Synthesis Of Complex Lipids Research Articles

Lipase Activity in the Larval Midgut of Rhynchophorus palmarum: Biochemical Characterization and the Effects of Reducing Agents.

Lipases have key roles in insect lipid acquisition, storage, and mobilization and are also fundamental to many physiological processes in insects. Lipids are an important component of insect diets, where they are hydrolyzed in the midgut lumen, absorbed, and used for the synthesis of complex lipids. The South American palm weevil Rhynchophorus palmarum is one of the most important pests on commercial palm plantations. However, there are few studies about lipid digestion for this insect. In this work, we have described the biochemical characterization of the lipase activity in the posterior midgut of the R. palmarum palm weevil. Lipase activity was highest between the temperatures of 37 °C and 45 °C and at pH 6.5. Lipase activity was also sensitive to variations in salt and calcium concentrations. Lipases have been described structurally as enzymes with the Ser-His-Asp Catalytic Triad, containing an active serine. The serine protease inhibitor PMSF (phenylmethane sulfonyl fluoride) inhibited the lipases from R. palmarum, demonstrating the importance of a serine residue for this activity. The ability of the lipases to hydrolyze p-Nitrophenyl esters with different chain lengths has revealed the activities of a broad range of substrates. The lipase activities of R. palmarum increased in the presence of reduced glutathione (GSH) and dithiothreitol (DTT), while in the presence of oxidized glutathione (GSSG), activities were drastically reduced. To our knowledge, this study has provided the first information about lipase activity in the R. palmarum palm weevil.

2-Deoxy-D-glucose treatment changes the Golgi apparatus architecture without blocking synthesis of complex lipids.

The classic Golgi apparatus organization, an arrangement of highly ordered cisternal stacks with tubular-vesicular membrane specializations on both sides, is the functional image of a continuous flow of contents and membranes with input, metabolization, and output in a dynamic steady state. In response to treatment with 2-deoxy-D-glucose (2-DG), which lowers the cellular ATP level by about 70% within minutes, this organization is rapidly replaced by tubular-glomerular membrane convolutes described as Golgi networks and bodies. 2-DG is a non-metabolizable glucose analogue and competitive inhibitor of glycolysis, which has become attractive in the context of therapeutic approaches for several kinds of tumors specifically targeting glycolysis in cancer. With the question of whether the functions of the Golgi apparatus in lipid synthesis would be influenced by the 2-DG-induced Golgi apparatus reorganization, we focused on lipid metabolism within the Golgi bodies. For this, we applied a fluorophore-labeled short-chain ceramide (BODIPY-Cer) in various combinations with 2-DG treatment to HepG2 cell cultures and followed uptake, enrichment and metabolization to higher ordered lipids. The cellular ATP status in each experiment was controlled with a bioluminescence assay, and the response of the Golgi apparatus was tracked by immunostaining of the trans-Golgi network protein TGN46. For electron microscopy, the fluorescent BODIPY-Cer signals were converted into electron-dense precipitates by photooxidation of diaminobenzidine (DAB); DAB precipitates labeled trans-Golgi areas in control cultures but also compartments at the periphery of the Golgi bodies formed in response to 2-DG treatment, thus indicating that concentration of ceramide takes place in spite of the Golgi apparatus reorganization. Lipid analyses by thin-layer chromatography (TLC) performed in parallel showed that BODIPY-Cer is not only concentrated in compartments of the 2-DG-induced Golgi bodies but is partly metabolized to BODIPY-sphingomyelin. Both, uptake and condensation of BODIPY-Cer and its conversion to complex lipids indicate that functions of the Golgi apparatus in the cellular lipid metabolism persist although the classic Golgi apparatus organization is abolished.

The clinical spectrum of inherited diseases involved in the synthesis and remodeling of complex lipids. A tentative overview.

Over one hundred diseases related to inherited defects of complex lipids synthesis and remodeling are now reported. Most of them were described within the last 5 years. New descriptions and phenotypes are expanding rapidly. While the associated clinical phenotype is currently difficult to outline, with only a few patients identified, it appears that all organs and systems may be affected. The main clinical presentations can be divided into (1) Diseases affecting the central and peripheral nervous system. Complex lipid synthesis disorders produce prominent motor manifestations due to upper and/or lower motoneuron degeneration. Motor signs are often complex, associated with other neurological and extra-neurological signs. Three neurological phenotypes, spastic paraparesis, neurodegeneration with brain iron accumulation and peripheral neuropathies, deserve special attention. Many apparently well clinically defined syndromes are not distinct entities, but rather clusters on a continuous spectrum, like for the PNPLA6-associated diseases, extending from Boucher-Neuhauser syndrome via Gordon Holmes syndrome to spastic ataxia and pure hereditary spastic paraplegia; (2) Muscular/cardiac presentations; (3) Skin symptoms mostly represented by syndromic (neurocutaneous) and non syndromic ichthyosis; (4) Retinal dystrophies with syndromic and non syndromic retinitis pigmentosa, Leber congenital amaurosis, cone rod dystrophy, Stargardt disease; (5) Congenital bone dysplasia and segmental overgrowth disorders with congenital lipomatosis; (6) Liver presentations characterized mainly by transient neonatal cholestatic jaundice and non alcoholic liver steatosis with hypertriglyceridemia; and (7) Renal and immune presentations. Lipidomics and molecular functional studies could help to elucidate the mechanism(s) of dominant versus recessive inheritance observed for the same gene in a growing number of these disorders.

The chain-flipping mechanism of ACP (acyl carrier protein)-dependent enzymes appears universal

ACPs (acyl carrier proteins) play essential roles in the synthesis of fatty acids, polyketides and non-ribosomal polypeptides. ACP function requires the modification of the protein by attachment of 4'-phosphopantetheine to a conserved serine residue. The phosphopantetheine thiol acts to tether the starting materials and intermediates as their thioesters. ACPs are small highly soluble proteins composed of four α-helices. The helices form a bundle that acts as a hydrophobic sleeve that sequesters the acyl chains and activated thioesters from solvent. However, in the synthesis of fatty acids and complex lipids the enzymes of the pathway must access the thioester and the proximal carbon atoms in order to perform the needed chemistry. How such access is provided without exposure of the acyl chains to solvent has been a longstanding question due to the lack of acyl-ACP-enzyme complexes, a situation generally attributed to the brevity of the interactions of acyl-ACPs with their cognate enzymes. As discussed in the present review the access question has now been answered by four recent crystal structures, each of which shows that the entire acyl chain plus the 4'-phosphopantetheine prosthetic group partitions from the ACP hydrophobic sleeve into a hydrophobic pocket or groove of the enzyme protein, a process termed chain flipping.

Regulation of neuron function and metabolism by acyl‐CoA hydrolysis (476.1)

Numerous neurological diseases are associated with dysregulated lipid metabolism; however, the regulation of fatty acid metabolism in neurons has been largely unexplored. The control of cellular fatty acid flux lies in the activation of fatty acids into acyl‐CoAs, and counterbalanced by the deactivation of acyl‐CoAs by acyl‐CoA thioesterases (ACOTs). ACOT7 is specifically and abundantly expressed in neurons, suggesting that it serves to antagonize fatty acid metabolism in neurons which have minimal capacity to oxidize or store lipids. To determine the role of ACOT7 in neural function and metabolism we have generated a nervous system specific ACOT7 knockout mouse, Acot7N‐/‐. Acot7N‐/‐ brains have increased fatty acid flux into complex lipid synthesis and increased phospholipid remodeling particularly under fasting conditions. The loss of neuronal ACOT7 causes physiological and behavioral abnormalities manifesting in a loss of loss of lean mass, hypermetabolism, hepatic steatosis, dyslipidemia, neurological dysfunction, neurodegeneration and behavioral hyperexcitability. The hyperexcitability of Acot7N‐/‐ mice is further exemplified by increased sensitivity to kainic acid induced seizures. These data show that ACOT7 mediated acyl‐CoA hydrolysis counter‐regulates fatty acid metabolism in neurons to protect against neurotoxicity and hyperexcitability.

Quantitative analysis of the green microalga Coccomyxa subellipsoidea C169 reveals homeostasis of complex lipid synthesis retained during nitrogen stress (605.14)

The inevitable decline of fossil fuel availability and the destructive escalation of atmospheric carbon dioxide concentration are pressing problems that scientific innovation can potentially address. Algae accumulate large amounts of oil when exposed to abiotic stress, enough to possibility use their cultivation to produce carbon‐neutral replacements of conventional transportation fuels. Unraveling the intricacies of triglyceride (TG) synthesis during abiotic stress is a means to generate genetic intervention strategies for improving lipid productivity. Our research is focused on understanding TG accumulation in the context of lipid biosynthesis for Coccomyxa subellipsoidea C169, a sequenced relative of Chlorella and a sensible proxy for biofuel production species. A thorough analysis of lipid synthesis was conducted including GC‐MS fatty acid profiles of isolated lipid pools and LC‐MS/MS quantification of membrane lipid (ML) and TG pools through a time course of N starvation. Stable isotopic labeling studies were done with 13CO2 and D2O to analyze the roles of photosynthesis and de novo synthesis from previously reduced carbon in TG synthesis. The data quantify an initial redistribution of membrane lipids into the TG pool followed by synthesis and incorporation of fatty acids into all complex lipid pools. Reproductive growth is inhibited by N stress, however lipid synthesis continues normally.Grant Funding Source: Supported by NSF‐EPSCoR, Nebraska Center for Algal Biology and Biotechnology

Atypical antipsychotics alter cholesterol and fatty acid metabolism in vitro

Haloperidol, a typical antipsychotic, has been shown to inhibit cholesterol biosynthesis by affecting Δ(7)-reductase, Δ(8,7)-isomerase, and Δ(14)-reductase activities, which results in the accumulation of different sterol intermediates. In the present work, we investigated the effects of atypical or second-generation antipsychotics (SGA), such as clozapine, risperidone, and ziprasidone, on intracellular lipid metabolism in different cell lines. All the SGAs tested inhibited cholesterol biosynthesis. Ziprasidone and risperidone had the same targets as haloperidol at inhibiting cholesterol biosynthesis, although with different relative activities (ziprasidone > haloperidol > risperidone). In contrast, clozapine mainly affected Δ(24)-reductase and Δ(8,7)-isomerase activities. These amphiphilic drugs also interfered with the LDL-derived cholesterol egress from the endosome/lysosome compartment, thus further reducing the cholesterol content in the endoplasmic reticulum. This triggered a homeostatic response with the stimulation of sterol regulatory element-binding protein (SREBP)-regulated gene expression. Treatment with SGAs also increased the synthesis of complex lipids (phospholipids and triacylglycerides). Once the antipsychotics were removed from the medium, a rebound in the cholesterol biosynthesis rate was detected, and the complex-lipid synthesis further increased. In this condition, apolipoprotein B secretion was also stimulated as demonstrated in HepG2 cells. These effects of SGAs on lipid homeostasis may be relevant in the metabolic side effects of antipsychotics, especially hypertriglyceridemia.

Ras-induced Up-regulation of CTP:Phosphocholine Cytidylyltransferase α Contributes to Malignant Transformation of Intestinal Epithelial Cells

Cancer cells have enhanced lipogenic capacity characterized by increased synthesis of fatty acids and complex lipids, including phosphatidylcholine (PC). As the rate-limiting enzyme in the CDP-choline pathway for PC synthesis, CTP:phosphocholine cytidylyltransferase α (CCTα) is implicated in the provision of membranes and bioactive lipids necessary of cell proliferation. In this study, we assessed the role of CCTα in malignant intestinal epithelial cells transformed with activated H-ras (IEC-ras). Three IEC-ras clones had significant up-regulation CCTα expression, but PC synthesis and in vitro activity of CCTα were similar to control IEC. RNA interference of CCTα in adherent IEC-ras did not affect PC synthesis, confirming that the enzyme was relatively inactive. However, CCTα silencing in ras-transformed IEC reduced anchorage-independent growth, a criterion for malignant transformation, as well as tumorigenicity in mice. Relative to their adherent counterparts, detached IEC-ras had increased PC synthesis that was attenuated by inducible CCTα silencing. Detachment of IEC-ras was accompanied by increased CCTα phosphorylation and cytosolic enzyme activity. We conclude that the expanded pool of CCTα in IEC-ras is activated by detachment. This provides the increased PC biosynthetic capacity that contributes to malignant transformation of intestinal epithelial cells when detached from the extracellular matrix.

Jean‐Marc Reyrat (29/04/1967–28/10/2009)

Jean‐Marc Reyrat (29/04/1967–28/10/2009)

Defining mycobacteria: Shared and specific genome features for different lifestyles

During the last decade, the combination of rapid whole genome sequencing capabilities, application of genetic and computational tools, and establishment of model systems for the study of a range of species for a spectrum of biological questions has enhanced our cumulative knowledge of mycobacteria in terms of their growth properties and requirements. The adaption of the corynebacterial surrogate system has simplified the study of cell wall biosynthetic machinery common to actinobacteria. Comparative genomics supported by experimentation reveals that superimposed on a common core of 'mycobacterial' gene set, pathogenic mycobacteria are endowed with multiple copies of several protein families that encode novel secretion and transport systems such as mce and esx; immunomodulators named PE/PPE proteins, and polyketide synthases for synthesis of complex lipids. The precise timing of expression, engagement and interactions involving one or more of these redundant proteins in their host environments likely play a role in the definition and differentiation of species and their disease phenotypes. Besides these, only a few species specific 'virulence' factors i.e., macromolecules have been discovered. Other subtleties may also arise from modifications of shared macromolecules. In contrast, to cope with the broad and changing growth conditions, their saprophytic relatives have larger genomes, in which the excess coding capacity is dedicated to transcriptional regulators, transporters for nutrients and toxic metabolites, biosynthesis of secondary metabolites and catabolic pathways. In this review, we present a sampling of the tools and techniques that are being implemented to tease apart aspects of physiology, phylogeny, ecology and pathology and illustrate the dominant genomic characteristics of representative species. The investigation of clinical isolates, natural disease states and discovery of new diagnostics, vaccines and drugs for existing and emerging mycobacterial diseases, particularly for multidrug resistant strains are the challenges in the coming decades.

Resveratrol inhibits fatty acid and triacylglycerol synthesis in rat hepatocytes

The putative role of resveratrol, a polyphenol present in grapes and other plants, in modulating dislypidemia, thus preventing cardiovascular diseases, is generally based on proliferating cell lines and in vivo studies in different pathological conditions. The aim of the present study was to investigate whether resveratrol plays a role on lipid biosynthesis in rat hepatocytes. The effect of resveratrol on total rate of fatty acid, cholesterol and complex lipid synthesis, assayed by the incorporation of [1-(14)C]acetate into these lipid fractions, was investigated in rat hepatocyte suspensions. Enzyme activities of acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) as well as 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA-R), pace-setting steps of de novo fatty acid and cholesterol synthesis, respectively, were in situ measured in digitonin-permeabilized hepatocytes. Resveratrol-treated hepatocytes exhibited a short-term (30 min) inhibition (IC(50) approximately 25 microm) of total fatty acid synthesis from [1-(14)C]acetate. Among neosynthesized fatty acids, palmitic acid formation was mainly reduced, thus suggesting that enzymatic step(s) of de novo fatty acid synthesis was affected by resveratrol. In digitonin-permeabilized hepatocytes, only ACC activity was noticeably reduced, while no change in FAS activity was observed. A noticeable resveratrol-induced reduction of label incorporation into triacylglycerols was also detected. Conversely, cholesterol synthesis and HMG-CoA-R activity were unaffected by resveratrol. Results here reported show that in isolated hepatocytes from normal rats a resveratrol-induced short-term inhibition of fatty acid and triacylglycerol synthesis occurs. This finding may represent a potential mechanism contributing to the reported hypolipidemic effect of resveratrol.

Shotgun lipidomics of neutral lipids as an enabling technology for elucidation of lipid-related diseases

Neutral lipids fulfill multiple specialized roles in cellular function. These roles include energy storage and utilization, the synthesis of complex lipids in cellular membranes, lipid second messengers for cellular signaling, and the modulation of membrane molecular dynamics. We have developed a novel mass spectrometric technology, now termed shotgun lipidomics, that can identify the types and amounts of thousands of lipids directly from extracts of biological samples. Shotgun lipidomics is well suited for the identification and measurement of the types and amounts of neutral lipid classes and individual molecular species through the use of multidimensional mass spectrometry. This review summarizes the basic principles underlying the use of shotgun lipidomics for the direct measurement of neutral lipids from extracts of biological tissues or fluids. Through exploiting the high information content inherent in shotgun lipidomics, this technology promises to greatly facilitate advances in our understanding of alterations in neutral lipid metabolism in health and disease.

Long-chain acyl-CoA synthetases and fatty acid channeling.

Thirteen homologous proteins comprise the long-chain acyl-CoA synthetase (ACSL), fatty acid transport protein (FATP), and bubblegum (ACSBG) subfamilies that activate long-chain and very-long-chain fatty acids to form acyl-CoAs. Gain- and loss-of-function studies show marked differences in the ability of these enzymes to channel fatty acids into different pathways of complex lipid synthesis. Further, the ability of the ACSLs and FATPs to enhance cellular FA uptake does not always require these proteins to be present on the plasma membrane; instead, FA uptake can be increased by enhancing its conversion to acyl-CoA and its metabolism in downstream pathways. Since altered fatty acid metabolism is a hallmark of numerous metabolic diseases and pathological conditions, the ACSL, FATP and ACSBG isoforms are likely to play important roles in disease etiology.

Development and Physiological Regulation of Intestinal Lipid Absorption. II. Dietary lipid absorption, complex lipid synthesis, and the intracellular packaging and secretion of chylomicrons

Research in dietary fat absorption has developed urgency because of the widely recognized epidemic of obesity in the United States. Despite its clinical importance, many controversies exist over some of the basic aspects of this process from the mechanisms of fatty acid uptake to the control of triacylglycerol export in chylomicrons. Recent advances have included the identification of a number of fatty acid transporters, the discovery of families of acyl-CoA synthetase long chains and acyltransferases, a physiological function for liver-fatty acid binding protein, and the characterization of the prechylomicron transport vesicle transporting chylomicrons from the endoplasmic reticulum to the Golgi.

In Vivo Functional Analyses of the Type II Acyl Carrier Proteins of Fatty Acid Biosynthesis

Acyl carrier protein (ACP) is a key component of the fatty acid synthesis pathways of both type I and type II synthesis systems. A large number of structure-function studies of various type II ACPs have been reported, but all are in vitro studies that assayed function or interaction of mutant ACPs with various enzymes of fatty acid synthesis or transfer. Hence in these studies functional properties of various mutant ACPs were assayed with only a subset of the many ACP-interacting proteins, which may not give an accurate overall view of the function of these proteins in vivo. This is especially so because Escherichia coli ACP has been reported to interact with several proteins that have no known roles in lipid metabolism. We therefore tested a large number of mutant derivatives of E. coli ACP carrying single amino acid substitutions for their abilities to restore growth to an E. coli strain carrying a temperature-sensitive mutation in acpP, the gene that encodes ACP. Many of these mutant proteins had previously been tested in vitro thus providing data for comparison with our results. We found that several mutant ACPs containing substitutions of ACP residues reported previously to be required for ACP function in vitro support normal growth of the acpP mutant strain. However, several mutant proteins reported to be severely defective in vitro failed to support growth of the acpP strain in vivo (or supported only weak growth). A collection of ACPs from diverse bacteria and from three eukaryotic organelles was also tested. All of the bacterial ACPs tested restored growth to the E. coli acpP mutant strain except those from two related bacteria, Enterococcus faecalis and Lactococcus lactis. Only one of the three eukaryotic organellar ACPs allowed growth. Strikingly the ACP is that of the apicoplast of Plasmodium falciparum (the protozoan that causes malaria). The fact that an ACP from a such diverse organism can replace AcpP function in E. coli suggests that some of the protein-protein interactions detected for AcpP may be not be essential for growth of E. coli.

Is Membrane Transport of FFA Mediated by Lipid, Protein, or Both?

![Figure][1] J. Patrick Kampf Torrey Pines Institute for Molecular Studies, San Diego, California ![Figure][1] Alan M. Kleinfeld Torrey Pines Institute for Molecular Studies, San Diego, California akleinfeld{at}tpims.org The mechanism of free fatty acid (FFA)

Extracellular glycerol regulates the cardiac energy balance in a working rat heart model

We reported previously that glycerol is a substrate for energy production in cardiomyocytes. Increasing glycerol availability results in increased glycerol uptake and its involvement in complex lipid biosynthesis and energy production. This study evaluated the relationship between glycerol supply, energy demand, and intermediary metabolism leading to energy production. The work was performed on isolated rat heart perfused in the working mode. Glycerol concentrations modeled the fasting (0.33 mM) and fed (3.33 mM) states. Cardiac energy demand was modeled by increasing heart rate from 350 to 450 beats/min (bpm). Increasing glycerol supply increased glycerol uptake from 1.4 (350 bpm) to 3.8 (450 bpm) and from 9.7 (350 bpm) to 34.2 (450 bpm) micro mol glycerol/heart in 30 min at 0.33 and 3.33 mM glycerol, respectively. At low glycerol supply, increasing heart rate did not influence the complex lipid synthesis. Conversely, high glycerol concentration increased the complex lipid synthesis by 5- and 30-fold at 350 and 450 bpm, respectively. Increasing glycerol supply and heart rate significantly increased glycerol oxidation rate. Moreover, increasing glycerol supply did not affect glucose oxidation but increased palmitate uptake and significantly decreased its beta-oxidation. Physiological concentrations of glycerol contribute to the cardiac intermediary metabolism, both for energy production and glycerolipid synthesis. Increasing energy demand enhances the requirement and use of glycerol. Glycerol contributes to the regulation of cardiac metabolism and energy balance, mainly by decreasing the contribution of fatty acid oxidation, and may thus represent a new factor in cardiac protection through the reduction of oxygen demand.

Thyroid hormone deficiency changes the distribution of oligodendrocyte/myelin markers during oligodendroglial differentiation in vitro

Myelination depends on the proper differentiation of oligodendrocytes and several factors may influence this event. For instance, thyroid hormone (T3) affects the timing of differentiation and regulates the expression of several enzymes involved in the synthesis of complex lipids and in the expression of some myelin structural proteins. We investigated the effect of T3 deficiency on oligodendroglial differentiation and in the distribution of oligodendrocyte/myelin proteins 2′3′-cyclic nucleotide 3′-phosphodiesterase (CNPase) and myelin basic protein (MBP). Oligodendroglial-enriched cultures were obtained from cerebra of neonate rats grown in a modified medium. The T3-deficient status was induced by using medium devoid of T3. We observed a delay, in T3-deficient cultures, in oligodendroglial maturation characterized by less extensive processes and membrane vellum than in controls. In control cultures, CNPase immunoreactivity was punctated, showing cell bodies and processes at earlier stages and redistribution to cytoskeleton vein-like structures in later stages. In T3-deficient cultures, CNPase remained in a punctated pattern and only at 10 days in vitro we observed CNPase redistribution to the presumptive cytoskeleton vein-like structures. MBP in control cultures was distributed through the whole cell body and processes whereas in T3-deficient cultures, MBP immunoreactivity was concentrated in the perinuclear region. These results reinforce the hypothesis that T3 is an important factor in oligodendrocyte differentiation, particularly regarding the distribution of myelin proteins.

22-Hydroxycholesterols regulate lipid metabolism differently than T0901317 in human myotubes

The nuclear liver X receptors (LXRα and β) are regulators of lipid and cholesterol metabolism. Oxysterols are known LXR ligands, but the functional role of hydroxycholesterols is at present unknown. In human myotubes, chronic exposure to the LXR ligand T0901317 promoted formation of diacylglycerol (DAG) and triacylglycerol (TAG), 22- R-hydroxycholesterol (22- R-HC) had no effect, and 22- S-hydroxycholesterol (22- S-HC) reduced the formation. In accordance with this, 22-HC and T0901317 regulated the expression of fatty acid transporter CD36, stearoyl-CoA desaturase-1, acyl-CoA synthetase long chain family member 1 and fatty acid synthase (FAS) differently; all genes were increased by T0901317, 22- R-HC did not change their expression level, while 22- S-HC reduced it. Transfection studies confirmed that the FAS promoter was activated by T0901317 and repressed by 22- S-HC through an LXR response element in the promoter. Both 22- R-HC and T0901317 increased gene expression of LXRα, sterol regulatory element-binding protein 1c and ATP-binding cassette transporter A1, while 22- S-HC had little effect. In summary, 22- R-HC regulated lipid metabolism and mRNA expression of some LXR target genes in human myotubes differently than T0901317. Moreover, 22- S-HC did not behave like an inactive ligand; it reduced synthesis of complex lipids and repressed certain genes involved in lipogenesis and lipid handling.

Eicosapentaenoic acid (20:5 n-3) increases fatty acid and glucose uptake in cultured human skeletal muscle cells

This study was conducted to evaluate the chronic effects of eicosapentaenoic acid (EPA) on fatty acid and glucose metabolism in human skeletal muscle cells. Uptake of [14C]oleate was increased >2-fold after preincubation of myotubes with 0.6 mM EPA for 24 h, and incorporation into various lipid classes showed that cellular triacylgycerol (TAG) and phospholipids were increased 2- to 3-fold compared with control cells. After exposure to oleic acid (OA), TAG was increased 2-fold. Insulin (100 nM) further increased the incorporation of [14C]oleate into all lipid classes for EPA-treated myotubes. Fatty acid beta-oxidation was unchanged, and complete oxidation (CO2) decreased in EPA-treated cells. Basal glucose transport and oxidation (CO2) were increased 2-fold after EPA, and insulin (100 nM) stimulated glucose transport and oxidation similarly in control and EPA-treated myotubes, whereas these responses to insulin were abolished after OA treatment. Lower concentrations of EPA (0.1 mM) also increased fatty acid and glucose uptake. CD36/FAT (fatty acid transporter) mRNA expression was increased after EPA and OA treatment compared with control cells. Moreover, GLUT1 expression was increased 2.5-fold by EPA, whereas GLUT4 expression was unchanged, and activities of the mitogen-activated protein kinase p38 and extracellular signal-regulated kinase were decreased after treatment with OA compared with EPA. Together, our data show that chronic exposure of myotubes to EPA promotes increased uptake and oxidation of glucose despite a markedly increased fatty acid uptake and synthesis of complex lipids.