catenin signaling. We hypothesized that HCy may similarly induce cardiac defects during gastrulation by targeting the Wnt-beta-catenin pathway. Because dietary FA supplementation protects from neural tube defects, we sought to determine whether FA also protects the embryonic heart from Li- or HCy-induced birth defects and whether the protection occurs by impacting Wnt signaling. Maternal elevation of HCy or Li on E6.75 induced defective heart and placental function on E15.5, as identified non-invasively using echocardiography. This functional analysis of HCy-exposed mouse hearts revealed defects in tricuspid and semilunar valves, together with altered myocardial thickness. A smaller embryo and placental size was observed in the treated groups. FA supplementation ameliorates the observed developmental errors in the Li- or HCy-exposed mouse embryos and normalized heart function. Molecular analysis of gene expression within the avian cardiogenic crescent determined that Li, HCy or Wnt3A suppress Wnt-modulated Hex (also known as Hhex) and Islet-1 (also known as Isl1) expression, and that FA protects from the gene misexpression that is induced by all three factors. Furthermore, myoinositol with FA synergistically enhances the protective effect. Although the specific molecular epigenetic control mechanisms remain to be defined, it appears that Li or HCy induction and FA protection of cardiac defects involve intimate control of the canonical Wnt pathway at a crucial time preceding, and during, early heart organogenesis.

Effects of Omega-3 Fatty Acids on Inflammatory Markers in Cerebrospinal Fluid and Plasma in Alzheimer’s Disease: The OmegAD Study

Background: ω-3 fatty acids (ω-3 FAs) found in dietary fish or fish oils are anti-inflammatory agents that may influence Alzheimer’s disease (AD). Objective: To study the effects of dietary ω-3 FA supplementation on inflammatory markers in cerebrospinal fluid (CSF) and plasma from patients with mild to moderate AD. Methods: Thirty-five patients (70.3 ± 8.2 years) were randomized to a daily intake of 2.3 g ω-3 FAs or placebo for 6 months. The inflammatory markers interleukin (IL)-6, tumour necrosis factor-α and soluble interleukin-1 receptor type II (sIL-1RII) were analysed in CSF and plasma at baseline and at 6 months. The AD markers tau-protein, hyperphosphorylated tau-protein and β-amyloid (Aβ_1–42) were assessed in CSF. High-sensitivity C-reactive protein was assessed in plasma. A possible relation to the APOE genotype was investigated. Results: There was no significant treatment effect of ω-3 FAs on inflammatory and AD biomarkers in CSF or on inflammatory markers in plasma, nor was there any relation with APOE. A significant correlation was observed at baseline between sIL-1RII and Aβ_1–42 levels in CSF. Conclusions: Treatment of AD patients with ω-3 FAs for 6 months did not influence inflammatory or biomarkers in CSF or plasma. The correlation between sIL-1RII and Aβ_1–42 may reflect the reciprocal interactions between IL-1 and Aβ peptides.

Omega-3 fatty acid supplementation decreases matrix metalloproteinase-9 production in relapsing-remitting multiple sclerosis ,

Objectives The primary objective was to evaluate the effect of omega-3 fatty acids (omega-3 FA) on matrix metalloproteinase-9 (MMP-9) production by immune cells in multiple sclerosis (MS). Quality of life, fatty acid levels, and safety were also evaluated. Materials and Methods Ten participants with relapsing-remitting MS (RRMS) received omega-3 FA supplementation (9.6 g/day fish oil) in an open-label study. Participants were evaluated at four time points, baseline, after 1 month of omega-3 FA supplementation, after 3 months of omega-3 FA supplementation, and after a 3-month wash out. Results Immune cell secretion of MMP-9 decreased by 58% after 3 months of omega-3 FA supplementation when compared with baseline levels ( p<0.01). This effect was coupled with a significant increase in omega-3 FA levels in red blood cell membranes. Conclusions Omega-3 FA significantly decreased MMP-9 levels in RRMS and may act as an immune-modulator that has potential therapeutic benefit in MS patients.