Showing papers in "Lipids in 2008"

Combining results of two GC separations partly achieves determination of all cis and trans 16:1, 18:1, 18:2 and 18:3 except CLA isomers of milk fat as demonstrated using Ag-ion SPE fractionation.

Abstract: Milk fat is a complex mixture of geometric and positional isomers of monounsaturated and polyunsaturated, including short-, long- and branch-chain fatty acids (FAs). There has been partial success to resolve this mixture of FAs using different GC temperature programs, or a combination of GC isothermal and temperature programs. To overcome the problem associated with overlapping isomers prior silver-ion separation was recommended. However, this procedure is time consuming and not practical for routine analysis. In addition, previous methods focused mainly on the trans and cis isomers of 18:1. The present method takes advantage of differences in the relative elution times between different types of FAs. The method involved analyzing each milk fat using the same highly polar 100-m capillary column and GC instrument, and conducting two separations using temperature programs that plateau at 175 and 150 °C. The relative shift among the geometric and positional isomers at these two temperature settings was enough to permit identification of most of the trans and cis 16:1, 18:1 and 20:1, the c/t-18:2 and the c/c/t-18:3 isomers found in milk fat. The identity of these FAs was confirmed by prior separation of the total fatty acid methyl esters (FAMEs) of milk fat using Ag+-SPE columns, and comparing the fractions to the total milk fat. The Ag+-SPE technique was modified to obtain pure saturated, trans- and cis-monounsaturated and diunsaturated FAMEs. By combining the results from these two separate GC analyses, knowing the elution order, it was possible to determine most of the geometric and positional isomers of 16:1, 18:1, 20:1, 18:2 and 18:3 without a prior silver-ion separation. Only few minor FAs could not be resolved, notable the conjugated linoleic acid isomers that still required the complimentary Ag+-HPLC separation. The two GC temperature programs have been successfully used to routinely analyze most FA isomers in total milk and beef fats in about 200 min without the use of prior silver-ion separations.

Comparison of Low Fat and Low Carbohydrate Diets on Circulating Fatty Acid Composition and Markers of Inflammation

Abstract: Abnormal distribution of plasma fatty acids and increased inflammation are prominent features of metabolic syndrome. We tested whether these components of metabolic syndrome, like dyslipidemia and glycemia, are responsive to carbohydrate restriction. Overweight men and women with atherogenic dyslipidemia consumed ad libitum diets very low in carbohydrate (VLCKD) (1504 kcal:%CHO:fat:protein = 12:59:28) or low in fat (LFD) (1478 kcal:%CHO:fat:protein = 56:24:20) for 12 weeks. In comparison to the LFD, the VLCKD resulted in an increased proportion of serum total n-6 PUFA, mainly attributed to a marked increase in arachidonate (20:4n-6), while its biosynthetic metabolic intermediates were decreased. The n-6/n-3 and arachidonic/eicosapentaenoic acid ratio also increased sharply. Total saturated fatty acids and 16:1n-7 were consistently decreased following the VLCKD. Both diets significantly decreased the concentration of several serum inflammatory markers, but there was an overall greater anti-inflammatory effect associated with the VLCKD, as evidenced by greater decreases in TNF-α, IL-6, IL-8, MCP-1, E-selectin, I-CAM, and PAI-1. Increased 20:4n-6 and the ratios of 20:4n-6/20:5n-3 and n-6/n-3 are commonly viewed as pro-inflammatory, but unexpectedly were consistently inversely associated with responses in inflammatory proteins. In summary, a very low carbohydrate diet resulted in profound alterations in fatty acid composition and reduced inflammation compared to a low fat diet.

SNPs of the FADS gene cluster are associated with polyunsaturated fatty acids in a cohort of patients with cardiovascular disease.

Abstract: Polymorphisms of the human Δ-5 (FADS1) and Δ-6 (FADS2) desaturase genes have been recently described to be associated with the level of several long-chain n-3 and n-6 polyunsaturated fatty acids (PUFAs) in serum phospholipids. We have genotyped 13 single nucleotide polymorphisms (SNPs) located on the FADS1–FADS2–FADS3 gene cluster (chromosome 11q12–13.1) in 658 Italian adults (78% males; mean age 59.7 ± 11.1 years) participating in the Verona Heart Project. Polymorphisms and statistically inferred haplotypes showed a strong association with arachidonic acid (C20:4n-6) levels in serum phospholipids and in erythrocyte cell membranes (rs174545 adjusted P value for multiple tests, P < 0.0001 and P < 0.0001, respectively). Other significant associations were observed for linoleic (C18:2n-6), alpha-linolenic (C18:3n-3) and eicosadienoic (C20:2n-6) acids. Minor allele homozygotes and heterozygotes were associated to higher levels of linoleic, alpha-linolenic, eicosadienoic and lower levels of arachidonic acid. No significant association was observed for stearidonic (C18:4n-3), eicosapentaenoic (C20:5n-3) and docosahexaenoic (C22:6n-3) acids levels. The observed strong association of FADS gene polymorphisms with the levels of arachidonic acid, which is a precursor of molecules involved in inflammation and immunity processes, suggests that SNPs of the FADS1 and FADS2 gene region are worth studying in diseases related to inflammatory conditions or alterations in the concentration of PUFAs.

Role of Fatty Acid Binding Proteins and Long Chain Fatty Acids in Modulating Nuclear Receptors and Gene Transcription

Abstract: Abnormal energy regulation may significantly contribute to the pathogenesis of obesity, diabetes mellitus, cardiovascular disease, and cancer. For rapid control of energy homeostasis, allosteric and posttranslational events activate or alter activity of key metabolic enzymes. For longer impact, transcriptional regulation is more effective, especially in response to nutrients such as long chain fatty acids (LCFA). Recent advances provide insights into how poorly water-soluble lipid nutrients [LCFA; retinoic acid (RA)] and their metabolites (long chain fatty acyl Coenzyme A, LCFA-CoA) reach nuclei, bind their cognate ligand-activated receptors, and regulate transcription for signaling lipid and glucose catabolism or storage: (i) while serum and cytoplasmic LCFA levels are in the 200 μM–mM range, real-time imaging recently revealed that LCFA and LCFA-CoA are also located within nuclei (nM range); (ii) sensitive fluorescence binding assays show that LCFA-activated nuclear receptors [peroxisome proliferator-activated receptor-α (PPARα) and hepatocyte nuclear factor 4α (HNF4α)] exhibit high affinity (low nM Kds) for LCFA (PPARα) and/or LCFA-CoA (PPARα, HNF4α)—in the same range as nuclear levels of these ligands; (iii) live and fixed cell immunolabeling and imaging revealed that some cytoplasmic lipid binding proteins [liver fatty acid binding protein (L-FABP), acyl CoA binding protein (ACBP), cellular retinoic acid binding protein-2 (CRABP-2)] enter nuclei, bind nuclear receptors (PPARα, HNF4α, CRABP-2), and activate transcription of genes in fatty acid and glucose metabolism; and (iv) studies with gene ablated mice provided physiological relevance of LCFA and LCFA-CoA binding proteins in nuclear signaling. This led to the hypothesis that cytoplasmic lipid binding proteins transfer and channel lipidic ligands into nuclei for initiating nuclear receptor transcriptional activity to provide new lipid nutrient signaling pathways that affect lipid and glucose catabolism and storage.

Stearidonic Acid-Enriched Soybean Oil Increased the Omega-3 Index, an Emerging Cardiovascular Risk Marker

Abstract: A plant source of omega-3 fatty acid (FA) that can raise tissue eicosapentaenoic acid (EPA) and/or docosahexaenoic acid (DHA) is needed. A soybean oil (SBO) containing approximately 20% stearidonic acid [SDA; the delta-6 desaturase product of alpha-linolenic acid (ALA)] derived from genetically modified soybeans is under development. This study compared the effects of EPA to SDA-SBO on erythrocyte EPA + DHA levels (the omega-3 index). Overweight healthy volunteers (n = 45) were randomized to SDA-SBO (24 ml/day providing ~3.7 g SDA) or to regular SBO (control group) without or with EPA ethyl esters (~1 g/day) for 16 weeks. Serum lipids, blood pressure, heart rate, platelet function and safety laboratory tests were measured along with the omega-3 index. A per-protocol analysis was conducted on 33 subjects (11 per group). Compared to baseline, average omega-3 index levels increased 19.5% in the SDA group and 25.4% in the EPA group (p < 0.05 for both, vs. control). DHA did not change in any group. Relative to EPA, SDA increased RBC EPA with about 17% efficiency. No other clinical endpoints were affected by SDA or EPA treatment (vs. control). In conclusion, SDA-enriched SBO significantly raised the omega-3 index. Since EPA supplementation has been shown to raise the omega-3 index and to lower risk for cardiac events, SDA-SBO may be a viable plant-based alternative for providing meaningful intakes of cardioprotective omega-3 FAs.

The percentage of n-3 highly unsaturated fatty acids in total HUFA as a biomarker for omega-3 fatty acid status in tissues.

Abstract: A blood biomarker of omega-3 fatty acid intake and tissue status could serve as a modifiable risk factor for cardiovascular disease. The percentage of omega-3 highly unsaturated fatty acid (HUFA ≥ 20 carbons and ≥3 double bonds) in the total HUFA pool (the n-3 HUFA score) was examined as a potential blood biomarker of omega-3 fatty acids in tissues. The fatty acid composition of total lipid extracts (TLE) and phospholipid (PL) fractions were determined for plasma and erythrocytes samples of human subjects (n = 20) and the n-3 HUFA score and the sum of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were compared. Omega-3 fatty acids in blood and tissues of rats (n = 31) and pigs (n = 48) were also determined and the associations were compared. The n-3 HUFA score is more consistent across plasma and erythrocytes, with strong correlations between TLE and PL in plasma (r = 0.93) and erythrocytes (r = 0.94). The n-3 HUFA score was less variable and blood levels correlated strongly with various animal tissues. The n-3 HUFA score is a useful blood biomarker that does not require the isolation of the PL class thereby supporting high throughput analyses. The strength of association between the n-3 HUFA score and disease risk needs to be examined.

Biological functions and metabolism of oleoylethanolamide.

Abstract: The present review is focused on the metabolism and the emerging roles of oleoylethanolamide (OEA) with emphasis on its effects on food intake control and lipid metabolism. The biological mechanism of action, including a non-genomic effect mediated through peroxisome proliferator-activated receptor alpha (PPAR-α) and transient receptor potential vanilloid type 1 (TRPV1) receptor, is discussed. The research related to fatty acid ethanolamides has been focused until recently on anandamide and its interaction with cannabinoid receptor subtype 1. The roles of other N-acyl ethanolamine fatty acid derivatives have been neglected until it was demonstrated that OEA can modulate food intake control through interaction with PPAR-α. Further investigations demonstrated that OEA modulates lipid and glucose metabolism, and recent study confirmed that OEA is an antagonist of TRVP1. It has been demonstrated that OEA has beneficial effects on health by inducing food intake control, lipid β-oxidation, body weight loss and analgesic effects. The investigation of the mechanism of action revealed that OEA activates PPAR-α and stimulates the vagal nerve through the capsaicin receptor TRPV1. Pre-clinical studies showed that OEA remains active when administered orally.

Dietary n-3 HUFA Affects Mitochondrial Fatty Acid β-Oxidation Capacity and Susceptibility to Oxidative Stress in Atlantic Salmon

Abstract: Atlantic salmon (Salmo salar) (90 g) were fed four different diets for 21 weeks (final weight 344 g). The levels of n-3 highly unsaturated fatty acids (HUFA) ranged from 11% of the total fatty acids (FA) in the low n-3 diet to 21% in the intermediate n-3 diet, to 55 and 58% in the high n-3 diets. The high n-3 diets were enriched with either docosahexaenoic acid (DHA) or eicosapentaenoic acid (EPA). Increasing dietary levels of n-3 HUFA led to increasing percentages (from 31 to 52%) of these FA in liver lipids. The group fed the highest level of DHA had higher expressions of peroxisome proliferator-activated receptor (PPAR) β and the FA β-oxidation genes acyl-CoA oxidase (ACO) and carnitine palmitoyltransferase (CPT)-II, compared to the low n-3 groups. The high n-3 groups had reduced activity of mitochondrial cytochrome c oxidase and β-oxidation capacity, together with increased activities of superoxide dismutase (SOD) and caspase-3 activities. In the group fed the highest level of n-3 HUFA, decreased percentages of major phospholipids (PL) in the mitochondrial and microsomal membranes of the liver were also apparent. The percentage of mitochondrial cardiolipin (Ptd2Gro) was 3.1 in the highest n-3 group compared to 6.6 in the intermediate group. These data clearly show an increased incidence of oxidative stress in the liver of fish fed the high n-3 diets.

CYP51 : A Major Drug Target in the Cytochrome P450 Superfamily

Abstract: The cytochrome P540 (CYP) superfamily currently includes about 9,000 proteins forming more than 800 families. The enzymes catalyze monooxygenation of a vast array of compounds and play essentially two roles. They provide biodefense (detoxification of xenobiotics, antibiotic production) and participate in biosynthesis of important endogenous molecules, particularly steroids. Based on these two roles, sterol 14|*alpha*|-demethylases (CYP51) belong to the second group of P450s. The CYP51 family, however, is very special as its members preserve strict functional conservation in enzyme activity in all biological kingdoms. At amino acid identity across the kingdoms as low as 25–30%, they all catalyze essentially the same three-step reaction of oxidative removal of the 14|*alpha*|-methyl group from the lanostane frame. This reaction is the required step in sterol biosynthesis of pathogenic microbes. We have shown that specific inhibition of protozoan CYP51 can potentially provide treatment for human trypanosomiases. Three sets of CYP51 inhibitors tested in vitro and in trypanosomal cells in this study include azoles [best results being 50% cell growth inhibition at <1 and at 1.3 μM for Trypanosoma cruzi (TC) and Trypanosoma brucei (TB), respectively], non-azole compounds (50% TC cell growth inhibition at 5 μM) and substrate analogs of the 14|*alpha*|-demethylase reaction. 32-Methylene cyclopropyl lanost-7-enol exhibited selectivity toward TC with 50% cell growth inhibition at 3 μM.

From molecular fossils of bacterial hopanoids to the formation of isoprene units: discovery and elucidation of the methylerythritol phosphate pathway.

Abstract: Investigations on the biosynthesis of bacterial triterpenoids of the hopane series led to the unexpected discovery of an alternative mevalonate independent pathway for the formation of isoprene units. Methylerythritol phosphate, already presenting the C5 branched isoprene skeleton, is the key intermediate. This pathway was independently characterized in ginkgo embryos for the formation of diterpenoids. It is present in most bacteria and in the plastids of all organisms belonging to phototrophic phyla. The key steps of the discovery and elucidation of this metabolic route are presented in this review.

Direct Microwave Transesterification of Fingertip Prick Blood Samples for Fatty Acid Determinations

Abstract: Omega-3 polyunsaturated fatty acid (PUFA) dietary intakes and tissue levels are positively associated with various health benefits. The development of cost efficient, high throughput methodologies would enable research in large clinical and population studies, and clinical fatty acid profiling. Microwave heating for the transesterification of blood fatty acids was examined. Samples were collected by venous puncture and fingertip prick onto chromatography paper. Aliquots of serum, plasma, erythrocytes and whole blood were prepared from venous blood. Boron trifluoride in methanol was used for transesterification but sample preparation and heating varied. Fatty acid determinations and markers of omega-3 fatty acid status including the sum of eicosapentaenoic acid and docosahexaenoic acid, the ratio of total n-3 PUFA to n-6 PUFA, and the percentage of n-3 highly unsaturated fatty acids (HUFA, ≥20 carbons and ≥3 carbon–carbon double bonds) in total HUFA were compared. Quantitative determinations indicate that microwave transesterification results in significantly lower estimates of monounsaturates and polyunsaturates, possibly through incomplete transesterification of triacylglycerols. However, qualitative estimates of omega-3 fatty acid status were relatively similar. Fingertip prick blood collection combined with direct transesterification by microwave may be a very rapid method to estimate omega-3 fatty acid status for selected applications.

The Identification of Mono-, Di-, Tri-, and Tetragalactosyl-diacylglycerols and their Natural Estolides in Oat Kernels

Abstract: Oat kernels were extracted with methanol, and glycolipid-enriched fractions were prepared using silica solid phase extraction. Using direct infusion electrospray ionization (ESI) tandem mass spectrometry (MS), high performance liquid chromatography (HPLC)-ESI-MS, and HPLC-atmospheric pressure chemical ionization (APCI)-MS, we confirmed previous reports that digalactosyldiacylglycerol (DGDG) was the most abundant glycolipid in oat kernels and confirmed a previous report of the presence of a DGDG mono-estolide in oat kernels. In the current study we also identified several additional natural galactolipid estolides: two new DGDG estolides (di- and tri-estolides), two trigalactosyldiacylglycerol (TriGDG) estolides (mono- and di-estolides), and one tetragalactosyldiacylglycerol (TetraGDG) estolide (mono-estolide). The levels of total galactolipid estolides in oat kernels were estimated to be about 29% of the total glycolipid fraction. To our knowledge, this report is the first evidence of natural di- and tri-estolides of polar lipids.

Fluorescence Techniques Using Dehydroergosterol to Study Cholesterol Trafficking

Abstract: Cholesterol itself has very few structural/chemical features suitable for real-time imaging in living cells. Thus, the advent of dehydroergosterol [ergosta-5,7,9(11),22-tetraen-3β-ol, DHE] the fluorescent sterol most structurally and functionally similar to cholesterol to date, has proven to be a major asset for real-time probing/elucidating the sterol environment and intracellular sterol trafficking in living organisms. DHE is a naturally occurring, fluorescent sterol analog that faithfully mimics many of the properties of cholesterol. Because these properties are very sensitive to sterol structure and degradation, such studies require the use of extremely pure (>98%) quantities of fluorescent sterol. DHE is readily bound by cholesterol-binding proteins, is incorporated into lipoproteins (from the diet of animals or by exchange in vitro), and for real-time imaging studies is easily incorporated into cultured cells where it co-distributes with endogenous sterol. Incorporation from an ethanolic stock solution to cell culture media is effective, but this process forms an aqueous dispersion of DHE crystals which can result in endocytic cellular uptake and distribution into lysosomes which is problematic in imaging DHE at the plasma membrane of living cells. In contrast, monomeric DHE can be incorporated from unilamellar vesicles by exchange/fusion with the plasma membrane or from DHE-methyl-β-cyclodextrin (DHE-MβCD) complexes by exchange with the plasma membrane. Both of the latter techniques can deliver large quantities of monomeric DHE with significant distribution into the plasma membrane. The properties and behavior of DHE in protein-binding, lipoproteins, model membranes, biological membranes, lipid rafts/caveolae, and real-time imaging in living cells indicate that this naturally occurring fluorescent sterol is a useful mimic for probing the properties of cholesterol in these systems.

Women and Smokers Have Elevated Urinary F2-Isoprostane Metabolites: A Novel Extraction and LC–MS Methodology

Abstract: F2-Isoprostanes (F2-IsoPs), regio- and stereoisomers of prostaglandin F2α (PGF2α), and urinary F2-IsoP metabolites including 2,3-dinor-5,6-dihydro-8-iso-PGF2α [2,3-dinor-8-iso-PGF1α (2,3-dinor-F1)] and 2,3 dinor-8-iso-PGF2α (2,3-dinor-F2), have all been used as biomarkers of oxidative stress. A novel method was developed to measure these biomarkers using a single solid phase extraction (SPE) cartridge, separation by HPLC, and detection by negative mode selected reaction monitoring (SRM) mass spectrometry (MS), using authentic standards of PGF2α; 8-iso-PGF2α; 2,3-dinor-F1 and 2,3-dinor-F2 to identify specific chromatographic peaks. The method was validated in a population of healthy, college-aged nonsmokers (n = 6 M/8F) and smokers (n = 6 M/5F). Urinary F2-IsoP concentrations were ~0.2–1.5 μg/g creatinine, 2,3-dinor-F1 was ~1–3 μg/g and 2,3-dinor-F2 was ~3–5 μg/g. Additional F2-IsoPs metabolites were identified using SRM. The sum of all urinary F2-IsoP metabolites was 50–100 μg/g creatinine indicating their greater abundance than F2-IsoPs. Women had higher F2-IsoP metabolite concentrations than did men (MANOVA, main effect P = 0.003); cigarette smokers had higher concentrations than did nonsmokers (main effect P = 0.036). For men or women, respectively, smokers had higher metabolite concentrations than did nonsmokers (P < 0.05). Thus, our method simultaneously allows measurement of urinary F2-IsoPs and their metabolites for the determination of oxidative stress.

Developmental, hormonal, and nutritional regulation of porcine adipose triglyceride lipase (ATGL).

Abstract: Adipose triglyceride lipase (ATGL) is a newly identified lipase. We report for the first time the porcine ATGL sequence and characterize ATGL gene and protein expression in vitro and in vivo. Adult pig tissue expresses ATGL at high levels in the white adipose and muscle tissue relative to other tested tissues. We show that within the white adipose tissue ATGL is expressed at higher levels in the adipocyte than in the stromal-vascular fraction. Additionally, ATGL expression increases dramatically in the subcutaneous adipose during adipose development and maturation, as well as during in vitro adipogenesis. Peroxisome proliferator-activated receptor gamma transcript levels increased concomitant with ATGL gene expression, suggesting a possible role in the regulation of ATGL by adipogenic regulators. In vitro treatment of differentiated primary pig preadipocytes with insulin and forskolin decreased ATGL gene expression in a dose-dependent manner, suggesting ATGL transcript levels are hormone sensitive. In vivo experimentation showed that calorie-restriction in gilts resulted in increased ATGL mRNA and protein levels in subcutaneous and peri-renal fat tissues. Our data demonstrate that ATGL expression reacts to hormonal stimuli and plays a role in catecholamine-induced lipolysis in porcine adipose tissue.

Fatty Acid Profile of Sunshine Bass: I. Profile Change is Affected by Initial Composition and Differs Among Tissues

Abstract: Fatty acid (FA) composition of fillet tissue can be tailored by transitioning fish from alternative lipid-based, low long-chain polyunsaturated fatty acid (LC-PUFA) grow-out feeds to high LC-PUFA "finishing" feeds. To address whether grow-out feed composition influences the responsiveness of fillet tissue to finishing, sunshine bass (SB, Morone chrysops x M. saxatilis) were reared to a submarketable size on grow-out feeds containing fish oil (FO) or a 50:50 blend of FO and coconut (CO), grapeseed (GO), linseed (LO), or poultry (PO) oil. For the final 8 weeks of the trial, fish were either maintained on assigned grow-out feeds or finished with the 100% FO feed. Production performance was unaffected by dietary lipid source, but fillet FA profile generally conformed to nutritional history. Regardless of grow-out regimen, finishing had a significant restorative effect on fillet FA composition; however, complete restoration of control levels of 20:5n-3, 22:6n-3, total LC-PUFA and n-3:n-6 FA ratio was achieved only among fish fed the CO-based grow-out feed. Saturated fatty acids (SFA) appear to be preferential catabolic substrates, whereas medium-chain and long-chain PUFA are selectively deposited in tissues. Provision of SFA in grow-out feeds appears to optimize selective FA metabolism and restoration of beneficial fillet FA profile during finishing.

Validation of a rapid measure of blood PUFA levels in humans.

Abstract: An assay involving a finger stick and filter paper blood spotting was developed to determine polyunsaturated fatty acid (PUFA) levels in blood. Capillary whole blood from a finger stick was blotted on antioxidant impregnated filter paper, air dried, saponified and methylated using sodium hydroxide and boron trifluoride in methanol. The method differed from those described previously because separation of plasma and red blood cells (RBCs) was not needed, thin-layer chromatography (TLC) was not required to separate phospholipids, initial extraction of lipids before transesterification was not necessary, and the fatty acid methyl ester (FAME) method was able to methylate steryl esters, free fatty acids, and sphingomyelins. Twenty-six subjects provided blood samples by finger stick and venipuncture. Levels of long-chain polyunsaturated fatty acids (LC-PUFA) from capillary whole blood were correlated with those from RBCs and PLs in venous blood (P < 0.001, R(2) ranged from 0.64 to 0.86). Although highly significant (P < 0.002), the R(2) values for the correlation between arachidonic acid (ARA) levels in capillary whole blood with ARA levels in RBCs and plasma phospholipids (PLs) were relatively lower (R(2) = 0.31-0.41, respectively). Results indicate that the described finger stick assay represents a fast, reliable method to measure specific LC-PUFA levels.

Regulation of Steroid Hormone Biosynthesis by the Cytoskeleton

Abstract: Steroid hormones are synthesized in response to signaling cascades initiated by the trophic peptide hormones derived from the anterior pituitary. The mechanisms by which these peptide hormones regulate steroid hormone production are multifaceted and include controlling the transcription of steroidogenic genes, regulating cholesterol (substrate) uptake and transport, modulating steroidogenic enzyme activity, and controlling electron availability. Cytoskeletal polymers such as microfilaments and microtubules have also been implicated in regulating steroidogenesis. Of note, steroidogenesis is a multi-step process that occurs in two organelles, the endoplasmic reticulum (ER) and the mitochondrion. However, the precise mechanism by which substrates are delivered back and forth between these two organelles is unknown. In this review we will discuss the role of components of the cytoskeleton in conferring optimal steroidogenic potential. Finally, we present data that identifying a novel mechanism by which sphingosine-1-phosphate induces mitochondrial trafficking to promote steroidogenesis.

Oral Administration of Dihomo-γ-Linolenic Acid Prevents Development of Atopic Dermatitis in NC/Nga Mice

Abstract: Disorders of the metabolism of essential fatty acids (EFAs) are related to atopic dermatitis (AD). Concentrations of dihomo-γ-linolenic acid (DGLA), an EFA, in the serum of AD patients are lower than those in healthy volunteers. Recently we developed a fermented DGLA oil, and examined whether oral administration of DGLA prevents development of dermatitis in NC/Nga mice, which spontaneously develop human AD-like skin lesions. NC/Nga mice were fed a diet either containing or not containing DGLA for 8 weeks under in air-uncontrolled conventional circumstances. Clinical skin severity scores were significantly lower in mice fed DGLA than in mice not fed it. Scratching behavior and plasma total IgE levels were also reduced in the DGLA group, in association with histological improvement. DGLA suppressed clinical severity of skin lesions dose-dependently, with an increase in DGLA contents in phospholipids of skin, spleen, and plasma. Discontinuation of DGLA administration resulted in the onset of dermatitis and a decrease in DGLA contents in skin, spleen, and plasma. These findings indicate that oral administration of DGLA effectively prevents the development of AD in NC/Nga mice, and that DGLA in phospholipids is a compound of key importance in the development and prevention of dermatitis.

Substrate Preferences of a Lysophosphatidylcholine Acyltransferase Highlight Its Role in Phospholipid Remodeling

Abstract: An important enzyme involved in phospholipid turnover is the acyl-CoA: lysophosphatidylcholine acyltransferase (LPCAT). Here, we report characterization of a newly discovered human LPCAT (LPCAT3), which has distinct substrate preferences strikingly consistent with a role in phosphatidylcholine (PtdCho) remodeling and modulating fatty acid composition of PtdCho. LPCAT3 prefers lysophosphatidylcholine (lysoPtdCho) with saturated fatty acid at the sn-1 position and exhibits acyl donor preference towards linoleoyl-CoA and arachidonoyl-CoA. Furthermore, LPCAT3 is active in mediating 1-O-alkyl-sn-glycero-3-phosphocholine acylation with long chain fatty acyl-CoAs to generate 1-O-alkyl-phosphatidylcholine, another very important constitute of mammalian membrane systems. These properties are precisely the known attributes of LPCAT previously ascribed to the isoform involved in Lands' cycle, and thus strongly suggest that LPCAT3 is involved in phospholipids remodeling to achieve appropriate membrane lipid fatty acid composition.

Estradiol Favors the Formation of Eicosapentaenoic Acid (20:5n-3) and n-3 Docosapentaenoic Acid (22:5n-3) from Alpha-Linolenic Acid (18:3n-3) in SH-SY5Y Neuroblastoma Cells

Abstract: Whether neurosteroids regulate the synthesis of long chain polyunsaturated fatty acids in brain cells is unknown. We examined the influence of 17-beta-estradiol (E2) on the capacity of SH-SY5Y cells supplemented with alpha-linolenic acid (ALA), to produce eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA). Cells were incubated for 24 or 72 h with ALA added alone or in combination with E2 (ALA + E2). Fatty acids were analyzed by gas chromatography of ethanolamine glycerophospholipids (EtnGpl) and phosphatidylcholine (PtdCho). Incubation for 24 h with ALA alone increased EPA and DPA in EtnGpl, by 330 and 430% compared to controls (P < 0.001) and DHA by only 10% (P < 0.05). Although DHA increased by 30% (P < 0.001) in ALA + E2-treated cells, the difference between the ALA and ALA + E2 treatments were not significant after 24 h (Anova-1, Fisher's test). After 72 h, EPA, DPA and DHA further increased in EtnGpl and PtdCho of cells supplemented with ALA or ALA + E2. Incubation for 72 h with ALA + E2 specifically increased EPA (+34% in EtnGpl, P < 0.001) and DPA (+15%, P < 0.001) compared to ALA alone. Thus, SH-SY5Y cells produced membrane EPA, DPA and DHA from supplemental ALA. The formation of DHA was limited, even in the presence of E2. E2 significantly favored EPA and DPA production in cells grown for 72 h. Enhanced synthesis of ALA-elongation products in neuroblastoma cells treated with E2 supports the hypothesis that neurosteroids could modulate the metabolism of PUFA.

Long-Chain Omega-3 Polyunsaturated Fatty Acids in the Blood of Children and Adolescents with Juvenile Bipolar Disorder

Abstract: Reduced long-chain omega-3 polyunsaturated fatty acids (LCn-3PUFA), including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), have been reported in adult patients suffering from depression and bipolar disorder (BD). LCn-3PUFA status has not previously been examined in children and adolescents with BD compared with healthy controls. Fifteen children and adolescents (9–18 years, M ± SD = 14.4 ± 3.48) diagnosed with juvenile bipolar disorder (JBD) and fifteen healthy age and sex-matched controls were assessed for dietary intake and fasting red blood cell (RBC) membrane concentrations of LCn-3PUFA. Fatty acid concentrations were compared between participants diagnosed with JBD and controls after controlling for dietary intake. RBC membrane concentrations of EPA and DHA were not significantly lower in participants diagnosed with JBD compared with healthy controls (M ± sem EPA = 3.37 ± 0.26 vs. 3.69 ± 0.27 µg/mL, P = 0.458; M ± sem DHA = 22.08 ± 2.23 vs. 24.61 ± 2.38 µg/mL, P = 0.528) after controlling for intake. Red blood cell DHA was negatively (r = −0.55; P = 0.044) related to clinician ratings of depression. Although lower RBC concentrations of LCn-3PUFA were explained by lower intakes in the current study, previous evidence has linked reduced LCn-3PUFA to the aetiology of BD. As RBC DHA was also negatively related to symptoms of depression, a randomised placebo-controlled study examining supplementation with LCn-3PUFA as an adjunct to standard pharmacotherapy appears warranted in this patient population.

Sesamin supplementation increases white muscle docosahexaenoic acid (DHA) levels in rainbow trout (Oncorhynchus mykiss) fed high alpha-linolenic acid (ALA) containing vegetable oil: metabolic actions.

Abstract: The effects of including an equi-mixture of sesamin and episesamin in fish diets based on vegetable oils of different fatty acid composition were examined. Sesamin/episesamin (hereafter named sesamin) was included at 0.58 g/100 g diet. The oil used in the feed was either a mixture of linseed and sunflower oils (6:4, by vol) or 100% linseed oil. Addition of sesamin increased the percentages of docosahexaenoic acid (DHA) in white muscle phospholipid and triacylglycerol fraction by up to 37% but the fatty acids in red muscle and liver were not affected. The expression of the peroxisome proliferator-activated receptor PPARα was significantly down regulated in the liver of the fish fed sesamin and mixed oil diet (P < 0.05). Sesamin and episesamin were detected in liver and muscle tissues of the fish that had been fed sesamin. Fish fed sesamin had elevated levels of total cytochrome P450 (CYP) enzymes and EROD activity in the liver, indicating an induction of CYP1A in this tissue. Our conclusion was that supplementation of fish feed with sesamin increased the proportions of DHA in the white muscle.

A whole body, in vivo, fatty acid balance method to quantify PUFA metabolism (desaturation, elongation and beta-oxidation) : erratum to : lipids (2007) 42 :1065-1071

Abstract: Currently there are several contrasting methods utilized for estimating elongation and desaturation of fatty acids and their general metabolism. The majority of these methods involve an ex vivo approach, requiring expensive and sophisticated equipment, likely to result in considerable variation in enzyme activity between and within species. In the present paper we introduce a further development of the whole-body fatty acid balance method for the estimation of the elongation and desaturation of fatty acids. This method though receiving considerable attention because of its simplicity and reliability has yet to be presented in detail. Theoretically, the whole-body fatty acid balance method can potentially be applied to any organism and requires little more than a gas chromatography unit for fatty acid analysis and elementary calculations. As such in this paper we attempt to spell out in detail the theoretical basis and the methods of application drawing specific examples. Using the present method it is possible to measure the fate of individual fatty acids towards desaturation, elongation and oxidation and calculate the elongase, Delta-6 desaturase and Delta-5 desaturase activities.

Genetic variation in ABC G5/G8 and NPC1L1 impact cholesterol response to plant sterols in hypercholesterolemic men.

Abstract: ATP-binding cassette hetero-dimeric transporters G5 and G8 (ABCG5/G8) have been postulated to mediate intestinal cholesterol efflux, whereas Niemann-Pick C1 Like 1 (NPC1L1) protein is believed to be essential for intestinal cholesterol influx The individual or combined genetic markers, such as single nuclear polymorphisms (SNPs), of these two transporter genes may explain inter-individual variations in plasma cholesterol response following plant sterol (PS) intervention The present study was aimed at investigating the association between ABCG5/G8 and NPC1L1 genotype SNPs with sterol absorption and corresponding plasma concentrations The study used a 4-week crossover design with 82 hypercholesterolemic men characterized by high vs low basal plasma PS concentrations consuming spreads with or without 2 g/day of PS For the ABCG8 1289 C > A (T400 K) polymorphism, the A allele carriers with high basal plasma PS concentrations demonstrated a 39-fold greater reduction (p G (L272L) and 3929 G > A (Y1291Y), individuals carrying mutant alleles showed a 24-fold greater (p < 005) reduction in LDL-C levels, compared to wild type counterparts Results suggest that genetic and metabolic biomarkers together may predict inter-individual lipid level responsiveness to PS-intervention, and thus could be useful in devising individualized cholesterol lowering strategies

Plasma Omega-3 Fatty Acid Response to a Fish Oil Supplement in the Healthy Elderly

Abstract: Little information is available concerning whether incorporation of dietary omega-3 fatty acids into plasma lipids changes during healthy aging. Elderly (74 +/- 4 years old) and young (24 +/- 2 years old) adults were given a fish oil supplement for 3 weeks that provided 680 mg/day of docosahexaenoic acid and 320 mg/day of eicosapentaenoic acid, followed by a 2 week wash-out period. Compliance was monitored by spiking the capsules with carbon-13 glucose, the excretion of which was measured in breath CO2. In response to the supplement, plasma docosahexaenoic acid rose 42% more in the elderly but eicosapentaenoic responded similarly in both groups. Despite raising docosahexaenoic acid intake by five to tenfold, the supplement did not raise plasma free docosahexaenoic acid (% or mg/dL) in either group. We conclude that healthy aging is accompanied by subtle but significant changes in DHA incorporation into plasma lipids.

Sesamin increases alpha-linolenic acid conversion to docosahexaenoic acid in atlantic salmon (Salmo salar L.) hepatocytes: role of altered gene expression.

Abstract: In vitro cultivated Atlantic salmon (Salmo salar L.), hepatocytes were incubated without or with a mixture of sesamin and episesamin in order to test for possible effects on lipid metabolism. Sesamin/episesamin exposure (0.05 mM, final concentration) led to increased elongation and desaturation of 14C 18:3n-3 to docosahexaenoic acid (14C 22:6n-3, DHA, P < 0.01) and down regulated gene expression of Δ6 and Δ5 desaturases compared to control treatment. Sesamin/episesamin further increased the hepatocytes capacity for fatty acid β-oxidation of 14C 18:3n-3 (P < 0.01) to the 14C acid soluble products, acetate, malate and oxaloacetate, in agreement with an increased gene expression of carnitine palmitoyltransferase I. Also the gene expression of cluster of differentiation 36 was upregulated and the expression of scavenger receptor type B, peroxisome proliferator-activated receptors α and γ were downregulated. The amount of triacylglycerols secreted by the cells tended to be lower in the sesamin/episesamin incubated hepatocytes than the control cells. This study shows that sesamin has favourable effects on lipid metabolism leading to increased level of DHA, which may be of interest for aquaculture use.

Fatty Acid Profile of Sunshine Bass: II. Profile Change Differs Among Fillet Lipid Classes

Abstract: Fatty acid (FA) profile of fish tissue mirrors dietary FA profile and changes in a time-dependent manner following a change in dietary FA composition. To determine whether FA profile change varies among lipid classes, we evaluated the FA composition of fillet cholesteryl esters (CE), phospholipids (PL), and triacylglycerols (TAG) of sunshine bass (SB, Morone chrysops x M. saxatilis) raised on feeds containing fish oil or 50:50 blend of fish oil and coconut, grapeseed, linseed, or poultry oil, with or without implementation of a finishing period (100% FO feed) prior to harvest. Each lipid class was associated with a generalized FA signature, irrespective of nutritional history: fillet PL was comprised largely of saturated FA (SFA), long-chain polyunsaturated FA (LC-PUFA), and total n-3 FA; fillet TAG was higher in MC-PUFA and total n-6 FA; and fillet CE was highest in monounsaturated FA (MUFA). Neutral lipids reflected dietary composition in a near-direct fashion; conversely, PL showed evidence of selectivity for MC- and LC-PUFA. Shorter-chain SFA were not strongly reflected within any lipid fraction, even when dietary availability was high, suggesting catabolism of these FA. FA metabolism in SB is apparently characterized by a division between saturated and unsaturated FA, whereby LC-PUFA are preferentially incorporated into tissues and SFA are preferentially oxidized for energy production. We demonstrated provision of SFA in grow-out feeds for SB, instead MC-PUFA which compete for tissue deposition, meets energy demands and allows for maximum inclusion of LC-PUFA within fillet lipids.

GCG-rich tea catechins are effective in lowering cholesterol and triglyceride concentrations in hyperlipidemic rats.

Abstract: The (-)-gallocatechin gallate (GCG) concentration in some tea beverages can account for as much as 50% of the total catechins, as a result of sterilization. The present study aims to examine the effects of GCG-rich tea catechins on hyperlipidemic rats and the mechanisms associated with regulating cholesterol metabolism in the liver. By performing heat epimerization of (-)-epigallocatechin gallate (EGCG), we manufactured a mixture of catechins that had a GCG content of approximately 50% (w/w). In sucrose-rich diet-induced hyperlipidemic rats, the GCG-rich tea catechins exhibited strong activity in reducing plasma cholesterol and triglyceride concentrations. Furthermore, the hepatic cholesterol and triglyceride concentrations that had increased as a result of the sucrose-rich diet were reduced due to GCG-rich tea catechins consumption. In order to investigate the hyperlipidemic mechanism of GCG-rich tea catechins, we examined the hepatic expressions of LDL receptor and HMG-CoA reductase in hyperlipidemic rats. We further evaluated the action of purified GCG on LDL receptor activity, which is a key contributor to the regulation of cholesterol concentrations. We found that purified GCG increased LDL receptor protein level and activity to a greater extent than EGCG. In conclusion, our study indicates that GCG-rich tea catechins in tea beverages may be effective in preventing hyperlipidemia by lowering plasma and hepatic cholesterol concentrations.

Significance of coprophagy for the fatty acid profile in body tissues of rabbits fed different diets.

Abstract: Four groups of eight New Zealand hybrid rabbits were fattened with ad libitum access to the following pelleted experimental diets: ryegrass meal or alfalfa meal fed either alone or with oats meal in a ratio of 1:1. After 25 weeks they were slaughtered and dissected. Fatty acid (FA) profiles of caecotrophs (re-ingested fermentation products of the caecum), perirenal adipose tissue and intramuscular fat in the Musculus quadriceps were determined. With high proportions of branched-chain FA (BFA) and trans FA, and increased proportions of saturated FA relative to the diets, the caecotroph FA profile showed a clear fingerprint of anaerobe microbial lipid metabolism including biohydrogenation. By contrast, the FA profiles of adipose and lean tissue comprised high proportions of polyunsaturated FA (PUFA), whilst BFA and trans FA occurred in much lower proportions compared to the caecotrophs. Thus, coprophagy did not substantially modify the FA composition of the tissues investigated. Use of forage-only diets, compared to the oats supplemented diets, led to extraordinary high proportions of n-3 PUFA (including 18:3 and long-chain n-3) in the fat of adipose (21.3 vs. 6.7%) and lean tissue (15.4 vs. 5.7%). The forage type diet (grass vs. alfalfa) had smaller effects on the FA profiles. Indications of diet effects on endogenous desaturation, chain elongation and differential distribution of functional FA between the two tissues investigated were found.