Incorporation Of Linoleic Acid Research Articles

Programming lipopeptide nanotherapeutics for tandem treatment of postsurgical infection and melanoma recurrence

Tumor recurrence and chronic bacterial infection constitute two major criteria in postsurgical intervention for malignant melanoma. One plausible strategy is the equipment of consolidation therapy after surgery, which relies on adjuvants to eliminate the residual tumor cells and inhibit bacterial growth. Until now, a number of proof-of-concept hybrid nanoadjuvants have been proposed to combat tumor recurrence and postsurgical bacterial infection, which may suffer from the potential bio-unsafety or involve complex design and synthesis. The batch-to-batch inconsistencies in drug composition further delay the clinical trials. To circumvent these issues, herein we develop a programmable strategy to generate lipopeptide nanotherapeutics with identical constitution for tandem intervention of postsurgical bacterial infection and cancer recurrence of melanoma. Increasing the number of hydrophobic linoleic acid within lipopeptides has been found to be a simple and practical strategy to improve the therapeutic outcomes for both tumor cells and bacteria. Self-assembled lipopeptide nanotherapeutics with two linoleic acid molecules possesses excellent antitumor activity and antimicrobial function toward both susceptible strains and drug-resistant bacteria. Arising from the incorporation of unsaturated linoleic acid, the unavoidable hemolysis of cationic peptide drugs was effectively alleviated. In vivo therapeutic abilities of postsurgical infection and tumor recurrence were investigated in BALB/c nude mice bearing a B16-F10 tumor model, with an incomplete surgical resection and in situ infection by methicillin-resistant Staphylococcus aureus (MRSA). Self-assembled lipopeptide nanotherapeutics could effectively inhibit cancer cell growth and bacterial infection, as well as promote wound healing. The easily scalable large-scale production, broad-spectrum antitumor and antibacterial bioactivities as well as fixed component endows lipopeptide nanotherapeutics as promising adjuvants for clinically postsurgical therapy of melanoma.

Recombinant PNPLA1 catalyzes the synthesis of acylceramides and acyl acids with selective incorporation of linoleic acid

Loss-of-function mutations in patatin-like phospholipase domain-containing protein 1 (PNPLA1) cause autosomal recessive congenital ichthyosis, and altered PNPLA1 activity is implicated in the pathogenesis of atopic dermatitis and other common skin diseases. To examine the hypothesis that PNPLA1 catalyzes the synthesis of acylceramides and acyl acids, we expressed and partially purified a soluble, truncated form of PNPLA1 in Escherichia coli, (PNPLA1trun) along with the related protein PNPLA2 (ATGL, adipose triglyceride lipase) and coactivator CGI-58. Liposomal substrates were incubated with recombinant enzymes for 0.5–24 h and products analyzed by HPLC-UV and LC-MS. Using trilinolein or dilinolein substrates, PNPLA1trun, like ATGLtrun, catalyzed lipolysis and acyltransferase reactions with 2–30% conversion into linoleic acid, monolinolein, and trilinolein. CGI-58 enhanced ATGL-catalyzed lipolysis as previously reported, but transacylase activity was not enhanced with ATGL or PNPLA1. In matching the proposed activity in vivo, PNPLA1 catalyzed acyl transfer from trilinolein and dilinolein donors to omega-hydroxy ceramide, omega-hydroxy glucosylceramide, and omega-hydroxy acid acceptors to form acylceramide, glucosyl-acylceramide, and acyl acid, respectively, albeit with only ∼0.05% conversion of the substrates. Notably, in experiments comparing dilinolein vs. diolein acyl donors, PNPLA1 transferred linoleate with 3:1 selectivity over oleate into acylceramide. These results support the role for PNPLA1 in the synthesis of acylceramides and acyl acids in epidermis and suggest that the enrichment of these lipids with linoleic acid could result from the substrate selectivity of PNPLA1.

Dietary Phospholipid-Bound Conjugated Linoleic Acid and Docosahexaenoic Acid Incorporation Into Fetal Liver and Brain Modulates Fatty Acid and N-Acylethanolamine Profiles.

We evaluated whether maternal intake of conjugated linoleic acid (CLA) and docosahexaenoic acid (DHA) in the phospholipid (PL) form (CLA-DHA PL) affects maternal and fetal brain and liver fatty acids (FAs) profile and the biosynthesis of FA-derived bioactive lipid mediators N-acylethanolamines (NAEs) involved in several neurophysiological functions. We fed rat dams during the first 2/3 of their pregnancy a CLA-DHA PL diet containing PL-bound 0.5% CLA and 0.2% DHA. FA and NAE profiles were analyzed in maternal and fetal liver and brain by Liquid Chromatography diode array detector (LC-DAD) and MS/MS in line. We found that CLA and DHA crossed the placenta and were readily incorporated into the fetal liver and brain. CLA metabolites were also found abundantly in fetal tissues. Changes in the FA profile induced by the CLA-DHA PL diet influenced the biosynthesis of NAE derived from arachidonic acid (ARA; N-arachidonoylethanolamine, AEA) and from DHA (N-docosahexaenoylethanolamine, DHEA). The latter has been previously shown to promote synaptogenesis and neuritogenesis. The reduced tissue n6/n3 ratio was associated to a significant decrease of AEA levels in the fetal and maternal liver and an increase of DHEA in the fetal and maternal liver and in the fetal brain. Maternal dietary CLA-DHA PL by promptly modifying fetal brain FA metabolism, and thereby, increasing DHEA, might represent an effective nutritional strategy to promote neurite growth and synaptogenesis and protect the offspring from neurological and psychiatric disorders with neuroinflammatory and neurodegenerative basis during the critical prenatal period.

Myeloid-Specific Deficiency of Long-Chain Acyl CoA Synthetase 4 Reduces Inflammation by Remodeling Phospholipids and Reducing Production of Arachidonic Acid-Derived Proinflammatory Lipid Mediators.

In response to infection or tissue damage, resident peritoneal macrophages (rpMACs) produce inflammatory lipid mediators from the polyunsaturated fatty acid (PUFA), arachidonic acid (AA). Long-chain acyl-CoA synthetase 4 (ACSL4) catalyzes the covalent addition of a CoA moiety to fatty acids, with a strong preference for AA and other PUFAs containing three or more double bonds. PUFA-CoA can be incorporated into phospholipids, which is the source of PUFA for lipid mediator synthesis. In this study, we demonstrated that deficiency of Acsl4 in mouse rpMACs resulted in a significant reduction of AA incorporated into all phospholipid classes and a reciprocal increase in incorporation of oleic acid and linoleic acid. After stimulation with opsonized zymosan (opZym), a diverse array of AA-derived lipid mediators, including leukotrienes, PGs, hydroxyeicosatetraenoic acids, and lipoxins, were produced and were significantly reduced in Acsl4-deficient rpMACs. The Acsl4-deficient rpMACs stimulated with opZym also demonstrated an acute reduction in mRNA expression of the inflammatory cytokines, Il6, Ccl2, Nos2, and Ccl5 When Acsl4-deficient rpMACs were incubated in vitro with the TLR4 agonist, LPS, the levels of leukotriene B4 and PGE2 were also significantly decreased. In LPS-induced peritonitis, mice with myeloid-specific Acsl4 deficiency had a significant reduction in leukotriene B4 and PGE2 levels in peritoneal exudates, which was coupled with reduced infiltration of neutrophils in the peritoneal cavity as compared with wild-type mice. Our data demonstrate that chronic deficiency of Acsl4 in rpMACs reduces the incorporation of AA into phospholipids, which reduces lipid mediator synthesis and inflammation.

Abstract 2581: Dietary fat influences pancreatic cancer progression by altering cell membrane lipid content to impact the PI3K/AKT pathway

Abstract Pancreatic cancer (PC) is one of the few cancers for which incidence and mortality continue to rise despite increasing knowledge of its etiology and risk factors. Amongst the latter, dietary patterns are significantly associated with PC risk. Due to its relatively slow progression, preventive strategies represent a simple means to improve outcomes. We are interested in understanding the influence of diet on PC prevention and have studied the effects of polyunsaturated fatty acids (PUFAs) in the progression of the disease both in vitro (via PC cell lines) and in vivo (via EL-KrasG12D and p48Cre/LSL-KrasG12D mouse models that recapitulate IPMN- and PanIN-like lesions, respectively). Our data demonstrate that dietary PUFAs are incorporated into plasma membrane phospholipids (PL) affecting signal transduction, particularly PI3K/AKT signaling, supporting the emergence of membrane-targeted therapies. We evaluated the effects of diets supplemented with omega-3 (ω-3) or omega-6 (ω-6) PUFAs on neoplastic lesion development. This work supports that ω-3 reduces while ω-6 accelerates lesion penetrance, tumor formation and proliferation associated with changes in pAKT. These results were also recapitulated in vitro to confirm that reduced activity of the PI3K/AKT pathway when incubated with Docosahexaenoic Acid (DHA), the primary component of the ω-3 enriched diet. This effect was abrogated by over-activation of the pathway when dosed with the main component of the ω-6 enriched diet, Linoleic Acid (LA). Since PI3K depends on its binding to PIP2 in the membrane for its activity, we next aimed to study if PUFAs were altering PL composition. By using IF and stably-transfected PC cells with a fluorescent translocation biosensor to monitor PIP2 and PIP3 lipids in the membrane, we discovered an effect of exogenous fatty acids expressed as a ratio of PIP2 to PIP3 in the plasma membrane. Enriched areas of membrane GFP staining (or PIP3) were observed in LA-treated Panc-1 cells. DHA treatment prevented PIP3 localization in the membrane of tumor cells, keeping the GFP signal diffuse in the cytoplasm. Using the PIP2 biosensor we sought to study the affinity of PI3K for PIP2 depending on the incorporation of DHA or LA in PIP2. DHA treatment reduced PI3K interaction (not total PI3K) with PIP2 resulting in lower levels of PIP3 in the membrane and reduced pAKT activation. Our results are encouraging because these PUFAs impinge on AKT, a downstream target of Kras which serves as a logical alternative to the elusive therapies for Kras and often toxic effects of PI3K/AKT pathway inhibition. Citation Format: Carolina Torres, Georgina Mancinelli, Emily Chen, Jose Cordoba-Chacon, Danielle Pins, Ronald McKinney, Sara Saeed, Karla Castellanos, Giulia Orsi, Megha Singhal, Sam Grimaldo, Poorna Chandra Rao Yalagala, Papasani Subbaiah, Cecilia Leal, Paul Grippo. Dietary fat influences pancreatic cancer progression by altering cell membrane lipid content to impact the PI3K/AKT pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2581.

Supplementing conjugated linoleic acid in breeder hens diet increased conjugated linoleic acid incorporation in liver and alters hepatic lipid metabolism in chick offspring.

This experiment was designed to investigate the effect of supplementing conjugated linoleic acid (CLA) in breeder hens diet on development and hepatic lipid metabolism of chick offspring. Hy-Line Brown breeder hens were allocated into two groups, supplemented with 0 (control (CT)) or 0·5 % CLA for 8 weeks. Offspring chicks were grouped according to the mother generation and fed for 7 d. CLA treatment had no significant influence on development, egg quality and fertility of breeder hens but darkened the egg yolks in shade and increased yolk sac mass compared with the CT group. Addition of CLA resulted in increased body mass and liver mass and decreased deposition of subcutaneous adipose tissue in chick offspring. The serum TAG and total cholesterol levels of chick offspring were decreased in CLA group. CLA treatment increased the incorporation of both CLA isomers (c9t11 and t10c12) in the liver of chick offspring, accompanied by the decreased hepatic TAG levels, related to the significant reduction of fatty acid synthase (FAS) and acetyl-CoA carboxylase (ACC) enzyme activities and the increased carnitine palmitoyltransferase-1 (CPT1) enzyme activity. Meanwhile, CLA treatment reduced the mRNA expression of genes related to fatty acid biosynthesis (FAS, ACC and sterol regulatory element-binding protein-1c) and induced the expression of genes related to β-oxidative (CPT1, AMP-activated protein kinase and PPARα) in chick offspring liver. In summary, the addition of CLA in breeder hens diet significantly increased the incorporation of CLA in the liver of chick offspring, which further regulate hepatic lipid metabolism.

Poly(butylene succinate) fibrous dressings containing natural antimicrobial agents

Poly(butylene succinate) (PBSU) is a biodegradable and biocompatible synthetic aliphatic polyester, which has been used extensively in packaging, catering and agriculture, and more recently in drug delivery and bone and cartilage repair. PBSU-based mats created by electrospinning show promise as wound dressing materials because of their good mechanical properties, high surface area-to-volume ratio and increased levels of porosity. In this work, we present the creation of antimicrobial PBSU fibrous mats through the incorporation of natural food grade agents via blend electrospinning. Three types of edible gums (namely arabic, karaya and tragacanth), two essential oils (coriander and lavender), and one free fatty acid (linoleic acid) were added to PBSU containing a chain extender and their effect on six clinically relevant pathogens was evaluated. Mats containing essential oils at the highest concentration studied (7% w/v) showed some antimicrobial behaviour against S. aureus, E. hirae and P. aeruginosa, whereas the incorporation of linoleic acid at both concentrations tested (3% and 5% w/v) gave a strong reaction against S. pyogenes. Gum arabic was the only gum that had a considerable impact on S. aureus. Furthermore, the three gums enhanced the mechanical properties of the polymer mats and brought them closer to those of the human skin, whilst all agents maintained the high biocompatibility of the PBSU mats when contacted with mouse fibroblasts. This work, for the first time, shows the great promise of PBSU blended fibres as a skin substitute and paves the way towards bioactive and cost effective wound dressings from renewable materials.

Fatty acyl availability modulates cardiolipin composition and alters mitochondrial function in HeLa cells

The molecular assembly of cells depends not only on the balance between anabolism and catabolism but to a large degree on the building blocks available in the environment. For cultured mammalian cells, this is largely determined by the composition of the applied growth medium. Here, we study the impact of lipids in the medium on mitochondrial membrane architecture and function by combining LC-MS/MS lipidomics and functional tests with lipid supplementation experiments in an otherwise serum-free and lipid-free cell culture model. We demonstrate that the composition of mitochondrial cardiolipins strongly depends on the lipid environment in cultured cells and favors the incorporation of essential linoleic acid over other fatty acids. Simultaneously, the mitochondrial respiratory complex I activity was altered, whereas the matrix-localized enzyme citrate synthase was unaffected. This raises the question on a link between membrane composition and respiratory control. In summary, we found a strong dependency of central mitochondrial features on the type of lipids contained in the growth medium. This underlines the importance of considering these factors when using and establishing cell culture models in biomedical research. In summary, we found a strong dependency of central mitochondrial features on the type of lipids contained in the growth medium.

Lowering the linoleic acid to alpha-linoleic acid ratio decreases the production of inflammatory mediators by cultured human endothelial cells

Alpha-linolenic acid (ALA) and linoleic acid (LA) are precursors for longer-chain more unsaturated fatty acids and for lipid signalling molecules that may influence inflammatory processes through a variety of mechanisms. The actions of LA and ALA may be divergent and interdependent. The aim of this study was to investigate the incorporation and metabolism of ALA and LA in cultured in EA.hy926 endothelial cells and the production of inflammatory mediators (VEGF, RANTES, ICAM-1, MCP-1, IL-6 and IL-8) by these cells when exposed to different concentrations of ALA, LA and ratios of LA:ALA. Human endothelial cells were cultured with either culture medium or culture medium supplemented with ALA, LA or various ratios of LA:ALA (1:4, 1:1, 4:1, 9:1 or 19:1) followed by 24 h TNF-α stimulation; the total concentration of ALA plus LA was kept constant at 100 μM. The incorporation and metabolism of ALA and LA was measured using gas chromatography. The production of inflammatory mediators in the supernatant was assessed using a Luminex Multi-Analyte kit. Both ALA and LA were incorporated and metabolised by the endothelial cells. Cells incubated with ALA had a statistically significantly lower production of VEGF, RANTES, ICAM-1, MCP-1 and IL-6 compared to cells incubated without additional ALA. LA was not found to exert pro-inflammatory effects. Cells incubated with low LA:ALA ratios had lower production of VEGF, RANTES, MCP-1 and IL-6 when compared with a LA:ALA ratio of 19:1. These findings suggest that a low LA:ALA ratio exerts anti-inflammatory effects by lowering the production VEGF, RANTES, ICAM-1, MCP-1 and IL-6 in TNF-α stimulated endothelial cells compared to a high ratio. These effects were likely mediated by ALA, but LA may also possess some anti-inflammatory effects.

Protein Modification by Endogenously Generated Lipid Electrophiles: Mitochondria as the Source and Target.

Determining the impact of lipid electrophile-mediated protein damage that occurs during oxidative stress requires a comprehensive analysis of electrophile targets adducted under pathophysiological conditions. Incorporation of ω-alkynyl linoleic acid into the phospholipids of macrophages prior to activation by Kdo2-lipid A, followed by protein extraction, click chemistry, and streptavidin affinity capture, enabled a systems-level survey of proteins adducted by lipid electrophiles generated endogenously during the inflammatory response. Results revealed a dramatic enrichment for membrane and mitochondrial proteins as targets for adduction. A marked decrease in adduction in the presence of MitoTEMPO demonstrated a primary role for mitochondrial superoxide in electrophile generation and indicated an important role for mitochondria as both a source and target of lipid electrophiles, a finding that has not been revealed by prior studies using exogenously provided electrophiles.

Toxicological aspects of interesterified fat: Brain damages in rats

In recent years, interesterified fat (IF) has been used to replace hydrogenated vegetable fat (HVF), rich in trans isomers, being found in processed foods. Studies involving IF have shown deleterious influences on the metabolic system, similarly to HVF, whereas no studies regarding its influence on the central nervous system (CNS) were performed. Rats from first generation born and maintained under supplementation (3g/Kg, p.o.) of soybean-oil or IF until adulthood were assessed on memory, biochemical and molecular markers in the hippocampus. IF group showed higher saturated fatty acids and linoleic acid and lower docosahexaenoic acid incorporation in the hippocampus. In addition, IF supplementation impaired short and long-term memory, which were related to increased reactive species generation and protein carbonyl levels, decreased catalase activity, BDNF and TrkB levels in the hippocampus. To the best of our knowledge, this is the first study to show that lifelong IF consumption may be related to brain oxidative damage, memory impairments and neurotrophins modifications, which collectively may be present indifferent neurological disorders. In fact, the use of IF in foods was intended to avoid damage from HVF consumption; however this substitute should be urgently reviewed, since this fat can be as harmful as trans fat.

Molecular Characterization of Two Lysophospholipid:acyl-CoA Acyltransferases Belonging to the MBOAT Family in Nicotiana benthamiana.

In the remodeling pathway for the synthesis of phosphatidylcholine (PC), acyl-CoA-dependent lysophosphatidylcholine (lysoPC) acyltransferase (LPCAT) catalyzes the reacylation of lysoPC. A number of genes encoding LPCATs have been cloned and characterized from several plants in recent years. Using Arabidopsis and other plant LPCAT sequences to screen the genome database of Nicotiana benthamiana, we identified two cDNAs encoding the putative tobacco LPCATs (NbLPCAT1 and NbLPCAT2). Both of them were predicted to encode a protein of 463 amino acids with high similarity to LPCATs from other plants. Protein sequence features such as the presence of at least eight putative transmembrane regions, four highly conserved signature motifs and several invariant residues indicate that NbLPCATs belong to the membrane bound O-acyltransferase family. Lysophospholipid acyltransferase activity of NbLPCATs was confirmed by testing lyso-platelet-activating factor (lysoPAF) sensitivity through heterologous expression of each full-length cDNA in a yeast mutant Y02431 (lca1△) disrupted in endogenous LPCAT enzyme activity. Analysis of fatty acid profiles of phospholipids from the NbLPCAT-expressing yeast mutant Y02431 cultures supplemented with polyunsaturated fatty acids suggested more incorporation of linoleic acid (18:2n6, LA) and α-linolenic acid (18:3n3, ALA) into PC compared to yeast mutant harbouring empty vector. In vitro enzymatic assay demonstrated that NbLPCAT1had high lysoPC acyltransferase activity with a clear preference for α-linolenoyl-CoA (18:3), while NbLPCAT2 showed a high lysophosphatidic acid (lysoPA) acyltransferase activity towards α-linolenoyl-CoA and a weak lysoPC acyltransferase activity. Tissue-specific expression analysis showed a ubiquitous expression of NbLPCAT1 and NbLPCAT2 in roots, stems, leaves, flowers and seeds, and a strong expression in developing flowers. This is the first report on the cloning and characterization of lysophospholipid acyltransferases from N. benthamiana.

Fatty Acid Binding Protein 3 Is Involved in n–3 and n–6 PUFA Transport in Mouse Trophoblasts

Background:Low placental fatty acid (FA) transport during the embryonic period has been suggested to result in fetal developmental disorders and various adult metabolic diseases, but the molecular mechanism by which FAs are transported through the placental unit remains largely unknown. Objective:The aim of this study was to examine the distribution and functional relevance of FA binding protein (FABP), a cellular chaperone of FAs, in the mouse placenta. Methods:We clarified the localization of FABPs and sought to examine their function in placental FA transport through the phenotypic analysis of Fabp3-knockout mice. Results:Four FABPs (FABP3, FABP4, FABP5, and FABP7) were expressed with spatial heterogeneity in the placenta, and FABP3 was dominantly localized to the trophoblast cells. In placentas from the Fabp3-knockout mice (both sexes), the transport coefficients for linoleic acid (LA) were significantly reduced compared with those from wild-type mice by 25% and 44% at embryonic day (E) 15.5 and E18.5, respectively, whereas those for α-linolenic acid (ALA) were reduced by 19% and 17%, respectively. The accumulation of LA (18% and 27% at E15.5 and E18.5) and ALA (16% at E15.5) was also significantly less in the Fabp3-knockout fetuses than in wild-type fetuses. In contrast, transport and accumulation of palmitic acid (PA) were unaffected and glucose uptake significantly increased by 23% in the gene-ablated mice compared with wild-type mice at E18.5. Incorporation of LA (51% and 52% at 1 and 60 min, respectively) and ALA (23% at 60 min), but not PA, was significantly less in FABP3-knockdown BeWo cells than in controls, whereas glucose uptake was significantly upregulated by 51%, 50%, 31%, and 33% at 1, 20, 40, and 60 min, respectively. Conclusions:Collectively FABP3 regulates n–3 (ω-3) and n–6 (ω-6) polyunsaturated FA transport in trophoblasts and plays a pivotal role in fetal development.

Formulation and characterization of nanostructured lipid carriers containing a mixed lipids core

The present study aimed to evaluate influences of mixed lipids and their proportions on formation and properties of nanostructured lipid carriers (NLCs). The physicochemical parameters of drug-free NLCs were characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM), differential scanning calorimetry (DSC), X-ray diffraction (XRD) and rheological measurements. In addition, a drug incorporation model was introduced into the NLCs for the application test. The hydrodynamic diameter (Rd) of the final NLCs (varying from 150nm to 350nm) was affected by diverse lipids core. DSC results showed that both onset and melting points of NLCs declined due to the addition of the liquid oil, and the liquid oil enabled the melting peak wider, indicating a reduced crystallinity of NLC products which almost coincided with the XRD analysis. Furthermore, rheological studies suggested stronger elasticity property than the viscosity property of the NLC systems. NLCs with high lipids content (20wt%) had good thixotropy. Conjugated linoleic acid incorporation study showed that the NLC formulation had the well-controlled release profile.

Regulation of Cardiolipin Remodeling in Human Lymphoblasts

BackgroundBarth Syndrome (BTHS) is a rare X‐linked genetic disorder caused by mutations in the TAZ gene. The TAZ gene product, tafazzin, is responsible for remodeling Cardiolipin (CL) with the necessary acyl species. TAZ mutations can result in cardiomyopathy. Monolysocardiolipin acyltransferase‐1 (MLCL AT‐1) is another enzyme capable of CL remodeling.ObjectiveTo examine if MLCL AT‐1 complements tafazzin in the remodeling of CL.MethodsLymphoblasts from normal or BTHS patients were transfected with TAZ and/or MLCL AT‐1 RNAi or an MLCL AT‐1 carrying plasmid. TAZ and MLCL AT‐1 gene expression were analyzed. Other analyses included measuring CL mass, MLCL AT‐1 enzyme activity and radiolabelling studies. Our studies also include using BN PAGE to analyse mitochondrial complexes in normal and BTHS lymphoblasts.ResultsMLCL AT‐1 gene expression increased when TAZ was knocked down. Expression of MLCL AT‐1 elevated CL levels, increased [1‐14C] Linoleic acid incorporation into CL and raised MLCL AT‐1 enzyme activity in normal and BTHS cells in which TAZ was knocked down. The double knockdown results are not lower than TAZ knockdown alone. Results show that MLCL AT‐1 over‐expression is able to elevate mitochondrial complex I formation in BTHS lymphoblasts.ConclusionMLCL AT‐1 over‐expression may serve as a potential therapeutic approach to treat BTHS.Funding:CIHR, Barth Syndrome Foundations of USA and Canada

Effect of Time and Fatty Acid Composition in Eggs of White Leghorn Hens Supplemented with Tuna Oil

The study evaluated the effect of time and fatty acid (FA) composition in eggs of laying hens supplemented with tuna oil (TO). Two hundred White Leghorn hens 30-week were divided into 5 treatments with 4 replicates of 10 birds each. Treatments were assigned randomly and consisted of 0%, 0.75%, 1.50%, 2.25% and 3% TO in commercial sorghum-soybean meal diets, in which the soybean oil was partially replaced. The experiment was conducted for 90 days. At days 28, 56 and 84 of the experiment, 10 eggs per replicate were collected for chemical analysis. Feed intake, egg production rate, egg weight, and feed conversion were not influenced by dietary treatment. FA content was significantly altered (P < 0.05) by TO, showing a progressive increase in egg n-3 FA (especially docosahexaenoic [DHA] and eicosapentaenoic [EPA] acids) when TO was added. Levels of EPA and DHA were higher (P < 0.05) in the egg lipids of TO fed hens than those in the control group. This was correlated (P < 0.05) with the concentration of TO in the diet. However, no significant differences were observed either in egg weight or percent of egg lay between groups. There was a high correlation (P < 0.05) between TO inclusion level in the diet and the incorporation of EPA (r = 0.96) or DHA (r = 0.92) into the egg yolk and an overall decrease in n-6 FA. There was a high correlation (P < 0.05) between TO inclusion level and the incorporation of linoleic acid (LA, r = －0.95) or arachidonic acid (AA, r = －0.96) into the egg yolk. The highest incorporation (P < 0.05) of total n-3 FA content in eggs was obtained with 3% TO/kg. This increase was proportional to TO inclusion levels in the diets (r = 0.95). The results indicate that the n-3 FA content in eggs can be increased by dietary supplementation with TO, and that TO can serve as a reasonable alternative feed ingredient in layer diets to produce a healthier choice of egg.

Linoleic acid stimulates neutral lipid accumulation in lipid droplets of maturing bovine oocytes

Linoleic acid (LA) is a polyunsaturated fatty acid present in high concentrations in bovine follicular fluid; when added to maturation culture media, it affects oocyte competence (depending on the type and concentration of LA used). To date, little is known about the effective level of incorporation of LA and there is apparently no information regarding its esterification into various lipid fractions of the oocyte and its effect on neutral lipid storage. Therefore, the objective was to assess the uptake and subcellular lipid distribution of LA by analyzing incorporation of radiolabeled LA into oocyte polar and neutral lipid classes. The effects of various concentrations of LA on the nuclear status and cytoplasmic lipid content of bovine oocytes matured in vitro was also analyzed, with particular emphasis on intermediate concentrations of LA. Neutral lipids stored in lipid droplets were quantified with a fluorescence approach. Linoleic acid at 9 and 43 μM did not affect the nuclear status of oocytes matured in vitro, and 100 μM LA inhibited germinal vesicle breakdown, resulting in a higher percentage of oocytes arrested at the germinal state (43.5 vs. 3.0 in controls; P < 0.05). Bovine oocytes actively incorporated LA from the maturation medium (83.4 pmol LA per 100 oocytes at 22 hours of incubation; P < 0.05) and metabolized it mainly into major lipid classes, e.g., triacylglycerols and phospholipids (61.1% and 29.3%, respectively). Supplementation of the maturation medium with LA increased triacylglycerol accumulation in cytoplasmic lipid droplets at all concentrations assayed (P < 0.05). In conclusion, LA added to a defined maturation medium at concentrations that did not alter the nuclear status of bovine oocytes matured in vitro (9 and 43 μM) improved their quality by increasing the content of neutral lipids stored in lipid droplets. By directing the free fatty acid (LA) to triacylglycerol synthesis pathways and increasing the degree of unsaturation of membrane phospholipids, the oocyte was protected from lipotoxic effects (with an expectation of improved cryotolerance).

Effect of Functional Buffalo Cheese on Fatty Acid Profile and Oxidative Status of Liver and Intestine of Mice

The objective of this study was to determine the influence of administration of buffalo dairy products on lipid content and conjugated linoleic acid (CLA) incorporation on liver and intestine of mice. Buffalo cheeses were selected according to nutritional properties and CLA content. Cheeses were previously manufactured using as adjunct culture bacteria with probiotic or technological properties. BALB/c mice were fed for 28 days, and then a single dose of 1,2-dimethylhydrazine (DMH) as oxidant agent was administered before the influence of diet and DMH on antioxidant status in tissues was evaluated. Mice fed buffalo cheese showed the highest body weight gain (P < .05). Polyunsaturated fatty acid (PUFA) content in foods was very different, but total PUFA incorporation was similar in mouse tissues. CLA was only detected in fat tissues of mice fed dairy products, with cis-9, trans-11 being the major isomer. A higher linolenic (C(18:3)) acid content was found in tissues of mice fed commercial diet (control group), and it was partially replaced by CLA in groups receiving buffalo milk or cheese. Lipoperoxides (thiobarbituric acid-reactive substances) were higher in tissues of the control group with or without DMH administration, and DMH had a cytotoxic effect on colon cells (P < .05). Superoxide dismutase (SOD) and catalase activities in liver and intestine were similar among animals, with a slight increase of SOD detected after DMH treatment. Consumption of buffalo dairy products did not affect the oxidative status of mice tissues even after DMH application. In the present study, a protective effect of buffalo cheese and milk on intestine cells was determined.

Epidermal FABP (FABP5) Regulates Keratinocyte Differentiation by 13(S)-HODE-Mediated Activation of the NF-κB Signaling Pathway

Fatty acid-binding proteins (FABPs) are postulated to serve as lipid shuttles that solubilize hydrophobic fatty acids and deliver them to appropriate intracellular sites. Epidermal FABP (E-FABP/FABP5) is predominantly expressed in keratinocytes and is overexpressed in the actively proliferating tissue characteristic of psoriasis and wound healing. In this study, we found decreased expression of the differentiation-specific proteins keratin 1, involucrin, and loricrin in E-FABP(-/-) keratinocytes relative to E-FABP(+/+) keratinocytes. We also determined that incorporation of linoleic acid was significantly reduced in E-FABP(-/-) keratinocytes. Although linoleic acid did not directly affect keratinocyte differentiation, keratin 1 expression was induced by the linoleic acid derivative 13(S)-hydroxyoctadecadienoic acid (13(S)-HODE), and this induction was concomitant with increased NF-κB activity. In E-FABP(-/-) keratinocytes, the expression of 13(S)-HODE and the subsequent induction of NF-κB activity was lower than in wild-type keratinocytes. The reduction of linoleic acid in E-FABP(-/-) keratinocytes led to decreased cellular 13(S)-HODE content, resulting in decreased keratin 1 expression through downregulation of NF-κB activity. The regulation of fatty acid metabolism by E-FABP during keratinocyte differentiation suggests that E-FABP may have a role in the pathogenesis of psoriasis.

Dietary conjugated linoleic acid isomers change the unsaturation degree of hepatic fatty acids in neutral lipids but not in polar lipids

The fatty acid composition of phospholipids plays a key role in the structural and functional properties of cellular membrane. In this study, it was hypothesized that conjugated linoleic acid (CLA) isomer supplementation changes the unsaturation degree of the fatty acids of neutral lipids (NLs) but not those of polar lipids (PLs). Thus, the main goal was to determine the pattern of fatty acid incorporation into hepatic PL and NL fractions. Wistar male rats were fed cis(c)9,trans(t)11 and t10,c12 CLA isomers, separately or as a mixture. Whereas the t10,c12 isomer incorporation in the PL fraction was similar when supplemented either individually or as a mixture, the c9,t11 isomer reached the highest values of incorporation when combined with t10,c12. In the PL fraction, the linoleic acid did not change; but the arachidonic acid decreased, especially in the rats given the mixture. Also in this fraction, the t10,c12 isomer, either separately or as a mixture, decreased the amounts of n-6 long-chain (LC) polyunsaturated fatty acids (PUFA) and increased those of the n-3 LC PUFA relative to the control. In the NL fraction, linoleic acid incorporation followed the diet composition, whereas the arachidonic acid was similar among treatments. Facing CLA isomer supplementation, the present study suggests that fatty acid incorporation into phospholipids, through the balance between n-6 and n-3 LC PUFA, is dependent upon maintaining the unsaturation degree of cellular membrane.