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Sample records for phagocytosis requires triacylglycerol

  1. Outer segment phagocytosis by cultured retinal pigment epithelial cells requires Gas6.

    Science.gov (United States)

    Hall, M O; Prieto, A L; Obin, M S; Abrams, T A; Burgess, B L; Heeb, M J; Agnew, B J

    2001-10-01

    The function and viability of vertebrate photoreceptors requires the daily phagocytosis of photoreceptor outer segments (OS) by the adjacent retinal pigment epithelium (RPE). We demonstrate here a critical role in this process for Gas6 and by implication one of its receptor protein tyrosine kinases (RTKs), Mertk (Mer). Gas6 specifically and selectively stimulates the phagocytosis of OS by normal cultured rat RPE cells. The magnitude of the response is dose-dependent and shows an absolute requirement for calcium. By contrast the Royal College of Surgeons (RCS) rat RPE cells, in which a mutation in the gene Mertk results in the expression of a truncated, non-functional receptor, does not respond to Gas6. These data strongly suggest that activation of Mertk by its ligand, Gas6, is the specific signaling pathway responsible for initiating the ingestion of shed OS. Moreover, photoreceptor degeneration in the RCS rat retina, which lacks Mertk, and in humans with a mutation in Mertk, strongly suggests that the Gas6/Mertk signaling pathway is essential for photoreceptor viability. We believe that this is the first demonstration of a specific function for Gas6 in the eye. Copyright 2001 Academic Press.

  2. DGAT enzymes are required for triacylglycerol synthesis and lipid droplets in adipocytes.

    Science.gov (United States)

    Harris, Charles A; Haas, Joel T; Streeper, Ryan S; Stone, Scot J; Kumari, Manju; Yang, Kui; Han, Xianlin; Brownell, Nicholas; Gross, Richard W; Zechner, Rudolf; Farese, Robert V

    2011-04-01

    The total contribution of the acyl CoA:diacylglycerol acyltransferase (DGAT) enzymes, DGAT1 and DGAT2, to mammalian triacylglycerol (TG) synthesis has not been determined. Similarly, whether DGAT enzymes are required for lipid droplet (LD) formation is unknown. In this study, we examined the requirement for DGAT enzymes in TG synthesis and LDs in differentiated adipocytes with genetic deletions of DGAT1 and DGAT2. Adipocytes with a single deletion of either enzyme were capable of TG synthesis and LD formation. In contrast, adipocytes with deletions of both DGATs were severely lacking in TG and did not have LDs, indicating that DGAT1 and DGAT2 account for nearly all TG synthesis in adipocytes and appear to be required for LD formation during adipogenesis. DGAT enzymes were not absolutely required for LD formation in mammalian cells, however; macrophages deficient in both DGAT enzymes were able to form LDs when incubated with cholesterol-rich lipoproteins. Although adipocytes lacking both DGATs had no TG or LDs, they were fully differentiated by multiple criteria. Our findings show that DGAT1 and DGAT2 account for the vast majority of TG synthesis in mice, and DGAT function is required for LDs in adipocytes, but not in all cell types.

  3. DGAT enzymes are required for triacylglycerol synthesis and lipid droplets in adipocytes[S

    Science.gov (United States)

    Harris, Charles A.; Haas, Joel T.; Streeper, Ryan S.; Stone, Scot J.; Kumari, Manju; Yang, Kui; Han, Xianlin; Brownell, Nicholas; Gross, Richard W.; Zechner, Rudolf; Farese, Robert V.

    2011-01-01

    The total contribution of the acyl CoA:diacylglycerol acyltransferase (DGAT) enzymes, DGAT1 and DGAT2, to mammalian triacylglycerol (TG) synthesis has not been determined. Similarly, whether DGAT enzymes are required for lipid droplet (LD) formation is unknown. In this study, we examined the requirement for DGAT enzymes in TG synthesis and LDs in differentiated adipocytes with genetic deletions of DGAT1 and DGAT2. Adipocytes with a single deletion of either enzyme were capable of TG synthesis and LD formation. In contrast, adipocytes with deletions of both DGATs were severely lacking in TG and did not have LDs, indicating that DGAT1 and DGAT2 account for nearly all TG synthesis in adipocytes and appear to be required for LD formation during adipogenesis. DGAT enzymes were not absolutely required for LD formation in mammalian cells, however; macrophages deficient in both DGAT enzymes were able to form LDs when incubated with cholesterol-rich lipoproteins. Although adipocytes lacking both DGATs had no TG or LDs, they were fully differentiated by multiple criteria. Our findings show that DGAT1 and DGAT2 account for the vast majority of TG synthesis in mice, and DGAT function is required for LDs in adipocytes, but not in all cell types. PMID:21317108

  4. Activation-Inactivation Cycling of Rab35 and ARF6 Is Required for Phagocytosis of Zymosan in RAW264 Macrophages

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    Youhei Egami

    2015-01-01

    Full Text Available Phagocytosis of zymosan by phagocytes is a widely used model of microbial recognition by the innate immune system. Live-cell imaging showed that fluorescent protein-fused Rab35 accumulated in the membranes of phagocytic cups and then dissociated from the membranes of newly formed phagosomes. By our novel pull-down assay for Rab35 activity, we found that Rab35 is deactivated immediately after zymosan internalization into the cells. Phagosome formation was inhibited in cells expressing the GDP- or GTP-locked Rab35 mutant. Moreover, the simultaneous expression of ACAP2—a Rab35 effector protein—with GTP-locked Rab35 or the expression of plasma membrane-targeted ACAP2 showed a marked inhibitory effect on phagocytosis through ARF6 inactivation by the GAP activity of ACAP2. ARF6, a substrate for ACAP2, was also localized on the phagocytic cups and dissociated from the membranes of internalized phagosomes. In support of the microscopic observations, ARF6-GTP pull-down experiments showed that ARF6 is transiently activated during phagosome formation. Furthermore, the expression of GDP- or GTP-locked ARF6 mutants also suppresses the uptake of zymosan. These data suggest that the activation-inactivation cycles of Rab35 and ARF6 are required for the uptake of zymosan and that ACAP2 is an important component that links Rab35/ARF6 signaling during phagocytosis of zymosan.

  5. The zipper mechanism in phagocytosis: energetic requirements and variability in phagocytic cup shape

    Directory of Open Access Journals (Sweden)

    Dart Anna E

    2010-11-01

    Full Text Available Abstract Background Phagocytosis is the fundamental cellular process by which eukaryotic cells bind and engulf particles by their cell membrane. Particle engulfment involves particle recognition by cell-surface receptors, signaling and remodeling of the actin cytoskeleton to guide the membrane around the particle in a zipper-like fashion. Despite the signaling complexity, phagocytosis also depends strongly on biophysical parameters, such as particle shape, and the need for actin-driven force generation remains poorly understood. Results Here, we propose a novel, three-dimensional and stochastic biophysical model of phagocytosis, and study the engulfment of particles of various sizes and shapes, including spiral and rod-shaped particles reminiscent of bacteria. Highly curved shapes are not taken up, in line with recent experimental results. Furthermore, we surprisingly find that even without actin-driven force generation, engulfment proceeds in a large regime of parameter values, albeit more slowly and with highly variable phagocytic cups. We experimentally confirm these predictions using fibroblasts, transfected with immunoreceptor FcγRIIa for engulfment of immunoglobulin G-opsonized particles. Specifically, we compare the wild-type receptor with a mutant receptor, unable to signal to the actin cytoskeleton. Based on the reconstruction of phagocytic cups from imaging data, we indeed show that cells are able to engulf small particles even without support from biological actin-driven processes. Conclusions This suggests that biochemical pathways render the evolutionary ancient process of phagocytic highly robust, allowing cells to engulf even very large particles. The particle-shape dependence of phagocytosis makes a systematic investigation of host-pathogen interactions and an efficient design of a vehicle for drug delivery possible.

  6. Stimulation of phagocytosis by sulforaphane

    International Nuclear Information System (INIS)

    Suganuma, Hiroyuki; Fahey, Jed W.; Bryan, Kelley E.; Healy, Zachary R.; Talalay, Paul

    2011-01-01

    Research highlights: → Sulforaphane stimulates the phagocytosis of RAW 264.7 macrophages under conditions of serum deprivation. → This effect does not require Nrf2-dependent induction of phase 2 genes. → Inactivation of macrophage migration inhibitory factor (MIF) by sulforaphane may be involved in stimulation of phagocytosis by sulforaphane. -- Abstract: Sulforaphane, a major isothiocyanate derived from cruciferous vegetables, protects living systems against electrophile toxicity, oxidative stress, inflammation, and radiation. A major protective mechanism is the induction of a network of endogenous cytoprotective (phase 2) genes that are regulated by transcription factor Nrf2. To obtain a more detailed understanding of the anti-inflammatory and immunomodulatory effects of sulforaphane, we evaluated its effect on the phagocytosis activity of RAW 264.7 murine macrophage-like cells by measuring the uptake of 2-μm diameter polystyrene beads. Sulforaphane raised the phagocytosis activity of RAW 264.7 cells but only in the absence or presence of low concentrations (1%) of fetal bovine serum. Higher serum concentrations depressed phagocytosis and abolished its stimulation by sulforaphane. This stimulation did not depend on the induction of Nrf2-regulated genes since it occurred in peritoneal macrophages of nrf2 -/- mice. Moreover, a potent triterpenoid inducer of Nrf2-dependent genes did not stimulate phagocytosis, whereas sulforaphane and another isothiocyanate (benzyl isothiocyanate) had comparable inducer potencies. It has been shown recently that sulforaphane is a potent and direct inactivator of macrophage migration inhibitory factor (MIF), an inflammatory cytokine. Moreover, the addition of recombinant MIF to RAW 264.7 cells attenuated phagocytosis, but sulforaphane-inactivated MIF did not affect phagocytosis. The inactivation of MIF may therefore be involved in the phagocytosis-enhancing activity of sulforaphane.

  7. Stimulation of phagocytosis by sulforaphane

    Energy Technology Data Exchange (ETDEWEB)

    Suganuma, Hiroyuki, E-mail: hsuganu1@jhmi.edu [Lewis B. and Dorothy Cullman Cancer Chemoprotection Center, Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, 725 N. Wolfe Street, Baltimore, MD 21205 (United States); Fahey, Jed W., E-mail: jfahey@jhmi.edu [Lewis B. and Dorothy Cullman Cancer Chemoprotection Center, Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, 725 N. Wolfe Street, Baltimore, MD 21205 (United States); Bryan, Kelley E., E-mail: kbryanm1@jhmi.edu [Lewis B. and Dorothy Cullman Cancer Chemoprotection Center, Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, 725 N. Wolfe Street, Baltimore, MD 21205 (United States); Healy, Zachary R., E-mail: zhealy1@jhmi.edu [Lewis B. and Dorothy Cullman Cancer Chemoprotection Center, Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, 725 N. Wolfe Street, Baltimore, MD 21205 (United States); Talalay, Paul, E-mail: ptalalay@jhmi.edu [Lewis B. and Dorothy Cullman Cancer Chemoprotection Center, Department of Pharmacology and Molecular Sciences, Johns Hopkins University School of Medicine, 725 N. Wolfe Street, Baltimore, MD 21205 (United States)

    2011-02-04

    Research highlights: {yields} Sulforaphane stimulates the phagocytosis of RAW 264.7 macrophages under conditions of serum deprivation. {yields} This effect does not require Nrf2-dependent induction of phase 2 genes. {yields} Inactivation of macrophage migration inhibitory factor (MIF) by sulforaphane may be involved in stimulation of phagocytosis by sulforaphane. -- Abstract: Sulforaphane, a major isothiocyanate derived from cruciferous vegetables, protects living systems against electrophile toxicity, oxidative stress, inflammation, and radiation. A major protective mechanism is the induction of a network of endogenous cytoprotective (phase 2) genes that are regulated by transcription factor Nrf2. To obtain a more detailed understanding of the anti-inflammatory and immunomodulatory effects of sulforaphane, we evaluated its effect on the phagocytosis activity of RAW 264.7 murine macrophage-like cells by measuring the uptake of 2-{mu}m diameter polystyrene beads. Sulforaphane raised the phagocytosis activity of RAW 264.7 cells but only in the absence or presence of low concentrations (1%) of fetal bovine serum. Higher serum concentrations depressed phagocytosis and abolished its stimulation by sulforaphane. This stimulation did not depend on the induction of Nrf2-regulated genes since it occurred in peritoneal macrophages of nrf2{sup -/-} mice. Moreover, a potent triterpenoid inducer of Nrf2-dependent genes did not stimulate phagocytosis, whereas sulforaphane and another isothiocyanate (benzyl isothiocyanate) had comparable inducer potencies. It has been shown recently that sulforaphane is a potent and direct inactivator of macrophage migration inhibitory factor (MIF), an inflammatory cytokine. Moreover, the addition of recombinant MIF to RAW 264.7 cells attenuated phagocytosis, but sulforaphane-inactivated MIF did not affect phagocytosis. The inactivation of MIF may therefore be involved in the phagocytosis-enhancing activity of sulforaphane.

  8. Syk-dependent tyrosine phosphorylation of 3BP2 is required for optimal FcRγ-mediated phagocytosis and chemokine expression in U937 cells.

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    Chihara, Kazuyasu; Kato, Yuji; Yoshiki, Hatsumi; Takeuchi, Kenji; Fujieda, Shigeharu; Sada, Kiyonao

    2017-09-13

    The adaptor protein c-Abl SH3 domain binding protein-2 (3BP2) is tyrosine phosphorylated by Syk in response to cross-linking of antigen receptors, which in turn activates various immune responses. Recently, a study using the mouse model of cherubism, a dominant inherited disorder caused by mutations in the gene encoding 3BP2, showed that 3BP2 is involved in the regulation of phagocytosis mediated by Fc receptor for IgG (FcγR) in macrophages. However, the molecular mechanisms underlying 3BP2-mediated regulation of phagocytosis and the physiological relevance of 3BP2 tyrosine phosphorylation remains elusive. In this study, we established various gene knockout U937 cell lines using the CRISPR/Cas9 system and found that 3BP2 is rapidly tyrosine phosphorylated by Syk in response to cross-linking of FcγRI. Depletion of 3BP2 caused significant reduction in the Fc receptor γ chain (FcRγ)-mediated phagocytosis in addition to the FcγRI-mediated induction of chemokine mRNA for IL-8, CCL3L3 and CCL4L2. Syk-dependent tyrosine phosphorylation of 3BP2 was required for overcoming these defects. Finally, we found that the PH and SH2 domains play important roles on FcγRI-mediated tyrosine phosphorylation of 3BP2 in HL-60 cells. Taken together, these results indicate that Syk-dependent tyrosine phosphorylation of 3BP2 is required for optimal FcRγ-mediated phagocytosis and chemokine expression.

  9. Sensing of triacylglycerol in the gut

    DEFF Research Database (Denmark)

    Kleberg, Karen; Jacobsen, Anne Katrine; Ferreira, Jozelia G

    2015-01-01

    KEY POINTS: Digestion is required for intestinal sensing of triacylglycerol in this behavioural model. The hydrolysis products of triacylglycerol, fatty acids and 2-monoacylglycerol, regulate feeding via separate mechanisms. Sensing of long-chain fatty acids, but not of 2-monoacylglycerol......, stimulated central dopaminergic signalling. Fatty acid chain length regulates behavioural responses to fatty acids. ABSTRACT: Sensing of dietary triacylglycerol in the proximal small intestine results in physiological, hormonal and behavioural responses. However, the exact physiological pathways linking...... intestinal fat sensing to food intake and the activation of brain circuits remain to be identified. In this study we examined the role of triacylglycerol digestion for intestinal fat sensing, and compared the effects of the triacylglycerol digestion products, fatty acids and 2-monoacylglycerol...

  10. Commensal Bacteria-Induced Inflammasome Activation in Mouse and Human Macrophages Is Dependent on Potassium Efflux but Does Not Require Phagocytosis or Bacterial Viability.

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    Kejie Chen

    Full Text Available Gut commensal bacteria contribute to the pathogenesis of inflammatory bowel disease, in part by activating the inflammasome and inducing secretion of interleukin-1ß (IL-1ß. Although much has been learned about inflammasome activation by bacterial pathogens, little is known about how commensals carry out this process. Accordingly, we investigated the mechanism of inflammasome activation by representative commensal bacteria, the Gram-positive Bifidobacterium longum subspecies infantis and the Gram-negative Bacteroides fragilis. B. infantis and B. fragilis induced IL-1ß secretion by primary mouse bone marrow-derived macrophages after overnight incubation. IL-1ß secretion also occurred in response to heat-killed bacteria and was only partly reduced when phagocytosis was inhibited with cytochalasin D. Similar results were obtained with a wild-type immortalized mouse macrophage cell line but neither B. infantis nor B. fragilis induced IL-1ß secretion in a mouse macrophage line lacking the nucleotide-binding/leucine-rich repeat pyrin domain containing 3 (NLRP3 inflammasome. IL-1ß secretion in response to B. infantis and B. fragilis was significantly reduced when the wild-type macrophage line was treated with inhibitors of potassium efflux, either increased extracellular potassium concentrations or the channel blocker ruthenium red. Both live and heat-killed B. infantis and B. fragilis also induced IL-1ß secretion by human macrophages (differentiated THP-1 cells or primary monocyte-derived macrophages after 4 hours of infection, and the secretion was inhibited by raised extracellular potassium and ruthenium red but not by cytochalasin D. Taken together, our findings indicate that the commensal bacteria B. infantis and B. fragilis activate the NLRP3 inflammasome in both mouse and human macrophages by a mechanism that involves potassium efflux and that does not require bacterial viability or phagocytosis.

  11. Downstream components of RhoA required for signal pathway of superoxide formation during phagocytosis of serum opsonized zymosans in macrophages.

    Science.gov (United States)

    Kim, Jun Sub; Kim, Jae Gyu; Jeon, Chan Young; Won, Ha Young; Moon, Mi Young; Seo, Ji Yeon; Kim, Jong Il; Kim, Jaebong; Lee, Jae Yong; Choi, Soo Young; Park, Jinseu; Yoon Park, Jung Han; Ha, Kwon Soo; Kim, Pyeung Hyeun; Park, Jae Bong

    2005-12-31

    Rac1 and Rac2 are essential for the control of oxidative burst catalyzed by NADPH oxidase. It was also documented that Rho is associated with the superoxide burst reaction during phagocytosis of serum- (SOZ) and IgG-opsonized zymosan particles (IOZ). In this study, we attempted to reveal the signal pathway components in the superoxide formation regulated by Rho GTPase. Tat-C3 blocked superoxide production, suggesting that RhoA is essentially involved in superoxide formation during phagocytosis of SOZ. Conversely SOZ activated both RhoA and Rac1/2. Inhibition of RhoA-activated kinase (ROCK), an important downstream effector of RhoA, by Y27632 and myosin light chain kinase (MLCK) by ML-7 abrogated superoxide production by SOZ. Extracellular signaling-regulated kinase (ERK)1/2 and p38 mitogen-activated protein kinase (MAPK) were activated during phagocytosis of SOZ, and Tat-C3 and SB203580 reduced ERK1/2 and p38 MAPK activation, suggesting that RhoA and p38 MAPK may be upstream regulators of ERK1/2. Inhibition of ERK1/2, p38 MAPK, phosphatidyl inositol 3-kinase did not block translocation of RhoA to membranes, suggesting that RhoA is upstream to these kinases. Inhibition of RhoA by Tat-C3 blocked phosphorylation of p47(PHOX). Taken together, RhoA, ROCK, p38MAPK, ERK1/2, and p47(PHOX) may be subsequently activated, leading to activation of NADPH oxidase to produce superoxide.

  12. Disruption of Sphingolipid Biosynthesis Blocks Phagocytosis of Candida albicans.

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    Fikadu G Tafesse

    2015-10-01

    Full Text Available The ability of phagocytes to clear pathogens is an essential attribute of the innate immune response. The role of signaling lipid molecules such as phosphoinositides is well established, but the role of membrane sphingolipids in phagocytosis is largely unknown. Using a genetic approach and small molecule inhibitors, we show that phagocytosis of Candida albicans requires an intact sphingolipid biosynthetic pathway. Blockade of serine-palmitoyltransferase (SPT and ceramide synthase-enzymes involved in sphingolipid biosynthesis- by myriocin and fumonisin B1, respectively, impaired phagocytosis by phagocytes. We used CRISPR/Cas9-mediated genome editing to generate Sptlc2-deficient DC2.4 dendritic cells, which lack serine palmitoyl transferase activity. Sptlc2-/- DC2.4 cells exhibited a stark defect in phagocytosis, were unable to bind fungal particles and failed to form a normal phagocytic cup to engulf C. albicans. Supplementing the growth media with GM1, the major ganglioside present at the cell surface, restored phagocytic activity of Sptlc2-/- DC2.4 cells. While overall membrane trafficking and endocytic pathways remained functional, Sptlc2-/- DC2.4 cells express reduced levels of the pattern recognition receptors Dectin-1 and TLR2 at the cell surface. Consistent with the in vitro data, compromised sphingolipid biosynthesis in mice sensitizes the animal to C. albicans infection. Sphingolipid biosynthesis is therefore critical for phagocytosis and in vivo clearance of C. albicans.

  13. Teaching Phagocytosis Using Flow Cytometry

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    John Boothby

    2009-12-01

    Full Text Available Investigative microbiology on protists in a basic teaching laboratory environment is limited by student skill level, ease of microbial culture and manipulation, instrumentation, and time. The flow cytometer is gaining use as a mainstream instrument in research and clinical laboratories, but has had minimal application in teaching laboratories. Although the cost of a flow cytometer is currently prohibitive for many microbiology teaching environments and the number of trained instructors and teaching materials is limited, in many ways the flow cytometer is an ideal instrument for teaching basic microbiology. We report here on a laboratory module to study phagocytosis in Tetrahymena sp. using flow cytometry in a basic microbiology teaching laboratory. Students and instructors found the flow cytometry data analysis program, Paint-A-GatePRO-TM, to be very intuitive and easy to learn within a short period of time. Assessment of student learning about Tetrahymena sp., phagocytosis, flow cytometry, and investigative microbiology using an inquiry-based format demonstrated an overall positive response from students.

  14. DGAT enzymes and triacylglycerol biosynthesis

    OpenAIRE

    Yen, Chi-Liang Eric; Stone, Scot J.; Koliwad, Suneil; Harris, Charles; Farese, Robert V.

    2008-01-01

    Triacylglycerols (triglycerides) (TGs) are the major storage molecules of metabolic energy and FAs in most living organisms. Excessive accumulation of TGs, however, is associated with human diseases, such as obesity, diabetes mellitus, and steatohepatitis. The final and the only committed step in the biosynthesis of TGs is catalyzed by acyl-CoA:diacylglycerol acyltransferase (DGAT) enzymes. The genes encoding two DGAT enzymes, DGAT1 and DGAT2, were identified in the past decade, ...

  15. Dietary control of equine plasma triacylglycerols

    NARCIS (Netherlands)

    Hallebeek, Johanna Maria

    2002-01-01

    The study of this thesis concerns the dietary influence on lipid metabolism in horses. The main issue is the effect of dietary medium chain triacylglycerols (MCT) on triacylglycerol metabolism. In certain conditions high-fat diets can be beneficial for horses. Diets rich in long-chain

  16. Intestinal absorption of specific structured triacylglycerols

    DEFF Research Database (Denmark)

    Mu, Huiling; Høy, Carl-Erik

    2001-01-01

    on a reversed-phase high performance Liquid chromatograph (RP-HPLC) and identified by atmospheric pressure chemical ionization mass spectrometry, The composition of triacylglycerols was quantified by RP-HPLC with evaporative Light scattering detection. The intact MLM-type triacylglycerols were detected...

  17. Triacylglycerol-based fuels: An evaluation

    Science.gov (United States)

    A variety of feedstocks exist and several processes have been developed to produce alternative diesel fuels from triacylglycerol-based materials, such as plant and algal oils as well as animal fats and used cooking oils. Biodiesel is obtained by transesterifying a triacylglycerol feedstock with an a...

  18. Integrins and small GTPases as modulators of phagocytosis.

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    Sayedyahossein, Samar; Dagnino, Lina

    2013-01-01

    Phagocytosis is the mechanism whereby cells engulf large particles. This process has long been recognized as a critical component of the innate immune response, which constitutes the organism's defense against microorganisms. In addition, phagocytic internalization of apoptotic cells or cell fragments plays important roles in tissue homeostasis and remodeling. Phagocytosis requires target interactions with receptors on the plasma membrane of the phagocytic cell. Integrins have been identified as important mediators of particle clearance, in addition to their well-established roles in cell adhesion, migration and mechanotransduction. Indeed, these ubiquitously expressed proteins impart phagocytic capacity to epithelial, endothelial and mesenchymal cell types. The importance of integrins in particle internalization is emphasized by the ability of microbial and viral pathogens to exploit their signaling pathways to invade host cells, and by the wide variety of disorders that arise from abnormalities in integrin-dependent phagocytic uptake. Copyright © 2013 Elsevier Inc. All rights reserved.

  19. Rapamycin-based inducible translocation systems for studying phagocytosis.

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    Bohdanowicz, Michal; Fairn, Gregory D

    2011-01-01

    Phagocytosis is an immune receptor-mediated process whereby cells engulf large particles. The process is dynamic and requires several localized factors acting in concert with and sequentially after the engagement of immune receptors to envelope the particle. Once the particle is internalized, the nascent -phagosome undergoes a series of events leading to its maturation to the microbicidal phagolysosome. Investigating these dynamic and temporally controlled series of events in live cells requires noninvasive methods. The ability to rapidly recruit the proteins of interest to the sites of phagocytosis or to nascent phagosomes would help dissect the regulatory mechanisms involved during phagocytosis. Here, we describe a general approach to express in RAW264.7 murine macrophages, a genetically encoded rapamycin--induced heterodimerization system. In the presence of rapamycin, tight association between FK506-binding protein (FKBP) and FKBP rapamycin-binding protein (FRB) is observed. Based on this principle, a synthetic system consisting of a targeting domain attached to FKBP can recruit a protein of interest fused to FRB upon the addition of rapamycin. Previously, this technique has been used to target lipid-modifying enzymes and small GTPases to the phagosome or plasma membrane. The recruitment of the FRB module can be monitored by fluorescent microscopy if a fluorescent protein is fused to the FRB sequence. While the focus of this chapter is on phagocytic events, this method can be employed to study any organelle of interest when the appropriate targeting sequence is used.

  20. DGAT enzymes and triacylglycerol biosynthesis

    Science.gov (United States)

    Yen, Chi-Liang Eric; Stone, Scot J.; Koliwad, Suneil; Harris, Charles; Farese, Robert V.

    2008-01-01

    Triacylglycerols (triglycerides) (TGs) are the major storage molecules of metabolic energy and FAs in most living organisms. Excessive accumulation of TGs, however, is associated with human diseases, such as obesity, diabetes mellitus, and steatohepatitis. The final and the only committed step in the biosynthesis of TGs is catalyzed by acyl-CoA:diacylglycerol acyltransferase (DGAT) enzymes. The genes encoding two DGAT enzymes, DGAT1 and DGAT2, were identified in the past decade, and the use of molecular tools, including mice deficient in either enzyme, has shed light on their functions. Although DGAT enzymes are involved in TG synthesis, they have distinct protein sequences and differ in their biochemical, cellular, and physiological functions. Both enzymes may be useful as therapeutic targets for diseases. Here we review the current knowledge of DGAT enzymes, focusing on new advances since the cloning of their genes, including possible roles in human health and diseases. PMID:18757836

  1. The digestion of dietary triacylglycerols

    DEFF Research Database (Denmark)

    Mu, Huiling; Høy, Carl-Erik

    2004-01-01

    Dietary triacylglycerols (TAGs) are the major lipid components in the human diet and they are carriers of energy as well as important fatty acids. Many factors affect the digestion and absorption of TAGs. Evidence is accumulating that, in addition to the overall fatty acid profile, the TAG......, or one may speculate additionally on the possibilities of modifying the structure of fats to affect their absorption and the distribution of the fatty acids in the body after digestion and uptake. In this review we will summarize diverse aspects of TAG digestion and absorption, as well as the influences...... of the fatty acid composition and the intramolecular structure of dietary TAGs on their digestion and absorption....

  2. Interaction between Mitochondrial Reactive Oxygen Species, Heme Oxygenase, and Nitric Oxide Synthase Stimulates Phagocytosis in Macrophages

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    Andrea Müllebner

    2018-01-01

    Full Text Available BackgroundMacrophages are cells of the innate immune system that populate every organ. They are required not only for defense against invading pathogens and tissue repair but also for maintenance of tissue homeostasis and iron homeostasis.AimThe aim of this study is to understand whether heme oxygenase (HO and nitric oxide synthase (NOS contribute to the regulation of nicotinamide adenine dinucleotide phosphate oxidase (NOX activity and phagocytosis, two key components of macrophage function.MethodsThis study was carried out using resting J774A.1 macrophages treated with hemin or vehicle. Activity of NOS, HO, or NOX was inhibited using specific inhibitors. Reactive oxygen species (ROS formation was determined by Amplex® red assay, and phagocytosis was measured using fluorescein isothiocyanate-labeled bacteria. In addition, we analyzed the fate of the intracellular heme by using electron spin resonance.ResultsWe show that both enzymes NOS and HO are essential for phagocytic activity of macrophages. NOS does not directly affect phagocytosis, but stimulates NOX activity via nitric oxide-triggered ROS production of mitochondria. Treatment of macrophages with hemin results in intracellular accumulation of ferrous heme and an inhibition of phagocytosis. In contrast to NOS, HO products, including carbon monoxide, neither clearly affect NOX activity nor clearly affect phagocytosis, but phagocytosis is accelerated by HO-mediated degradation of heme.ConclusionBoth enzymes contribute to the bactericidal activity of macrophages independently, by controlling different pathways.

  3. Phagocytosis in phosphate chromium (III) suspensions

    International Nuclear Information System (INIS)

    Cruz-Arencibia, Jorge; Fano Machín, Yoiz; Cruz-Morales, Ahmed; Tamayo Fuente, Radamés; Morín-Zorrilla, José

    2015-01-01

    Phagocytosis in vivo and in vitro of a suspension of chromic phosphate (III) labeled with 51 Cr and 32 P is studied. The radioactive particles dispersed in a media of 2 % gelatin in acetate buffer pH 4-4.5 have a predominant size of 0.8 μm and 5 μm. According with biodistribution experiments in rats after 30 minutes near the 80 % of radioactivity is registered in the liver, probably associated with phagocytosis of the particles by liver Kupffer cells. Is also showed that the suspension particles are phagocytized in vitro by mouse peritoneal macrophages. This facts indicate that the studied suspension have appropriate characteristics to be used in radiosynoviorthesis according to the principal action mechanism described for this procedure, particles phagocytosis by cells present in the inflamed synovium. (author)

  4. Autophagy and phagocytosis converge for better vision.

    Science.gov (United States)

    Ferguson, Thomas A; Green, Douglas R

    2014-01-01

    The retinal pigment epithelium (RPE) is a single layer of nonregenerating cells essential to homeostasis in the retina and the preservation of vision. While the RPE perform a number of important functions, 2 essential processes are phagocytosis, which removes the most distal tips of the photoreceptors to support disk renewal, and the visual cycle, which maintains the supply of chromophore for regeneration of photo-bleached visual pigments. We recently reported that these processes are linked by a noncanonical form of autophagy termed LC3-associated phagocytosis (LAP) in which components of the autophagy pathway are co-opted by phagocytosis to recover vitamin A in support of optimal vision. Here we summarize these findings.

  5. Determination of phagocytosis in periodontitis postjuvenilis using 14C

    International Nuclear Information System (INIS)

    Zietek, M.

    1990-01-01

    In 20 patients with periodontitis and 10 healthy controls phagocytosis was determined using 14 C-labelled Staphylococcus aureus. The decrease of phagocytosis found in the diseased patients was significant

  6. Rat Neutrophil Phagocytosis Following Feed Restriction

    Czech Academy of Sciences Publication Activity Database

    Slapničková, Martina; Berger, J.

    2002-01-01

    Roč. 11, č. 3 (2002), s. 172-177 ISSN 0938-7714 Institutional research plan: CEZ:AV0Z5052915 Keywords : circulating neutrophil * diet restriction * phagocytosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 0.167, year: 2001

  7. Dietary carbohydrates and triacylglycerol metabolism.

    Science.gov (United States)

    Roche, H M

    1999-02-01

    There is a growing body of scientific evidence which demonstrates that plasma triacylglycerol (TAG) concentration, especially in the postprandial state, is an important risk factor in relation to the development of CHD. Postprandial hypertriacylglycerolaemia is associated with a number of adverse metabolic risk factors, including the preponderance of small dense LDL, low HDL-cholesterol concentrations and elevated factor VII activity. Traditionally, a low-fat high-carbohydrate diet was used to prevent CHD because it effectively reduces plasma cholesterol concentrations, but this dietary regimen increases plasma TAG concentrations and reduces HDL-cholesterol concentrations. There is substantial epidemiological evidence which demonstrates that high plasma TAG and low plasma HDL concentrations are associated with an increased risk of CHD. Thus, there is reason for concern that the adverse effects of low-fat high-carbohydrate diets on TAG and HDL may counteract or negate the beneficial effect of reducing LDL-cholesterol concentrations. Although there have been no prospective studies to investigate whether reduced fat intake has an adverse effect on CHD, there is strong epidemiological evidence that reducing total fat intake is not protective against CHD. On the other hand, high-fat diets predispose to obesity, and central obesity adversely affects TAG metabolism. There is substantial evidence that in free-living situations low-fat high-carbohydrate diets lead to weight loss, which in turn will correct insulin resistance and plasma TAG metabolism. Clearly there is a need for prospective studies to resolve the issue as to whether low-fat high-carbohydrate diets play an adverse or beneficial role in relation to the development of CHD.

  8. Regioisomeric and enantiomeric analysis of triacylglycerols

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Pádrová, K.; Sigler, Karel

    2017-01-01

    Roč. 524, SI (2017), s. 3-12 ISSN 0003-2697 R&D Projects: GA ČR GA14-00227S Institutional support: RVO:61388971 Keywords : Regioisomers * Enantiomers * Triacylglycerols Subject RIV: EE - Microbiology, Virology OBOR OECD: Microbiology Impact factor: 2.334, year: 2016

  9. Modifications of the metabolic pathways of lipid and triacylglycerol production in microalgae

    Directory of Open Access Journals (Sweden)

    Yu Wei-Luen

    2011-11-01

    Full Text Available Abstract Microalgae have presented themselves as a strong candidate to replace diminishing oil reserves as a source of lipids for biofuels. Here we describe successful modifications of terrestrial plant lipid content which increase overall lipid production or shift the balance of lipid production towards lipid varieties more useful for biofuel production. Our discussion ranges from the biosynthetic pathways and rate limiting steps of triacylglycerol formation to enzymes required for the formation of triacylglycerol containing exotic lipids. Secondarily, we discuss techniques for genetic engineering and modification of various microalgae which can be combined with insights gained from research in higher plants to aid in the creation of production strains of microalgae.

  10. Phagocytosis in Teleosts. Implications of the New Cells Involved

    Directory of Open Access Journals (Sweden)

    María Ángeles Esteban

    2015-12-01

    Full Text Available Phagocytosis is the process by which cells engulf some solid particles to form internal vesicles known as phagosomes. Phagocytosis is in fact a specific form of endocytosis involving the vesicular interiorization of particles. Phagocytosis is essentially a defensive reaction against infection and invasion of the body by foreign substances and, in the immune system, phagocytosis is a major mechanism used to remove pathogens and/or cell debris. For these reasons, phagocytosis in vertebrates has been recognized as a critical component of the innate and adaptive immune responses to pathogens. Furthermore, more recent studies have revealed that phagocytosis is also crucial for tissue homeostasis and remodeling. Professional phagocytes in teleosts are monocyte/macrophages, granulocytes and dendritic cells. Nevertheless, in recent years phagocytic properties have also been attributed to teleost lymphocytes and thrombocytes. The possible implications of such cells on this important biological process, new factors affecting phagocytosis, evasion of phagocytosis or new forms of phagocytosis will be considered and discussed.

  11. [Update views on the theory of phagocytosis].

    Science.gov (United States)

    Freĭdlin, I S

    2008-01-01

    Developer of the phagocytosis theory I.I Mechnikov forecasted the most fruitful directions of its development. Macrophages express on the plasma membranes broad spectrum of receptors, which mediate their interaction with altered organism's own components as well as with exogenous agents, including various microorganisms. Recognition leads to changes of expression of surface molecules, enhancement of phagocytic activity as well as production and secretion of cytokines, presentation functions, signaling and genes expression. This reflected on maintenance of homeostasis, as well as on host defense effectiveness, including mechanisms of innate and adaptive immunity.

  12. Essential role of integrin-linked kinase in regulation of phagocytosis in keratinocytes.

    Science.gov (United States)

    Sayedyahossein, Samar; Nini, Lylia; Irvine, Timothy S; Dagnino, Lina

    2012-10-01

    Phagocytic melanosome uptake by epidermal keratinocytes is a central protective mechanism against damage induced by ultraviolet radiation. Phagocytosis requires formation of pseudopodia via actin cytoskeleton rearrangements. Integrin-linked kinase (ILK) is an important modulator of actin cytoskeletal dynamics. We have examined the role of ILK in regulation of phagocytosis, using epidermal keratinocytes isolated from mice with epidermis-restricted Ilk gene inactivation. ILK-deficient cells exhibited severely impaired capacity to engulf fluorescent microspheres in response to stimulation of the keratinocyte growth factor (KGF) receptor or the protease-activated receptor-2. KGF induced ERK phosphorylation in ILK-expressing and ILK-deficient cells, suggesting that ILK is not essential for KGF receptor signaling. In contrast, KGF promoted activation of Rac1 and formation of pseudopodia in ILK-expressing, but not in ILK-deficient cells. Rac1-deficient keratinocytes also showed substantially impaired phagocytic ability, underlining the importance of ILK-dependent Rac1 function for particle engulfment. Finally, cross-modulation of KGF receptors by integrins may be another important element, as integrin β1-deficient keratinocytes also fail to show significant phagocytosis in response to KGF. Thus, we have identified a novel signaling pathway essential for phagocytosis in keratinocytes, which involves ILK-dependent activation of Rac1 in response to KGF, resulting in the formation of pseudopodia and particle uptake.

  13. Air-drying of cells, the novel conditions for stimulated synthesis of triacylglycerol in a Green Alga, Chlorella kessleri.

    Directory of Open Access Journals (Sweden)

    Takuma Shiratake

    Full Text Available Triacylglycerol is used for the production of commodities including food oils and biodiesel fuel. Microalgae can accumulate triacylglycerol under adverse environmental conditions such as nitrogen-starvation. This study explored the possibility of air-drying of green algal cells as a novel and simple protocol for enhancement of their triacylglycerol content. Chlorella kessleri cells were fixed on the surface of a glass fibre filter and then subjected to air-drying with light illumination. The dry cell weight, on a filter, increased by 2.7-fold in 96 h, the corresponding chlorophyll content ranging from 1.0 to 1.3-fold the initial one. Concomitantly, the triacylglycerol content remarkably increased to 70.3 mole% of fatty acids and 15.9% (w/w, relative to total fatty acids and dry cell weight, respectively, like in cells starved of nitrogen. Reduction of the stress of air-drying by placing the glass filter on a filter paper soaked in H2O lowered the fatty acid content of triacylglycerol to 26.4 mole% as to total fatty acids. Moreover, replacement of the H2O with culture medium further decreased the fatty acid content of triacylglycerol to 12.2 mole%. It thus seemed that severe dehydration is required for full induction of triacylglycerol synthesis, and that nutritional depletion as well as dehydration are crucial environmental factors. Meanwhile, air-drying of Chlamydomonas reinhardtii cells increased the triacylglycerol content to only 37.9 mole% of fatty acids and 4.8% (w/w, relative to total fatty acids and dry cell weight, respectively, and a marked decrease in the chlorophyll content, on a filter, of 33%. Air-drying thus has an impact on triacylglycerol synthesis in C. reinhardtii also, however, the effect is considerably limited, owing probably to instability of the photosynthetic machinery. This air-drying protocol could be useful for the development of a system for industrial production of triacylglycerol with appropriate selection of the

  14. Desaturation of excess intramyocellular triacylglycerol in obesity

    DEFF Research Database (Denmark)

    Haugaard, S B; Madsbad, S; Mu, Huiling

    2010-01-01

    , however, was increased twofold in obese women compared to obese men (Pfasting glucose (P...OBJECTIVE: Excess intramyocellular triacylglycerol (IMTG), found especially in obese women, is slowly metabolized and, therefore, prone to longer exposure to intracellular desaturases. Accordingly, it was hypothesized that IMTG content correlates inversely with IMTG fatty acid (FA) saturation...... in sedentary subjects. In addition, it was validated if IMTG palmitic acid is associated with insulin resistance as suggested earlier. DESIGN: Cross-sectional human study. SUBJECTS: In skeletal muscle biopsies, which were obtained from sedentary subjects (34 women, age 48+/-2 years (27 obese including 7 type 2...

  15. Changes in the triacylglycerol content of flaxseeds during ...

    African Journals Online (AJOL)

    Changes in the composition of triacylglycerols (TAGs) of three varieties of flaxseeds (H52, O116 and P129) during development were investigated using non-aqueous reversed phase liquid chromatography coupled to atmospheric pressure photoionization-mass spectrometry (LC-APPI-MS). 17 species of triacylglycerols ...

  16. Cortactin and phagocytosis in isolated Sertoli cells

    Directory of Open Access Journals (Sweden)

    Wolski Katja M

    2005-12-01

    Full Text Available Abstract Background Cortactin, an actin binding protein, has been associated with Sertoli cell ectoplasmic specializations in vivo, based on its immunolocalization around the heads of elongated spermatids, but not previously identified in isolated Sertoli cells. In an in vitro model of Sertoli cell-spermatid binding, cortactin was identified around debris and dead germ cells. Based on this observation, we hypothesized that this actin binding protein may be associated with a non-junction-related physiological function, such as phagocytosis. The purpose of this study was to identify the presence and distribution of cortactin in isolated rat Sertoli cells active in phagocytic activity following the addition of 0.8 μm latex beads. Results Sertoli cell monocultures were incubated with or without follicle stimulating hormone (FSH; 0.1 μg/ml in the presence or absence of cytochalasin D (2 μM, as an actin disrupter. Cortactin was identified by standard immunostaining with anti-cortactin, clone 4F11 (Upstate after incubation times of 15 min, 2 hr, and 24 hr with or without beads. Cells exposed to no hormone and no beads appeared to have a ubiquitous distribution of cortactin throughout the cytoplasm. In the presence of cytochalasin D, cortactin immunostaining was punctate and distributed in a pattern similar to that reported for actin in cells exposed to cytochalasin D. Sertoli cells not exposed to FSH, but activated with beads, did not show cortactin immunostaining around the phagocytized beads at any of the time periods. FSH exposure did not alter the distribution of cortactin within Sertoli cells, even when phagocytic activity was upregulated by the presence of beads. Conclusion Results of this study suggest cortactin is not associated with peripheralized actin at junctional or phagocytic sites. Further studies are necessary to clarify the role of cortactin in Sertoli cells.

  17. Effects of microparticle size and Fc density on macrophage phagocytosis.

    Directory of Open Access Journals (Sweden)

    Patricia Pacheco

    Full Text Available Controlled induction of phagocytosis in macrophages offers the ability to therapeutically regulate the immune system as well as improve delivery of chemicals or biologicals for immune processing. Maximizing particle uptake by macrophages through Fc receptor-mediated phagocytosis could lead to new delivery mechanisms in drug or vaccine development. Fc ligand density and particle size were examined independently and in combination in order to optimize and tune the phagocytosis of opsonized microparticles. We show the internalization efficiency of small polystyrene particles (0.5 µm to 2 µm is significantly affected by changes in Fc ligand density, while particles greater than 2 µm show little correlation between internalization and Fc density. We found that while macrophages can efficiently phagocytose a large number of smaller particles, the total volume of phagocytosed particles is maximized through the non-specific uptake of larger microparticles. Therefore, larger microparticles may be more efficient at delivering a greater therapeutic payload to macrophages, but smaller opsonized microparticles can deliver bio-active substances to a greater percentage of the macrophage population. This study is the first to treat as independent variables the physical and biological properties of Fc density and microparticle size that initiate macrophage phagocytosis. Defining the physical and biological parameters that affect phagocytosis efficiency will lead to improved methods of microparticle delivery to macrophages.

  18. Aliphatic alcohols in spirits inhibit phagocytosis by human monocytes.

    Science.gov (United States)

    Pál, László; Árnyas, Ervin M; Bujdosó, Orsolya; Baranyi, Gergő; Rácz, Gábor; Ádány, Róza; McKee, Martin; Szűcs, Sándor

    2015-04-01

    A large volume of alcoholic beverages containing aliphatic alcohols is consumed worldwide. Previous studies have confirmed the presence of ethanol-induced immunosuppression in heavy drinkers, thereby increasing susceptibility to infectious diseases. However, the aliphatic alcohols contained in alcoholic beverages might also impair immune cell function, thereby contributing to a further decrease in microbicidal activity. Previous research has shown that aliphatic alcohols inhibit phagocytosis by granulocytes but their effect on human monocytes has not been studied. This is important as they play a crucial role in engulfment and killing of pathogenic microorganisms and a decrease in their phagocytic activity could lead to impaired antimicrobial defence in heavy drinkers. The aim of this study was to measure monocyte phagocytosis following their treatment with those aliphatic alcohols detected in alcoholic beverages. Monocytes were separated from human peripheral blood and phagocytosis of opsonized zymosan particles by monocytes treated with ethanol and aliphatic alcohols individually and in combination was determined. It was shown that these alcohols could suppress the phagocytic activity of monocytes in a concentration-dependent manner and when combined with ethanol, they caused a further decrease in phagocytosis. Due to their additive effects, it is possible that they may inhibit phagocytosis in a clinically meaningful way in alcoholics and episodic heavy drinkers thereby contribute to their increased susceptibility to infectious diseases. However, further research is needed to address this question.

  19. Triacylglycerol and melting profiles of milk fat from several species

    NARCIS (Netherlands)

    Smiddy, M.A.; Huppertz, T.; Ruth, van S.M.

    2012-01-01

    Gas chromatography and differential scanning calorimetry (DSC) were used to differentiate the fats of cow, goat, sheep, water buffalo, donkey, horse and camel milk (n = 20 for each species). Principal component analysis of triacylglycerol (TAG) composition allowed classification into groups

  20. Progranulin increases phagocytosis by retinal pigment epithelial cells in culture.

    Science.gov (United States)

    Murase, Hiromi; Tsuruma, Kazuhiro; Kuse, Yoshiki; Shimazawa, Masamitsu; Hara, Hideaki

    2017-12-01

    Retinal pigment epithelium (RPE) cells take part in retinal preservation, such as phagocytizing the shed photoreceptor outer segments (POS), every day. The incomplete phagocytic function accelerates RPE degeneration and formation of the toxic by-product lipofuscin. Excessive lipofuscin accumulation is characteristic of various blinding diseases in the human eye. Progranulin is a cysteine-rich protein that has multiple biological activities, and it has a high presence in the retina. Progranulin has been recognized to be involved in macrophage phagocytosis in the brain. The purpose of this study is to determine whether progranulin influences phagocytosis by RPE cells. All experiments were performed on primary human RPE (hRPE) cells in culture. pHrodo was used to label the isolated porcine POS, and quantification of pHrodo fluorescence was used to determine the degree of phagocytosis. Western blotting and immunohistochemistry of key proteins involved in phagocytosis were used to clarify the mechanism of progranulin. Progranulin increased RPE phagocytosis in hydrogen peroxide-treated and nontreated RPE cells. The phosphorylated form of Mer tyrosine kinase, which is important for POS internalization, was significantly increased in the progranulin-exposed cells. This increase was attenuated by SU11274, an inhibitor of hepatic growth factor receptor. Under the oxidative stress condition, exposure to progranulin led to an approximately twofold increase in integrin alpha-v, which is associated with the first step in recognition of POS by RPE cells. These results suggest that progranulin could be an effective stimulator for RPE phagocytosis and could repair RPE function. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  1. Phagocytosis escape by a Staphylococcus aureus protein that connects complement and coagulation proteins at the bacterial surface.

    Directory of Open Access Journals (Sweden)

    Ya-Ping Ko

    Full Text Available Upon contact with human plasma, bacteria are rapidly recognized by the complement system that labels their surface for uptake and clearance by phagocytic cells. Staphylococcus aureus secretes the 16 kD Extracellular fibrinogen binding protein (Efb that binds two different plasma proteins using separate domains: the Efb N-terminus binds to fibrinogen, while the C-terminus binds complement C3. In this study, we show that Efb blocks phagocytosis of S. aureus by human neutrophils. In vitro, we demonstrate that Efb blocks phagocytosis in plasma and in human whole blood. Using a mouse peritonitis model we show that Efb effectively blocks phagocytosis in vivo, either as a purified protein or when produced endogenously by S. aureus. Mutational analysis revealed that Efb requires both its fibrinogen and complement binding residues for phagocytic escape. Using confocal and transmission electron microscopy we show that Efb attracts fibrinogen to the surface of complement-labeled S. aureus generating a 'capsule'-like shield. This thick layer of fibrinogen shields both surface-bound C3b and antibodies from recognition by phagocytic receptors. This information is critical for future vaccination attempts, since opsonizing antibodies may not function in the presence of Efb. Altogether we discover that Efb from S. aureus uniquely escapes phagocytosis by forming a bridge between a complement and coagulation protein.

  2. Bovine-associated CNS species resist phagocytosis differently

    Science.gov (United States)

    2013-01-01

    Background Coagulase-negative staphylococci (CNS) cause usually subclinical or mild clinical bovine mastitis, which often remains persistent. Symptoms are usually mild, mostly only comprising slight changes in the appearance of milk and possibly slight swelling. However, clinical mastitis with severe signs has also been reported. The reasons for the differences in clinical expression are largely unknown. Macrophages play an important role in the innate immunity of the udder. This study examined phagocytosis and killing by mouse macrophage cells of three CNS species: Staphylococcus chromogenes (15 isolates), Staphylococcus agnetis (6 isolates) and Staphylococcus simulans (15 isolates). Staphylococcus aureus (7 isolates) was also included as a control. Results All the studied CNS species were phagocytosed by macrophages, but S. simulans resisted phagocytosis more effectively than the other CNS species. Only S. chromogenes was substantially killed by macrophages. Significant variations between isolates were seen in both phagocytosis and killing by macrophages and were more common in the killing assays. Significant differences between single CNS species and S. aureus were observed in both assays. Conclusion This study demonstrated that differences in the phagocytosis and killing of mastitis-causing staphylococci by macrophages exist at both the species and isolate level. PMID:24207012

  3. Multiple sclerosis : Mechanisms of myelin phagocytosis and lesion expansion

    NARCIS (Netherlands)

    Hendrickx, D.A.E.

    2018-01-01

    Multiple sclerosis (MS) is characterized by immune activation and focal demyelination in the central nervous system. The aim of this thesis was to gain more insight into the mechanisms of myelin phagocytosis by resident microglia and infiltrating macrophages. We first evaluated the expression of the

  4. DMPD: Complement-mediated phagocytosis--the role of Syk. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 16754322 Complement-mediated phagocytosis--the role of Syk. Tohyama Y, Yamamura H. ...IUBMB Life. 2006 May-Jun;58(5-6):304-8. (.png) (.svg) (.html) (.csml) Show Complement-mediated phagocytosis-...-the role of Syk. PubmedID 16754322 Title Complement-mediated phagocytosis--the role of Syk. Authors Tohyama

  5. Complement receptor-3 negatively regulates the phagocytosis of degenerated myelin through tyrosine kinase Syk and cofilin

    Directory of Open Access Journals (Sweden)

    Hadas Smadar

    2012-07-01

    Full Text Available Abstract Background Intact myelin, which normally surrounds axons, breaks down in Wallerian degeneration following axonal injury and during neurodegenerative diseases such as multiple sclerosis. Clearance of degenerated myelin by phagocytosis is essential since myelin impedes repair and exacerbates damage. CR3 (complement receptor-3 is a principal phagocytic receptor in myelin phagocytosis. We studied how tyrosine kinase Syk (spleen tyrosine kinase and cofilin control phagocytosis of degenerated myelin by CR3 in microglia and macrophages. Syk is a non-receptor tyrosine kinase that CR3 recruits to convey cellular functions. Cofilin is an actin-depolymerizing protein that controls F-actin (filamentous actin remodeling (i.e., disassembly and reassembly by shifting between active unphosphorylated and inactive phosphorylated states. Results Syk was continuously activated during prolonged phagocytosis. Phagocytosis increased when Syk activity and expression were reduced, suggesting that normally Syk down regulates CR3-mediated myelin phagocytosis. Levels of inactive p-cofilin (phosphorylated cofilin decreased transiently during prolonged phagocytosis. In contrast, p-cofilin levels decreased continuously when Syk activity and expression were continuously reduced, suggesting that normally Syk advances the inactive state of cofilin. Observations also revealed inverse relationships between levels of phagocytosis and levels of inactive p-cofilin, suggesting that active unphosphorylated cofilin advances phagocytosis. Active cofilin could advance phagocytosis by promoting F-actin remodeling, which supports the production of membrane protrusions (e.g., filopodia, which, as we also revealed, are instrumental in myelin phagocytosis. Conclusions CR3 both activates and downregulates myelin phagocytosis at the same time. Activation was previously documented. We presently demonstrate that downregulation is mediated through Syk, which advances the inactive

  6. Phagocytosis-induced /sup 45/calcium efflux in polymorphonuclear leucocytes

    Energy Technology Data Exchange (ETDEWEB)

    Barthelemy, A; Schell-Frederick, E [Brussels Univ. (Belgium). Institut de Recherche Interdisciplinaire; Paridaens, R [Brussels Univ. (Belgium). Faculte de Medicine

    1977-10-15

    The role of calcium ions in regulating the structure and function of non-muscle cells is a subject of intense study. Several lines of evidence that calcium may be essential in the function of polymorphonuclear leuocytes (PMNL) and an important control element in the process of phagocytosis. Direct studies of calcium distribution and fluxes have only recently been undertaken. To our knowledge, no report of calcium movements during normal phagocytosis has been published. In the context of an overall study of calcium dynamics in the PMNL, we report here initial studies on /sup 45/Ca efflux in prelabelled guinea pig PMNL. The results demonstrate the energy-dependence of resting calcium efflux and an increased efflux upon addition of phagocytic particles which is not dependent on particle internalization.

  7. Lipopolysaccharide O-antigen prevents phagocytosis of Vibrio anguillarum by rainbow trout (Oncorhynchus mykiss skin epithelial cells.

    Directory of Open Access Journals (Sweden)

    Kristoffer Lindell

    Full Text Available Colonization of host tissues is a first step taken by many pathogens during the initial stages of infection. Despite the impact of bacterial disease on wild and farmed fish, only a few direct studies have characterized bacterial factors required for colonization of fish tissues. In this study, using live-cell and confocal microscopy, rainbow trout skin epithelial cells, the main structural component of the skin epidermis, were demonstrated to phagocytize bacteria. Mutant analyses showed that the fish pathogen Vibrio anguillarum required the lipopolysaccharide O-antigen to evade phagocytosis and that O-antigen transport required the putative wzm-wzt-wbhA operon, which encodes two ABC polysaccharide transporter proteins and a methyltransferase. Pretreatment of the epithelial cells with mannose prevented phagocytosis of V. anguillarum suggesting that a mannose receptor is involved in the uptake process. In addition, the O-antigen transport mutants could not colonize the skin but they did colonize the intestines of rainbow trout. The O-antigen polysaccharides were also shown to aid resistance to the antimicrobial factors, lysozyme and polymyxin B. In summary, rainbow trout skin epithelial cells play a role in the fish innate immunity by clearing bacteria from the skin epidermis. In defense, V. anguillarum utilizes O-antigen polysaccharides to evade phagocytosis by the epithelial cells allowing it to colonize rapidly fish skin tissues.

  8. Lipopolysaccharide O-Antigen Prevents Phagocytosis of Vibrio anguillarum by Rainbow Trout (Oncorhynchus mykiss) Skin Epithelial Cells

    Science.gov (United States)

    Lindell, Kristoffer; Fahlgren, Anna; Hjerde, Erik; Willassen, Nils-Peder; Fällman, Maria; Milton, Debra L.

    2012-01-01

    Colonization of host tissues is a first step taken by many pathogens during the initial stages of infection. Despite the impact of bacterial disease on wild and farmed fish, only a few direct studies have characterized bacterial factors required for colonization of fish tissues. In this study, using live-cell and confocal microscopy, rainbow trout skin epithelial cells, the main structural component of the skin epidermis, were demonstrated to phagocytize bacteria. Mutant analyses showed that the fish pathogen Vibrio anguillarum required the lipopolysaccharide O-antigen to evade phagocytosis and that O-antigen transport required the putative wzm-wzt-wbhA operon, which encodes two ABC polysaccharide transporter proteins and a methyltransferase. Pretreatment of the epithelial cells with mannose prevented phagocytosis of V. anguillarum suggesting that a mannose receptor is involved in the uptake process. In addition, the O-antigen transport mutants could not colonize the skin but they did colonize the intestines of rainbow trout. The O-antigen polysaccharides were also shown to aid resistance to the antimicrobial factors, lysozyme and polymyxin B. In summary, rainbow trout skin epithelial cells play a role in the fish innate immunity by clearing bacteria from the skin epidermis. In defense, V. anguillarum utilizes O-antigen polysaccharides to evade phagocytosis by the epithelial cells allowing it to colonize rapidly fish skin tissues. PMID:22662189

  9. Modulation of hepatic reticuloendothelial system phagocytosis by pancreatic hormones.

    Science.gov (United States)

    Cornell, R P; McClellan, C C

    1982-12-01

    Experiments were conducted to determine the influence of the pancreatic hormones insulin, glucagon, and somatostatin on reticuloendothelial system (RES) phagocytosis both in vivo and in the isolated perfused livers of rats. Chronic pancreatic hormonal treatment consisted of twice daily injections SC of NPH insulin with doses ranging from 0.75 U on day 1 to 9.0 U on day 13 and unchanged doses of glucagon (200 micrograms) and somatostatin (50 micrograms). Chronic treatment with insulin significantly depressed by 48% intravascular phagocytosis of colloidal carbon administered IV at a dose of 8 mg/100 g, while glucagon and somatostatin stimulated macrophage endocytic function by 32% and 26%, respectively, compared to the control value. Acute treatment with the three pancreatic hormones at 30 min prior to carbon administration similarly produced insulin depression as well as glucagon and somatostatin stimulation of RES phagocytosis. Addition of the three hormones at near physiologic concentrations (20 ng/ml for insulin, 10 ng/ml for glucagon, and 5 ng/ml for somatostatin) to the recirculating perfusate of isolated perfused rat livers simultaneous with 24 mg of colloidal carbon likewise resulted in phagocytic reduction after insulin and enhancement after glucagon and somatostatin. Experiments involving insulin in vitro with isolated perfused livers as well as glucose replacement therapy concomitant with insulin in vivo demonstrated that hypoglycemia is not necessary for phagocytic depression by insulin while severe hypoglycemia in the perfusion medium is sufficient to depress carbon uptake by isolated perfused livers independent of insulin. Both pancreatic hormones and the level of glycemia seem to be important in modulating hepatic reticuloendothelial system phagocytosis.

  10. How cells engulf: a review of theoretical approaches to phagocytosis

    Science.gov (United States)

    Richards, David M.; Endres, Robert G.

    2017-12-01

    Phagocytosis is a fascinating process whereby a cell surrounds and engulfs particles such as bacteria and dead cells. This is crucial both for single-cell organisms (as a way of acquiring nutrients) and as part of the immune system (to destroy foreign invaders). This whole process is hugely complex and involves multiple coordinated events such as membrane remodelling, receptor motion, cytoskeleton reorganisation and intracellular signalling. Because of this, phagocytosis is an excellent system for theoretical study, benefiting from biophysical approaches combined with mathematical modelling. Here, we review these theoretical approaches and discuss the recent mathematical and computational models, including models based on receptors, models focusing on the forces involved, and models employing energetic considerations. Along the way, we highlight a beautiful connection to the physics of phase transitions, consider the role of stochasticity, and examine links between phagocytosis and other types of endocytosis. We cover the recently discovered multistage nature of phagocytosis, showing that the size of the phagocytic cup grows in distinct stages, with an initial slow stage followed by a much quicker second stage starting around half engulfment. We also address the issue of target shape dependence, which is relevant to both pathogen infection and drug delivery, covering both one-dimensional and two-dimensional results. Throughout, we pay particular attention to recent experimental techniques that continue to inform the theoretical studies and provide a means to test model predictions. Finally, we discuss population models, connections to other biological processes, and how physics and modelling will continue to play a key role in future work in this area.

  11. Some Observations on Carbon Nano tubes Susceptibility to Cell Phagocytosis

    International Nuclear Information System (INIS)

    Fraczek-Szczypta, A.; Menaszek, E.; Blazewicz, S.; Menaszek, E.

    2011-01-01

    The aim of this study was to assess the influence of different types of carbon nano tubes (CNTs) on cell phagocytosis. Three kinds of carbon nano tubes: single-walled carbon nano horns (SWCNHs), multi walled carbon nano tubes (MWCNTs), and ultra-long single-walled carbon nano tubes (ULSWCNTs) before and after additional chemical functionalization were seeded with macrophage cell culture. Prior to biological testing, the CNTs were subjected to dispersion process with the use of phosphate buffered solution (PBS) and PBS containing surfactant (Tween 20) or dimethyl sulfoxide (DMSO). The results indicate that the cells interaction with an individual nano tube is entirely different as compared to CNTs in the form of aggregate. The presence of the surfactant favors the CNTs dispersion in culture media and facilitates phagocytosis process, while it has disadvantageous influence on cells morphology. The cells phagocytosis is a more effective for MWCNTs and SWCNHs after their chemical functionalization. Moreover, these nano tubes were well dispersed in culture media without using DMSO or surfactant. The functionalized carbon nano tubes were easily dispersed in pure PBS and seeded with cells

  12. Characterization of myosin light chain in shrimp hemocytic phagocytosis.

    Science.gov (United States)

    Han, Fang; Wang, Zhiyong; Wang, Xiaoqing

    2010-11-01

    Myosin light chain, a well-known cytoskeleton gene, regulates multiple processes that are involved in material transport, muscle shrink and cell division. However, its function in phagocytosis against invading pathogens in crustacean remains unknown. In this investigation, a myosin light chain gene was obtained from Marsupenaeus japonicus shrimp. The full-length cDNA of this gene was of 766 bp and an open reading frame (ORF) of 462 bp encoding a polypeptide of 153 amino acids. The myosin light chain protein was expressed in Escherichia coli and purified. Subsequently the specific antibody was raised using the purified GST fusion protein. As revealed by immuno-electron microscopy, the myosin light chain protein was only expressed in the dark bands of muscle. In the present study, the myosin light chain gene was up-regulated in the WSSV-resistant shrimp as revealed by real-time PCR and western blot. And the phagocytic percentage and phagocytic index using FITC-labeled Vibrio parahemolyticus were remarkably increased in the WSSV-resistant shrimp, suggesting that the myosin light chain protein was essential in hemocytic phagocytosis. On the other hand, RNAi assays indicated that the phagocytic percentage and phagocytic index were significantly decreased when the myosin light chain gene was silenced by sequence-specific siRNA. These findings suggested that myosin light chain protein was involved in the regulation of hemocytic phagocytosis of shrimp. Copyright 2010 Elsevier Ltd. All rights reserved.

  13. Separation of Enantiomeric Triacylglycerols by Chiral-Phase

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Sigler, Karel

    2014-01-01

    Roč. 49, č. 2014 (2014), s. 1251-1260 ISSN 0024-4201 R&D Projects: GA ČR(CZ) GAP503/11/0215 Institutional support: RVO:61388971 Keywords : triacylglycerols * atmospheric pressure Subject RIV: EE - Microbiology, Virology Impact factor: 1.854, year: 2014

  14. High triacylglycerol turnover rate in human skeletal muscle

    DEFF Research Database (Denmark)

    Sacchetti, Massimo; Saltin, Bengt; Olsen, David B

    2004-01-01

    In the present study we investigated the relationship between plasma fatty acids (FA) and intramuscular triacylglycerol (IMTAG) kinetics of healthy volunteers. With this aim [U-(13)C]-palmitate was infused for 10 h and FA kinetics determined across the leg. In addition, the rate of FA incorporation...

  15. Identification of characteristic fatty acids to quantify triacylglycerols in microalgae

    Directory of Open Access Journals (Sweden)

    Peili eShen

    2016-02-01

    Full Text Available The fatty acid profiles of lipids from microalgae are unique. Polyunsaturated fatty acids are generally enriched in polar lipids, whereas saturated and monounsaturated fatty acids constitute the majority of fatty acids in triacylglycerols (TAG. Each species has characteristic fatty acids, and their content is positively or negatively correlated with TAGs. The marine oleaginous diatom Phaeodactylum tricornutum was used as the paradigm to determine the quantitative relationship between TAG and characteristic fatty acid content. Fatty acid profiles and TAG content of Phaeodactylum tricornutum were determined in a time course. C16:0/C16:1 and eicosapentaenoic acid (EPA, C20:5n3 were identified as characteristic fatty acids in TAGs and polar lipids, respectively. The percentage of those characteristic fatty acids in total fatty acids had a significant linear relationship with TAG content and thus the correlation coefficient presenting r2 were 0.96, 0.94 and 0.97 respectively. The fatty acid-based method for TAG quantification could also be applied to other microalgae such as Nannochloropsis oceanica in which the r2 of C16:0, EPA were 0.94, 0.97 respectively and Chlorella pyrenoidosa, whose r2 value correspondingly between C18:1, C18:3 and TAG content were 0.91, 0.99 as well. This characteristic fatty acid-based method provided a distinct way to quantify TAGs in microalgae, by which TAGs could be measured precisely by immediate transesterification from wet biomass rather than using conventional methods. This procedure simplified the operation and required smaller samples than conventional methods.

  16. Improved Triacylglycerol Production in Acinetobacter baylyi ADP1 by Metabolic Engineering

    Directory of Open Access Journals (Sweden)

    Karp Matti

    2011-05-01

    Full Text Available Abstract Background Triacylglycerols are used in various purposes including food applications, cosmetics, oleochemicals and biofuels. Currently the main sources for triacylglycerol are vegetable oils, and microbial triacylglycerol has been suggested as an alternative for these. Due to the low production rates and yields of microbial processes, the role of metabolic engineering has become more significant. As a robust model organism for genetic and metabolic studies, and for the natural capability to produce triacylglycerol, Acinetobacter baylyi ADP1 serves as an excellent organism for modelling the effects of metabolic engineering for energy molecule biosynthesis. Results Beneficial gene deletions regarding triacylglycerol production were screened by computational means exploiting the metabolic model of ADP1. Four deletions, acr1, poxB, dgkA, and a triacylglycerol lipase were chosen to be studied experimentally both separately and concurrently by constructing a knock-out strain (MT with three of the deletions. Improvements in triacylglycerol production were observed: the strain MT produced 5.6 fold more triacylglycerol (mg/g cell dry weight compared to the wild type strain, and the proportion of triacylglycerol in total lipids was increased by 8-fold. Conclusions In silico predictions of beneficial gene deletions were verified experimentally. The chosen single and multiple gene deletions affected beneficially the natural triacylglycerol metabolism of A. baylyi ADP1. This study demonstrates the importance of single gene deletions in triacylglycerol metabolism, and proposes Acinetobacter sp. ADP1 as a model system for bioenergetic studies regarding metabolic engineering.

  17. Collectin-11 Is an Important Modulator of Retinal Pigment Epithelial Cell Phagocytosis and Cytokine Production.

    Science.gov (United States)

    Dong, Xia; Wu, Weiju; Ma, Liang; Liu, Chengfei; Bhuckory, Mohajeet B; Wang, Liping; Nandrot, Emeline F; Xu, Heping; Li, Ke; Liu, Yizhi; Zhou, Wuding

    2017-01-01

    In this paper, we report previously unknown roles for collectin-11 (CL-11, a soluble C-type lectin) in modulating the retinal pigment epithelial (RPE) cell functions of phagocytosis and cytokine production. We found that CL-11 and its carbohydrate ligand are expressed in both the murine and human neural retina; these resemble each other in terms of RPE and photoreceptor cells. Functional analysis of murine RPE cells showed that CL-11 facilitates the opsonophagocytosis of photoreceptor outer segments and apoptotic cells, and also upregulates IL-10 production. Mechanistic analysis revealed that calreticulin on the RPE cells is required for CL-11-mediated opsonophagocytosis whereas signal-regulatory protein α and mannosyl residues on the cells are involved in the CL-11-mediated upregulation of IL-10 production. This study is the first to demonstrate the role of CL-11 and the molecular mechanisms involved in modulating RPE cell phagocytosis and cytokine production. It provides a new insight into retinal health and disease and has implications for other phagocytic cells. © 2017 S. Karger AG, Basel.

  18. The kinetics of phagocytosis of 198Au colloids ''in vitro''

    International Nuclear Information System (INIS)

    Astorri, N.L.; Bergoc, R.M.; Bianchin, A.M.; Caro, R.A.; Ihlo, J.E.; Rivera, E.S.

    1982-01-01

    The kinetics of the phagocytosis of 198-Au colloids by macrophages ''in vitro'' was studied by incubating during 5 hours phagocytic cells from the liver and the spleen of Wistar rats with colloidal radiogold particles, in the presence of an adequate culture medium (TC-199 with 10 per cent of Bovine Fetal Serum). In each experiment, the number of colloidal gold particles offered to each phatocytic cell, (Au) 0 and the mean rate of phagocytosis v, were calculated. The latter value was determined by measuring the radioactivity incorporated into the phagocytic cells during the incubation; it was expressed as the number of phagocytized colloidal gold particles per cell per minute. The values of log v = f [log (Au) 0 ] were plotted. The Lineweaver-Burk analysis of the results demonstrates that the kinetics of the phagocytosis of colloidal radiogold particles ''in vitro'' follows a model similar to Michaelis-Menten equations for enzyme reactions. The values of the substratum constant Ks and maximun velocity Vm were obtained by the regression analysis of the 1/v vs. 1/(Au) 0 graph. Vm was equal to 9.44 x 10 and 1.63 x 10 phagocytized colloidal gold particles per cell per minute for liver and spleen macrophages, respectively. Ks was equal to 6.01 x 10 9 and 8.02 x 10 8 colloidal gold particles per cell for liver and spleen macrophages, respectively. The significance of these differences is discussed and attributed mainly to a change of the specific engulfment rate constant. (author) [es

  19. Influence of UV-radiation on granulocytic phagocytosis in vitro

    International Nuclear Information System (INIS)

    Walther, T.; Rytter, M.; Gast, W.; Haustein, U.F.

    1987-01-01

    The influence of UV radiation on the vitality, the performance of phagocytosis and the ability to reduce nitro-blue tetrazolium test (NBT) by human granulocytes was investigated in vitro. Already by low doses of UVA (8% UVB) the percentage of phagocytizing granulocytes was decreased more distinctly than their cell vitality. The number of ingested Candida albicans particles was 4.5 particles per granulocyte in the controls. It was reduced to about 1.4 particles per cell by UV radiation independent of the dosis applied. On the other hand the ability of granulocytes to reduce NBT intracellularly remained completely unchanged. (author)

  20. Structured triacylglycerol containing behenic and oleic acids suppresses triacylglycerol absorption and prevents obesity in rats

    Directory of Open Access Journals (Sweden)

    Takamatsu Kiyoharu

    2010-07-01

    Full Text Available Abstract Background Dietary 1(3-behenoyl-2,3(1-dioleoyl-rac-glycerol (BOO has been reported to inhibit pancreatic lipase activity in vitro and suppress postprandial hypertriacylglycerolemia in humans. In the present study, the anti-obesity activities of BOO and its inhibitory effects on lymphatic triacylglycerol (TAG absorption were investigated in rats. Methods In Experiment 1, rats were fed either BOO or soybean oil (SO diet for 6 weeks. In the BOO diet, 20% of SO was replaced with an experimental oil rich in BOO. In Experiments 2 and 3, rats cannulated in the thoracic duct were administered an emulsions containing trioleoylglycerol (OOO or an oil mixture (OOO:BOO, 9:1. Tri[1-14C]oleoylglycerol (14C-OOO was added to the emulsions administered in Experiment 3. Results No observable differences were detected in food intake or body weight gain between the BOO and SO groups in Experiment 1. Plasma and liver TAG concentrations and visceral fat weights were significantly lower in the BOO group than in the SO group. The apparent absorption rate of fat was significantly lower in the BOO group than in the SO group. In Experiment 2, the lymphatic recovery of oleic and behenic acids was significantly lower at 5 and 6 h after BOO administration than after OOO administration. In Experiment 3, the lymphatic recovery of 14C-OOO was significantly lower at 5 and 6 h after BOO administration than after OOO administration. Conclusions These results suggest that BOO prevents deposition of visceral fat and hepatic TAG by lowering and delaying intestinal absorption of TAG.

  1. Fat-specific protein 27 regulates storage of triacylglycerol

    DEFF Research Database (Denmark)

    Keller, P.; Petrie, J.T.; Rose, P. De

    2008-01-01

    FSP27 (fat-specific protein 27) is a member of the cell death-inducing DNA fragmentation factor-alpha-like effector (CIDE) family. Although Cidea and Cideb were initially characterized as activators of apoptosis, recent studies have demonstrated important metabolic roles for these proteins...... in several cell types without induction of adipocyte genes. Increased triacylglycerol is likely due to decreased beta-oxidation of nonesterified fatty acids. Altered flux of fatty acids into triacylglycerol may be a direct effect of FSP27 function, which is localized to lipid droplets in 293T cells and 3T3-L...... decreases with total fat mass but is not associated with measures of insulin resistance (e.g. homeostasis model assessment). Together, these data indicate that FSP27 binds to lipid droplets and regulates their enlargement Udgivelsesdato: 2008/5/23...

  2. The Haemophilus ducreyi LspA1 protein inhibits phagocytosis by using a new mechanism involving activation of C-terminal Src kinase.

    Science.gov (United States)

    Dodd, Dana A; Worth, Randall G; Rosen, Michael K; Grinstein, Sergio; van Oers, Nicolai S C; Hansen, Eric J

    2014-05-20

    Haemophilus ducreyi causes chancroid, a sexually transmitted infection. A primary means by which this pathogen causes disease involves eluding phagocytosis; however, the molecular basis for this escape mechanism has been poorly understood. Here, we report that the LspA virulence factors of H. ducreyi inhibit phagocytosis by stimulating the catalytic activity of C-terminal Src kinase (Csk), which itself inhibits Src family protein tyrosine kinases (SFKs) that promote phagocytosis. Inhibitory activity could be localized to a 37-kDa domain (designated YL2) of the 456-kDa LspA1 protein. The YL2 domain impaired ingestion of IgG-opsonized targets and decreased levels of active SFKs when expressed in mammalian cells. YL2 contains tyrosine residues in two EPIYG motifs that are phosphorylated in mammalian cells. These tyrosine residues were essential for YL2-based inhibition of phagocytosis. Csk was identified as the predominant mammalian protein interacting with YL2, and a dominant-negative Csk rescued phagocytosis in the presence of YL2. Purified Csk phosphorylated the tyrosines in the YL2 EPIYG motifs. Phosphorylated YL2 increased Csk catalytic activity, resulting in positive feedback, such that YL2 can be phosphorylated by the same kinase that it activates. Finally, we found that the Helicobacter pylori CagA protein also inhibited phagocytosis in a Csk-dependent manner, raising the possibility that this may be a general mechanism among diverse bacteria. Harnessing Csk to subvert the Fcγ receptor (FcγR)-mediated phagocytic pathway represents a new bacterial mechanism for circumventing a crucial component of the innate immune response and may potentially affect other SFK-involved cellular pathways. Phagocytosis is a critical component of the immune system that enables pathogens to be contained and cleared. A number of bacterial pathogens have developed specific strategies to either physically evade phagocytosis or block the intracellular signaling required for

  3. Triacylglycerols in some underutilised tropical seed oils 1. Systematic studies of ten oils

    International Nuclear Information System (INIS)

    Adebowale, K.O.; Adebowale, Y.A.; Nicholson, G.

    2002-05-01

    Triacylglycerols composition of ten lesser known and underutilised tropical seed oils have been determined. The seed oils include Monodora tenuifolia, Monodora myristica, Colocynthis citrullus, Cyperus esculentus, Cucumeropsis edulis, Andenopus breviflorus, Telfairia occidentalis, Blighia sapida, Antiaris africana and Sesame indicum. In the Moreaceae family (M. tenuifolia, M. myristica) the dominant triacylglycerol are OPO/POO, LLO, OOL, and OOO. They accounted for over 60% of the total triacylglycerol content in the oils. In the Cyperaceae family (C. esculentus), OPP/POO, POL and OOO accounted for over 80% of the total triacylglycerol content. In the Cucurbitaceae family, SSP was the dominant triacylglycerol specie in A. breviflorus, while OPO/POO and OOO were the dominant species in C. citrullus and C. edulis. Blighia sapida recorded a different distribution of triacylglycerols composition. PLL occurred at the highest concentration, while other high molecular weight triacylglycerols were also identified in the oil. They include, SSA, OSA, LSA, OAA and LLA. Analysis of A. antiaris oil showed a different pattern in the distribution of the triacylglycerols. LaLaM, MMLa and LaLaLa accounted for about 80% of the total triacylglycerols. This result reflected the fatty acid composition of the oil. Lauric acid (C12:0) and Myristic acid (C14:0) accounted for 71.5% of the total fatty acid. The possible use of the oils as cocoa butter equivalents CBEs and cocoa butter substitutes CBSs are discussed. (author)

  4. In vitro phagocytosis of several Candida berkhout species by murine leukocytes.

    Science.gov (United States)

    Fontenla de Petrino, S E; Bibas Bonet de Jorrat, M E; Sirena, A

    1985-03-01

    In vitro phagocytosis of thirteen Candida berkhout species by rat leukocytes was studied to assess a possible correlation between pathogenicity and phagocytosis Yeast-leukocyte suspensions were mixed up for 3 h and phagocytic index, germ-tube formation and leukocyte candidacidal activity were evaluated. Highest values for phagocytosis were reached in all cases at the end of the first hour. Leukocyte candidacidal activity was absent. Only C. albicans produced germ-tubes. The various phagocytosis indices were determined depending on the Candida species assayed. Under these conditions, the more pathogenic species presented the lower indices of phagocytosis. It is determined that the in vitro phagocytic index may bear a close relationship with the pathogenicity of the Candida berkhout.

  5. Wip1-dependent modulation of macrophage migration and phagocytosis

    DEFF Research Database (Denmark)

    Tang, Yiting; Pan, Bing; Zhou, Xin

    2017-01-01

    Macrophage accumulation within the vascular wall is a hallmark of atherosclerosis. Controlling macrophage conversion into foam cells remains a major challenge for treatment of atherosclerotic diseases. Here, we show that Wip1, a member of the PP2C family of Ser/Thr protein phosphatases, modulates...... macrophage migration and phagocytosis associated with atherosclerotic plaque formation. Wip1 deficiency increases migratory and phagocytic activities of the macrophage under stress conditions. Enhanced migration of Wip1-/- macrophages is mediated by Rac1-GTPase and PI3K/AKT signalling pathways. Elevated...... phagocytic ability of Wip1-/- macrophages is linked to CD36 plasma membrane recruitment that is regulated by AMPK activity. Our study identifies Wip1 as an intrinsic negative regulator of macrophage chemotaxis. We propose that Wip1-dependent control of macrophage function may provide avenues for preventing...

  6. Buoyant triacylglycerol-filled green algae and methods therefor

    Science.gov (United States)

    Goodenough, Ursula; Goodson, Carrie

    2015-04-14

    Cultures of Chlamydomonas are disclosed comprising greater than 340 mg/l triacylglycerols (TAG). The cultures can include buoyant Chlamydomonas. Methods of forming the cultures are also disclosed. In some embodiments, these methods comprise providing Chlamydomonas growing in log phase in a first culture medium comprising a nitrogen source and acetate, replacing the first culture medium with a second medium comprising acetate but no nitrogen source, and subsequently supplementing the second medium with additional acetate. In some embodiments, a culture can comprise at least 1,300 mg/l triacyglycerols. In some embodiments, cultures can be used to produce a biofuel such as biodiesel.

  7. Induction of Live Cell Phagocytosis by a Specific Combination of Inflammatory Stimuli

    Directory of Open Access Journals (Sweden)

    Takamasa Ishidome

    2017-08-01

    Full Text Available Conditions of severe hyper-inflammation can lead to uncontrolled activation of macrophages, and the ensuing phagocytosis of live cells. However, relationships between inflammatory stimuli and uncontrolled phagocytosis of live cells by macrophages are poorly understood. To identify mediators of this process, we established phagocytosis assays of live cells by stimulating macrophages with CpG DNA, interferon-γ, and anti-interleukin-10 receptor antibody. In this model, various cell surface receptors were upregulated on macrophages, and phagocytosis of live cells was induced in a Rac1-dependent manner. Subsequent inhibition of the ICAM-1, VCAM-1, and both of these receptors abolished in vitro and in vivo phagocytosis of live T cells, myeloid cells, and B cells, respectively. Specifically, the reduction in lymphocyte numbers due to in vivo activation of macrophages was ameliorated in Icam-1-deficient mice. In addition, overexpression of ICAM-1 or VCAM-1 in non-phagocytic NIH3T3 cells led to active phagocytosis of live cells. These data indicate molecular mechanisms underlying live cell phagocytosis induced by hyper-inflammation, and this experimental model will be useful to clarify the pathophysiological mechanisms of hemophagocytosis and to indicate therapeutic targets.

  8. Phagocytosis of gram-negative bacteria by a unique CD14-dependent mechanism.

    Science.gov (United States)

    Schiff, D E; Kline, L; Soldau, K; Lee, J D; Pugin, J; Tobias, P S; Ulevitch, R J

    1997-12-01

    THP-1-derived cell lines were stably transfected with constructs encoding glycophosphatidylinositol (GPI)-anchored or transmembrane forms of human CD14. CD14 expression was associated with enhanced phagocytosis of serum (heat-inactivated)-opsonized Escherichia coli (opEc). Both the GPI-anchored and transmembrane forms of CD14 supported phagocytosis of opEc equally well. Lipopolysaccharide-binding protein (LBP) played a role in CD14-dependent phagocytosis as evidenced by inhibition of CD14-dependent phagocytosis of opEc with anti-LBP monoclonal antibody (mAb) and by enhanced phagocytosis of E. coli opsonized with purified LBP. CD14-dependent phagocytosis was inhibited by a phosphatidylinositol (PI) 3-kinase inhibitor (wortmannin) and a protein tyrosine kinase inhibitor (tyrphostin 23) but not a protein kinase C inhibitor (bisindolyl-maleimide) or a divalent cation chelator (ethylenediaminetetraacetate). Anti-LBP mAb 18G4 and anti-CD14 mAb 18E12 were used to differentiate between the pathways involved in CD14-dependent phagocytosis and CD14-dependent cell activation. F(ab')2 fragments of 18G4, a mAb to LBP that does not block cell activation, inhibited ingestion of opEc by THP1-wtCD14 cells. 18E12 (an anti-CD14 mAb that does not block LPS binding to CD14 but does inhibit CD14-dependent cell activation) did not inhibit phagocytosis of LBP-opEc by THP1-wtCD14 cells. Furthermore, CD14-dependent phagocytosis was not inhibited by anti-CD18 (CR3 and CR4 beta-chain) or anti-Fcgamma receptor mAb.

  9. A versatile, high through-put, bead-based phagocytosis assay for Plasmodium falciparum

    DEFF Research Database (Denmark)

    Lloyd, Yukie M.; Ngati, Elise P.; Salanti, Ali

    2017-01-01

    Antibody-mediated phagocytosis is an important immune effector mechanism against Plasmodium falciparum-infected erythrocytes (IE); however, current phagocytosis assays use IE collected from infected individuals or from in vitro cultures of P. falciparum, making them prone to high variation....... A simple, high-throughput flow cytometric assay was developed that uses THP-1 cells and fluorescent beads covalently-coupled with the malarial antigen VAR2CSA. The assay is highly repeatable, provides both the overall percent phagocytosis and semi-quantitates the number of antigen-coupled beads...

  10. EFFECTS OF PHYSICAL EXERCISES ON TRIACYLGLYCEROL LEVEL IN SKELETAL MUSCLES IN DIETARY-INDUCED OBESE RATS

    Directory of Open Access Journals (Sweden)

    I. Yu. Yakimovich

    2014-01-01

    Full Text Available The accumulation of triacylglycerol in peripheral tissues is one of mechanisms of insulin resistance. This paper presents the investigation of the influence of aerobic and anaerobic physical exercises on triacylglycerol level in skeletal muscles and on insulin resistance in dietary-induced obese rats. It is estimated that a high-energy (HE diet causes the accumulation of triacylglycerols in skeletal muscles that leads to high resistance to insulin. Aerobic and anaerobic physical exercises reduce the level of triacylglycerols in skeletal  muscles  and  raise  sensitivity to  insulin  in  obese  rats.  Physical  exercises  raise  the  level  of triacylglycerols in skeletal muscles in standard-diet rats that probably is the adaptation to high energy expenditure, but does not lead to high insulin resistance.

  11. Opsonic Phagocytosis in Chronic Obstructive Pulmonary Disease is Enhanced by Nrf2 Agonists.

    NARCIS (Netherlands)

    Bewley, Martin A; Budd, Richard C; Ryan, Eilise; Cole, Joby; Collini, Paul; Marshall, Jennifer; Kolsum, Umme; Beech, Gussie; Emes, Richard D; Tcherniaeva, Irina; Berbers, Guy A M; Walmsley, Sarah R; Donaldson, Gavin; Wedzicha, Jadwiga A; Kilty, Iain; Rumsey, William; Sanchez, Yolanda; Brightling, Christopher E; Donnelly, Louise E; Barnes, Peter J; Singh, Dave; Whyte, Moira K B; Dockrell, David H

    2018-01-01

    Previous studies have identified defects in bacterial phagocytosis by alveolar macrophages (AM) in patients with chronic obstructive pulmonary disease (COPD) but the mechanisms and clinical consequences remain incompletely defined.

  12. Overexpression of Enterococcus faecalis elr operon protects from phagocytosis.

    Science.gov (United States)

    Cortes-Perez, Naima G; Dumoulin, Romain; Gaubert, Stéphane; Lacoux, Caroline; Bugli, Francesca; Martin, Rebeca; Chat, Sophie; Piquand, Kevin; Meylheuc, Thierry; Langella, Philippe; Sanguinetti, Maurizio; Posteraro, Brunella; Rigottier-Gois, Lionel; Serror, Pascale

    2015-05-25

    Mechanisms underlying the transition from commensalism to virulence in Enterococcus faecalis are not fully understood. We previously identified the enterococcal leucine-rich protein A (ElrA) as a virulence factor of E. faecalis. The elrA gene is part of an operon that comprises four other ORFs encoding putative surface proteins of unknown function. In this work, we compared the susceptibility to phagocytosis of three E. faecalis strains, including a wild-type (WT), a ΔelrA strain, and a strain overexpressing the whole elr operon in order to understand the role of this operon in E. faecalis virulence. While both WT and ΔelrA strains were efficiently phagocytized by RAW 264.7 mouse macrophages, the elr operon-overexpressing strain showed a decreased capability to be internalized by the phagocytic cells. Consistently, the strain overexpressing elr operon was less adherent to macrophages than the WT strain, suggesting that overexpression of the elr operon could confer E. faecalis with additional anti-adhesion properties. In addition, increased virulence of the elr operon-overexpressing strain was shown in a mouse peritonitis model. Altogether, our results indicate that overexpression of the elr operon facilitates the E. faecalis escape from host immune defenses.

  13. Immunomodulation by gadolinium chloride-induced Kupffer cell phagocytosis blockade

    International Nuclear Information System (INIS)

    Lazar, G.; Husztik, E.; Kiss, I.; Szakacs, J.; Olah, J.

    1998-01-01

    Gadolinium chloride (GdCl 3 ), a rare earth metal salt, depresses macrophage activity, and is commonly used to study the physiology of the reticuloendothelial system. In the present work, the effect of GdCl 3 -induced Kupffer cell blockade on the humoral immune response in mice to sheep red blood cells (SRBC) was investigated. Kupffer cell phagocytosis blockade was found to increase both the primary and secondary immune responses to SRBC. The primary immune response was significantly augmented in animals injected intravenously with GdCl 3 2, 3 or 4 days before injection of the cellular antigen, but GdCl 3 injected 7 days before the antigen did not modify the immune response. Increased secondary humoral immune responses were also observed. When GdCl 3 was injected 2 days before the second dose of antigen, the numbers of both IgM and IgG-producing plaque forming cells were augmented. GdCl 3 injected 2 days before the first dose of SRBC did not modify the humoral immune response. Earlier studies with 51 Cr-labelled foreign red blood cells suggested that the augmentation of the humoral immune response in GdCl 3 -pretreated mice is a consequence of the spillover of the antigen from the liver into the spleen and other extrahepatic reticuloendothelial organs. (orig.)

  14. Phagocytosis and Inflammation: Exploring the effects of the components of E?cigarette vapor on macrophages

    OpenAIRE

    Ween, Miranda P.; Whittall, Jonathan J.; Hamon, Rhys; Reynolds, Paul N.; Hodge, Sandra J.

    2017-01-01

    Abstract E?cigarettes are perceived as harmless; however, evidence of their safety is lacking. New data suggests E?cigarettes discharge a range of compounds capable of physiological damage to users. We previously established that cigarette smoke caused defective alveolar macrophage phagocytosis. The present study compared the effect E?cigarette of components; E?liquid flavors, nicotine, vegetable glycerine, and propylene glycol on phagocytosis, proinflammatory cytokine secretion, and phagocyt...

  15. CEACAM3-mediated phagocytosis of human-specific bacterial pathogens involves the adaptor molecule Nck

    OpenAIRE

    Peterson, Lisa

    2010-01-01

    Carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) are exploited by human-specific pathogens to anchor themselves to or invade host cells. Interestingly, human granulocytes express a specific isoform, CEACAM3, that can direct efficient, opsonin-independent phagocytosis of CEACAM-binding Neisseria, Moraxella and Haemophilus species. As opsonin-independent phagocytosis of CEACAM-binding Neisseria depends on Src-family protein tyrosine kinase (PTK) phosphorylation of the CEACAM3 ...

  16. Alveolar macrophage phagocytosis is enhanced after blunt chest trauma and alters the posttraumatic mediator release.

    Science.gov (United States)

    Seitz, Daniel H; Palmer, Annette; Niesler, Ulrike; Fröba, Janine S; Heidemann, Vera; Rittlinger, Anne; Braumüller, Sonja T; Zhou, Shaoxia; Gebhard, Florian; Knöferl, Markus W

    2011-12-01

    Blunt chest trauma is known to induce a pulmonary invasion of short-lived polymorphonuclear neutrophils and apoptosis of alveolar epithelial type 2 (AT2) cells. Apoptotic cells are removed by alveolar macrophages (AMΦ). We hypothesized that chest trauma alters the phagocytic response of AMΦ as well as the mediator release of AMΦ during phagocytosis. To study this, male Sprague-Dawley rats were subjected to blunt chest trauma. Phagocytosis assays were performed in AMΦ isolated 2 or 24 h after trauma with apoptotic cells or opsonized beads. Phagocytosis of apoptotic AT2 cells by unstimulated AMΦ was significantly increased 2 h after trauma. At 24 h, AMΦ from traumatized animals, stimulated with phorbol-12-myristate-13-acetate, ingested significantly more apoptotic polymorphonuclear neutrophils than AMΦ from sham animals. Alveolar macrophages after trauma released significantly higher levels of tumor necrosis factor α, macrophage inflammatory protein 1α, and cytokine-induced neutrophil chemoattractant 1 when they incorporated latex beads, but significantly lower levels of interleukin 1β and macrophage inflammatory protein 1α when they ingested apoptotic cells. In vivo, phagocytosis of intratracheally instilled latex beads was decreased in traumatized rats. The bronchoalveolar lavage concentrations of the phagocytosis-supporting surfactant proteins A and D after blunt chest trauma were slightly decreased, whereas surfactant protein D mRNA expression in AT2 cells was significantly increased after 2 h. These findings indicate that chest trauma augments the phagocytosis of apoptotic cells by AMΦ. Phagocytosis of opsonized beads enhances and ingestion of apoptotic cells downregulates the immunologic response following lung contusion. Our data emphasize the important role of phagocytosis during posttraumatic inflammation after lung contusion.

  17. Antibiotic-Enhanced Phagocytosis of ’Borrelia recurrentis’ by Blood Polymorphonuclear Leukocytes.

    Science.gov (United States)

    1979-11-30

    hours after Butler 7 institution of antibiotic treatment. Polymorphonuclear leukocytes are known to release endogenous pyrogen after phagocytosis of...other bacteria (6), and endogenous pyrogen may be one of the mediators of the rigor and temperature rise in the Jarisch-Herxheimer reaction (2). Release...the pathogenesis of fever. XII. The effect of phagocytosis on the release of endogenous pyrogen by polymorphonuclear leukocytes. J. Exp. Med. 119:715

  18. Fc-receptor-mediated phagocytosis is regulated by mechanical properties of the target

    Science.gov (United States)

    Beningo, Karen A.; Wang, Yu-li

    2002-01-01

    Phagocytosis is an actin-based process used by macrophages to clear particles greater than 0.5 microm in diameter. In addition to its role in immunological responses, phagocytosis is also necessary for tissue remodeling and repair. To prevent catastrophic autoimmune reactions, phagocytosis must be tightly regulated. It is commonly assumed that the recognition/selection of phagocytic targets is based solely upon receptor-ligand binding. Here we report an important new criterion, that mechanical parameters of the target can dramatically affect the efficiency of phagocytosis. When presented with particles of identical chemical properties but different rigidity, macrophages showed a strong preference to engulf rigid objects. Furthermore, phagocytosis of soft particles can be stimulated with the microinjection of constitutively active Rac1 but not RhoA, and with lysophosphatidic acid, an agent known to activate the small GTP-binding proteins of the Rho family. These data suggest a Rac1-dependent mechanosensory mechanism for phagocytosis, which probably plays an important role in a number of physiological and pathological processes from embryonic development to autoimmune diseases.

  19. Influence of apolipoprotein A-V on the metabolic fate of triacylglycerol.

    Science.gov (United States)

    Sharma, Vineeta; Forte, Trudy M; Ryan, Robert O

    2013-04-01

    Apolipoprotein (apo) A-V functions to modulate intracellular and extracellular triacylglycerol metabolism. The present review addresses molecular mechanisms underlying these effects. The relevance of apoA-V to human disease conditions is illustrated by the strong correlation between single nucleotide polymorphisms in APOA5, elevated plasma triacylglycerol and dyslipidemic disease. Despite undergoing processing for secretion from hepatocytes, a portion of apoA-V escapes this destiny and accumulates as a component of cytosolic lipid droplets. Expression of recombinant apoA-V in hepatocarcinoma cells results in increased lipid droplet size and number at the expense of triacylglycerol secretion.ApoA-V modulates atherosclerosis in hypercholesterolemic apoE null mice. ApoE null/human apoA-V transgenic mice had reduced levels of triacylglycerol and cholesterol in plasma along with decreased aortic lesion size. ApoA-V modulates triacylglycerol metabolic fate. Following its synthesis, apoA-V enters the endoplasmic reticulum and associates with membrane defects created by triacylglycerol accumulation. Association of apoA-V with endoplasmic reticulum membrane defects promotes nascent lipid droplets budding toward the cytosol. Despite its low concentration in plasma (∼150 ng/ml), apoA-V modulates lipoprotein metabolism by binding to glycosylphosphatidylinositol-anchored high-density lipoprotein binding protein 1. This interaction effectively localizes triacylglycerol-rich lipoproteins in the vicinity of glycosylphosphatidylinositol-anchored high-density lipoprotein binding protein1's other ligand, lipoprotein lipase.

  20. Thematic review series: glycerolipids. DGAT enzymes and triacylglycerol biosynthesis.

    Science.gov (United States)

    Yen, Chi-Liang Eric; Stone, Scot J; Koliwad, Suneil; Harris, Charles; Farese, Robert V

    2008-11-01

    Triacylglycerols (triglycerides) (TGs) are the major storage molecules of metabolic energy and FAs in most living organisms. Excessive accumulation of TGs, however, is associated with human diseases, such as obesity, diabetes mellitus, and steatohepatitis. The final and the only committed step in the biosynthesis of TGs is catalyzed by acyl-CoA:diacylglycerol acyltransferase (DGAT) enzymes. The genes encoding two DGAT enzymes, DGAT1 and DGAT2, were identified in the past decade, and the use of molecular tools, including mice deficient in either enzyme, has shed light on their functions. Although DGAT enzymes are involved in TG synthesis, they have distinct protein sequences and differ in their biochemical, cellular, and physiological functions. Both enzymes may be useful as therapeutic targets for diseases. Here we review the current knowledge of DGAT enzymes, focusing on new advances since the cloning of their genes, including possible roles in human health and diseases.

  1. Metabolic engineering of Saccharomyces cerevisiae for overproduction of triacylglycerols

    DEFF Research Database (Denmark)

    Ferreira, Raphael; Teixeira, Paulo Goncalves; Gossing, Michael

    2018-01-01

    Triacylglycerols (TAGs) are valuable versatile compounds that can be used as metabolites for nutrition and health, as well as feedstocks for biofuel production. Although Saccharomyces cerevisiae is the favored microbial cell factory for industrial production of biochemicals, it does not produce...... large amounts of lipids and TAGs comprise only ~1% of its cell dry weight. Here, we engineered S. cerevisiae to reorient its metabolism for overproduction of TAGs, by regulating lipid droplet associated-proteins involved in TAG synthesis and hydrolysis. We implemented a push-and-pull strategy...... PXA1 led to accumulation of  254 mg∙gCDW−1. The TAG levels achieved here are the highest titer reported in S. cerevisiae, reaching 27.4% of the maximum theoretical yield in minimal medium with 2% glucose. This work shows the potential of using an industrially established and robust yeast species...

  2. Desaturation of excess intramyocellular triacylglycerol in obesity: implications for glycemic control

    DEFF Research Database (Denmark)

    Haugaard, S B; Madsbad, S; Mu, Huiling

    2010-01-01

    Excess intramyocellular triacylglycerol (IMTG), found especially in obese women, is slowly metabolized and, therefore, prone to longer exposure to intracellular desaturases. Accordingly, it was hypothesized that IMTG content correlates inversely with IMTG fatty acid (FA) saturation in sedentary...

  3. Microglial Phagocytosis and Its Regulation: A Therapeutic Target in Parkinson’s Disease?

    Directory of Open Access Journals (Sweden)

    Elzbieta Janda

    2018-04-01

    Full Text Available The role of phagocytosis in the neuroprotective function of microglia has been appreciated for a long time, but only more recently a dysregulation of this process has been recognized in Parkinson’s disease (PD. Indeed, microglia play several critical roles in central nervous system (CNS, such as clearance of dying neurons and pathogens as well as immunomodulation, and to fulfill these complex tasks they engage distinct phenotypes. Regulation of phenotypic plasticity and phagocytosis in microglia can be impaired by defects in molecular machinery regulating critical homeostatic mechanisms, including autophagy. Here, we briefly summarize current knowledge on molecular mechanisms of microglia phagocytosis, and the neuro-pathological role of microglia in PD. Then we focus more in detail on the possible functional role of microglial phagocytosis in the pathogenesis and progression of PD. Evidence in support of either a beneficial or deleterious role of phagocytosis in dopaminergic degeneration is reported. Altered expression of target-recognizing receptors and lysosomal receptor CD68, as well as the emerging determinant role of α-synuclein (α-SYN in phagocytic function is discussed. We finally discuss the rationale to consider phagocytic processes as a therapeutic target to prevent or slow down dopaminergic degeneration.

  4. Overload training inhibits phagocytosis and ROS generation of peritoneal macrophages: role of IGF-1 and MGF.

    Science.gov (United States)

    Xiao, Weihua; Chen, Peijie; Wang, Ru; Dong, Jingmei

    2013-01-01

    We tested the hypothesis that overload training inhibits the phagocytosis and the reactive oxygen species (ROS) generation of peritoneal macrophages (Mϕs), and that insulin-like growth factor-1(IGF-1) and mechano-growth factor (MGF) produced by macrophages may contribute to this process. Rats were randomized to two groups, sedentary control group (n = 10) and overload training group (n = 10). The rats of overload training group were subjected to 11 weeks of experimental training protocol. Blood sample was used to determine the content of hemoglobin, testosterone, and corticosterone. The phagocytosis and the ROS generation of Mϕs were measured by the uptake of neutral red and the flow cytometry, respectively. IGF-1 and MGF mRNA levels in Mϕs were determined by real-time PCR. In addition, we evaluated the effects of IGF-1 and MGF peptide on phagocytosis and ROS generation of Mϕs in vitro. The data showed that overload training significantly decreased the body weight (19.3 %, P overload training significantly decreased the phagocytosis (27 %, P overload training group increased significantly compared with the control group (21-fold and 92-fold, respectively; P overload training inhibits the phagocytosis and the ROS generation of peritoneal macrophages, and that MGF produced by macrophages may play a key role in this process. This may represent a novel mechanism of immune suppression induced by overload training.

  5. Aliphatic alcohol contaminants of illegally produced spirits inhibit phagocytosis by human granulocytes.

    Science.gov (United States)

    Pál, László; Árnyas, Ervin M; Tóth, Béla; Ádám, Balázs; Rácz, Gábor; Ádány, Róza; McKee, Martin; Szűcs, Sándor

    2013-04-01

    Unregulated production of spirits in many countries leads to products containing appreciable levels of aliphatic alcohols (AAs) and is the main source of human exposure to these substances worldwide. Previous studies have confirmed that alcohol abuse can lead to ethanol-induced immunosuppression and thereby increased susceptibility to infectious diseases. Granulocytes, as professional phagocytic cells, play a crucial role in engulfment and killing of pathogenic microorganisms. Thus, a decrease in their phagocytic activity has been invoked as a factor in the impaired antimicrobial defense observed in alcoholics. However, AAs consumed as contaminants of illicit spirits may also influence phagocytosis, thereby contributing to a further decrease in microbicidal activity but, so far, this has not been studied. Therefore, the aim of this study was to measure granulocyte phagocytosis following treatment of granulocytes with those higher alcohols found in illegal spirits. Granulocytes were isolated from human peripheral blood. Then phagocytosis of opsonized zymosan particles by granulocytes treated with AAs individually and in combination was determined. These alcohols inhibited phagocytosis in a concentration-dependent manner and at lower concentrations when combined than when tested individually. Due to their synergistic effects, it is possible that, in combination with ethanol, they may inhibit phagocytosis in a clinically meaningful way in episodic heavy drinkers.

  6. Conjugated linoleic acid-rich soy oil triacylglycerol identification.

    Science.gov (United States)

    Lall, Rahul K; Proctor, Andrew; Jain, Vishal P; Lay, Jackson O

    2009-03-11

    Conjugated linoleic acid (CLA)-rich soy oil has been produced by soy oil linoleic acid (LA) photoisomerization, but CLA-rich oil triacylglycerol (TAG) characterization was not described. Therefore, the objectives were to identify and quantify new TAG fractions in CLA-rich oil by nonaqueous reversed-phase high-performance liquid chromatography (NARP-HPLC). Analytical NARP-HPLC with an acetonitrile/dichloromethane (ACN/DCM) gradient and an evaporating light scattering detector/ultraviolet (ELSD/UV) detector was used. New TAG peaks from LA-containing TAGs were observed. The LnLL, LLL, LLO, and LLP (Ln, linolenic; L, linoleic; O, oleic; and P, palmitic) peaks reduced after isomerization with an increase in adjacent peaks that coeluted with LnLnO, LnLO, LnOO, and LnPP. The newly formed peaks were wider than those of the original oil and absorbed at 233 nm, suggesting the possibility of various CLA containing TAGs. The HPLC profile showed five fractions of mixed TAGs, and fatty acid analysis showed that CLA isomers were found predominately in fractions 2 and 3, which originally contained most LA. The CLA isomers were 70-80% trans,trans and 20-30% cis,trans and trans,cis.

  7. A novel assay of DGAT activity based on high temperature GC/MS of triacylglycerol.

    Science.gov (United States)

    Greer, Michael S; Zhou, Ting; Weselake, Randall J

    2014-08-01

    Diacylglycerol acyltransferase (DGAT) catalyzes the final step in the acyl-CoA-dependent biosynthesis of triacylglycerol (TAG), a high-energy compound composed of three fatty acids esterified to a glycerol backbone. In vitro DGAT assays, which are usually conducted with radiolabeled substrate using microsomal fractions, have been useful in identifying compounds and genetic modifications that affect DGAT activity. Here, we describe a high-temperature gas chromatography (GC)/mass spectrometry (MS)-based method for monitoring molecular species of TAG produced by the catalytic action of microsomal DGAT. This method circumvents the need for radiolabeled or modified substrates, and only requires a simple lipid extraction prior to GC. The utility of the method is demonstrated using a recombinant type-1 Brassica napus DGAT produced in a strain of Saccharomyces cerevisae that is deficient in TAG synthesis. The GC/MS-based assay of DGAT activity was strongly correlated with the typical in vitro assay of the enzyme using [1-(14)C] acyl-CoA as an acyl donor. In addition to determining DGAT activity, the method is also useful for determining substrate specificity and selectivity properties of the enzyme.

  8. Target of rapamycin (TOR) plays a critical role in triacylglycerol accumulation in microalgae.

    Science.gov (United States)

    Imamura, Sousuke; Kawase, Yasuko; Kobayashi, Ikki; Sone, Toshiyuki; Era, Atsuko; Miyagishima, Shin-Ya; Shimojima, Mie; Ohta, Hiroyuki; Tanaka, Kan

    2015-10-01

    Most microalgae produce triacylglycerol (TAG) under stress conditions such as nitrogen depletion, but the underlying molecular mechanism remains unclear. In this study, we focused on the role of target of rapamycin (TOR) in TAG accumulation. TOR is a serine/threonine protein kinase that is highly conserved and plays pivotal roles in nitrogen and other signaling pathways in eukaryotes. We previously constructed a rapamycin-susceptible Cyanidioschyzon merolae, a unicellular red alga, by expressing yeast FKBP12 protein to evaluate the results of TOR inhibition (Imamura et al. in Biochem Biophys Res Commun 439:264-269, 2013). By using this strain, we here report that rapamycin-induced TOR inhibition results in accumulation of cytoplasmic lipid droplets containing TAG. Transcripts for TAG synthesis-related genes, such as glycerol-3-phosphate acyltransferase and acyl-CoA:diacylglycerol acyltransferase (DGAT), were increased by rapamycin treatment. We also found that fatty acid synthase-dependent de novo fatty acid synthesis was required for the accumulation of lipid droplets. Induction of TAG and up-regulation of DGAT gene expression by rapamycin were similarly observed in the unicellular green alga, Chlamydomonas reinhardtii. These results suggest the general involvement of TOR signaling in TAG accumulation in divergent microalgae.

  9. Triacylglycerol Storage in Lipid Droplets in Procyclic Trypanosoma brucei.

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    Stefan Allmann

    Full Text Available Carbon storage is likely to enable adaptation of trypanosomes to nutritional challenges or bottlenecks during their stage development and migration in the tsetse. Lipid droplets are candidates for this function. This report shows that feeding of T. brucei with oleate results in a 4-5 fold increase in the number of lipid droplets, as quantified by confocal fluorescence microscopy and by flow cytometry of BODIPY 493/503-stained cells. The triacylglycerol (TAG content also increased 4-5 fold, and labeled oleate is incorporated into TAG. Fatty acid carbon can thus be stored as TAG in lipid droplets under physiological growth conditions in procyclic T. brucei. β-oxidation has been suggested as a possible catabolic pathway for lipids in T. brucei. A single candidate gene, TFEα1 with coding capacity for a subunit of the trifunctional enzyme complex was identified. TFEα1 is expressed in procyclic T. brucei and present in glycosomal proteomes, Unexpectedly, a TFEα1 gene knock-out mutant still expressed wild-type levels of previously reported NADP-dependent 3-hydroxyacyl-CoA dehydrogenase activity, and therefore, another gene encodes this enzymatic activity. Homozygous Δtfeα1/Δtfeα1 null mutant cells show a normal growth rate and an unchanged glycosomal proteome in procyclic T. brucei. The decay kinetics of accumulated lipid droplets upon oleate withdrawal can be fully accounted for by the dilution effect of cell division in wild-type and Δtfeα1/Δtfeα1 cells. The absence of net catabolism of stored TAG in procyclic T. brucei, even under strictly glucose-free conditions, does not formally exclude a flux through TAG, in which biosynthesis equals catabolism. Also, the possibility remains that TAG catabolism is completely repressed by other carbon sources in culture media or developmentally activated in post-procyclic stages in the tsetse.

  10. A Simple Microscopy Assay to Teach the Processes of Phagocytosis and Exocytosis

    Science.gov (United States)

    Gray, Ross; Gray, Andrew; Fite, Jessica L.; Jordan, Renee; Stark, Sarah; Naylor, Kari

    2012-01-01

    Phagocytosis and exocytosis are two cellular processes involving membrane dynamics. While it is easy to understand the purpose of these processes, it can be extremely difficult for students to comprehend the actual mechanisms. As membrane dynamics play a significant role in many cellular processes ranging from cell signaling to cell division to…

  11. Hemocyte-mediated phagocytosis differs between honey bee (Apis mellifera worker castes.

    Directory of Open Access Journals (Sweden)

    Eva Marit Hystad

    Full Text Available Honey bees as other insects rely on the innate immune system for protection against diseases. The innate immune system includes the circulating hemocytes (immune cells that clear pathogens from hemolymph (blood by phagocytosis, nodulation or encapsulation. Honey bee hemocyte numbers have been linked to hemolymph levels of vitellogenin. Vitellogenin is a multifunctional protein with immune-supportive functions identified in a range of species, including the honey bee. Hemocyte numbers can increase via mitosis, and this recruitment process can be important for immune system function and maintenance. Here, we tested if hemocyte mediated phagocytosis differs among the physiologically different honey bee worker castes (nurses, foragers and winter bees, and study possible interactions with vitellogenin and hemocyte recruitment. To this end, we adapted phagocytosis assays, which-together with confocal microscopy and flow cytometry-allow qualitative and quantitative assessment of hemocyte performance. We found that nurses are more efficient in phagocytic uptake than both foragers and winter bees. We detected vitellogenin within the hemocytes, and found that winter bees have the highest numbers of vitellogenin-positive hemocytes. Connections between phagocytosis, hemocyte-vitellogenin and mitosis were worker caste dependent. Our results demonstrate that the phagocytic performance of immune cells differs significantly between honey bee worker castes, and support increased immune competence in nurses as compared to forager bees. Our data, moreover, provides support for roles of vitellogenin in hemocyte activity.

  12. Phagocytosis and Inflammation: Exploring the effects of the components of E-cigarette vapor on macrophages.

    Science.gov (United States)

    Ween, Miranda P; Whittall, Jonathan J; Hamon, Rhys; Reynolds, Paul N; Hodge, Sandra J

    2017-08-01

    E-cigarettes are perceived as harmless; however, evidence of their safety is lacking. New data suggests E-cigarettes discharge a range of compounds capable of physiological damage to users. We previously established that cigarette smoke caused defective alveolar macrophage phagocytosis. The present study compared the effect E-cigarette of components; E-liquid flavors, nicotine, vegetable glycerine, and propylene glycol on phagocytosis, proinflammatory cytokine secretion, and phagocytic recognition molecule expression using differentiated THP-1 macrophages. Similar to CSE, phagocytosis of NTHi bacteria was significantly decreased by E-liquid flavoring (11.65-15.75%) versus control (27.01%). Nicotine also decreased phagocytosis (15.26%). E-liquid, nicotine, and E-liquid+ nicotine reduced phagocytic recognition molecules; SR-A1 and TLR-2. IL-8 secretion increased with flavor and nicotine, while TNF α , IL-1 β , IL-6, MIP-1 α , MIP-1 β , and MCP-1 decreased after exposure to most flavors and nicotine. PG, VG, or PG:VG mix also induced a decrease in MIP-1 α and MIP-1 β We conclude that E-cigarettes can cause macrophage phagocytic dysfunction, expression of phagocytic recognition receptors and cytokine secretion pathways. As such, E-cigarettes should be treated with caution by users, especially those who are nonsmokers. © 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.

  13. Hemocyte-mediated phagocytosis differs between honey bee (Apis mellifera) worker castes.

    Science.gov (United States)

    Hystad, Eva Marit; Salmela, Heli; Amdam, Gro Vang; Münch, Daniel

    2017-01-01

    Honey bees as other insects rely on the innate immune system for protection against diseases. The innate immune system includes the circulating hemocytes (immune cells) that clear pathogens from hemolymph (blood) by phagocytosis, nodulation or encapsulation. Honey bee hemocyte numbers have been linked to hemolymph levels of vitellogenin. Vitellogenin is a multifunctional protein with immune-supportive functions identified in a range of species, including the honey bee. Hemocyte numbers can increase via mitosis, and this recruitment process can be important for immune system function and maintenance. Here, we tested if hemocyte mediated phagocytosis differs among the physiologically different honey bee worker castes (nurses, foragers and winter bees), and study possible interactions with vitellogenin and hemocyte recruitment. To this end, we adapted phagocytosis assays, which-together with confocal microscopy and flow cytometry-allow qualitative and quantitative assessment of hemocyte performance. We found that nurses are more efficient in phagocytic uptake than both foragers and winter bees. We detected vitellogenin within the hemocytes, and found that winter bees have the highest numbers of vitellogenin-positive hemocytes. Connections between phagocytosis, hemocyte-vitellogenin and mitosis were worker caste dependent. Our results demonstrate that the phagocytic performance of immune cells differs significantly between honey bee worker castes, and support increased immune competence in nurses as compared to forager bees. Our data, moreover, provides support for roles of vitellogenin in hemocyte activity.

  14. The adaptor molecule Nck localizes the WAVE complex to promote actin polymerization during CEACAM3-mediated phagocytosis of bacteria.

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    Stefan Pils

    Full Text Available BACKGROUND: CEACAM3 is a granulocyte receptor mediating the opsonin-independent recognition and phagocytosis of human-restricted CEACAM-binding bacteria. CEACAM3 function depends on an intracellular immunoreceptor tyrosine-based activation motif (ITAM-like sequence that is tyrosine phosphorylated by Src family kinases upon receptor engagement. The phosphorylated ITAM-like sequence triggers GTP-loading of Rac by directly associating with the guanine nucleotide exchange factor (GEF Vav. Rac stimulation in turn is critical for actin cytoskeleton rearrangements that generate lamellipodial protrusions and lead to bacterial uptake. PRINCIPAL FINDINGS: In our present study we provide biochemical and microscopic evidence that the adaptor proteins Nck1 and Nck2, but not CrkL, Grb2 or SLP-76, bind to tyrosine phosphorylated CEACAM3. The association is phosphorylation-dependent and requires the Nck SH2 domain. Overexpression of the isolated Nck1 SH2 domain, RNAi-mediated knock-down of Nck1, or genetic deletion of Nck1 and Nck2 interfere with CEACAM3-mediated bacterial internalization and with the formation of lamellipodial protrusions. Nck is constitutively associated with WAVE2 and directs the actin nucleation promoting WAVE complex to tyrosine phosphorylated CEACAM3. In turn, dominant-negative WAVE2 as well as shRNA-mediated knock-down of WAVE2 or the WAVE-complex component Nap1 reduce internalization of bacteria. CONCLUSIONS: Our results provide novel mechanistic insight into CEACAM3-initiated phagocytosis. We suggest that the CEACAM3 ITAM-like sequence is optimized to co-ordinate a minimal set of cellular factors needed to efficiently trigger actin-based lamellipodial protrusions and rapid pathogen engulfment.

  15. STUDIES ON THE PATHOGENESIS OF FEVER. 13. THE EFFECT OF PHAGOCYTOSIS ON THE RELEASE OF ENDOGENOUS PYROGEN BY POLYMORPHONUCLEAR LEUKOCYTES.

    Science.gov (United States)

    BERLIN, R D; WOOD, W B

    1964-05-01

    1. Phagocytosis promotes the release of endogenous pyrogen from polymorphonuclear leucocytes. 2. The release of pyrogen, though initiated by the phagocytic event, is not synchronous with it. 3. The postphagocytic release mechanism is not inhibited by sodium fluoride and, therefore, appears not to require continued production of energy by the cell. 4. The release process, on the other hand, is inhibited by arsenite, suggesting the participation of one or more sulfhydryl-dependent enzymes in the over-all reaction. 5. Particle for particle, the ingestion of heat-killed rough pneumococci causes the release of approximately 100 times as much pyrogen as the ingestion of polystyrene beads of the same size. 6. The pyrogen release mechanism of polymorphonuclear leucocytes separated directly from blood, unlike that of granulocytes in acute inflammatory exudates, is not readily activated by incubation of the cells in K-free saline. Despite this difference, both blood and exudate leucocytes following phagocytosis release large amounts of pyrogen, even in the presence of K(+). The fact that the postphagocytic reaction is uninhibited by the concentrations of K(+) which are present in plasma and extracellular fluids, suggests that this mechanism of pyrogen release may well operate in vivo. 7. As might be expected from the foregoing observations, the intravenous injection of a sufficiently large number of heat-killed pneumococci causes fever in the intact host. Intravenously injected polystyrene beads, on the other hand, are significantly less pyrogenic. Evidence is presented to support the conclusion that the fever in both instances is caused by pyrogen released from the circulating leucocytes which have phagocyted the injected particles. 8. The possible relationships of these findings to the pathogenesis of fevers caused by acute bacterial infections are discussed.

  16. Diclofenac enhances proinflammatory cytokine-induced phagocytosis of cultured microglia via nitric oxide production

    International Nuclear Information System (INIS)

    Kakita, Hiroki; Aoyama, Mineyoshi; Nagaya, Yoshiaki; Asai, Hayato; Hussein, Mohamed Hamed; Suzuki, Mieko; Kato, Shin; Saitoh, Shinji; Asai, Kiyofumi

    2013-01-01

    Influenza-associated encephalopathy (IAE) is a central nervous system complication with a high mortality rate, which is increased significantly by the non-steroidal anti-inflammatory drug diclofenac sodium (DCF). In the present study, we investigated the effects of DCF on brain immune cells (i.e. microglia) stimulated with three proinflammatory cytokines, namely tumor necrosis factor-α, interleukin-1β, and interferon-γ. Similar to previous findings in astrocytes, all three cytokines induced the expression of inducible NO synthase (iNOS), as well as NO production, in microglia. The addition of DCF to the culture system augmented iNOS expression and NO production. Immunocytochemical analysis and the phagocytosis assay revealed that cytokine treatment induced morphological changes to and phagocytosis by the microglia. The addition of DCF to the culture system enhanced microglial activation, as well as the phagocytic activity of cytokine-stimulated microglia. Inhibitors of nuclear factor (NF)-κB inhibited iNOS gene expression in cytokine-stimulated microglia with or without DCF, suggesting that the NF-κB pathway is one of the main signaling pathways involved. The iNOS inhibitor N G -monomethyl-L-arginine (L-NMMA) reduced both cytokine-induced phagocytosis and phagocytosis induced by the combination of cytokines plus DCF. Furthermore, the NO donor sodium nitroprusside induced phagocytosis, indicating that NO production is a key regulator of microglial phagocytosis. In conclusion, DCF acts synergistically with proinflammatory cytokines to increase the production of NO in microglia, leading to phagocytic activity of the activated microglia. These findings, together with previous observations regarding astrocytes, may explain the significant increase in mortality of IAE patients treated with DCF. - Highlights: ► Influenza-associated encephalopathy (IAE) is associated with a high mortality rate. ► Hyperimmunization in the brain is believed to be responsible for IAE

  17. LRRK2 kinase inhibition prevents pathological microglial phagocytosis in response to HIV-1 Tat protein

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    Marker Daniel F

    2012-11-01

    Full Text Available Abstract Background Human Immunodeficiency Virus-1 (HIV-1 associated neurocognitive disorders (HANDs are accompanied by significant morbidity, which persists despite the use of combined antiretroviral therapy (cART. While activated microglia play a role in pathogenesis, changes in their immune effector functions, including phagocytosis and proinflammatory signaling pathways, are not well understood. We have identified leucine-rich repeat kinase 2 (LRRK2 as a novel regulator of microglial phagocytosis and activation in an in vitro model of HANDs, and hypothesize that LRRK2 kinase inhibition will attenuate microglial activation during HANDs. Methods We treated BV-2 immortalized mouse microglia cells with the HIV-1 trans activator of transcription (Tat protein in the absence or presence of LRRK2 kinase inhibitor (LRRK2i. We used Western blot, qRT-PCR, immunocytochemistry and latex bead engulfment assays to analyze LRRK2 protein levels, proinflammatory cytokine and phagocytosis receptor expression, LRRK2 cellular distribution and phagocytosis, respectively. Finally, we utilized ex vivo microfluidic chambers containing primary hippocampal neurons and BV-2 microglia cells to investigate microglial phagocytosis of neuronal axons. Results We found that Tat-treatment of BV-2 cells induced kinase activity associated phosphorylation of serine 935 on LRRK2 and caused the formation of cytoplasmic LRRK2 inclusions. LRRK2i decreased Tat-induced phosphorylation of serine 935 on LRRK2 and inhibited the formation of Tat-induced cytoplasmic LRRK2 inclusions. LRRK2i also decreased Tat-induced process extension in BV-2 cells. Furthermore, LRRK2i attenuated Tat-induced cytokine expression and latex bead engulfment. We examined relevant cellular targets in microfluidic chambers and found that Tat-treated BV-2 microglia cells cleared axonal arbor and engulfed neuronal elements, whereas saline treated controls did not. LRRK2i was found to protect axons in the presence

  18. Diclofenac enhances proinflammatory cytokine-induced phagocytosis of cultured microglia via nitric oxide production

    Energy Technology Data Exchange (ETDEWEB)

    Kakita, Hiroki [Department of Molecular Neurobiology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Department of Pediatrics and Neonatology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Aoyama, Mineyoshi, E-mail: ao.mine@med.nagoya-cu.ac.jp [Department of Molecular Neurobiology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Nagaya, Yoshiaki; Asai, Hayato [Department of Molecular Neurobiology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Department of Pediatrics and Neonatology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Hussein, Mohamed Hamed [Neonatal Intensive Care Unit, Pediatric Hospital, Cairo University, Cairo 11559 (Egypt); Maternal and Child Health Department, VACSERA, 51 Wizaret El-Zeraa-Agouza, Giza 22311 (Egypt); Suzuki, Mieko [Department of Molecular Neurobiology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Kato, Shin [Department of Molecular Neurobiology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Department of Pediatrics and Neonatology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Saitoh, Shinji [Department of Pediatrics and Neonatology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Asai, Kiyofumi [Department of Molecular Neurobiology, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan)

    2013-04-15

    Influenza-associated encephalopathy (IAE) is a central nervous system complication with a high mortality rate, which is increased significantly by the non-steroidal anti-inflammatory drug diclofenac sodium (DCF). In the present study, we investigated the effects of DCF on brain immune cells (i.e. microglia) stimulated with three proinflammatory cytokines, namely tumor necrosis factor-α, interleukin-1β, and interferon-γ. Similar to previous findings in astrocytes, all three cytokines induced the expression of inducible NO synthase (iNOS), as well as NO production, in microglia. The addition of DCF to the culture system augmented iNOS expression and NO production. Immunocytochemical analysis and the phagocytosis assay revealed that cytokine treatment induced morphological changes to and phagocytosis by the microglia. The addition of DCF to the culture system enhanced microglial activation, as well as the phagocytic activity of cytokine-stimulated microglia. Inhibitors of nuclear factor (NF)-κB inhibited iNOS gene expression in cytokine-stimulated microglia with or without DCF, suggesting that the NF-κB pathway is one of the main signaling pathways involved. The iNOS inhibitor N{sup G}-monomethyl-L-arginine (L-NMMA) reduced both cytokine-induced phagocytosis and phagocytosis induced by the combination of cytokines plus DCF. Furthermore, the NO donor sodium nitroprusside induced phagocytosis, indicating that NO production is a key regulator of microglial phagocytosis. In conclusion, DCF acts synergistically with proinflammatory cytokines to increase the production of NO in microglia, leading to phagocytic activity of the activated microglia. These findings, together with previous observations regarding astrocytes, may explain the significant increase in mortality of IAE patients treated with DCF. - Highlights: ► Influenza-associated encephalopathy (IAE) is associated with a high mortality rate. ► Hyperimmunization in the brain is believed to be responsible for

  19. Roles of Acyl-CoA:Diacylglycerol Acyltransferases 1 and 2 in Triacylglycerol Synthesis and Secretion in Primary Hepatocytes.

    Science.gov (United States)

    Li, Chen; Li, Lena; Lian, Jihong; Watts, Russell; Nelson, Randal; Goodwin, Bryan; Lehner, Richard

    2015-05-01

    Very low-density lipoprotein assembly and secretion are regulated by the availability of triacylglycerol. Although compelling evidence indicates that the majority of triacylglycerol in very low-density lipoprotein is derived from re-esterification of lipolytic products released by endoplasmic reticulum-associated lipases, little is known about roles of acyl-CoA:diacylglycerol acyltransferases (DGATs) in this process. We aimed to investigate the contribution of DGAT1 and DGAT2 in lipid metabolism and lipoprotein secretion in primary mouse and human hepatocytes. We used highly selective small-molecule inhibitors of DGAT1 and DGAT2, and we tracked storage and secretion of lipids synthesized de novo from [(3)H]acetic acid and from exogenously supplied [(3)H]oleic acid. Inactivation of individual DGAT activity did not affect incorporation of either radiolabeled precursor into intracellular triacylglycerol, whereas combined inactivation of both DGATs severely attenuated triacylglycerol synthesis. However, inhibition of DGAT2 augmented fatty acid oxidation, whereas inhibition of DGAT1 increased triacylglycerol secretion, suggesting preferential channeling of separate DGAT-derived triacylglycerol pools to distinct metabolic pathways. Inactivation of DGAT2 impaired cytosolic lipid droplet expansion, whereas DGAT1 inactivation promoted large lipid droplet formation. Moreover, inactivation of DGAT2 attenuated expression of lipogenic genes. Finally, triacylglycerol secretion was significantly reduced on DGAT2 inhibition without altering extracellular apolipoprotein B levels. Our data suggest that DGAT1 and DGAT2 can compensate for each other to synthesize triacylglycerol, but triacylglycerol synthesized by DGAT1 is preferentially channeled to oxidation, whereas DGAT2 synthesizes triacylglycerol destined for very low-density lipoprotein assembly. © 2015 American Heart Association, Inc.

  20. Synthesis of cocoa butter triacylglycerols using a model acidolysis system

    Directory of Open Access Journals (Sweden)

    Göǧüş, Fahrettin

    2008-12-01

    Full Text Available The effects of reaction parameters such as substrate mole ratio, reaction temperature, enzyme load, water content and reaction time were studied in a model enzymatic acidolysis system. Palmitic and stearic acids were incorporated into triolein (OOO under the catalysis of sn-1,3 specific lipase to produce the three major triacylglycerols (TAGs in cocoa butter (CB, namely, 1,3-dipalmitoyl-2-oleoyl--glycerol (POP, 1(3-palmitoyl-3(1-stearoyl-2-oleoyl-glycerol (POS and 1,3distearoyl-2-oleoyl-glycerol (SOS. TAG contents of the reaction products were analyzed by High Performance Liquid Chromatograph (HPLC.The best results (15.2% POP, 30.4% POS, 15.2 % SOS were obtained at 1:3:3 (OOO:palmitic acid:stearic acid substrate mole ratio and reaction parameters: time 10 h, temperature 45 oC, enzyme load 20 %, water content 5 %. The results obtained in this model system might be used for the optimization and application of lipase catalyzed acidolysis reactions in natural systems to produce cocoa butter equivalents (CBEs.El efecto de parámetros de reacción como el ratio molar de sustratos, carga de enzima, contenido de agua o temperatura de reacción fueron estudiados en un sistema modelo de acidolisis para la síntesis de triglicéridos disaturados. Los ácidos grasos palmítico y esteárico se incorporaron a la molécula de trioleína (OOO mediante la catálisis con una li-pasa sn-1,3 específica para producir los tres triglicéridos (TAGs mayoritarios de la manteca de cacao (CB: 1,3-dipalmitil-2-oleoil glicerol (POP, 1(3-palmitil-3(1-estearil-2glicerol (POS and 1,3-diestearil-2-oleoil glicerol (SOS. Los TAGs producidos en cada reacción se analizaron por cromatografía líquida de alta eficacia (HPLC. Los mejores resultados (15.2 %, POP, 30.4 % POS, 15.2 % SOS se obtuvieron empleando la relación molar de sustratos 1:3:3 (OOO: ácido palmítico:ácido esteárico y los parámetros de reacción: tiempo 10h, temperatura 45 °C, carga de enzima 20

  1. Involvement of triacylglycerol in the metabolism of fatty acids by cultured neuroblastoma and glioma cells

    International Nuclear Information System (INIS)

    Cook, H.W.; Clarke, J.T.; Spence, M.W.

    1982-01-01

    The metabolism (chain elongation, desaturation, and incorporation into complex lipids) of thirteen different radiolabeled fatty acids and acetate was examined in N1E-115 neuroblastoma and C-6 glioma cell lines in culture. During 6-hr incubations, all fatty acids were extensively (14-80%) esterified to complex lipids, mainly choline phosphoglycerides and triacylglycerol. With trienoic and tetraenoic substrates, inositol and ethanolamine phosphoglycerides also contained up to 30% of the labeled fatty acids; plasmalogen contained up to half of the label in the ethanolamine phosphoglyceride fraction of neuroblastoma cells. Chain elongation and delta 9, delta 6, and delta 5 desaturation occurred in both cell lines; delta 4 desaturation was not observed. Seemingly anomalous utilization of arachidic acid and some selectivity based on the geometric configuration of double bonds was observed. These studies indicate that these cell lines are capable of modulating cellular membrane composition by a combination of selective exclusion and removal of inappropriate acyl chains and of modification of other acyl chains by desaturation and chain elongation. The time courses and patterns of modification and incorporation of exogenous substrates into phospholipids and triacylglycerol suggest that exogenous unsaturated fatty acid may be incorporated into triacylglycerol and later released for further metabolism and incorporation into phospholipids. This supports a role for triacylglycerol in the synthesis of membrane complex lipids in cell lines derived from neural tissue

  2. Enantiomeric separation of triacylglycerols containing polyunsaturated fatty acids with 18 carbon atoms

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Nedbalová, L.; Sigler, Karel

    2016-01-01

    Roč. 1467, October (2016), s. 261-269 ISSN 0021-9673 R&D Projects: GA ČR GA14-00227S Institutional support: RVO:61388971 Keywords : Chiral triacylglycerols * Enantiomers * Regioisomers Subject RIV: EE - Microbiology, Virology Impact factor: 3.981, year: 2016

  3. Regulation of the Yeast Triacylglycerol Lipase Tgl3p by Formation of Nonpolar Lipids*

    Science.gov (United States)

    Schmidt, Claudia; Athenstaedt, Karin; Koch, Barbara; Ploier, Birgit; Daum, Günther

    2013-01-01

    Tgl3p, the major triacylglycerol lipase of the yeast Saccharomyces cerevisiae, is a component of lipid droplets but is also present in the endoplasmic reticulum in a minor amount. Recently, it was shown that this enzyme can also serve as a lysophospholipid acyltransferase (Rajakumari, S., and Daum, G. (2010) Mol. Biol. Cell 21, 501–510). Here, we describe the effects of the presence/absence of triacylglycerols and lipid droplets on the functionality of Tgl3p. In a dga1Δlro1Δare1Δare2Δ quadruple mutant lacking all four triacylglycerol- and steryl ester-synthesizing acyltransferases and consequently the lipid droplets, the gene expression of TGL3 was only slightly altered. In contrast, protein level and stability of Tgl3p were markedly reduced in the absence of lipid droplets. Under these conditions, the enzyme was localized to the endoplasmic reticulum. Even the lack of the substrate, triacylglycerol, affected stability and localization of Tgl3p to some extent. Interestingly, Tgl3p present in the endoplasmic reticulum seems to lack lipolytic as well as acyltransferase activity as shown by enzymatic analysis and lipid profiling. Thus, we propose that the activity of Tgl3p is restricted to lipid droplets, whereas the endoplasmic reticulum may serve as a parking lot for this enzyme. PMID:23673660

  4. Protein and energy metabolism of young male Wistar rats fed conjugated linoleic acid as structured triacylglycerol

    DEFF Research Database (Denmark)

    Jørgensen, H.; Hansen, C. H.; Mu, Huiling

    2010-01-01

    Twelve 4-week-old male Wistar rats weighing 100 g were fed diets semi-ad libitum for 22 d containing either 1.5% conjugated linoleic acid (CLA-diet) or high oleic sunflower oil (Control-diet). The CLA was structured triacylglycerol with predominantly cis-9, trans-11 and trans-10, cis-12 fatty acid...

  5. Lymphatic recovery of exogenous oleic acid in rats on long chain or specific structured triacylglycerol diets

    DEFF Research Database (Denmark)

    Vistisen, Bodil; Mu, Huiling; Høy, Carl-Erik

    2006-01-01

    Specific structured triacylglycerols, MLM (M = medium-chain fatty acid, L = long-chain fatty acid), rapidly deliver energy and long-chain fatty acids to the body and are used for longer periods in human enteral feeding. In the present study rats were fed diets of 10 wt% MLM or LLL (L = oleic acid......% and 45%, respectively). However, the recovery of exogenous 18:1 n-9 was higher after a single bolus of MLM compared with a bolus of LLL in rats on the MLM diet (40% and 24%, respectively, P = 0.009). The recovery of lymphatic 18:1 n-9 of the LLL bolus tended to depend on the diet triacylglycerol...... structure and composition (P = 0.07). This study demonstrated that with a diet containing specific structured triacylglycerol, the lymphatic recovery of 18:1 n-9 after a single bolus of fat was dependent on the triacylglycerol structure of the bolus. This indicates that the lymphatic recovery of long...

  6. Isolated hyperglycaemia does not increase VLDL-triacylglycerol secretion in type 1 diabetic men

    DEFF Research Database (Denmark)

    Johansen, Rakel Fuglsang; Søndergaard, Esben; Sørensen, Lars Peter

    2015-01-01

    AIMS/HYPOTHESIS: In type 1 diabetes, abnormalities of both glucose and lipoprotein metabolism are seen. The relationship between these factors is not understood, but studies indicate that hyperglycaemia may increase hepatic VLDL-triacylglycerol (VLDL-TG) secretion and reduce VLDL-TG fatty acid...

  7. Postprandial triacylglycerol uptake in the legs is increased during exercise and post-exercise recovery

    DEFF Research Database (Denmark)

    Enevoldsen, L H; Simonsen, L; Bülow, J

    2005-01-01

    plasma triacylglycerol (TAG) uptake and clearance in the exercising legs immediately and for at least 4 h post-exercise, while food intake per se did not change leg plasma TAG uptake or clearance for up to 6 h. It is hypothesized that the effect of exercise on leg plasma TAG metabolism is a result...

  8. Enzymatic and fluorometric determination of triacylglycerols in cow milk and other opaque matrices

    DEFF Research Database (Denmark)

    Larsen, Torben; Larsen, Mette Krogh; Friggens, Nic

    2011-01-01

    The fat content of milk is an important indication of quality, both economically and physiologically. Existing analytical methods of milk fat are based on physical determination, i.e. gravimetric determination or near infrared spectroscopy (IR). Triacylglycerols (TAG) constitute up to 98% of tota...

  9. The stereospecific triacylglycerol structures and fatty acid profiles of human milk and infant formulas

    DEFF Research Database (Denmark)

    Straarup, Ellen Marie; Lauritzen, L.; Færk, Jan

    2006-01-01

    Background: The stereospecific structures of the triacylglycerol molecules in human milk differ from that of cow's milk and vegetable oils, which are the fat sources used in infant formula. In human milk, palmitic acid (16:0) is predominantly esterified in the sn2 position, whereas vegetable oils...

  10. Lipidemic effects of an interesterified mixture of butter, medium-chain triacylglycerol and safflower oils

    DEFF Research Database (Denmark)

    Mascioli, E.A.; McLennan, C.E.; Schaefer, E.J.

    1999-01-01

    The objective of this study was to determine if the positional structure of dietary triacylglycerol affected lipidemic responses. Thirty healthyadults (16 men and 14 postmenopausal women) with low-density lipoprotein cholesterol (LDL-C) concentrations >3.37 mM (130 mg/dL)enrolled in a prospective...

  11. Triacylglycerol infusion improves exercise endurance in patients with mitochondrial myopathy due to complex I deficiency

    NARCIS (Netherlands)

    Roef, MJ; de Meer, K; Reijngoud, DJ; Straver, HWHC; de Barse, M; Kalhan, SC; Berger, R

    Background: A high-fat diet has been recommended for the treatment of patients with mitochondrial myopathy due to complex I (NADH dehydrogenase) deficiency (CID). Objective: This study evaluated the effects of intravenous infusion of isoenergetic amounts of triacylglycerol or glucose on substrate

  12. Muscle triacylglycerol and hormone-sensitive lipase activity in untrained and trained human muscles

    DEFF Research Database (Denmark)

    Helge, Jørn Wulff; Biba, Taus O; Galbo, Henrik

    2006-01-01

    During exercise, triacylglycerol (TG) is recruited in skeletal muscles. We hypothesized that both muscle hormone-sensitive lipase (HSL) activity and TG recruitment would be higher in trained than in untrained subjects in response to prolonged exercise. Healthy male subjects (26 +/- 1 years, body ...

  13. The transport of DDT from chylomicrons to adipocytes does not mimic triacylglycerol transport

    Science.gov (United States)

    Kohan, Alison B.; Vandersall, Abbey E.; Yang, Qing; Xu, Min; Jandacek, Ronald J.; Tso, Patrick

    2012-01-01

    Despite being banned in the U.S., organochlorine toxins such as DDT are frequently detected in human adipose tissue. The main route of exposure is through the consumption of contaminated foods and subsequent intestinal packaging of DDT into chylomicrons. These chylomicrons, which also contain dietary triacylglycerol (TG), are delivered directly to peripheral tissues without first being metabolized by the liver. The physiological process by which these compounds are delivered from chylomicrons to adipose is not well understood, but is clinically relevant since it bypasses first-pass metabolism. Based on its highly lipophilic nature, it has been assumed that DDT is transferred to peripheral tissues similar to TG; however, this has not been measured. Here, we use the lymph fistula rat to isolate chylomicrons containing both DDT and TG. These chylomicrons are the in vivo DDT delivery vehicle. Using 3T3-L1 adipocytes, we investigated the rate at which DDT transfers from chylomicrons to adipocytes, and mediators of this process. This novel approach closely approximates the in vivo DDT exposure route. We show that: 1) DDT repartitions from chylomicrons to adipocytes, 2) this transport does not require hydrolysis of TG within the chylomicron, and is stimulated by the inhibition of LPL, 3) albumin does not inhibit DDT uptake, 4) DDT dissolved in DMSO does not appropriately mimic in vivo DDT transport; and most importantly, 5) DDT uptake from chylomicrons does not mimic the uptake of TG from the same particles. Understanding these factors is important for designing interventions for human populations exposed to DDT. PMID:22885168

  14. Diversity and Versatility of Phagocytosis: Roles in Innate Immunity, Tissue Remodeling, and Homeostasis.

    Science.gov (United States)

    Lim, Justin J; Grinstein, Sergio; Roth, Ziv

    2017-01-01

    Phagocytosis, a critical early event in the microbicidal response of neutrophils, is now appreciated to serve multiple functions in a variety of cell types. Professional phagocytes play a central role in innate immunity by eliminating pathogenic bacteria, fungi and malignant cells, and contribute to adaptive immunity by presenting antigens to lymphocytes. In addition, phagocytes play a part in tissue remodeling and maintain overall homeostasis by disposing of apoptotic cells, a task shared by non-professional phagocytes, often of epithelial origin. This functional versatility is supported by a vast array of receptors capable of recognizing a striking variety of foreign and endogenous ligands. Here we present an abbreviated overview of the different types of phagocytes, their varied modes of signaling and particle engulfment, and the multiple physiological roles of phagocytosis.

  15. Depression of efficiency of neutrophils for Candida albicans phagocytosis in personnel working in radiation field

    International Nuclear Information System (INIS)

    Hassan, A.A.

    2000-01-01

    The neutrophil functions, chemotaxis (direct and random migration), phagocytosis using Candida albicans (percent, index), phagocytosis by NBT (percent, score) and adherence were studied on 55 persons working in radiation field (group I) and 40 persons as control (group II). The effect of radiation on blood picture of persons working in this field with special references to leucocytic counts and neutrophil functions was studied. White and red cells counts were 6.275 +- 1.723 and 5.475 +- 1.039 (group I) and 6.440 +- 1.556, 4.704 +- 0.734 for group II, respectively with no significant difference, while in neutrophil function there was a statistically significant difference in all functions between two groups (P < 0.01). This indicates the importance of neutrophil functions in following up persons working in radiation field

  16. Influence of Corynebacterium parvum on the phagocytosis of /sup 198/Au colloids in rats

    Energy Technology Data Exchange (ETDEWEB)

    Bergoc, R.M.; Bianchin, A.M.; Caro, R.A.; Ihlo, J.E.; Rivera, E.S. (Buenos Aires Univ. Nacional (Argentina). Facultad de Farmacia y Bioquimica)

    1982-07-01

    The kinetics of the phagocytosis of gelatin-protected /sup 198/Au colloids in Wistar rats treated with Corynebacterium Parvum (CBP), was studied in order to explain its mechanism of immunomodulation. A previously developed extracorporeal blood circulation technique was used. The changes in the rate of phagocytosis, v, after the administration of CBP, for a dose of the /sup 198/Au colloid smaller or higher than the substratum constant, were studied. In the first case, no significant changes of v were observed; in the second case, significant increases of v were determined, which reached a maximum 6 days after the CBP administration. The kinetic analysis of the obtained data indicates that the action of CBP is exerted on the stage of the entrance of the colloidal particle into the reticuloendothelial cell.

  17. Influence of Corynebacterium parvum on the phagocytosis of 198Au colloids in rats

    International Nuclear Information System (INIS)

    Bergoc, R.M.; Bianchin, A.M.; Caro, R.A.; Ihlo, J.E.; Rivera, E.S.

    1982-01-01

    The kinetics of the phagocytosis of gelatin-protected 198 Au colloids in Wistar rats treated with Corynebacterium Parvum (CBP), was studied in order to explain its mechanism of immunomodulation. A previously developed extracorporeal blood circulation technique was used. The changes in the rate of phagocytosis, v, after the administration of CBP, for a dose of the 198 Au colloid smaller or higher than the substratum constant, were studied. In the first case, no significant changes of v were observed; in the second case, significant increases of v were determined, which reached a maximum 6 days after the CBP administration. The kinetic analysis of the obtained data indicates that the action of CBP is exerted on the stage of the entrance of the colloidal particle into the reticuloendothelial cell. (author) [es

  18. Neisseria meningitidis and Escherichia coli are protected from leukocyte phagocytosis by binding to erythrocyte complement receptor 1 in human blood

    DEFF Research Database (Denmark)

    Brekke, O. L.; Hellerud, B. C.; Christiansen, D.

    2011-01-01

    The initial interaction of Gram-negative bacteria with erythrocytes and its implications on leukocyte phagocytosis and oxidative burst in human whole blood were examined. Alexa-labeled Escherichia coli, wild-type H44/76 N. meningitidis and the H44/76lpxA lipopolysaccharide (LPS)-deficient mutant...... antagonist (C5aRa) and a complement receptor 1 (CR1)-blocking antibody (3D9) were examined. Most bacteria (80%) immediately bound to erythrocytes. The binding gradually declined over time, with a parallel increase in phagocytosis. Complement inhibition with compstatin reduced erythrocyte binding...... and bacterial C3 opsonization. In contrast, the C5aRa efficiently reduced phagocytosis, but did not affect the binding of bacteria to erythrocytes. The anti-CR1 blocking mAb dose-dependently reduced bacterial binding to erythrocytes to nil, with subsequent increased phagocytosis and oxidative burst. LPS had...

  19. Inhaled corticosteroid treatment for 6?months was not sufficient to normalize phagocytosis in asthmatic children

    OpenAIRE

    da Silva-Martins, Carmen L?via Faria; Couto, Shirley Claudino; Muniz-Junqueira, Maria Imaculada

    2013-01-01

    Background Corticosteroids are the first-line therapy for asthma; however, the effect of corticosteroids on the innate immune system remains unclear. This study?s objective was to evaluate the effect of inhaled corticosteroid therapy (ICT) on phagocytic functions. Methods To evaluate the impact of ICT, the phagocytosis of Saccharomyces cerevisiae by blood monocytes and neutrophils and the production of superoxide anions were assessed before and after three and six months of ICT treatment in 5...

  20. Phagocytosis by Thrombocytes is a Conserved Innate Immune Mechanism in Lower Vertebrates

    OpenAIRE

    Nagasawa, Takahiro; Nakayasu, Chihaya; Rieger, Aja M.; Barreda, Daniel R.; Somamoto, Tomonori; Nakao, Miki

    2014-01-01

    Thrombocytes, nucleated hemostatic blood cells of non-mammalian vertebrates, are regarded as the functional equivalent of anucleated mammalian platelets. Additional immune functions, including phagocytosis, have also been suggested for thrombocytes, but no conclusive molecular or cellular experimental evidence for their potential ingestion and clearance of infiltrating microbes has been provided till date. In the present study, we demonstrate the active phagocytic ability of thrombocytes in l...

  1. ITAM-like signalling for efficient phagocytosis : The paradigm of the granulocyte receptor CEACAM3

    OpenAIRE

    Pils, Stefan

    2010-01-01

    Human CEACAM3 is a tailor-made receptor of the innate immune system to fight pathogens exploiting epithelial CEACAM-family members for colonisation and invasion of their host. Previous studies established CEACAM3 as the receptor facilitating rapid phagocytosis and elimination of N. gonorrhoeae by human granulocytes. The studies reported here set out to shed light on the evolution of this highly specialised receptor and the associated signalling machinery.CEACAM3 arose from exon shuffling afte...

  2. Viral Inhibition of Bacterial Phagocytosis by Human Macrophages: Redundant Role of CD36.

    Directory of Open Access Journals (Sweden)

    Grace E Cooper

    Full Text Available Macrophages are essential to maintaining lung homoeostasis and recent work has demonstrated that influenza-infected lung macrophages downregulate their expression of the scavenger receptor CD36. This receptor has also been shown to be involved in phagocytosis of Streptococcus pneumoniae, a primary agent associated with pneumonia secondary to viral infection. The aim of this study was to investigate the role of CD36 in the effects of viral infection on macrophage phagocytic function. Human monocyte-derived macrophages (MDM were exposed to H3N2 X31 influenza virus, M37 respiratory syncytial virus (RSV or UV-irradiated virus. No infection of MDM was seen upon exposure to UV-irradiated virus but incubation with live X31 or M37 resulted in significant levels of viral detection by flow cytometry or RT-PCR respectively. Infection resulted in significantly diminished uptake of S. pneumoniae by MDM and significantly decreased expression of CD36 at both the cell surface and mRNA level. Concurrently, there was a significant increase in IFNβ gene expression in response to infection and we observed a significant decrease in bacterial phagocytosis (p = 0.031 and CD36 gene expression (p = 0.031 by MDM cultured for 24 h in 50IU/ml IFNβ. Knockdown of CD36 by siRNA resulted in decreased phagocytosis, but this was mimicked by transfection reagent alone. When MDM were incubated with CD36 blocking antibodies no effect on phagocytic ability was observed. These data indicate that autologous IFNβ production by virally-infected cells can inhibit bacterial phagocytosis, but that decreased CD36 expression by these cells does not play a major role in this functional deficiency.

  3. Alginate-Derived Oligosaccharide Inhibits Neuroinflammation and Promotes Microglial Phagocytosis of β-Amyloid

    Directory of Open Access Journals (Sweden)

    Rui Zhou

    2015-09-01

    Full Text Available Alginate from marine brown algae has been widely applied in biotechnology. In this work, the effects of alginate-derived oligosaccharide (AdO on lipopolysaccharide (LPS/β-amyloid (Aβ-induced neuroinflammation and microglial phagocytosis of Aβ were studied. We found that pretreatment of BV2 microglia with AdO prior to LPS/Aβ stimulation led to a significant inhibition of production of nitric oxide (NO and prostaglandin E2 (PGE2, expression of inducible nitric oxide synthase (iNOS and cyclooxygenase-2 (COX-2 and secretion of proinflammatory cytokines. We further demonstrated that AdO remarkably attenuated the LPS-activated overexpression of toll-like receptor 4 (TLR4 and nuclear factor (NF-κB in BV2 cells. In addition to the impressive inhibitory effect on neuroinflammation, we also found that AdO promoted the phagocytosis of Aβ through its interaction with TLR4 in microglia. Our results suggested that AdO exerted the inhibitory effect on neuroinflammation and the promotion effect on microglial phagocytosis, indicating its potential as a nutraceutical or therapeutic agent for neurodegenerative diseases, particularly Alzheimer’s disease (AD.

  4. Phagocytosis by macrophages mediated by receptors for denatured proteins - dependence on tyrosine protein kinases

    Directory of Open Access Journals (Sweden)

    M.R. Hespanhol

    2002-03-01

    Full Text Available Previous studies have demonstrated that some components of the leukocyte cell membrane, CR3 (Mac-1, CD11b/CD18 and p150/95, are able to bind to denatured proteins. Thus, it is of interest to know which effector functions of these cells can be triggered by these receptors when they interact with particles or surfaces covered with denatured proteins. In the present study we analyzed their possible role as mediators of phagocytosis of red cells covered with denatured bovine serum albumin (BSA by mouse peritoneal macrophages. We observed that a macrophages are able to recognize (bind to these red cells, b this interaction can be inhibited by denatured BSA in the fluid phase, c there is no phagocytosis of these particles by normal macrophages, d phagocytosis mediated by denatured BSA can be, however, effectively triggered in inflammatory macrophages induced by glycogen or in macrophages activated in vivo with LPS, and e this phagocytic capacity is strongly dependent on the activity of tyrosine protein kinases in its signal transduction pathway, as demonstrated by using three kinds of enzyme inhibitors (genistein, quercetin and herbimycin A.

  5. Rab20 regulates phagosome maturation in RAW264 macrophages during Fc gamma receptor-mediated phagocytosis.

    Directory of Open Access Journals (Sweden)

    Youhei Egami

    Full Text Available Rab20, a member of the Rab GTPase family, is known to be involved in membrane trafficking, however its implication in FcγR-mediated phagocytosis is unclear. We examined the spatiotemporal localization of Rab20 during phagocytosis of IgG-opsonized erythrocytes (IgG-Es in RAW264 macrophages. By the live-cell imaging of fluorescent protein-fused Rab20, it was shown that Rab20 was transiently associated with the phagosomal membranes. During the early stage of phagosome formation, Rab20 was not localized on the membranes of phagocytic cups, but was gradually recruited to the newly formed phagosomes. Although Rab20 was colocalized with Rab5 to some extent, the association of Rab20 with the phagosomes persisted even after the loss of Rab5 from the phagosomal membranes. Then, Rab20 was colocalized with Rab7 and Lamp1, late endosomal/lysosomal markers, on the internalized phagosomes. Moreover, our analysis of Rab20 mutant expression revealed that the maturation of phagosomes was significantly delayed in cells expressing the GDP-bound mutant Rab20-T19N. These data suggest that Rab20 is an important component of phagosome and regulates the phagosome maturation during FcγR-mediated phagocytosis.

  6. Effects of pentachlorophenol on survival of earthworms (Lumbricus terrestris) and phagocytosis by their immunoactive coelomocytes

    Energy Technology Data Exchange (ETDEWEB)

    Giggleman, M.A.; Fitzpatrick, L.C.; Goven, A.J. [Univ. of North Texas, Denton, TX (United States); Venables, B.J. [TRAC Labs., Denton, TX (United States)

    1998-12-01

    Earthworms, Lumbricus terrestris, exposed for 96 h to filter paper saturated with five nominal concentrations of pentachlorophenol, exhibited a 50% lethal concentration (LC50) of 25.0 {micro}g PCP/cm{sup 2} and corresponding whole worm body burden-based 50% lethal dose (LD50) of 877.7 {micro}g PCP/g dry mass. Linear regression modeling showed that worms increased body concentrations (BC = {micro}g PCP/g dry tissue mass) with increasing exposure concentrations (EC) according to BC = 113.5 + 29.5EC. Phagocytosis of yeast cells by immunoactive coelomocytes was suppressed only at body concentrations (863.3 {micro}g PCP/g dry mass) that approximated the calculated LD50 and overlapped those demonstrating lethality, indicating a sharp transition between sublethal and lethal toxicity. An exposure concentration of 15 {micro}g PCP/cm{sup 2} produced significant suppression of phagocytosis of yeast cells by immunoactive coelomocytes. However, the average measured body burden from this group approximated the estimated LD50, indicating a sharp toxic response slope. Exposure to 10 {micro}g PCP/cm{sup 2} with a corresponding body concentration of 501.3 {micro}g PCP/g dry mass did not affect phagocytosis. The importance of body burden data is emphasized.

  7. A novel flow cytometric assay for measurement of In Vivo pulmonary neutrophil phagocytosis

    Directory of Open Access Journals (Sweden)

    Gentry-Nielsen Martha J

    2006-07-01

    Full Text Available Abstract Background Phagocytosis assays are traditionally performed in vitro using polymorphonuclear leukocytes (PMNs isolated from peripheral blood or the peritoneum and heat-killed, pre-opsonized organisms. These assays may not adequately mimic the environment within the infected lung. Our laboratory therefore has developed a flow cytometric in vivo phagocytosis assay that enables quantification of PMN phagocytosis of viable bacteria within the lungs of rats. In these studies, rats are injected transtracheally with lipopolysaccharide (LPS to recruit PMNs to their lungs. They are then infected with live 5(-and 6 carboxyfluorescein diacetate succinimidyl ester (CFDA/SE labeled type 3 Streptococcus pneumoniae. Bronchoalveolar lavage is performed and resident alveolar macrophages and recruited PMNs are labeled with monoclonal antibodies specific for surface epitopes on each cell type. Three color flow cytometry is utilized to identify the cell types, quantify recruitment, and determine uptake of the labeled bacteria. Results The viability of the alveolar macrophages and PMNs isolated from the lavage fluid was >95%. The values of the percentage of PMNs in the lavage fluid as well as the percentage of PMNs associated with CFSE-labeled S. pneumoniae as measured through flow cytometry showed a high degree of correlation with the results from manual counting of cytospin slides. Conclusion This assay is suitable for measuring bacterial uptake within the infected lung. It can be adapted for use with other organisms and/or animal model systems.

  8. CD47 limits antibody dependent phagocytosis against non-malignant B cells.

    Science.gov (United States)

    Gallagher, Sandra; Turman, Sean; Lekstrom, Kristen; Wilson, Susan; Herbst, Ronald; Wang, Yue

    2017-05-01

    Recent studies have demonstrated the importance of CD47 in protecting malignant B cells from antibody dependent cellular phagocytosis (ADCP). Combined treatment of anti-CD47 and -CD20 antibodies synergistically augment elimination of tumor B cells in xenograft mouse models. This has led to the development of novel reagents that can potentially enhance killing of malignant B cells in patients. B cell depleting therapy is also a promising treatment for autoimmune patients. In the current study, we aimed to investigate whether or not CD47 protects non-malignant B cells from ADCP. We show that CD47 is expressed on all B cells in mice, with the highest level on plasma cells in bone marrow and spleen. Although its expression is dispensable for B cell development in mice, CD47 on B cells limits antibody mediated phagocytosis. B cell depletion following in vivo anti-CD19 treatment is more efficient in CD47-/- mice than in wild type mice. In vitro, both naïve and activated B cells from CD47-/- mice are more sensitive to ADCP than wild type B cells. Lastly, we show in an ADCP assay that blocking CD47 can enhance anti-CD19 antibody mediated phagocytosis of wild type B cells. These results suggest that in addition to its already demonstrated benefit in cancer, targeting CD47 may be used as an adjunct in combination with B cell depletion antibodies for treatment of autoimmune diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Red palm oil-supplemented and biofortified cassava gari increase the carotenoid and retinyl palmitate concentrations of triacylglycerol-rich plasma in women.

    Science.gov (United States)

    Zhu, Chenghao; Cai, Yimeng; Gertz, Erik R; La Frano, Michael R; Burnett, Dustin J; Burri, Betty J

    2015-11-01

    Boiled biofortified cassava containing β-carotene can increase retinyl palmitate in triacylglycerol-rich plasma. Thus, it might alleviate vitamin A deficiency. Cassava requires extensive preparation to decrease its level of cyanogenic glucosides, which can be fatal. Garification is a popular method of preparing cassava that removes cyanogen glucosides. Our objective was to compare the effectiveness of biofortified gari to gari prepared with red palm oil. The study was a randomized crossover trial in 8 American women. Three gari preparations separated by 2-week washout periods were consumed. Treatments (containing 200-225.9 g gari) were as follows: biofortified gari (containing 1 mg β-carotene), red palm oil-fortified gari (1 mg β-carotene), and unfortified gari with a 0.3-mg retinyl palmitate reference dose. Blood was collected 6 times from -0.5 to 9.5 hours after ingestion. Triacylglycerol-rich plasma was separated by ultracentrifugation and analyzed by high-performance liquid chromatography (HPLC) with diode array detection. Area under the curve for β-carotene, α-carotene, and retinyl palmitate increased after the fortified meals were fed (P palm oil treatment was greater than that induced by the biofortified treatment (P palm oil and biofortified gari, respectively. These results show that both treatments increased β-carotene, α-carotene, and retinyl palmitate in triacylglycerol-rich plasma concentrations in healthy well-nourished adult women, supporting our hypothesis that both interventions could support efforts to alleviate vitamin A deficiency. Published by Elsevier Inc.

  10. Relevance of splenic tissue preservation to bacteria phagocytosis

    International Nuclear Information System (INIS)

    Marques, Ruy Garcia; Petroianu, Andy; Oliveira, Marcia Betania Nunes de; Bernardo Filho, Mario

    2002-01-01

    Objective: Being the major lymphoid organ of human body, spleen performs critical immunological functions such as bacteria depuration from bloodstream and precocious antibody production against various antigenic particles. Spleen, liver and lung accounts for more than 95% of phagocytic activity in humans. Methods: We used intravenous inoculation of Escherichia coli AB 1157 in Wistar rats in order to evaluate this function. Young and adult rats of both sexes were submitted to total splenectomy and compared to animals not submitted to any surgical procedure. Sixteen weeks later, rats of both groups were challenged with a suspension of E. coli labeled with Tc- 99 m, and, after twenty minutes, they were killed. Liver, lung, spleen and a blood clot sample were removed for radioactivity determination. The statistical study was performed with Student.s t test. Results: There were no significant differences in the radioactivity uptake obtained from young and adult splenectomized rats. However, these rats were associated with higher levels of blood counts than animals of the control group (p<0.0001) due to a larger bacteria remnant in the bloodstream. Conclusion: This finding suggests that some failure in mononuclear phagocytic system occurs in spleen absence adaptation, ratifying the necessity of development of alternative surgical techniques to total splenectomy in patients requiring spleen removal. (author)

  11. Decoupling internalization, acidification and phagosomal-endosomal/lysosomal fusion during phagocytosis of InlA coated beads in epithelial cells.

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    Craig D Blanchette

    Full Text Available BACKGROUND: Phagocytosis has been extensively examined in 'professional' phagocytic cells using pH sensitive dyes. However, in many of the previous studies, a separation between the end of internalization, beginning of acidification and completion of phagosomal-endosomal/lysosomal fusion was not clearly established. In addition, very little work has been done to systematically examine phagosomal maturation in 'non-professional' phagocytic cells. Therefore, in this study, we developed a simple method to measure and decouple particle internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in Madin-Darby Canine Kidney (MDCK and Caco-2 epithelial cells. METHODOLOGY/PRINCIPAL FINDINGS: Our method was developed using a pathogen mimetic system consisting of polystyrene beads coated with Internalin A (InlA, a membrane surface protein from Listeria monocytogenes known to trigger receptor-mediated phagocytosis. We were able to independently measure the rates of internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in epithelial cells by combining the InlA-coated beads (InlA-beads with antibody quenching, a pH sensitive dye and an endosomal/lysosomal dye. By performing these independent measurements under identical experimental conditions, we were able to decouple the three processes and establish time scales for each. In a separate set of experiments, we exploited the phagosomal acidification process to demonstrate an additional, real-time method for tracking bead binding, internalization and phagosomal acidification. CONCLUSIONS/SIGNIFICANCE: Using this method, we found that the time scales for internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion ranged from 23-32 min, 3-4 min and 74-120 min, respectively, for MDCK and Caco-2 epithelial cells. Both the static and real-time methods developed here are expected to be readily and broadly applicable, as they simply

  12. Synthesis of structured triacylglycerols containing caproic acid by lipase-catalyzed acidolysis: Optimization by response surface methodology

    DEFF Research Database (Denmark)

    Zhou, D.Q.; Xu, Xuebing; Mu, Huiling

    2001-01-01

    Production in a batch reactor with a solvent-free system of structured triacylglycerols containing short-chain fatty acids by Lipozyme RM IM-catalyzed acidolysis between rapeseed oil and caproic acid was optimized using response surface methodology (RSM). Reaction time (t(r)), substrate ratio (S......-r = 2-6 mol/mol; and W-c = 2-12 wt %. The biocatalyst was Lipozyme RM IM, in which Rhizomucor miehei lipase is immobilized on a resin. The incorporation of caproic acid into rapeseed oil was the main monitoring response. In addition, the contents of mono-incorporated structured triacylglycerols and di......-incorporated structured triacylglycerols were also evaluated. The optimal reaction conditions for the incorporation of caproic acid and the content of di-incorporated structured triacylglycerols were as follows: t(r) = 17 h; 8, = 5; E-1 = 14 wt %; W-c = 10 wt %; T-e = 65 degreesC. At these conditions, products with 55...

  13. CD14-dependent monocyte isolation enhances phagocytosis of listeria monocytogenes by proinflammatory, GM-CSF-derived macrophages.

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    Caroline Neu

    Full Text Available Macrophages are an important line of defence against invading pathogens. Human macrophages derived by different methods were tested for their suitability as models to investigate Listeria monocytogenes (Lm infection and compared to macrophage-like THP-1 cells. Human primary monocytes were isolated by either positive or negative immunomagnetic selection and differentiated in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF or macrophage colony-stimulating factor (M-CSF into pro- or anti-inflammatory macrophages, respectively. Regardless of the isolation method, GM-CSF-derived macrophages (GM-Mφ stained positive for CD206 and M-CSF-derived macrophages (M-Mφ for CD163. THP-1 cells did not express CD206 or CD163 following incubation with PMA, M- or GM-CSF alone or in combination. Upon infection with Lm, all primary macrophages showed good survival at high multiplicities of infection whereas viability of THP-1 was severely reduced even at lower bacterial numbers. M-Mφ generally showed high phagocytosis of Lm. Strikingly, phagocytosis of Lm by GM-Mφ was markedly influenced by the method used for isolation of monocytes. GM-Mφ derived from negatively isolated monocytes showed low phagocytosis of Lm whereas GM-Mφ generated from positively selected monocytes displayed high phagocytosis of Lm. Moreover, incubation with CD14 antibody was sufficient to enhance phagocytosis of Lm by GM-Mφ generated from negatively isolated monocytes. By contrast, non-specific phagocytosis of latex beads by GM-Mφ was not influenced by treatment with CD14 antibody. Furthermore, phagocytosis of Lactococcus lactis, Escherichia coli, human cytomegalovirus and the protozoan parasite Leishmania major by GM-Mφ was not enhanced upon treatment with CD14 antibody indicating that this effect is specific for Lm. Based on these observations, we propose macrophages derived by ex vivo differentiation of negatively selected human primary monocytes as the most

  14. 76 FR 55264 - Lipase, Triacylglycerol; Exemption From the Requirement of a Tolerance

    Science.gov (United States)

    2011-09-07

    .... Potentially affected entities may include, but are not limited to: Crop production (NAICS code 111). Animal production (NAICS code 112). Food manufacturing (NAICS code 311). Pesticide manufacturing (NAICS code 32532... home uses, such as components of laundry detergents and food contact surface sanitizing solutions, are...

  15. The in vitro biocompatibility and macrophage phagocytosis of Mg17Al12 phase in Mg-Al-Zn alloys.

    Science.gov (United States)

    Liu, Chen; He, Peng; Wan, Peng; Li, Mei; Wang, Kehong; Tan, Lili; Zhang, Yu; Yang, Ke

    2015-07-01

    Mg alloys are gaining interest for applications as biodegradable medical implant, including Mg-Al-Zn series alloys with good combination of mechanical properties and reasonable corrosion resistance. However, whether the existence of second phase particles in the alloys exerts influence on the biocompatibility is still not clear. A deeper understanding of how the particles regulate specific biological responses is becoming a crucial requirement for their subsequent biomedical application. In this work, the in vitro biocompatibility of Mg17Al12 as a common second phase in biodegradable Mg-Al-Zn alloys was investigated via hemolysis, cytotoxicity, cell proliferation, and cell adhesion tests. Moreover, osteogenic differentiation was evaluated by the extracellular matrix mineralization assay. The Mg17Al12 particles were also prepared to simulate the real situation of second phase in the in vivo environment in order to estimate the cellular response in macrophages to the Mg17Al12 particles. The experimental results indicated that no hemolysis was found and an excellent cytocompatibility was also proved for the Mg17Al12 second phase when co-cultured with L929 cells, MC3T3-E1 cells and BMSCs. Macrophage phagocytosis co-culture test revealed that Mg17Al12 particles exerted no harmful effect on RAW264.7 macrophages and could be phagocytized by the RAW264.7 cells. Furthermore, the possible inflammatory reaction and metabolic way for Mg17Al12 phase were also discussed in detail. © 2014 Wiley Periodicals, Inc.

  16. Influence of ionizing irradiation to enzymatic metabolism and concentration of triacylglycerols in serum of chicken

    International Nuclear Information System (INIS)

    Novakova, J.; Danova, D.; Benova, K.; Toropila, M.; Dvorak, P.

    2006-01-01

    The investigation of the effect of ionizing irradiation helps to develop methods of protection against its negative influences. In the study we investigated the effect of low-dose ionizing irradiation on the organism of chicken. We investigated changes of activity of aspartate aminotransferase and alanine aminotransferase and changes of concentration of triacylglycerols in time gap 1, 4, 14 and 25 days after expose with a single whole-body gamma irradiation of 3 Gy. The activity of aspartate aminotransferase was significantly increased on day of 1. after irradiation and concentration of triacylglycerols was significantly increased on day of 14. and 25. after irradiation. From this study we can see by this relatively low dose of irradiation in view of higher radioresistance of chickens the organism of poultry sensitive responds with visible changes of selected biochemical parameters. (authors)

  17. Advances in silver ion chromatography for the analysis of fatty acids and triacylglycerols-2001 to 2011.

    Science.gov (United States)

    Momchilova, Svetlana M; Nikolova-Damyanova, Boryana M

    2012-01-01

    An effort is made to critically present the achievements in silver ion chromatography during the last decade. Novelties in columns, mobile-phase compositions and detectors are described. Recent applications of silver ion chromatography in the analysis of fatty acids and triacylglycerols are presented while stressing novel analytical strategies or new objects. The tendencies in the application of the method in complementary ways with reversed-phase chromatography, chiral chromatography and, especially, mass detection are outlined.

  18. Kinetic studies of Candida parapsilosis phagocytosis by macrophages and detection of intracellular survival mechanisms

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    Renata eToth

    2014-11-01

    Full Text Available Even though the number of Candida infections due to non-albicans species like C. parapsilosis has been increasing, little is known about their pathomechanisms. Certain aspects of C. parapsilosis and host interactions have already been investigated; however we lack information about the innate cellular responses towards this species. The aim of our project was to dissect and compare the phagocytosis of C. parapsilosis to C. albicans and to another Candida species C. glabrata by murine and human macrophages by live cell video microscopy. We broke down the phagocytic process into three stages: macrophage migration, engulfment of fungal cells and host cell killing after the uptake. Our results showed increased macrophage migration towards C. parapsilosis and we observed differences during the engulfment processes when comparing the three species. The engulfment time of C. parapsilosis was comparable to that of C. albicans regardless of the pseudohypha length and spatial orientation relative to phagocytes, while the rate of host cell killing and the overall uptake regarding C. parapsilosis showed similarities mainly with C. glabrata. Furthermore, we observed difference between human and murine phagocytes in the uptake of C. parapsilosis. UV-treatment of fungal cells had varied effects on phagocytosis dependent upon which Candida strain was used. Besides statistical analysis, live cell imaging videos showed that this species similarly to the other two also has the ability to survive in host cells via the following mechanisms: yeast replication, and pseudohypha growth inside of phagocytes, exocytosis of fungal cells and also abortion of host cell mitosis following the uptake. According to our knowledge this is the first study that provides a thorough examination of C. parapsilosis phagocytosis and reports intracellular survival mechanisms associated with this species.

  19. Extracellular vesicles modulate host-microbe responses by altering TLR2 activity and phagocytosis.

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    Jeroen van Bergenhenegouwen

    Full Text Available Oral delivery of Gram positive bacteria, often derived from the genera Lactobacillus or Bifidobacterium, can modulate immune function. Although the exact mechanisms remain unclear, immunomodulatory effects may be elicited through the direct interaction of these bacteria with the intestinal epithelium or resident dendritic cell (DC populations. We analyzed the immune activation properties of Lactobacilli and Bifidobacterium species and made the surprising observation that cellular responses in vitro were differentially influenced by the presence of serum, specifically the extracellular vesicle (EV fraction. In contrast to the tested Lactobacilli species, tested Bifidobacterium species induce TLR2/6 activity which is inhibited by the presence of EVs. Using specific TLR ligands, EVs were found to enhance cellular TLR2/1 and TLR4 responses while TLR2/6 responses were suppressed. No effect could be observed on cellular TLR5 responses. We determined that EVs play a role in bacterial aggregation, suggesting that EVs interact with bacterial surfaces. EVs were found to slightly enhance DC phagocytosis of Bifidobacterium breve whereas phagocytosis of Lactobacillus rhamnosus was virtually absent upon serum EV depletion. DC uptake of a non-microbial substance (dextran was not affected by the different serum fractions suggesting that EVs do not interfere with DC phagocytic capacity but rather modify the DC-microbe interaction. Depending on the microbe, combined effects of EVs on TLR activity and phagocytosis result in a differential proinflammatory DC cytokine release. Overall, these data suggest that EVs play a yet unrecognized role in host-microbe responses, not by interfering in recipient cellular responses but via attachment to, or scavenging of, microbe-associated molecular patterns. EVs can be found in any tissue or bodily fluid, therefore insights into EV-microbe interactions are important in understanding the mechanism of action of potential

  20. Cigarette Smoke Exposure Inhibits Bacterial Killing via TFEB-Mediated Autophagy Impairment and Resulting Phagocytosis Defect

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    Garrett Pehote

    2017-01-01

    Full Text Available Introduction. Cigarette smoke (CS exposure is the leading risk factor for COPD-emphysema pathogenesis. A common characteristic of COPD is impaired phagocytosis that causes frequent exacerbations in patients leading to increased morbidity. However, the underlying mechanism is unclear. Hence, we investigated if CS exposure causes autophagy impairment as a mechanism for diminished bacterial clearance via phagocytosis by utilizing murine macrophages (RAW264.7 cells and Pseudomonas aeruginosa (PA01-GFP as an experimental model. Methods. Briefly, RAW cells were treated with cigarette smoke extract (CSE, chloroquine (autophagy inhibitor, TFEB-shRNA, CFTR(inh-172, and/or fisetin prior to bacterial infection for functional analysis. Results. Bacterial clearance of PA01-GFP was significantly impaired while its survival was promoted by CSE (p<0.01, autophagy inhibition (p<0.05; p<0.01, TFEB knockdown (p<0.01; p<0.001, and inhibition of CFTR function (p<0.001; p<0.01 in comparison to the control group(s that was significantly recovered by autophagy-inducing antioxidant drug, fisetin, treatment (p<0.05; p<0.01; and p<0.001. Moreover, investigations into other pharmacological properties of fisetin show that it has significant mucolytic and bactericidal activities (p<0.01; p<0.001, which warrants further investigation. Conclusions. Our data suggests that CS-mediated autophagy impairment as a critical mechanism involved in the resulting phagocytic defect, as well as the therapeutic potential of autophagy-inducing drugs in restoring is CS-impaired phagocytosis.

  1. Polysaccharides from Ganoderma lucidum attenuate microglia-mediated neuroinflammation and modulate microglial phagocytosis and behavioural response.

    Science.gov (United States)

    Cai, Qing; Li, Yuanyuan; Pei, Gang

    2017-03-24

    Ganoderma lucidum (GL) has been widely used in Asian countries for hundreds of years to promote health and longevity. The pharmacological functions of which had been classified, including the activation of innate immune responses, suppression of tumour and modulation of cell proliferations. Effective fractions of Ganoderma lucidum polysaccharides (GLP) had already been reported to regulate the immune system. Nevertheless, the role of GLP in the microglia-mediated neuroinflammation has not been sufficiently elucidated. Further, GLP effect on microglial behavioural modulations in correlation with the inflammatory responses remains to be unravelled. The aim of this work was to quantitatively analyse the contributions of GLP on microglia. The BV2 microglia and primary mouse microglia were stimulated by lipopolysaccharides (LPS) and amyloid beta 42 (Aβ 42 ) oligomer, respectively. Investigation on the effect of GLP was carried by quantitative determination of the microglial pro- and anti-inflammatory cytokine expressions and behavioural modulations including migration, morphology and phagocytosis. Analysis of microglial morphology and phagocytosis modulations was confirmed in the zebrafish brain. Quantitative results revealed that GLP down-regulates LPS- or Aβ-induced pro-inflammatory cytokines and promotes anti-inflammatory cytokine expressions in BV-2 and primary microglia. In addition, GLP attenuates inflammation-related microglial migration, morphological alterations and phagocytosis probabilities. We also showed that modulations of microglial behavioural responses were associated with MCP-1 and C1q expressions. Overall, our study provides an insight into the GLP regulation of LPS- and Aβ-induced neuroinflammation and serves an implication that the neuroprotective function of GLP might be achieved through modulation of microglial inflammatory and behavioural responses.

  2. Scavenger Receptor C Mediates Phagocytosis of White Spot Syndrome Virus and Restricts Virus Proliferation in Shrimp

    Science.gov (United States)

    Yang, Ming-Chong; Shi, Xiu-Zhen; Yang, Hui-Ting; Sun, Jie-Jie; Xu, Ling; Wang, Xian-Wei; Zhao, Xiao-Fan

    2016-01-01

    Scavenger receptors are an important class of pattern recognition receptors that play several important roles in host defense against pathogens. The class C scavenger receptors (SRCs) have only been identified in a few invertebrates, and their role in the immune response against viruses is seldom studied. In this study, we firstly identified an SRC from kuruma shrimp, Marsupenaeus japonicus, designated MjSRC, which was significantly upregulated after white spot syndrome virus (WSSV) challenge at the mRNA and protein levels in hemocytes. The quantity of WSSV increased in shrimp after knockdown of MjSRC, compared with the controls. Furthermore, overexpression of MjSRC led to enhanced WSSV elimination via phagocytosis by hemocytes. Pull-down and co-immunoprecipitation assays demonstrated the interaction between MjSRC and the WSSV envelope protein. Electron microscopy observation indicated that the colloidal gold-labeled extracellular domain of MjSRC was located on the outer surface of WSSV. MjSRC formed a trimer and was internalized into the cytoplasm after WSSV challenge, and the internalization was strongly inhibited after knockdown of Mjβ-arrestin2. Further studies found that Mjβ-arrestin2 interacted with the intracellular domain of MjSRC and induced the internalization of WSSV in a clathrin-dependent manner. WSSV were co-localized with lysosomes in hemocytes and the WSSV quantity in shrimp increased after injection of lysosome inhibitor, chloroquine. Collectively, this study demonstrated that MjSRC recognized WSSV via its extracellular domain and invoked hemocyte phagocytosis to restrict WSSV systemic infection. This is the first study to report an SRC as a pattern recognition receptor promoting phagocytosis of a virus. PMID:28027319

  3. Biosynthesis of triacylglycerols containing very long chain monounsaturated acyl moieties in developing seeds

    International Nuclear Information System (INIS)

    Fehling, E.; Murphy, D.J.; Mukherjee, K.D.

    1990-01-01

    Particulate (15,000g) fractions from developing seeds of honesty (Lunaria annua L.) and mustard (Sinapis alba L.) synthesize radioactive very long chain monounsaturated fatty acids (gadoleic, erucic, and nervonic) from [1- 14 C]oleoyl-CoA and malonyl-CoA or from oleoyl-CoA and [2- 14 C]malonyl-CoA. The very long chain monounsaturated fatty acids are rapidly channeled to triacylglycerols and other acyl lipids without intermediate accumulation of their CoA thioesters. When [1- 14 C]oleoyl-CoA is used as the radioactive substrate, phosphatidylcholines and other phospholipids are most extensively radiolabeled by oleoyl moieties rather than by very long chain monounsaturated acyl moieties. When [2- 14 C]malonyl-CoA is used as the radioactive substrate, no radioactive oleic acid is formed and the newly synthesized very long chain monounsaturated fatty acids are extensively incorporated into phosphatidylcholines and other phospholipids as well as triacylglycerols. The pattern of labeling of the key intermediates of the Kennedy pathway, e.g. lysophosphatidic acids, phosphatidic acids, and diacylglycerols by the newly synthesized very long chain monounsaturated fatty acids is consistent with the operation of this pathway in the biosynthesis of triacylglycerols

  4. Non-degradable contrast agent with selective phagocytosis for cellular and hepatic magnetic resonance imaging

    International Nuclear Information System (INIS)

    Chen, Fei-Yan; Gu, Zhe-Jia; Zhao, Dawen; Tang, Qun

    2015-01-01

    Degradation is the long-existing toxic issue of metal-containing inorganic medicine. In this paper, we fully investigated the degradation of dextran-coated KMnF 3 nanocube in the in vitro and in vivo surroundings. Different from the general decomposing and ion releasing events, this special agent is resistant to acidic environment, as well as ion exchange. Non-degradability was proved by simulated and real cellular experiments. Moreover, it can be engulfed in the macrophage cells and kept stable in the lysosome. Due to its stability and highly selective phagocytosis, implanted liver cancer can be clearly visualized after administration

  5. Impaired neonatal macrophage phagocytosis is not explained by overproduction of prostaglandin E2

    Directory of Open Access Journals (Sweden)

    Ballinger Megan N

    2011-12-01

    Full Text Available Abstract Background Neonates and young infants manifest increased susceptibility to bacterial, viral and fungal lung infections. Previous work has identified a role for eicosanoids in mediating host defense functions of macrophages. This study examines the relationship between alveolar macrophage (AM host defense and production of lipid mediators during the neonatal period compared to adult AMs. Methods AMs were harvested from young (day 7 and day 14 and adult (~10 week rats. The functionality of these cells was assessed by examining their ability to phagocytose opsonized targets, produce cytokines, eicosanoids and intracellular cAMP measured by enzyme immunoassays, and gene expression of proteins, enzymes and receptors essential for eicosanoid generation and phagocytosis measured by real time RT-PCR. Results AMs from young animals (day 7 and 14 were defective in their ability to phagocytose opsonized targets and produce tumor necrosis factor (TNF- α. In addition, young AMs produce more prostaglandin (PG E2, a suppressor of host defense, and less leukotriene (LT B4, a promoter of host defense. Young AMs express higher levels of enzymes responsible for the production of PGE2 and LTB4; however, there was no change in the expression of E prostanoid (EP receptors or LT receptors. Despite the similar EP profiles, young AMs are more responsive to PGE2 as evidenced by their increased production of the important second messenger, cyclic AMP. In addition, young AMs express higher levels of PDE3B and lower levels of PDE4C compared to adult AMs. However, even though the young AMs produced a skewed eicosanoid profile, neither the inhibition of PGE2 by aspirin nor the addition of exogenous LTB4 rescued the defective opsonized phagocytosis. Examination of a receptor responsible for mediating opsonized phagocytosis showed a significant decrease in the gene expression levels of the Fcgamma receptor in young (day 7 AMs compared to adult AMs. Conclusion These

  6. Non-degradable contrast agent with selective phagocytosis for cellular and hepatic magnetic resonance imaging

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Fei-Yan [Nanchang University, College of Chemistry (China); Gu, Zhe-Jia [Nanchang University, Institute for Advanced Study (China); Zhao, Dawen [UT Southwestern Medical Center, Department of Radiology (United States); Tang, Qun, E-mail: tangqun@ncu.edu.cn [Nanchang University, Institute for Advanced Study (China)

    2015-09-15

    Degradation is the long-existing toxic issue of metal-containing inorganic medicine. In this paper, we fully investigated the degradation of dextran-coated KMnF{sub 3} nanocube in the in vitro and in vivo surroundings. Different from the general decomposing and ion releasing events, this special agent is resistant to acidic environment, as well as ion exchange. Non-degradability was proved by simulated and real cellular experiments. Moreover, it can be engulfed in the macrophage cells and kept stable in the lysosome. Due to its stability and highly selective phagocytosis, implanted liver cancer can be clearly visualized after administration.

  7. Determination of phagocytosis of 32P-labeled Staphylococcus aureus by bovine polymorphonuclear leukocytes

    International Nuclear Information System (INIS)

    Dulin, A.M.; Paape, M.J.; Weinland, B.T.

    1984-01-01

    A procedure for the measurement of phagocytosis by bovine polymorphonuclear leukocytes (PMN) of 32 P-labeled Staphylococcus aureus was modified so that a larger number of samples could be compared in a single run, and smaller volumes of sample, PMN, and 32 P-labeled S aureus could be used. Results were highly reproducible, with a coefficient of variation between duplicate determinations of less than or equal to 2%. Lysostaphin was prepared from the supernatant of S staphylolyticus and was compared with a commercially available preparation. Effects of lysostaphin on PMN and influence of incubation media on release of 32 P from 32 P-labeled S aureus by lysostaphin were examined

  8. Ability of Staphylococcus aureus coagulase genotypes to resist neutrophil bactericidal activity and phagocytosis

    DEFF Research Database (Denmark)

    Aarestrup, Frank Møller; Scott, N. L.; Sordillo, L. M.

    1994-01-01

    This study investigated the functional capabilities of neutrophils against different Staphylococcus aureus genotypes isolated from cows with mastitis. Six strains of S. aureus were chosen for use in the study, two with a common genotype, two with an intermediate genotype, and two with a rare......; rare type, 10.5/cell). These findings suggest that one of the reasons for the variation in prevalence of different genotypes of S. aureus in the mammary gland is due to the superior ability of some types to resist phagocytosis and/or killing by bovine neutrophils...

  9. Clearing the corpses: regulatory mechanisms, novel tools, and therapeutic potential of harnessing microglial phagocytosis in the diseased brain

    Directory of Open Access Journals (Sweden)

    Irune Diaz-Aparicio

    2016-01-01

    Full Text Available Apoptosis is a widespread phenomenon that occurs in the brain in both physiological and pathological conditions. Dead cells must be quickly removed to avoid the further toxic effects they exert in the parenchyma, a process executed by microglia, the brain professional phagocytes. Although phagocytosis is critical to maintain tissue homeostasis, it has long been either overlooked or indirectly assessed based on microglial morphology, expression of classical activation markers, or engulfment of artificial phagocytic targets in vitro. Nevertheless, these indirect methods present several limitations and, thus, direct observation and quantification of microglial phagocytosis is still necessary to fully grasp its relevance in the diseased brain. To overcome these caveats and obtain a comprehensive, quantitative picture of microglial phagocytosis we have developed a novel set of parameters. These parameters have allowed us to identify the different strategies utilized by microglia to cope with apoptotic challenges induced by excitotoxicity or inflammation. In contrast, we discovered that in mouse and human epilepsy microglia failed to find and engulf apoptotic cells, resulting in accumulation of debris and inflammation. Herein, we advocate that the efficiency of microglial phagocytosis should be routinely tested in neurodegenerative and neurological disorders, in order to determine the extent to which it contributes to apoptosis and inflammation found in these conditions. Finally, our findings point towards enhancing microglial phagocytosis as a novel therapeutic strategy to control tissue damage and inflammation, and accelerate recovery in brain diseases.

  10. Myelin down-regulates myelin phagocytosis by microglia and macrophages through interactions between CD47 on myelin and SIRPα (signal regulatory protein-α on phagocytes

    Directory of Open Access Journals (Sweden)

    Reichert Fanny

    2011-03-01

    Full Text Available Abstract Background Traumatic injury to axons produces breakdown of axons and myelin at the site of the lesion and then further distal to this where Wallerian degeneration develops. The rapid removal of degenerated myelin by phagocytosis is advantageous for repair since molecules in myelin impede regeneration of severed axons. Thus, revealing mechanisms that regulate myelin phagocytosis by macrophages and microglia is important. We hypothesize that myelin regulates its own phagocytosis by simultaneous activation and down-regulation of microglial and macrophage responses. Activation follows myelin binding to receptors that mediate its phagocytosis (e.g. complement receptor-3, which has been previously studied. Down-regulation, which we test here, follows binding of myelin CD47 to the immune inhibitory receptor SIRPα (signal regulatory protein-α on macrophages and microglia. Methods CD47 and SIRPα expression was studied by confocal immunofluorescence microscopy, and myelin phagocytosis by ELISA. Results We first document that myelin, oligodendrocytes and Schwann cells express CD47 without SIRPα and further confirm that microglia and macrophages express both CD47 and SIRPα. Thus, CD47 on myelin can bind to and subsequently activate SIRPα on phagocytes, a prerequisite for CD47/SIRPα-dependent down-regulation of CD47+/+ myelin phagocytosis by itself. We then demonstrate that phagocytosis of CD47+/+ myelin is augmented when binding between myelin CD47 and SIRPα on phagocytes is blocked by mAbs against CD47 and SIRPα, indicating that down-regulation of phagocytosis indeed depends on CD47-SIRPα binding. Further, phagocytosis in serum-free medium of CD47+/+ myelin is augmented after knocking down SIRPα levels (SIRPα-KD in phagocytes by lentiviral infection with SIRPα-shRNA, whereas phagocytosis of myelin that lacks CD47 (CD47-/- is not. Thus, myelin CD47 produces SIRPα-dependent down-regulation of CD47+/+ myelin phagocytosis in phagocytes

  11. Peripheral blood leukocyte phagocytosis and respiratory response to certain macromolecular substances in the ABCC-JNIH adult health study, Hiroshima

    Energy Technology Data Exchange (ETDEWEB)

    Barerras, R F; Finch, S C

    1974-01-01

    The functional integrity of the peripheral blood leukocytes of 10 heavily exposed subjects and 10 matched controls was evaluated by measuring oxygen consumption following the addition of latex particles and E. coli endotoxin, and measuring sensitized and unsensitized starch granule phagocytosis. There was no significant difference in the responses of leukocytes from exposed subjects and controls following the addition of latex particles. The ultimate response of leukocytes from exposed subjects to the stimulus of E. coli endotoxin was comparable to that of the controls. Depressed early response in 5 of 10 exposed and 1 of 10 control subjects was observed. Interpretation is unclear. No evidence of radiation-related impairment of starch granule phagocytosis was observed. Results fail to demonstrate any late radiation-related functional impairment of peripheral blood leukocytes during phagocytosis. The serum-related changes were inconstant and probably were of little significance. (auth)

  12. Intraocular pressure elevation precedes a phagocytosis decline in a model of pigmentary glaucoma [version 2; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Yalong Dang

    2018-04-01

    Full Text Available Background: Outflow regulation and phagocytosis are key functions of the trabecular meshwork (TM, but it is not clear how the two are related in secondary open angle glaucomas characterized by an increased particle load. We hypothesized that diminished TM phagocytosis is not the primary cause of early ocular hypertension and recreated pigment dispersion in a porcine ex vivo model. Methods: Sixteen porcine anterior chamber cultures received a continuous infusion of pigment granules (Pg, while 16 additional anterior chambers served as controls (C. Pressure transducers recorded the intraocular pressure (IOP. The phagocytic capacity of the trabecular meshwork was determined by fluorescent microspheres. Results: The baseline IOPs in Pg and C were similar (P=0.82. A significant IOP elevation occurred in Pg at 48, 120, and 180 hours (all P0.05. Conclusions: In this porcine model of pigmentary glaucoma, an IOP elevation occurs much earlier than when phagocytosis fails, suggesting that two separate mechanisms might be at work.

  13. Phagocytosis by Thrombocytes is a Conserved Innate Immune Mechanism in Lower Vertebrates.

    Science.gov (United States)

    Nagasawa, Takahiro; Nakayasu, Chihaya; Rieger, Aja M; Barreda, Daniel R; Somamoto, Tomonori; Nakao, Miki

    2014-01-01

    Thrombocytes, nucleated hemostatic blood cells of non-mammalian vertebrates, are regarded as the functional equivalent of anucleated mammalian platelets. Additional immune functions, including phagocytosis, have also been suggested for thrombocytes, but no conclusive molecular or cellular experimental evidence for their potential ingestion and clearance of infiltrating microbes has been provided till date. In the present study, we demonstrate the active phagocytic ability of thrombocytes in lower vertebrates using teleost fishes and amphibian models. Ex vivo, common carp thrombocytes were able to ingest live bacteria as well as latex beads (0.5-3 μm in diameter) and kill the bacteria. In vivo, we found that thrombocytes represented nearly half of the phagocyte population in the common carp total peripheral blood leukocyte pool. Phagocytosis efficiency was further enhanced by serum opsonization. Particle internalization led to phagolysosome fusion and killing of internalized bacteria, pointing to a robust ability for microbe elimination. We find that this potent phagocytic activity is shared across teleost (Paralichthys olivaceus) and amphibian (Xenopus laevis) models examined, implying its conservation throughout the lower vertebrate lineage. Our results provide novel insights into the dual nature of thrombocytes in the immune and homeostatic response and further provide a deeper understanding of the potential immune function of mammalian platelets based on the conserved and vestigial functions.

  14. Phagocytosis by thrombocytes is a conserved innate immune mechanism in lower vertebrates

    Directory of Open Access Journals (Sweden)

    Takahiro eNagasawa

    2014-09-01

    Full Text Available Thrombocytes, nucleated hemostatic blood cells of nonmammalian vertebrates, are regarded as the functional equivalent of anucleated mammalian platelets. Additional immune functions, including phagocytosis, have also been suggested for thrombocytes, but no conclusive molecular or cellular experimental evidence for their potential ingestion and clearance of infiltrating microbes has been provided till date. In the present study, we demonstrate the active phagocytic ability of thrombocytes in lower vertebrates using teleost fishes and amphibian models. Ex vivo, common carp thrombocytes were able to ingest live bacteria as well as latex beads (0.5–3 μm in diameter and kill the bacteria. In vivo, we found that thrombocytes represented nearly half of the phagocyte population in the common carp total peripheral blood leukocyte pool. Phagocytosis efficiency was further enhanced by serum opsonization. Particle internalization led to phagolysosome fusion and killing of internalized bacteria, pointing to a robust ability for microbe elimination. We find that this potent phagocytic activity is shared across teleost (Paralichthys olivaceus and amphibian (Xenopus laevis models examined, implying its conservation throughout the lower vertebrate lineage. Our results provide novel insights into the dual nature of thrombocytes in the immune and homeostatic response and further provide a deeper understanding of the potential immune function of mammalian platelets based on the conserved and vestigial functions.

  15. Brucella abortus-activated microglia induce neuronal death through primary phagocytosis.

    Science.gov (United States)

    Rodríguez, Ana M; Delpino, M Victoria; Miraglia, M Cruz; Costa Franco, Miriam M; Barrionuevo, Paula; Dennis, Vida A; Oliveira, Sergio C; Giambartolomei, Guillermo H

    2017-07-01

    Inflammation has long been implicated as a contributor to pathogenesis in neurobrucellosis. Many of the associated neurocognitive symptoms of neurobrucellosis may be the result of neuronal dysfunction resulting from the inflammatory response induced by Brucella abortus infection in the central nervous system. In this manuscript, we describe an immune mechanism for inflammatory activation of microglia that leads to neuronal death upon B. abortus infection. B. abortus was unable to infect or harm primary cultures of mouse neurons. However, when neurons were co-cultured with microglia and infected with B. abortus significant neuronal loss occurred. This phenomenon was dependent on TLR2 activation by Brucella lipoproteins. Neuronal death was not due to apoptosis, but it was dependent on the microglial release of nitric oxide (NO). B. abortus infection stimulated microglial proliferation, phagocytic activity and engulfment of neurons. NO secreted by B. abortus-activated microglia induced neuronal exposure of the "eat-me" signal phosphatidylserine (PS). Blocking of PS-binding to protein milk fat globule epidermal growth factor-8 (MFG-E8) or microglial vitronectin receptor-MFG-E8 interaction was sufficient to prevent neuronal loss by inhibiting microglial phagocytosis without affecting their activation. Taken together, our results indicate that B. abortus is not directly toxic to neurons; rather, these cells become distressed and are killed by phagocytosis in the inflammatory surroundings generated by infected microglia. Neuronal loss induced by B. abortus-activated microglia may explain, in part, the neurological deficits observed during neurobrucellosis. © 2017 Wiley Periodicals, Inc.

  16. Effects of aflibercept on primary RPE cells: toxicity, wound healing, uptake and phagocytosis.

    Science.gov (United States)

    Klettner, Alexa; Tahmaz, Nihat; Dithmer, Michaela; Richert, Elisabeth; Roider, Johann

    2014-10-01

    Anti-VEGF treatment is the therapy of choice in age-related macular degeneration, and is also applied in diabetic macular oedema or retinal vein occlusion. Recently, the fusion protein, aflibercept, has been approved for therapeutic use. In this study, we investigate the effects of aflibercept on primary RPE cells. Primary RPE cells were prepared from freshly slaughtered pigs' eyes. The impact of aflibercept on cell viability was investigated with MTT and trypan blue exclusion assay. The influence of aflibercept on wound healing was assessed with a scratch assay. Intracellular uptake of aflibercept was investigated in immunohistochemistry and its influence on phagocytosis with a phagocytosis assay using opsonised latex beads. Aflibercept displays no cytotoxicity on RPE cells but impairs its wound healing ability. It is taken up into RPE cells and can be intracellularly detected for at least 7 days. Intracellular aflibercept impairs the phagocytic capacity of RPE cells. Aflibercept interferes with the physiology of RPE cells, as it is taken up into RPE cells, which is accompanied by a reduction of the phagocytic ability. Additionally, it impairs the wound healing capacity of RPE cells. These effects on the physiology of RPE cells may indicate possible side effects. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

  17. Macrophage phagocytosis alters the MRI signal of ferumoxytol-labeled mesenchymal stromal cells in cartilage defects

    Science.gov (United States)

    Nejadnik, Hossein; Lenkov, Olga; Gassert, Florian; Fretwell, Deborah; Lam, Isaac; Daldrup-Link, Heike E.

    2016-05-01

    Human mesenchymal stem cells (hMSCs) are a promising tool for cartilage regeneration in arthritic joints. hMSC labeling with iron oxide nanoparticles enables non-invasive in vivo monitoring of transplanted cells in cartilage defects with MR imaging. Since graft failure leads to macrophage phagocytosis of apoptotic cells, we evaluated in vitro and in vivo whether nanoparticle-labeled hMSCs show distinct MR signal characteristics before and after phagocytosis by macrophages. We found that apoptotic nanoparticle-labeled hMSCs were phagocytosed by macrophages while viable nanoparticle-labeled hMSCs were not. Serial MRI scans of hMSC transplants in arthritic joints of recipient rats showed that the iron signal of apoptotic, nanoparticle-labeled hMSCs engulfed by macrophages disappeared faster compared to viable hMSCs. This corresponded to poor cartilage repair outcomes of the apoptotic hMSC transplants. Therefore, rapid decline of iron MRI signal at the transplant site can indicate cell death and predict incomplete defect repair weeks later. Currently, hMSC graft failure can be only diagnosed by lack of cartilage defect repair several months after cell transplantation. The described imaging signs can diagnose hMSC transplant failure more readily, which could enable timely re-interventions and avoid unnecessary follow up studies of lost transplants.

  18. Identification of intracellular phospholipases A2 in the human eye: involvement in phagocytosis of photoreceptor outer segments

    DEFF Research Database (Denmark)

    Kolko, Miriam; Wang, Jinmei; Zhan, Chen

    2007-01-01

    PURPOSE: To identify intracellular phospholipases A(2) (PLA(2)) in the human retina and to explore the role of these enzymes in human retinal pigment epithelium (RPE) phagocytosis of photoreceptor outer segments (POS). METHODS: PCR amplification and Western blot analysis were used to identify m......)-VIA activity was found to be specifically increased 12 hours after ARPE-19 cells were fed with POS. Finally, RPE phagocytosis was inhibited by the iPLA(2)-VIA inhibitor bromoenol lactone. CONCLUSIONS: Various intracellular PLA(2) subtypes are present in the human retina. iPLA(2)-VIA may play...

  19. Analysis of conjugated linoleic acid-enriched triacylglycerol mixtures by isocratic silver-ion high-performance liquid chromatography.

    Science.gov (United States)

    Adlof, R O; Menzel, A; Dorovska-Taran, V

    2002-04-12

    Silver-ion HPLC (Ag-HPLC) was applied to the fractionation of a triacylglycerol (TAG) sample enriched (>80%) with conjugated linoleic acid (CLA). After conversion of the TAGs to fatty acid methyl esters using sodium methoxide in methanol, Ag-HPLC (dual-column; isocratic solvent system of 0.1% acetonitrile in hexane; UV detection at 233 nm) was used to determine the CLA isomer distribution (50:50 mixture of 9c 11t- and 10t,12c-18:2). Three or four Ag-HPLC columns connected in series (0.6-1.0% acetonitrile in hexane as solvent; UV detection at 206 nm) were used to analyze the sample in TAG form. Elution times for CLA-enriched TAGs averaged 30 min or less. Isocratic solvent conditions were used to eliminate the solvent equilibration times (often 30 min or more) required between sample injections when solvent programming is used. The ratio of TAGs containing three vs. only two CLA molecules was found to be approximately 3 to 1. Ag-HPLC has thus been shown to be a useful method for rapidly analyzing not only CLA isomers as esters, but also in the TAG form.

  20. Involvement of Tiam1, RhoG and ELMO2/ILK in Rac1-mediated phagocytosis in human trabecular meshwork cells.

    Science.gov (United States)

    Peotter, Jennifer L; Phillips, Jenny; Tong, Tiegang; Dimeo, Kaylee; Gonzalez, Jose M; Peters, Donna M

    2016-10-01

    We previously demonstrated that an αvβ5 integrin/FAK- mediated pathway regulated the phagocytic properties of human trabecular meshwork (HTM) cells. Here we demonstrate that this process is mediated by Rac-1 and a previously unreported signaling pathway that utilizes the Tiam1 as well as a novel ILK/RhoG/ELMO2 signaling pathway. Phagocytosis in both a TM-1 cell line and normal HTM cells was mediated by Rac1 and could be significantly decreased by >75% using the Rac1 inhibitor EHop-016. Knockdown of Rac1 in TM-1 cells also inhibited phagocytosis by 40% whereas overexpression of a constitutively active Rac1 or stimulation with PDGF increased phagocytosis by 83% and 32% respectively. Tiam1 was involved in regulating phagocytosis. Knockdown of Tiam1 inhibited phagocytosis by 72% while overexpression of Tiam1 C1199 increased phagocytosis by 75%. Other upstream effectors of Rac1 found to be involved included ELMO2, RhoG, and ILK. Knockdowns of ELMO2, ILK, and RhoG caused a reduction in phagocytosis by 51%, 55% and 46% respectively. In contrast, knockdown of Vav2 and Dock1 or overexpression of Vav2 Y159/172F did not cause a significant change in phagocytosis. These data suggest a novel link between Tiam1 and RhoG/ILK /ELMO2 pathway as upstream effectors of the Rac1-mediated phagocytic process in TM cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. P2X receptor-dependent erythrocyte damage by α-hemolysin from Escherichia coli triggers phagocytosis by THP-1 cells

    DEFF Research Database (Denmark)

    Fagerberg, Steen Kåre; Skals, Marianne Gerberg; Leipziger, Jens Georg

    2013-01-01

    , which is known to be a keen trigger for phagocytosis. We hypothesize that exposure to HlyA elicits removal of the damaged erythrocytes by phagocytic cells. Cultured THP-1 cells as a model for erythrocytal phagocytosis was verified by a variety of methods, including live cell imaging. We consistently...

  2. The assessment of serum-mediated phagocytosis of necrotic material by polymorphonuclear leukocytes to diagnose and predict the clinical features of systemic lupus erythematosus

    DEFF Research Database (Denmark)

    Compagno, Michele; Gullstrand, Birgitta; Jacobsen, Søren

    2016-01-01

    BACKGROUND: Serum-mediated phagocytosis of antibody- and complement-opsonized necrotic cell material (NCM) by polymorphonuclear leukocytes can be quantified by using a flow cytometry-based assay. The phagocytosis of necrotic cell material (PNC) assay parallels the well-known lupus erythematosus c...

  3. n-3 PUFA Esterified to Glycerol or as Ethyl Esters Reduce Non-Fasting Plasma Triacylglycerol in Subjects with Hypertriglyceridemia

    DEFF Research Database (Denmark)

    Hedengran, Anne; Szecsi, Pal B; Dyerberg, Jørn

    2015-01-01

    To date, treatment of hypertriglyceridemia with long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) has been investigated solely in fasting and postprandial subjects. However, non-fasting triacylglycerols are more strongly associated with risk of cardiovascular disease. The objective of this st......To date, treatment of hypertriglyceridemia with long-chain n-3 polyunsaturated fatty acids (n-3 PUFA) has been investigated solely in fasting and postprandial subjects. However, non-fasting triacylglycerols are more strongly associated with risk of cardiovascular disease. The objective...... of this study was to investigate the effect of long-chain n-3 PUFA on non-fasting triacylglycerol levels and to compare the effects of n-3 PUFA formulated as acylglycerol (AG-PUFA) or ethyl esters (EE-PUFA). The study was a double-blinded randomized placebo-controlled interventional trial, and included 120...... subjects with non-fasting plasma triacylglycerol levels of 1.7-5.65 mmol/L (150-500 mg/dL). The participants received approximately 3 g/day of AG-PUFA, EE-PUFA, or placebo for a period of eight weeks. The levels of non-fasting plasma triacylglycerols decreased 28 % in the AG-PUFA group and 22 % in the EE...

  4. Antibody-mediated phagocytosis contributes to the anti-tumor activity of the therapeutic antibody daratumumab in lymphoma and multiple myeloma

    DEFF Research Database (Denmark)

    Overdijk, M. B.; Verploegen, S.; Bogels, M.

    2015-01-01

    Daratumumab (DARA) is a human CD38-specific IgG1 antibody that is in clinical development for the treatment of multiple myeloma (MM). The potential for IgG1 antibodies to induce macrophage-mediated phagocytosis, in combination with the known presence of macrophages in the tumor microenvironment...... in MM and other hematological tumors, led us to investigate the contribution of antibody-dependent, macrophage-mediated phagocytosis to DARA's mechanism of action. Live cell imaging revealed that DARA efficiently induced macrophage-mediated phagocytosis, in which individual macrophages rapidly...... and sequentially engulfed multiple tumor cells. DARA-dependent phagocytosis by mouse and human macrophages was also observed in an in vitro flow cytometry assay, using a range of MM and Burkitt's lymphoma cell lines. Phagocytosis contributed to DARA's anti-tumor activity in vivo, in both a subcutaneous...

  5. SUCROSE TRANSPORTER 5 supplies Arabidopsis embryos with biotin and affects triacylglycerol accumulation

    Science.gov (United States)

    Pommerrenig, Benjamin; Popko, Jennifer; Heilmann, Mareike; Schulmeister, Sylwia; Dietel, Katharina; Schmitt, Bianca; Stadler, Ruth; Feussner, Ivo; Sauer, Norbert

    2013-01-01

    The Arabidopsis SUC5 protein represents a classical sucrose/H+ symporter. Functional analyses previously revealed that SUC5 also transports biotin, an essential co-factor for fatty acid synthesis. However, evidence for a dual role in transport of the structurally unrelated compounds sucrose and biotin in plants was lacking. Here we show that SUC5 localizes to the plasma membrane, and that the SUC5 gene is expressed in developing embryos, confirming the role of the SUC5 protein as substrate carrier across apoplastic barriers in seeds. We show that transport of biotin but not of sucrose across these barriers is impaired in suc5 mutant embryos. In addition, we show that SUC5 is essential for the delivery of biotin into the embryo of biotin biosynthesis-defective mutants (bio1 and bio2). We compared embryo and seedling development as well as triacylglycerol accumulation and fatty acid composition in seeds of single mutants (suc5, bio1 or bio2), double mutants (suc5 bio1 and suc5 bio2) and wild-type plants. Although suc5 mutants were like the wild-type, bio1 and bio2 mutants showed developmental defects and reduced triacylglycerol contents. In suc5 bio1 and suc5 bio2 double mutants, developmental defects were severely increased and the triacylglycerol content was reduced to a greater extent in comparison to the single mutants. Supplementation with externally applied biotin helped to reduce symptoms in both single and double mutants, but the efficacy of supplementation was significantly lower in double than in single mutants, showing that transport of biotin into the embryo is lower in the absence of SUC5. PMID:23031218

  6. The effect of dietary sugars on triacylglycerol metabolism in subjects at increased risk of metabolic syndrome.

    OpenAIRE

    Marino, Andrea

    2016-01-01

    Background: High sugar diet may increase plasma triacylglycerol (TG) levels and cause dyslipidaemia, resulting in a higher cardiometabolic risk. High sugar intake may also promote the accumulation of ectopic fat in the liver. Objectives: To determine the effect of two isocaloric diets, low and high in extrinsic sugars (6% or 26% total energy respectively corresponding to the lower and upper 2.5th percentile of the intake in men aged 40-65 in the UK) but with the same total carbohydrate co...

  7. Lymphatic fatty acids in canine with pharmaceutical formulations containing structured triacylglycerols

    DEFF Research Database (Denmark)

    Holm, R.; Porsgaard, Trine; Porter, C.J.H.

    2006-01-01

    The intramolecular structure of dietary triacylglycerols (TAG) influences absorption. In this study, two different pharmaceutical formulations were compared containing TAG differing in fatty acid profiles and intramolecular structures: LML and MLM, where M represented medium-chain fatty acids (MCFA...... was generally higher than from the animals dosed with the MLM vehicle; however, statistically significant differences were only found for 18:0 and 18:3n-3. In conclusion, these results indicated that the fatty acid profile and intramolecular structure of administered TAG influenced the absorption of fatty acids...

  8. Triacylglycerol Accumulation is not primarily affected in Myotubes established from Type 2 Diabetic Subjects

    DEFF Research Database (Denmark)

    Gaster, Michael; Beck-Nielsen, Henning

    2006-01-01

    In the present study, we investigated triacylglycerol (TAG) accumulation, glucose and fatty acid (FA) uptake, and glycogen synthesis (GS) in human myotubes from healthy, lean, and obese subjects with and without type 2 diabetes (T2D), exposed to increasing palmitate (PA) and oleate (OA...... uptake (P0.05). These results indicate that (1) TAG accumulation is not primarily affected in skeletal muscle tissue of obese and T2D; (2) induced inhibition of oxidative phosphorylation is followed by TAG accumulation...... in skeletal muscle of obese and T2D subjects is adaptive....

  9. Tick Thioester-Containing Proteins and Phagocytosis Do Not Affect Transmission of Borrelia afzelii from the Competent Vector Ixodes ricinus

    Czech Academy of Sciences Publication Activity Database

    Urbanová, V.; Hajdušek, O.; Hönig Mondeková, Helena; Šíma, R.; Kopáček, P.

    2017-01-01

    Roč. 7, MAR 16 (2017), s. 1-16, č. článku 73. ISSN 2235-2988 Institutional support: RVO:61388971 Keywords : Borrelia * complement * phagocytosis Subject RIV: EE - Microbiology, Virology OBOR OECD: Microbiology Impact factor: 4.300, year: 2016

  10. Effects of early-life lead exposure on oxidative status and phagocytosis activity in great tits (Parus major)

    NARCIS (Netherlands)

    Rainio, Miia J.; Eeva, Tapio; Lilley, Thomas; Stauffer, Janina; Ruuskanen, Suvi

    2015-01-01

    Abstract Lead is a highly poisonous metal with a very long half-life, distributing throughout the body in blood and accumulating primarily in bones and kidney. We studied the short and long-term effects of early-life lead exposure on antioxidant defense and phagocytosis activity in a small passerine

  11. Cryptococcus neoformans Is Internalized by Receptor-Mediated or ‘Triggered’ Phagocytosis, Dependent on Actin Recruitment

    Science.gov (United States)

    Guerra, Caroline Rezende; Seabra, Sergio Henrique; de Souza, Wanderley; Rozental, Sonia

    2014-01-01

    Cryptococcosis by the encapsulated yeast Cryptococcus neoformans affects mostly immunocompromised individuals and is a frequent neurological complication in AIDS patients. Recent studies support the idea that intracellular survival of Cryptococcus yeast cells is important for the pathogenesis of cryptococcosis. However, the initial steps of Cryptococcus internalization by host cells remain poorly understood. Here, we investigate the mechanism of Cryptococcus neoformans phagocytosis by peritoneal macrophages using confocal and electron microscopy techniques, as well as flow cytometry quantification, evaluating the importance of fungal capsule production and of host cell cytoskeletal elements for fungal phagocytosis. Electron microscopy analyses revealed that capsular and acapsular strains of C. neoformans are internalized by macrophages via both ‘zipper’ (receptor-mediated) and ‘trigger’ (membrane ruffle-dependent) phagocytosis mechanisms. Actin filaments surrounded phagosomes of capsular and acapsular yeasts, and the actin depolymerizing drugs cytochalasin D and latrunculin B inhibited yeast internalization and actin recruitment to the phagosome area. In contrast, nocodazole and paclitaxel, inhibitors of microtubule dynamics decreased internalization but did not prevent actin recruitment to the site of phagocytosis. Our results show that different uptake mechanisms, dependent on both actin and tubulin dynamics occur during yeast internalization by macrophages, and that capsule production does not affect the mode of Cryptococcus uptake by host cells. PMID:24586631

  12. Protein C Inhibitor (PCI) Binds to Phosphatidylserine Exposing Cells with Implications in the Phagocytosis of Apoptotic Cells and Activated Platelets

    Science.gov (United States)

    Rieger, Daniela; Assinger, Alice; Einfinger, Katrin; Sokolikova, Barbora; Geiger, Margarethe

    2014-01-01

    Protein C Inhibitor (PCI) is a secreted serine protease inhibitor, belonging to the family of serpins. In addition to activated protein C PCI inactivates several other proteases of the coagulation and fibrinolytic systems, suggesting a regulatory role in hemostasis. Glycosaminoglycans and certain negatively charged phospholipids, like phosphatidylserine, bind to PCI and modulate its activity. Phosphatidylerine (PS) is exposed on the surface of apoptotic cells and known as a phagocytosis marker. We hypothesized that PCI might bind to PS exposed on apoptotic cells and thereby influence their removal by phagocytosis. Using Jurkat T-lymphocytes and U937 myeloid cells, we show here that PCI binds to apoptotic cells to a similar extent at the same sites as Annexin V, but in a different manner as compared to live cells (defined spots on ∼10–30% of cells). PCI dose dependently decreased phagocytosis of apoptotic Jurkat cells by U937 macrophages. Moreover, the phagocytosis of PS exposing, activated platelets by human blood derived monocytes declined in the presence of PCI. In U937 cells the expression of PCI as well as the surface binding of PCI increased with time of phorbol ester treatment/macrophage differentiation. The results of this study suggest a role of PCI not only for the function and/or maturation of macrophages, but also as a negative regulator of apoptotic cell and activated platelets removal. PMID:25000564

  13. Melanin targets LC3-associated phagocytosis (LAP): A novel pathogenetic mechanism in fungal disease.

    Science.gov (United States)

    Chamilos, Georgios; Akoumianaki, Tonia; Kyrmizi, Irene; Brakhage, Axel; Beauvais, Anne; Latge, Jean-Paul

    2016-05-03

    Intracellular swelling of conidia of the major human airborne fungal pathogen Aspergillus fumigatus results in surface exposure of immunostimulatory pathogen-associated molecular patterns (PAMPs) and triggers activation of a specialized autophagy pathway called LC3-associated phagocytosis (LAP) to promote fungal killing. We have recently discovered that, apart from PAMPs exposure, cell wall melanin removal during germination of A. fumigatus is a prerequisite for activation of LAP. Importantly, melanin promotes fungal pathogenicity via targeting LAP, as a melanin-deficient A. fumigatus mutant restores its virulence upon conditional inactivation of Atg5 in hematopoietic cells of mice. Mechanistically, fungal cell wall melanin selectively excludes the CYBA/p22phox subunit of NADPH oxidase from the phagosome to inhibit LAP, without interfering with signaling regulating cytokine responses. Notably, inhibition of LAP is a general property of melanin pigments, a finding with broad physiological implications.

  14. Plasma triacylglycerol and coagulation factor concentrations predict the anticoagulant effect of dietary fish oil in overweight subjects

    DEFF Research Database (Denmark)

    Vanschoonbeek, Kristof; Feijge, Marion A H; Saris, Wim H M

    2007-01-01

    fish-oil effects. In study 1, 54 overweight subjects consumed 3.1 g (n-3) PUFA daily. In study 2, which involved 42 overweight patients with type 2 diabetes, 20 subjects consumed (n-3) PUFA, whereas 22 others ingested a preparation rich in (n-6) PUFA. Tissue factor-induced thrombin generation (thrombin...... potential) was determined as an integrated measure of plasma coagulant activity. In both studies, multivariate analysis indicated a strong clustering of fasting concentrations of triacylglycerols, prothrombin, factor V, factor VII, and factor X with one another at baseline. This cluster of factors......-induced lowering of triacylglycerol and coagulation factor V, VII, and X concentrations, and thrombin generation. We conclude that high fasting triacylglycerol concentrations predict high procoagulant activity and a lowering of thrombin potential with dietary fish oil....

  15. In situ alcoholysis of triacylglycerols by application of switchable-polarity solvents. A new derivatization procedure for the gas-chromatographic analysis of vegetable oils.

    Science.gov (United States)

    Saliu, Francesco; Orlandi, Marco

    2013-10-01

    We describe a new use of switchable-polarity solvents for the simultaneous derivatization and extraction of triacylglycerols from vegetable oils before gas-chromatographic analysis. Different equimolecular mixtures of the commercially available amidine 1,8-diazabicyclo[5.4.0]undec-7-ene and n-alkyl alcohols were tested. Triolein was used as a model compound. Very good results were achieved by using butanol (recovery of butyl oleate was 89 ± 4%). The procedure was applied for the characterization of the fatty acid profile of different vegetable oils. No statistically significant differences from the results obtained with the application of two traditional methods were evidenced. Moreover, the use of switchable-polarity solvents showed many advantages: owing to the basicity of the amidines, no catalyst was required; the transterification reaction was conducted under mild conditions, one step and in situ; no particular matrix interferences were evidenced; the solvent was recovered.

  16. Intravenous immunglobulin binds beta amyloid and modifies its aggregation, neurotoxicity and microglial phagocytosis in vitro.

    Directory of Open Access Journals (Sweden)

    Susann Cattepoel

    Full Text Available Intravenous Immunoglobulin (IVIG has been proposed as a potential therapeutic for Alzheimer's disease (AD and its efficacy is currently being tested in mild-to-moderate AD. Earlier studies reported the presence of anti-amyloid beta (Aβ antibodies in IVIG. These observations led to clinical studies investigating the potential role of IVIG as a therapeutic agent in AD. Also, IVIG is known to mediate beneficial effects in chronic inflammatory and autoimmune conditions by interfering with various pathological processes. Therefore, we investigated the effects of IVIG and purified polyclonal Aβ-specific antibodies (pAbs-Aβ on aggregation, toxicity and phagocytosis of Aβ in vitro, thus elucidating some of the potential mechanisms of action of IVIG in AD patients. We report that both IVIG and pAbs-Aβ specifically bound to Aβ and inhibited its aggregation in a dose-dependent manner as measured by Thioflavin T assay. Additionally, IVIG and the purified pAbs-Aβ inhibited Aβ-induced neurotoxicity in the SH-SY5Y human neuroblastoma cell line and prevented Aβ binding to rat primary cortical neurons. Interestingly, IVIG and pAbs-Aβ also increased the number of phagocytosing cells as well as the amount of phagocytosed fibrillar Aβ by BV-2 microglia. Phagocytosis of Aβ depended on receptor-mediated endocytosis and was accompanied by upregulation of CD11b expression. Importantly, we could also show that Privigen dose-dependently reversed Aβ-mediated LTP inhibition in mouse hippocampal slices. Therefore, our in vitro results suggest that IVIG may have an impact on different processes involved in AD pathogenesis, thereby promoting further understanding of the effects of IVIG observed in clinical studies.

  17. The coffee diterpene cafestol increases plasma triacylglycerol by increasing the production rate of large VLDL apolipoprotein B in healthy normolipidemic subjects

    NARCIS (Netherlands)

    Roos, de B.; Caslake, M.J.; Stalenhoef, A.F.H.; Bedford, D.; Demacker, P.N.; Katan, M.B.; Packard, C.J.

    2001-01-01

    Background: Cafestol is a diterpene in unfiltered coffee that raises plasma triacylglycerol in humans. Objective: We studied whether cafestol increases plasma triacylglycerol by increasing the production rate or by decreasing the fractional catabolic rate of VLDL1 [Svedberg flotation unit (Sf)

  18. Modifications of the C terminus affect functionality and stability of yeast triacylglycerol lipase Tgl3p.

    Science.gov (United States)

    Koch, Barbara; Schmidt, Claudia; Ploier, Birgit; Daum, Günther

    2014-07-11

    Lipid droplets are specific organelles for the storage of triacylglycerols and steryl esters. They are surrounded by a phospholipid monolayer with a small but specific set of proteins embedded. Assembly and insertion of proteins into this surface membrane is an intriguing question of lipid droplet biology. To address this question we studied the topology of Tgl3p, the major triacylglycerol lipase of the yeast Saccharomyces cerevisiae, on lipid droplets. Employing the method of limited proteolysis of lipid droplet surface proteins, we found that the C terminus of Tgl3p faces the inside of the organelle, whereas the N terminus is exposed at the cytosolic side of lipid droplets. Detailed analysis of the C terminus revealed a stretch of seven amino acids that are critical for protein stability and functionality. The negative charge of two aspartate residues within this stretch is crucial for lipase activity of Tgl3p. A portion of Tgl3p, which is located to the endoplasmic reticulum, exhibits a different topology. In the phospholipid bilayer of the endoplasmic reticulum the C terminus faces the cytosol, which results in instability of the protein. Thus, the topology of Tgl3p is important for its function and strongly dependent on the membrane environment. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  19. Examination of triacylglycerol biosynthetic pathways via de novo transcriptomic and proteomic analyses in an unsequenced microalga.

    Directory of Open Access Journals (Sweden)

    Michael T Guarnieri

    Full Text Available Biofuels derived from algal lipids represent an opportunity to dramatically impact the global energy demand for transportation fuels. Systems biology analyses of oleaginous algae could greatly accelerate the commercialization of algal-derived biofuels by elucidating the key components involved in lipid productivity and leading to the initiation of hypothesis-driven strain-improvement strategies. However, higher-level systems biology analyses, such as transcriptomics and proteomics, are highly dependent upon available genomic sequence data, and the lack of these data has hindered the pursuit of such analyses for many oleaginous microalgae. In order to examine the triacylglycerol biosynthetic pathway in the unsequenced oleaginous microalga, Chlorella vulgaris, we have established a strategy with which to bypass the necessity for genomic sequence information by using the transcriptome as a guide. Our results indicate an upregulation of both fatty acid and triacylglycerol biosynthetic machinery under oil-accumulating conditions, and demonstrate the utility of a de novo assembled transcriptome as a search model for proteomic analysis of an unsequenced microalga.

  20. Overall glycemic index and glycemic load of vegan diets in relation to plasma lipoproteins and triacylglycerols.

    Science.gov (United States)

    Waldmann, Annika; Ströhle, Alexander; Koschizke, Jochen W; Leitzmann, Claus; Hahn, Andreas

    2007-01-01

    To investigate the overall glycemic index (GI), glycemic load (GL), and intake of dietary fiber, and to examine the associations between these factors and plasma lipoproteins and triacylglycerols in adult vegans in the German Vegan Study (GVS). Cross-sectional study, Germany. Healthy men (n = 67) and women (n = 87), who fulfilled the study criteria (vegan diet for >or=1 year prior to study start; minimum age of 18 years; no pregnancy/childbirth during the last 12 months) and who participated in all study segments. The average dietary GL of the GVS population was 144, and the average GI was 51.4. The adjusted geometric mean total, HDL, and LDL cholesterol concentrations decreased across the increasing quartiles of GL, carbohydrate and dietary fiber intake. The associations between total cholesterol, HDL cholesterol, LDL cholesterol and GL density and GI were inconsistent. Also, associations between GI, GL, the intake of carbohydrates, and triacylglycerol concentration were not observed. Fiber-rich vegan diets are characterized by a low GI and a low to moderate GL. The data do not support the hypothesis that a carbohydrate-rich diet per se is associated with unfavorable effects on triaclyglycerols that would be predicted to increase the risk of coronary heart disease. Copyright (c) 2007 S. Karger AG, Basel.

  1. Developmental co-expression of small molecular weight apolipoprotein B synthesis and triacylglycerol secretion

    International Nuclear Information System (INIS)

    Coleman, R.A.; Haynes, E.B.; Sand, T.M.; Davis, R.A.

    1987-01-01

    The development of the liver's ability to coordinately express the synthesis and secretion of the two major components of very low density lipoproteins (VLDL): triacylglycerol (TG) and apolipoprotein B (apo B) was examined in cultured hepatocytes obtained from fetal, suckling and adult rats. Hepatocytes from fetal and suckling rats synthesized and secreted TG at rates lower than that displayed by adult cells. When TG synthesis was equalized by adding oleic acid to the culture medium, fetal cells still secreted only 39% as much TG as did adult cells. To determine the basis for the apparent defect in VLDL assembly/secretion displayed by fetal cells, the synthesis and secretion of [ 35 S]methionine-labeled apo B was quantified by immunoprecipitation. Although adult and fetal cells synthesized and secreted large molecular weight apo B at similar rates, the synthesis and secretion of small molecular weight apo B was 2-fold greater in adult cells. These data suggest that the ability to assemble/secrete VLDL triacylglycerol varies in parallel with the developmental expression of small molecular weight apo B. Furthermore, these studies show the usefulness of the cultured rat hepatocyte model for examining the ontogeny and regulation of VLDL assembly/secretion

  2. Comparison of diet-induced thermogenesis of foods containing medium- versus long-chain triacylglycerols.

    Science.gov (United States)

    Kasai, Michio; Nosaka, Naohisa; Maki, Hideaki; Suzuki, Yoshie; Takeuchi, Hiroyuki; Aoyama, Toshiaki; Ohra, Atsushi; Harada, Youji; Okazaki, Mitsuko; Kondo, Kazuo

    2002-12-01

    The purpose of this study was to investigate the effect of 5-10 g of medium-chain triacylglycerols (MCT) on diet-induced thermogenesis in healthy humans. The study compared diet-induced thermogenesis after ingestion of test foods containing MCT and long-chain triacylglycerols (LCT), using a double-blind, crossover design. Eight male and eight female subjects participated in study 1 and study 2, respectively. In both studies, the LCT was a blend of rapeseed oil and soybean oil. In study 1, the liquid meals contained 10 g MCT (10M), a mixture of 5 g MCT and 5 g LCT (5M5L), and 10 g LCT (10L). In study 2, the subjects were given a meal (sandwich and clear soup) with the mayonnaise or margarine containing 5 g of MCT or LCT. Postprandial energy expenditure was measured by indirect calorimetry before and during the 6 h after ingestion of the test meals. Diet-induced thermogenesis was significantly greater after 5M5L and 10M Ingestion as compared to 10L ingestion. Ingestion of the mayonnaise or margarine containing 5 g MCT caused significantly larger diet-induced thermogenesis as compared to that of LCT. These results suggest that, in healthy humans, the intake of 5-10 g of MCT causes larger diet-induced thermogenesis than that of LCT, irrespective of the form of meal containing the MCT.

  3. Regioisomers of octanoic acid-containing structured triacylglycerols analyzed by tandem mass spectrometry using ammonia negative ion chemical ionization

    DEFF Research Database (Denmark)

    Kurvinen, J.P.; Mu, Huiling; Kallio, H.

    2001-01-01

    Tandem mass spectrometry based on ammonia negative ion chemical ionization and sample introduction via direct exposure probe was applied to analysis of regioisomeric structures of octanoic acid containing structured triacylglycerols (TAG) of type MML, MLM, MLL, and LML (M, medium-chain fatty acid...

  4. Triacylglycerol infusion does not improve hyperlactemia in resting patients with mitochondrial myopathy due to complex I deficiency

    NARCIS (Netherlands)

    Roef, MJ; de Meer, K; Reijngoud, DJ; Straver, HWHC; de Barse, M; Kalhan, SC; Berger, R

    Background: A high-fat diet has been recommended for correction of biochemical abnormalities and muscle energy state in patients with complex I (NADH dehydrogenase) deficiency (CID). Objective: This study evaluated the effects of intravenous infusion of isoenergetic amounts of triacylglycerol or

  5. Effect of rumen-protected choline on performance, blood metabolites, and hepatic triacylglycerols of periparturient dairy cattle

    NARCIS (Netherlands)

    Zom, R.L.G.; Baal, van J.; Goselink, R.M.A.; Bakker, J.A.; Veth, M.J.; Vuuren, van A.M.

    2011-01-01

    The effects of a dietary supplement of rumen-protected choline on feed intake, milk yield, milk composition, blood metabolites, and hepatic triacylglycerol were evaluated in periparturient dairy cows. Thirty-eight multiparous cows were blocked into 19 pairs and then randomly allocated to either one

  6. Central GLP-1 receptor signalling accelerates plasma clearance of triacylglycerol and glucose by activating brown adipose tissue in mice

    NARCIS (Netherlands)

    Kooijman, Sander; Wang, Yanan; Parlevliet, Edwin T.; Boon, Mariëtte R.; Edelschaap, David; Snaterse, Gido; Pijl, Hanno; Romijn, Johannes A.; Rensen, Patrick C. N.

    2015-01-01

    Glucagon-like peptide 1 (GLP-1) receptor (GLP-1R) agonism, used in the treatment of type 2 diabetes, has recently been shown to increase thermogenesis via the brain. As brown adipose tissue (BAT) produces heat by burning triacylglycerol (TG) and takes up glucose for de novo lipogenesis, the aim of

  7. Comparative analysis of triacylglycerols from different Stichococcus strains by RP-HPLC/APCI-MS and chiral HPLC

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Nedbalová, L.; Sigler, Karel

    2015-01-01

    Roč. 27, č. 2 (2015), s. 685-696 ISSN 0921-8971 R&D Projects: GA ČR(CZ) GAP503/11/0215 Institutional support: RVO:61388971 Keywords : Green alga * Stichococcus bacillaris * Triacylglycerols Subject RIV: EE - Microbiology, Virology Impact factor: 2.372, year: 2015

  8. Phagocytosis (cannibalism) of apoptotic neutrophils by tumor cells in gastric micropapillary carcinomas.

    Science.gov (United States)

    Barresi, Valeria; Branca, Giovanni; Ieni, Antonio; Rigoli, Luciana; Tuccari, Giovanni; Caruso, Rosario Alberto

    2015-05-14

    cytoplasmic vacuoles of tumor cells. These data suggest phagocytosis (cannibalism) of apoptotic neutrophils by micropapillary tumor cells. Tumor cell cannibalism is usually found in aggressive tumors with anaplastic morphology. Our data extend these observations to gastric micropapillary carcinoma: a tumor histotype analogously characterized by aggressive behavior and poor prognosis. The results are of interest because they raise the intriguing possibility that neutrophil cannibalism by tumor cells may be one of the mechanisms favoring tumor growth in gastric micropapillary carcinomas. This is the first study showing phagocytosis (cannibalism) of apoptotic neutrophils by tumor cells in gastric micropapillary carcinomas.

  9. Microglial phagocytosis induced by fibrillar β-amyloid is attenuated by oligomeric β-amyloid: implications for Alzheimer's disease

    Directory of Open Access Journals (Sweden)

    Lin Nan

    2011-06-01

    Full Text Available Abstract Background Reactive microglia are associated with β-amyloid (Aβ deposit and clearance in Alzhiemer's Disease (AD. Paradoxically, entocranial resident microglia fail to trigger an effective phagocytic response to clear Aβ deposits although they mainly exist in an "activated" state. Oligomeric Aβ (oAβ, a recent target in the pathogenesis of AD, can induce more potent neurotoxicity when compared with fibrillar Aβ (fAβ. However, the role of the different Aβ forms in microglial phagocytosis, induction of inflammation and oxidation, and subsequent regulation of phagocytic receptor system, remain unclear. Results We demonstrated that Aβ(1-42 fibrils, not Aβ(1-42 oligomers, increased the microglial phagocytosis. Intriguingly, the pretreatment of microglia with oAβ(1-42 not only attenuated fAβ(1-42-triggered classical phagocytic response to fluorescent microspheres but also significantly inhibited phagocytosis of fluorescent labeled fAβ(1-42. Compared with the fAβ(1-42 treatment, the oAβ(1-42 treatment resulted in a rapid and transient increase in interleukin 1β (IL-1β level and produced higher levels of tumor necrosis factor-α (TNF-α, nitric oxide (NO, prostaglandin E2 (PGE2 and intracellular superoxide anion (SOA. The further results demonstrated that microglial phagocytosis was negatively correlated with inflammatory mediators in this process and that the capacity of phagocytosis in fAβ(1-42-induced microglia was decreased by IL-1β, lippolysaccharide (LPS and tert-butyl hydroperoxide (t-BHP. The decreased phagocytosis could be relieved by pyrrolidone dithiocarbamate (PDTC, a nuclear factor-κB (NF-κB inhibitor, and N-acetyl-L-cysteine (NAC, a free radical scavenger. These results suggest that the oAβ-impaired phagocytosis is mediated through inflammation and oxidative stress-mediated mechanism in microglial cells. Furthermore, oAβ(1-42 stimulation reduced the mRNA expression of CD36, integrin β1 (Itgb1, and Ig

  10. A chloroplast pathway for the de novo biosynthesis of triacylglycerol in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Fan, J.; Xu, C.; Andre, C.

    2011-06-23

    Neutral lipid metabolism has been extensively studied in yeast, plants and mammals. In contrast, little information is available regarding the biochemical pathway, enzymes and regulatory factors involved in the biosynthesis of triacylglycerol (TAG) in microalgae. In the conventional TAG biosynthetic pathway widely accepted for yeast, plants and mammals, TAG is assembled in the endoplasmic reticulum (ER) from its immediate precursor diacylglycerol (DAG) made by ER-specific acyltransferases, and is deposited exclusively in lipid droplets in the cytosol. Here, we demonstrated that the unicellular microalga Chlamydomonas reinhardtii employs a distinct pathway that uses DAG derived almost exclusively from the chloroplast to produce TAG. This unique TAG biosynthesis pathway is largely dependent on de novo fatty acid synthesis, and the TAG formed in this pathway is stored in lipid droplets in both the chloroplast and the cytosol. These findings have wide implications for understanding TAG biosynthesis and storage and other areas of lipid metabolism in microalgae and other organisms.

  11. Characterization of non-endcapped polymeric ODS column for the separation of triacylglycerol positional isomers.

    Science.gov (United States)

    Gotoh, Naohiro; Matsumoto, Yumiko; Yuji, Hiromi; Nagai, Toshiharu; Mizobe, Hoyo; Ichioka, Kenji; Kuroda, Ikuma; Noguchi, Noriko; Wada, Shun

    2010-01-01

    The characteristics of a non-endcapped polymeric ODS column for the resolution of triacylglycerol positional isomers (TAG-PI) were examined using a recycle HPLC-atmospheric pressure chemical ionization/mass spectrometry system. A pair of TAG-PI containing saturated fatty acids at least 12 carbons was separated. Except for TAG-PI containing elaidic acid, pairs of TAG-PI containing three unsaturated fatty acids were not separated, even by recycle runs. These results indicate that the resolution of TAG-PI on a non-endcapped polymeric ODS stationary phase is realized by the recognition of the linear structure of the fatty acid and the binding position of the saturated fatty acid in TAG-PI. Chain length was also an important factor for resolution. This method may be a useful and simple for measuring the abundance ratio of TAG-PI containing saturated fatty acids in natural oils.

  12. TOR (target of rapamycin) is a key regulator of triacylglycerol accumulation in microalgae.

    Science.gov (United States)

    Imamura, Sousuke; Kawase, Yasuko; Kobayashi, Ikki; Shimojima, Mie; Ohta, Hiroyuki; Tanaka, Kan

    2016-01-01

    Most microalgae abundantly accumulate lipid droplets (LDs) containing triacylglycerols (TAGs) under several stress conditions, but the underlying molecular mechanism of this accumulation remains unclear. In a recent study, we found that inhibition of TOR (target of rapamycin), a highly conserved protein kinase of eukaryotes, by rapamycin resulted in TAG accumulation in microalgae, indicating that TOR negatively regulates TAG accumulation. Here, we show that formation of intracellular LDs and TAG accumulation were also induced in the unicellular green alga Chlamydomonas reinhardtii after exposure to Torin1 or AZD8055, which are novel TOR inhibitors that inhibit TOR activity in a manner different from rapamycin. These results supported quite well our previous conclusion that TOR is a central regulator of TAG accumulation in microalgae.

  13. Effect of Bothrops alternatus snake venom on macrophage phagocytosis and superoxide production: participation of protein kinase C

    Directory of Open Access Journals (Sweden)

    SS Setubal

    2011-01-01

    Full Text Available Envenomations caused by different species of Bothrops snakes result in severe local tissue damage, hemorrhage, pain, myonecrosis, and inflammation with a significant leukocyte accumulation at the bite site. However, the activation state of leukocytes is still unclear. According to clinical cases and experimental work, the local effects observed in envenenomation by Bothrops alternatus are mainly the appearance of edema, hemorrhage, and necrosis. In this study we investigated the ability of Bothrops alternatus crude venom to induce macrophage activation. At 6 to 100 ¼g/mL, BaV is not toxic to thioglycollate-elicited macrophages; at 3 and 6 ¼g/mL, it did not interfere in macrophage adhesion or detachment. Moreover, at concentrations of 1.5, 3, and 6 ¼g/mL the venom induced an increase in phagocytosis via complement receptor one hour after incubation. Pharmacological treatment of thioglycollate-elicited macrophages with staurosporine, a protein kinase (PKC inhibitor, abolished phagocytosis, suggesting that PKC may be involved in the increase of serum-opsonized zymosan phagocytosis induced by BaV. Moreover, BaV also induced the production of anion superoxide (O2_ by thioglycollate-elicited macrophages. This BaV stimulated superoxide production was abolished after treating the cells with staurosporine, indicating that PKC is an important signaling pathway for the production of this radical. Based on these results, we suggest that phagocytosis and reactive oxygen species are involved in the pathogenesis of local tissue damage characteristic of Bothrops spp. envenomations.

  14. Rapid Quantification of Low-Viscosity Acetyl-Triacylglycerols Using Electrospray Ionization Mass Spectrometry.

    Science.gov (United States)

    Bansal, Sunil; Durrett, Timothy P

    2016-09-01

    Acetyl-triacylglycerols (acetyl-TAG) possess an sn-3 acetate group, which confers useful chemical and physical properties to these unusual triacylglycerols (TAG). Current methods for quantification of acetyl-TAG are time consuming and do not provide any information on the molecular species profile. Electrospray ionization mass spectrometry (ESI-MS)-based methods can overcome these drawbacks. However, the ESI-MS signal intensity for TAG depends on the aliphatic chain length and unsaturation index of the molecule. Therefore response factors for different molecular species need to be determined before any quantification. The effects of the chain length and the number of double-bonds of the sn-1/2 acyl groups on the signal intensity for the neutral loss of short chain length sn-3 groups were quantified using a series of synthesized sn-3 specific structured TAG. The signal intensity for the neutral loss of the sn-3 acyl group was found to negatively correlated with the aliphatic chain length and unsaturation index of the sn-1/2 acyl groups. The signal intensity of the neutral loss of the sn-3 acyl group was also negatively correlated with the size of that chain. Further, the position of the group undergoing neutral loss was also important, with the signal from an sn-2 acyl group much lower than that from one located at sn-3. Response factors obtained from these analyses were used to develop a method for the absolute quantification of acetyl-TAG. The increased sensitivity of this ESI-MS-based approach allowed successful quantification of acetyl-TAG in various biological settings, including the products of in vitro enzyme activity assays.

  15. Impaired phagocytosis in localized aggressive periodontitis: rescue by Resolvin E1.

    Directory of Open Access Journals (Sweden)

    Gabrielle Fredman

    Full Text Available Resolution of inflammation is an active temporally orchestrated process demonstrated by the biosynthesis of novel proresolving mediators. Dysregulation of resolution pathways may underlie prevalent human inflammatory diseases such as cardiovascular diseases and periodontitis. Localized Aggressive Periodontitis (LAP is an early onset, rapidly progressing form of inflammatory periodontal disease. Here, we report increased surface P-selectin on circulating LAP platelets, and elevated integrin (CD18 surface expression on neutrophils and monocytes compared to healthy, asymptomatic controls. Significantly more platelet-neutrophil and platelet-monocyte aggregates were identified in circulating whole blood of LAP patients compared with asymptomatic controls. LAP whole blood generates increased pro-inflammatory LTB4 with addition of divalent cation ionophore A23187 (5 µM and significantly less, 15-HETE, 12-HETE, 14-HDHA, and lipoxin A(4. Macrophages from LAP subjects exhibit reduced phagocytosis. The pro-resolving lipid mediator, Resolvin E1 (0.1-100 nM, rescues the impaired phagocytic activity in LAP macrophages. These abnormalities suggest compromised resolution pathways, which may contribute to persistent inflammation resulting in establishment of a chronic inflammatory lesion and periodontal disease progression.

  16. Elasticity of nanoparticles influences their blood circulation, phagocytosis, endocytosis, and targeting.

    Science.gov (United States)

    Anselmo, Aaron C; Zhang, Mengwen; Kumar, Sunny; Vogus, Douglas R; Menegatti, Stefano; Helgeson, Matthew E; Mitragotri, Samir

    2015-03-24

    The impact of physical and chemical modifications of nanoparticles on their biological function has been systemically investigated and exploited to improve their circulation and targeting. However, the impact of nanoparticles' flexibility (i.e., elastic modulus) on their function has been explored to a far lesser extent, and the potential benefits of tuning nanoparticle elasticity are not clear. Here, we describe a method to synthesize polyethylene glycol (PEG)-based hydrogel nanoparticles of uniform size (200 nm) with elastic moduli ranging from 0.255 to 3000 kPa. These particles are used to investigate the role of particle elasticity on key functions including blood circulation time, biodistribution, antibody-mediated targeting, endocytosis, and phagocytosis. Our results demonstrate that softer nanoparticles (10 kPa) offer enhanced circulation and subsequently enhanced targeting compared to harder nanoparticles (3000 kPa) in vivo. Furthermore, in vitro experiments show that softer nanoparticles exhibit significantly reduced cellular uptake in immune cells (J774 macrophages), endothelial cells (bEnd.3), and cancer cells (4T1). Tuning nanoparticle elasticity potentially offers a method to improve the biological fate of nanoparticles by offering enhanced circulation, reduced immune system uptake, and improved targeting.

  17. Flow cytometric quantitation of phagocytosis in heparinized complete blood with latex particles and Candida albicans

    Directory of Open Access Journals (Sweden)

    Jesús M. Egido

    1997-12-01

    Full Text Available We report a rapid method for the flow cytometric quantitation of phagocytosis in heparinized complete peripherial blood (HCPB, using commercially available phycoerythrin-conjugated latex particles of 1µm diameter. The method is faster and shows greater reproducibility than Bjerknes' (1984 standard technique using propidium iodide-stained Candida albicans, conventionally applied to the leukocytic layer of peripherial blood but here modified for HCPB. We also report a modification of Bjerknes' Intracellular Killing Test to allow its application to HCPB.Se da cuenta de un método rápido para la cuantización del flujo citométrico de la fagocitosis en sangre periférica completamente heparinizada (HCPB, mediante la utilización de partículas de látex phycoerythrin-conjugadas de 1µm de diámetro disponibles comercialmente. El método es más rápido y presenta mayor reproducibilidad que la técnica estandar de Bjerknes' (1984 utilizando propidium iodide-teñida Candida albicans, aplicada convencionalmente a la capa leucocitica de sangre periférica pero modificada por HCPB. Tambien damos cuenta de una modificación de Bjerknes' Intracellular Killing Test para permitir su aplicación a HCPB.

  18. Stage-specific sampling by pattern recognition receptors during Candida albicans phagocytosis.

    Directory of Open Access Journals (Sweden)

    Sigrid E M Heinsbroek

    2008-11-01

    Full Text Available Candida albicans is a medically important pathogen, and recognition by innate immune cells is critical for its clearance. Although a number of pattern recognition receptors have been shown to be involved in recognition and phagocytosis of this fungus, the relative role of these receptors has not been formally examined. In this paper, we have investigated the contribution of the mannose receptor, Dectin-1, and complement receptor 3; and we have demonstrated that Dectin-1 is the main non-opsonic receptor involved in fungal uptake. However, both Dectin-1 and complement receptor 3 were found to accumulate at the site of uptake, while mannose receptor accumulated on C. albicans phagosomes at later stages. These results suggest a potential role for MR in phagosome sampling; and, accordingly, MR deficiency led to a reduction in TNF-alpha and MCP-1 production in response to C. albicans uptake. Our data suggest that pattern recognition receptors sample the fungal phagosome in a sequential fashion.

  19. Igg Subclasses Targeting the Flagella of Salmonella enterica Serovar Typhimurium Can Mediate Phagocytosis and Bacterial Killing

    Science.gov (United States)

    Goh, Yun Shan; Armour, Kathryn L; Clark, Michael R; Grant, Andrew J; Mastroeni, Pietro

    2016-01-01

    Invasive non-typhoidal Salmonella are a common cause of invasive disease in immuno-compromised individuals and in children. Multi-drug resistance poses challenges to disease control, with a critical need for effective vaccines. Flagellin is an attractive vaccine candidate due to surface exposure and high epitope copy number, but its potential as a target for opsonophacytic antibodies is unclear. We examined the effect of targeting flagella with different classes of IgG on the interaction between Salmonella Typhimurium and a human phagocyte-like cell line, THP-1. We tagged the FliC flagellar protein with a foreign CD52 mimotope (TSSPSAD) and bacteria were opsonized with a panel of humanised CD52 antibodies with the same antigen-binding V-region, but different constant regions. We found that IgG binding to flagella increases bacterial phagocytosis and reduces viable intracellular bacterial numbers. Opsonisation with IgG3, followed by IgG1, IgG4, and IgG2, resulted in the highest level of bacterial uptake and in the highest reduction in the intracellular load of viable bacteria. Taken together, our data provide proof-of-principle evidence that targeting flagella with antibodies can increase the antibacterial function of host cells, with IgG3 being the most potent subclass. These data will assist the rational design of urgently needed, optimised vaccines against iNTS disease. PMID:27366588

  20. Roles of phagocytosis activating protein (PAP) in Aeromonas hydrophila infected Cyprinus carpio.

    Science.gov (United States)

    Wonglapsuwan, Monwadee; Kongmee, Pataraporn; Suanyuk, Naraid; Chotigeat, Wilaiwan

    2016-06-01

    Cyprinus carpio (koi) is one of the most popular ornamental fish. A major problem for C. carpio farming is bacterial infections especially by Aeromonas hydrophila. Previously studies had shown that the Phagocytosis Activating Protein (PAP) gene was involved in the innate immune response of animals. Therefore, we attempted to identify a role for the PAP gene in the immunology of C. carpio. The expression of the PAP was found in C. carpio whole blood and increased when the fish were stimulated by inactivated A. hydrophila. In addition, PAP-phMGFP DNA was injected as an immunostimulant. The survival rate and the phagocytic index were significantly increased in the A. hydrophila infected fish that received the PAP-phMGFP DNA immunostimulant. A chitosan-PAP-phMGFP nanoparticle was then developed and feeded into fish which infected with A. hydrophila. These fish had a significantly lower mortality rate than the control. Therefore, this research confirmed a key role for PAP in protection fish from bacterial infection and the chitosan-PAP-phMGFP nanoparticle could be a good prototype for fish immunostimulant in the future. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Antibody dependent cellular phagocytosis by macrophages is a novel mechanism of action of elotuzumab.

    Science.gov (United States)

    Kurdi, Ahmed T; Glavey, Siobhan V; Bezman, Natalie A; Jhatakia, Amy; Guerriero, Jennifer L; Manier, Salomon; Moschetta, Michele; Mishima, Yuji; Roccaro, Aldo; Detappe, Alexandre; Liu, Chia-Jen; Sacco, Antonio; Huynh, Daisy; Tai, Yu-Tzu; Robbins, Michael D; Azzi, Jamil; Ghobrial, Irene M

    2018-04-13

    Elotuzumab, a recently approved antibody for the treatment of multiple myeloma (MM), has been shown to stimulate Fcγ receptor (FcγR)-mediated antibody-dependent cellular cytotoxicity (ADCC) by natural killer (NK) cells towards myeloma cells. The modulatory effects of elotuzumab on other effector cells in the tumor microenvironment, however, has not been fully explored. Antibody dependent cellular phagocytosis (ADCP) is a mechanism by which macrophages contribute to anti-tumor potency of monoclonal antibodies. Herein, we studied the NK cell independent effect of elotuzumab on tumor associated macrophages (TAMs) using a xenograft tumor model deficient in NK and adaptive immune cells. We demonstrate significant anti-tumor efficacy of single agent elotuzumab in immunocompromised xenograft models of multiple myeloma, which is in part mediated by Fc-FcγR interaction of elotuzumab with macrophages. Elotuzumab is shown in this study to induce phenotypic activation of macrophages in-vivo and mediates ADCP of myeloma cells though a FcγR dependent manner in-vitro. Together, these findings propose a novel immune mediated mechanism by which elotuzumab exerts anti-myeloma activity and helps to provide rationale for combination therapies that can enhance macrophage activity. Copyright ©2018, American Association for Cancer Research.

  2. Glycoengineering of therapeutic antibodies enhances monocyte/macrophage-mediated phagocytosis and cytotoxicity.

    Science.gov (United States)

    Herter, Sylvia; Birk, Martina C; Klein, Christian; Gerdes, Christian; Umana, Pablo; Bacac, Marina

    2014-03-01

    Therapeutic Abs possess several clinically relevant mechanisms of action including perturbation of tumor cell signaling, activation of complement-dependent cytotoxicity, Ab-dependent cellular cytotoxicity (ADCC), Ab-dependent cellular phagocytosis (ADCP), and induction of adaptive immunity. In view of the important role of phagocytic lineage cells in the mechanism of action of therapeutic Abs, we analyzed FcγR receptor-dependent effector functions of monocytes and macrophages triggered by glycoengineered (GE) Abs (having enhanced FcγRIIIa [CD16a] binding affinity) versus their wild-type (WT) counterparts under different experimental conditions. We first defined the precise FcγR repertoire on classical and nonclassical intermediate monocytes--M1 and M2c macrophage populations. We further show that WT and GE Abs display comparable binding and induce similar effector functions (ADCC and ADCP) in the absence of nonspecific, endogenous IgGs. However, in the presence of these IgGs (i.e., in a situation that more closely mimics physiologic conditions), GE Abs display significantly superior binding and promote stronger monocyte and macrophage activity. These data show that in addition to enhancing CD16a-dependent NK cell cytotoxicity, glycoengineering also enhances monocyte and macrophage phagocytic and cytotoxic activities through enhanced binding to CD16a under conditions that more closely resemble the physiologic setting.

  3. Enhanced antibody-dependent cellular phagocytosis by chimeric monoclonal antibodies with tandemly repeated Fc domains.

    Science.gov (United States)

    Nagashima, Hiroaki; Ootsubo, Michiko; Fukazawa, Mizuki; Motoi, Sotaro; Konakahara, Shu; Masuho, Yasuhiko

    2011-04-01

    We previously reported that chimeric monoclonal antibodies (mAbs) with tandemly repeated Fc domains, which were developed by introducing tandem repeats of Fc domains downstream of 2 Fab domains, augmented binding avidities for all Fcγ receptors, resulting in enhanced antibody (Ab)-dependent cellular cytotoxicity. Here we investigated regarding Ab-dependent cellular phagocytosis (ADCP) mediated by these chimeric mAbs, which is considered one of the most important mechanisms that kills tumor cells, using two-color flow cytometric methods. ADCP mediated by T3-Ab, a chimeric mAb with 3 tandemly repeated Fc domains, was 5 times more potent than that by native anti-CD20 M-Ab (M-Ab hereafter). Furthermore, T3-Ab-mediated ADCP was resistant to competitive inhibition by intravenous Ig (IVIG), although M-Ab-mediated ADCP decreased in the presence of IVIG. An Fcγ receptor-blocking study demonstrated that T3-Ab mediated ADCP via both FcγRIA and FcγRIIA, whereas M-Ab mediated ADCP exclusively via FcγRIA. These results suggest that chimeric mAbs with tandemly repeated Fc domains enhance ADCP as well as ADCC, and that Fc multimerization may significantly enhance the efficacy of therapeutic Abs. Copyright © 2010 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  4. Phagocytosis, bacterial killing, and cytokine activation of circulating blood neutrophils in horses with severe equine asthma and control horses.

    Science.gov (United States)

    Vanderstock, Johanne M; Lecours, Marie-Pier; Lavoie-Lamoureux, Annouck; Gottschalk, Marcelo; Segura, Mariela; Lavoie, Jean-Pierre; Jean, Daniel

    2018-04-01

    OBJECTIVE To evaluate in vitro phagocytosis and bactericidal activity of circulating blood neutrophils in horses with severe equine asthma and control horses and to determine whether circulating blood neutrophils in horses with severe equine asthma have an increase in expression of the proinflammatory cytokine tumor necrosis factor (TNF)-α and the chemokine interleukin (IL)-8 and a decrease in expression of the anti-inflammatory cytokine IL-10 in response to bacteria. ANIMALS 6 horses with severe equine asthma and 6 control horses. PROCEDURES Circulating blood neutrophils were isolated from horses with severe equine asthma and control horses. Phagocytosis was evaluated by use of flow cytometry. Bactericidal activity of circulating blood neutrophils was assessed by use of Streptococcus equi and Streptococcus zooepidemicus as targets, whereas the cytokine mRNA response was assessed by use of a quantitative PCR assay. RESULTS Circulating blood neutrophils from horses with severe equine asthma had significantly lower bactericidal activity toward S zooepidemicus but not toward S equi, compared with results for control horses. Phagocytosis and mRNA expression of TNF-α, IL-8, and IL-10 were not different between groups. CONCLUSIONS AND CLINCAL RELEVANCE Impairment of bactericidal activity of circulating blood neutrophils in horses with severe equine asthma could contribute to an increased susceptibility to infections.

  5. Source and role of diacylglycerol formed during phagocytosis of opsonized yeast particles and associated respiratory burst in human neutrophils

    International Nuclear Information System (INIS)

    Della Bianca, V.; Grzeskowiak, M.; Lissandrini, D.; Rossi, F.

    1991-01-01

    The results presented in this paper demonstrate that in human neutrophils phagocytosis of C3b/bi and IgG-opsonized yeast particles is associated with activation of phospholipase D and that this reaction is the main source of diglycerides. The demonstration is based upon the following findings: (1) the challenge of neutrophils with these opsonized particles was followed by a rapid formation of [3H]alkyl-phosphatidic acid [( 3H]alkyl-PA) and [3H]alkyl-diglyceride [( 3H]alkyl-DG) in cells labeled with [3H]alkyl-lyso-phosphatidylcholine; (2) in the presence of ethanol [3H]alkyl-phosphatidylethanol was formed, and accumulation of [3H]alkyl-PA and [3H]alkyl-DG was depressed; (3) propranolol, by inhibiting the dephosphorylation of [3H]alkyl-PA, completely inhibited the accumulation of [3H]alkyl-DG and depressed by about 75% the formation of diglyceride mass. Evidence is also presented that phagocytosis of C3b/bi and IgG-opsonized yeast particles and associated respiratory burst can take place independently of diglyceride formation and of the activity of this second messenger on protein kinase C. In fact: (a) propranolol while completely inhibited the formation of diglyceride mass did not modify either the phagocytosis or respiratory burst; (b) these two processes were insensitive to staurosporine

  6. Quantitation of triacylglycerols in edible oils by off-line comprehensive two-dimensional liquid chromatography-atmospheric pressure chemical ionization mass spectrometry using a single column.

    Science.gov (United States)

    Wei, Fang; Hu, Na; Lv, Xin; Dong, Xu-Yan; Chen, Hong

    2015-07-24

    In this investigation, off-line comprehensive two-dimensional liquid chromatography-atmospheric pressure chemical ionization mass spectrometry using a single column has been applied for the identification and quantification of triacylglycerols in edible oils. A novel mixed-mode phenyl-hexyl chromatographic column was employed in this off-line two-dimensional separation system. The phenyl-hexyl column combined the features of traditional C18 and silver-ion columns, which could provide hydrophobic interactions with triacylglycerols under acetonitrile conditions and can offer π-π interactions with triacylglycerols under methanol conditions. When compared with traditional off-line comprehensive two-dimensional liquid chromatography employing two different chromatographic columns (C18 and silver-ion column) and using elution solvents comprised of two phases (reversed-phase/normal-phase) for triacylglycerols separation, the novel off-line comprehensive two-dimensional liquid chromatography using a single column can be achieved by simply altering the mobile phase between acetonitrile and methanol, which exhibited a much higher selectivity for the separation of triacylglycerols with great efficiency and rapid speed. In addition, an approach based on the use of response factor with atmospheric pressure chemical ionization mass spectrometry has been developed for triacylglycerols quantification. Due to the differences between saturated and unsaturated acyl chains, the use of response factors significantly improves the quantitation of triacylglycerols. This two-dimensional liquid chromatography-mass spectrometry system was successfully applied for the profiling of triacylglycerols in soybean oils, peanut oils and lord oils. A total of 68 triacylglycerols including 40 triacylglycerols in soybean oils, 50 triacylglycerols in peanut oils and 44 triacylglycerols in lord oils have been identified and quantified. The liquid chromatography-mass spectrometry data were analyzed

  7. Reversible flow of cholesteryl ester between high-density lipoproteins and triacylglycerol-rich particles is modulated by the fatty acid composition and concentration of triacylglycerols

    Directory of Open Access Journals (Sweden)

    E.C.R. Quintão

    2010-12-01

    Full Text Available We determined the influence of fasting (FAST and feeding (FED on cholesteryl ester (CE flow between high-density lipoproteins (HDL and plasma apoB-lipoprotein and triacylglycerol (TG-rich emulsions (EM prepared with TG-fatty acids (FAs. TG-FAs of varying chain lengths and degrees of unsaturation were tested in the presence of a plasma fraction at d > 1.21 g/mL as the source of CE transfer protein. The transfer of CE from HDL to FED was greater than to FAST TG-rich acceptor lipoproteins, 18% and 14%, respectively. However, percent CE transfer from HDL to apoB-containing lipoproteins was similar for FED and FAST HDL. The CE transfer from HDL to EM depended on the EM TG-FA chain length. Furthermore, the chain length of the monounsaturated TG-containing EM showed a significant positive correlation of the CE transfer from HDL to EM (r = 0.81, P < 0.0001 and a negative correlation from EM to HDL (r = -041, P = 0.0088. Regarding the degree of EM TG-FAs unsaturation, among EMs containing C18, the CE transfer was lower from HDL to C18:2 compared to C18:1 and C18:3, 17.7%, 20.7%, and 20%, respectively. However, the CE transfer from EMs to HDL was higher to C18:2 than to C18:1 and C18:3, 83.7%, 51.2%, and 46.3%, respectively. Thus, the EM FA composition was found to be the rate-limiting factor regulating the transfer of CE from HDL. Consequently, the net transfer of CE between HDL and TG-rich particles depends on the specific arrangement of the TG acyl chains in the lipoprotein particle core.

  8. Hypoglycemic depression of hepatic phagocytosis in vivo and in the in situ perfused rat liver.

    Science.gov (United States)

    Kober, P M; Filkins, J P

    1981-01-01

    Depression of the phagocytic function of the reticuloendothelial system (RES) during endotoxic hypoglycemia has been implicated in the pathogenesis of endotoxin shock. The present study evaluated the in vivo effects of hypoglycemia on RES function and assessed the effects of an vivo bout of hypoglycemia on phagocytosis in the in situ perfused rat liver. Hypoglycemia was produced in male Holtzman rats using either 1 U of regular insulin (RI) (ILETIN, Lilly) or 0.75 U of long-acting insulin (LAI) (85% LENTE/15% ULTRALENTE, Lilly). RES function was quantitated by intravascular clearance of 8 mg/100 gm body weight colloidal carbon (CC). Two hr after RI and 2.5 hr after LAI, the intravascular halftimes of CC clearance were 19 +/- 2 min (N = 22) and 18 +/- 1 min (N = 19), respectively, as compared to control, 11.3 +/- 0.4 min (N = 53, P less than 0.001). The corresponding plasma glucose (PG) levels were 95 +/- 2 mg/dl in control, 14.4 +/- 0.9 for the RI group, and 17 +/- 1 for LAI. Two hr after RI, livers were perfused for 10 min in situ with 50 mg/liter CC in saline 5% rat serum. PG for control liver donors were 90 +/- 3 mg/dl, while those for hypoglycemic liver donors were 15 +/- 2. CC uptake was decreased from 22 micrograms/min/gm liver in the control (+ serum, n = 19) to 11 +/- 2 in hypoglycemia livers (N = 6); no effect of serum on hypoglycemic depression of the RES was seen. There were no differences in flow rates in the 2 groups. These results indicate that hypoglycemia directly impairs RES function and that the in vivo depression of intravascular clearance is not related to either the presence or absence of serum factors or total hepatic blood flow. Thus, the characteristic hypoglycemia of endotoxin shock may contribute to RES depression and the lethal shock syndrome.

  9. Non-opsonic phagocytosis of Legionella pneumophila by macrophages is mediated by phosphatidylinositol 3-kinase.

    Directory of Open Access Journals (Sweden)

    Souvenir D Tachado

    Full Text Available BACKGROUND: Legionella pneumophila, is an intracellular pathogen that causes Legionnaires' disease in humans, a potentially lethal pneumonia. L. pneumophila has the ability to enter and replicate in the host and is essential for pathogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Phagocytosis was measured by cell invasion assays. Construction of PI3K mutant by PCR cloning and expression of dominant negative mutant was detected by Western blot. PI3K activity was measured by 32P labeling and detection of phospholipids products by thin layer chromatography. Infection of macrophages with virulent L. pneumophila stimulated the formation of phosphatidylinositol 3-phosphate (PIP3, a phosphorylated lipid product of PI3K whereas two structurally distinct phosphatidylinositol 3 kinase (PI3K inhibitors, wortmannin and LY294002, reduced L. pneumophila entry into macrophages in a dose-dependent fashion. Furthermore, PI3K activation led to Akt stimulation, a serine/threonine kinase, which was also inhibited by wortmannin and LY294002. In contrast, PI3K and protein kinase B (PKB/Akt activities were lower in macrophages infected with an avirulent bacterial strain. Only virulent L. pneumophila increased lipid kinase activity present in immunoprecipitates of the p85alpha subunit of class I PI3K and tyrosine phosphorylated proteins. In addition, macrophages expressing a specific dominant negative mutant of PI3K reduced L. pneumophila entry into these cells. CONCLUSION/SIGNIFICANCE: Entry of L. pneumophila is mediated by PI3K/Akt signaling pathway. These results suggest an important role for PI3K and Akt in the L. pneumophila infection process. They point to possible novel strategies for undermining L. pneumophila host uptake and reducing pathogenesis of Legionnaires' disease.

  10. Cyclic GMP-dependent protein kinase II is necessary for macrophage M1 polarization and phagocytosis via toll-like receptor 2.

    Science.gov (United States)

    Liao, Wei-Ting; You, Huey-Ling; Li, Changgui; Chang, Jan-Gowth; Chang, Shun-Jen; Chen, Chung-Jen

    2015-05-01

    Cyclic GMP-dependent protein kinase II (cGKII; PRKG2) phosphorylates a variety of biological targets and has been identified as a gout-susceptible gene. However, the regulatory role of cGKII in triggering gout disease has yet to be clarified. Thus, we plan to explore the specific function of cGKII in macrophages related to gout disease. By using cGKII gene knockdown method, we detected macrophage M1/M2 polarization, phagocytosis, and their responses to stimulation by monosodium urate (MSU). cGKII was highly expressed in M1 phenotype, but not in M2, and cGKII knockdown significantly inhibited macrophage M1 polarization by decreasing M1 chemokine markers (CXCL10 and CCL2) and downregulating phagocytosis function. We further identified that cGKII-associated phagocytosis was mediated by upregulating toll-like receptor 2 (TLR2) expression, but not by TLR4. Mimicking gout condition by MSU treatments, we found that MSU alone induced cGKII and TLR2 expression with increased M1 polarization markers and phagocytosis activity. It means that cGKII knockdown significantly inhibited this MSU-induced cGKII-TLR2-phagocytosis axis. Our study showed that cGKII plays a key role in M1 polarization, especially in TLR2-mediated phagocytosis under MSU exposure. The findings provide evidence for the possible role of cGKII as an inflammation exciter in gout disease. Gout-susceptible gene cGKII is necessary for macrophage M1 polarization. cGKII regulates M1 phagocytosis function via TLR2. Monosodium urate treatments increase cGKII expression and related function. This study reveals the role of cGKII in enhancing gouty inflammatory responses.

  11. CLA supplementation and aerobic exercise lower blood triacylglycerol, but have no effect on peak oxygen uptake or cardiorespiratory fatigue thresholds.

    Science.gov (United States)

    Jenkins, Nathaniel D M; Buckner, Samuel L; Cochrane, Kristen C; Bergstrom, Haley C; Goldsmith, Jacob A; Weir, Joseph P; Housh, Terry J; Cramer, Joel T

    2014-09-01

    This study examined the effects of 6 weeks of conjugated linoleic acid (CLA) supplementation and moderate aerobic exercise on peak oxygen uptake (VO2 peak), the gas exchange threshold (GET), the respiratory compensation point (RCP), and serum concentrations of cholesterol, triacylglycerol, and glucose in humans. Thirty-four untrained to moderately trained men (mean ± SD; age = 21.5 ± 2.8 years; mass = 77.2 ± 9.5 kg) completed this double-blind, placebo controlled study and were randomly assigned to either a CLA (Clarinol A-80; n = 18) or placebo (PLA; sunflower oil; n = 16) group. Prior to and following 6 weeks of aerobic training (50% VO2 peak for 30 min, twice per week) and supplementation (5.63 g of total CLA isomers [of which 2.67 g was c9, t11 and 2.67 g was t10, c12] or 7.35 g high oleic sunflower oil per day), each participant completed an incremental cycle ergometer test to exhaustion to determine their [Formula: see text] peak, GET, and RCP and fasted blood draws were performed to measure serum concentrations of cholesterol, triacylglycerol, and glucose. Serum triacylglycerol concentrations were lower (p interactions (p 0.05) between the CLA and PLA groups. GET and RCP increased (p < 0.05) from pre- to post-training for both the CLA and PLA groups. Overall, these data suggested that CLA and aerobic exercise may have synergistic, blood triacylglycerol lowering effects, although CLA may be ineffective for enhancing aerobic exercise performance in conjunction with a 6-week aerobic exercise training program in college-age men.

  12. Two-Stage Enzymatic Preparation of Eicosapentaenoic Acid (EPA) And Docosahexaenoic Acid (DHA) Enriched Fish Oil Triacylglycerols.

    Science.gov (United States)

    Zhang, Zhen; Liu, Fang; Ma, Xiang; Huang, Huihua; Wang, Yong

    2018-01-10

    Fish oil products in the form of triacylglycerols generally have relatively low contents of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and so it is of potential research and industrial interest to enrich the related contents in commercial products. Thereby an economical and efficient two-stage preparation of EPA and DHA enriched fish oil triacylglycerols is proposed in this study. The first stage was the partial hydrolysis of fish oil by only 0.2 wt.‰ AY "Amano" 400SD which led to increases of EPA and DHA contents in acylglycerols from 19.30 and 13.09 wt % to 25.95 and 22.06 wt %, respectively. Subsequently, products of the first stage were subjected to transesterification with EPA and DHA enriched fatty acid ethyl esters (EDEE) as the second stage to afford EPA and DHA enriched fish oil triacylglycerols by using as low as 2 wt % Novozyme 435. EDEEs prepared from fish oil ethyl ester, and recycled DHA and EPA, respectively, were applied in this stage. Final products prepared with two different sources of EDEEs were composed of 97.62 and 95.92 wt % of triacylglycerols, respectively, with EPA and DHA contents of 28.20 and 21.41 wt % for the former and 25.61 and 17.40 wt % for the latter. Results not only demonstrate this two-stage process's capability and industrial value for enriching EPA and DHA in fish oil products, but also offer new opportunities for the development of fortified fish oil products.

  13. Early dietary intervention with structured triacylglycerols containing docosahexaenoic acid. Effect on brain, liver, and adipose tissue lipids

    DEFF Research Database (Denmark)

    Christensen, Merete Myrup; Høy, Carl-Erik

    1997-01-01

    Newborn rats were fed liquid diets containing 7 wt% fat in which 3.8% of the total fatty acids were 22:6n-3. The fats were either a specificstructured oil with 22:6n-3 mostly located in the sn-2 position or a randomized oil with 22:6n-3 equally distributed in the triacylglycerol (TAG)molecules. T...

  14. Ultrasound-assisted extraction (UAE) and solvent extraction of papaya seed oil: yield, fatty acid composition and triacylglycerol profile.

    Science.gov (United States)

    Samaram, Shadi; Mirhosseini, Hamed; Tan, Chin Ping; Ghazali, Hasanah Mohd

    2013-10-10

    The main objective of the current work was to evaluate the suitability of ultrasound-assisted extraction (UAE) for the recovery of oil from papaya seed as compared to conventional extraction techniques (i.e., Soxhlet extraction (SXE) and solvent extraction (SE)). In the present study, the recovery yield, fatty acid composition and triacylglycerol profile of papaya seed oil obtained from different extraction methods and conditions were compared. Results indicated that both solvent extraction (SE, 12 h/25 °C) and ultrasound-assisted extraction (UAE) methods recovered relatively high yields (79.1% and 76.1% of total oil content, respectively). Analysis of fatty acid composition revealed that the predominant fatty acids in papaya seed oil were oleic (18:1, 70.5%-74.7%), palmitic (16:0, 14.9%-17.9%), stearic (18:0, 4.50%-5.25%), and linoleic acid (18:2, 3.63%-4.6%). Moreover, the most abundant triacylglycerols of papaya seed oil were triolein (OOO), palmitoyl diolein (POO) and stearoyl oleoyl linolein (SOL). In this study, ultrasound-assisted extraction (UAE) significantly (p < 0.05) influenced the triacylglycerol profile of papaya seed oil, but no significant differences were observed in the fatty acid composition of papaya seed oil extracted by different extraction methods (SXE, SE and UAE) and conditions.

  15. Ultrasound-Assisted Extraction (UAE and Solvent Extraction of Papaya Seed Oil: Yield, Fatty Acid Composition and Triacylglycerol Profile

    Directory of Open Access Journals (Sweden)

    Hasanah Mohd Ghazali

    2013-10-01

    Full Text Available The main objective of the current work was to evaluate the suitability of ultrasound-assisted extraction (UAE for the recovery of oil from papaya seed as compared to conventional extraction techniques (i.e., Soxhlet extraction (SXE and solvent extraction (SE. In the present study, the recovery yield, fatty acid composition and triacylglycerol profile of papaya seed oil obtained from different extraction methods and conditions were compared. Results indicated that both solvent extraction (SE, 12 h/25 °C and ultrasound-assisted extraction (UAE methods recovered relatively high yields (79.1% and 76.1% of total oil content, respectively. Analysis of fatty acid composition revealed that the predominant fatty acids in papaya seed oil were oleic (18:1, 70.5%–74.7%, palmitic (16:0, 14.9%–17.9%, stearic (18:0, 4.50%–5.25%, and linoleic acid (18:2, 3.63%–4.6%. Moreover, the most abundant triacylglycerols of papaya seed oil were triolein (OOO, palmitoyl diolein (POO and stearoyl oleoyl linolein (SOL. In this study, ultrasound-assisted extraction (UAE significantly (p < 0.05 influenced the triacylglycerol profile of papaya seed oil, but no significant differences were observed in the fatty acid composition of papaya seed oil extracted by different extraction methods (SXE, SE and UAE and conditions.

  16. Coordination of storage lipid synthesis and membrane biogenesis: evidence for cross-talk between triacylglycerol metabolism and phosphatidylinositol synthesis.

    Science.gov (United States)

    Gaspar, Maria L; Hofbauer, Harald F; Kohlwein, Sepp D; Henry, Susan A

    2011-01-21

    Despite the importance of triacylglycerols (TAG) and steryl esters (SE) in phospholipid synthesis in cells transitioning from stationary-phase into active growth, there is no direct evidence for their requirement in synthesis of phosphatidylinositol (PI) or other membrane phospholipids in logarithmically growing yeast cells. We report that the dga1Δlro1Δare1Δare2Δ strain, which lacks the ability to synthesize both TAG and SE, is not able to sustain normal growth in the absence of inositol (Ino(-) phenotype) at 37 °C especially when choline is present. Unlike many other strains exhibiting an Ino(-) phenotype, the dga1Δlro1Δare1Δare2Δ strain does not display a defect in INO1 expression. However, the mutant exhibits slow recovery of PI content compared with wild type cells upon reintroduction of inositol into logarithmically growing cultures. The tgl3Δtgl4Δtgl5Δ strain, which is able to synthesize TAG but unable to mobilize it, also exhibits attenuated PI formation under these conditions. However, unlike dga1Δlro1Δare1Δare2Δ, the tgl3Δtgl4Δtgl5Δ strain does not display an Ino(-) phenotype, indicating that failure to mobilize TAG is not fully responsible for the growth defect of the dga1Δlro1Δare1Δare2Δ strain in the absence of inositol. Moreover, synthesis of phospholipids, especially PI, is dramatically reduced in the dga1Δlro1Δare1Δare2Δ strain even when it is grown continuously in the presence of inositol. The mutant also utilizes a greater proportion of newly synthesized PI than wild type for the synthesis of inositol-containing sphingolipids, especially in the absence of inositol. Thus, we conclude that storage lipid synthesis actively influences membrane phospholipid metabolism in logarithmically growing cells.

  17. Effect of UVB irradiation of the blood on cellular volume adherence and phagocytosis in probands and patients with multiple sclerosis

    International Nuclear Information System (INIS)

    Mix, E.; Jenssen, H.L.; Lehmitz, R.; Buddenhagen, F.; Hitzschke, B.; Richter, M.

    1988-01-01

    UVB-induced changes of blood cell properties were investigated in 12 multiple sclerosis (MS) patients and in 10 healthy volunteers. The mean cell volume (MCV) was determined by electronic sizing, the granulocyte and lymphocyte adherence was estimated in a capillary assay, and the phagocytic activity of granulocytes was measured in a test system based on the incorporation of opsonized baker's yeast. In MS patients the MCV of red cells and lymphocytes decreased rapidly within 6 UVB treatments. In contrast, the reduction of the granulocyte volume was delayed (between the 6th and 12th UVB). In the control group the mean value of the red cell and lymphocyte MCV remained rather unaffected. There was a slight rise of the granulocyte volume after the 6th UVB. The only significant change of adherence was an increase of granulocyte adherence in MS patients. Untreated patients had a significantly enhanced phagocytic activity in comparison to the control group. 6 UVB treatments induced a singificant reduction of the phagocytic activity in MS patients. However, subsequently the percentage of phagocytizing cells increased again, whereas the particle uptake per cell continued to decrease. In the control group only minor UVB-induced changes of phagocytosis were observed. The in vitro UV irradiation caused an enhanced phagocytosis in the majority of cases in both controls and MS patients. In general, under the UVB treatment all parameters examined changed in the sense of a normalisation, in that the measured values reached a new level lying between the extreme pretreatment values accompanied by a reduced standard deviation. The effect of UVB was more pronounced in MS patients when compared with normal control. This could result from an enhanced sensitivity to the influence of UVB of pathologically altered cells in MS patients. The monitoring of the MCV of red cells and lymphocytes as well as the repeated testing of granulocyte phagocytosis are recommended for supportion of therapy

  18. Pathogen-Specific Binding Soluble Down Syndrome Cell Adhesion Molecule (Dscam Regulates Phagocytosis via Membrane-Bound Dscam in Crab

    Directory of Open Access Journals (Sweden)

    Xue-Jie Li

    2018-04-01

    Full Text Available The Down syndrome cell adhesion molecule (Dscam gene is an extraordinary example of diversity that can produce thousands of isoforms and has so far been found only in insects and crustaceans. Cumulative evidence indicates that Dscam may contribute to the mechanistic foundations of specific immune responses in insects. However, the mechanism and functions of Dscam in relation to pathogens and immunity remain largely unknown. In this study, we identified the genome organization and alternative Dscam exons from Chinese mitten crab, Eriocheir sinensis. These variants, designated EsDscam, potentially produce 30,600 isoforms due to three alternatively spliced immunoglobulin (Ig domains and a transmembrane domain. EsDscam was significantly upregulated after bacterial challenge at both mRNA and protein levels. Moreover, bacterial specific EsDscam isoforms were found to bind specifically with the original bacteria to facilitate efficient clearance. Furthermore, bacteria-specific binding of soluble EsDscam via the complete Ig1–Ig4 domain significantly enhanced elimination of the original bacteria via phagocytosis by hemocytes; this function was abolished by partial Ig1–Ig4 domain truncation. Further studies showed that knockdown of membrane-bound EsDscam inhibited the ability of EsDscam with the same extracellular region to promote bacterial phagocytosis. Immunocytochemistry indicated colocalization of the soluble and membrane-bound forms of EsDscam at the hemocyte surface. Far-Western and coimmunoprecipitation assays demonstrated homotypic interactions between EsDscam isoforms. This study provides insights into a mechanism by which soluble Dscam regulates hemocyte phagocytosis via bacteria-specific binding and specific interactions with membrane-bound Dscam as a phagocytic receptor.

  19. Using an improved phagocytosis assay to evaluate the effect of HIV on specific antibodies to pregnancy-associated malaria.

    Science.gov (United States)

    Ataíde, Ricardo; Hasang, Wina; Wilson, Danny W; Beeson, James G; Mwapasa, Victor; Molyneux, Malcolm E; Meshnick, Steven R; Rogerson, Stephen J

    2010-05-25

    Pregnant women residing in malaria endemic areas are highly susceptible to Plasmodium falciparum malaria, particularly during their first pregnancy, resulting in low birth weight babies and maternal anaemia. This susceptibility is associated with placental sequestration of parasitised red blood cells expressing pregnancy-specific variant surface antigens. Acquisition of antibodies against these variant surface antigens may protect women and their offspring. Functions of such antibodies may include prevention of placental sequestration or opsonisation of parasitised cells for phagocytic clearance. Here we report the development and optimisation of a new high-throughput flow cytometry-based phagocytosis assay using undifferentiated Thp-1 cells to quantitate the amount of opsonizing antibody in patient sera, and apply this assay to measure the impact of HIV on the levels of antibodies to a pregnancy malaria-associated parasite line in a cohort of Malawian primigravid women. The assay showed high reproducibility, with inter-experimental correlation of r(2) = 0.99. In primigravid women, concurrent malaria infection was associated with significantly increased antibodies, whereas HIV decreased the ability to acquire opsonising antibodies (Mann-Whitney ranksum: p = 0.013). This decrease was correlated with HIV-induced immunosuppression, with women with less than 350 x 10(6) CD4+ T- cells/L having less opsonising antibodies (coef: -11.95,P = 0.002). Levels of antibodies were not associated with protection from low birth weight or anaemia. This flow cytometry-based phagocytosis assay proved to be efficient and accurate for the measurement of Fc-receptor mediated phagocytosis-inducing antibodies in large cohorts. HIV was found to affect mainly the acquisition of antibodies to pregnancy-specific malaria in primigravidae. Further studies of the relationship between opsonising antibodies to malaria in pregnancy and HIV are indicated.

  20. Analysis of co-crystallized free phytosterols with triacylglycerols as a functional food ingredient.

    Science.gov (United States)

    Acevedo, Nuria C; Franchetti, Danielle

    2016-07-01

    This research focuses on the analysis of mixtures of free phytosterols (FPSs) with fully hydrogenated soybean oil (FHSO):soybean oil (SO) mixtures as a potential zero-trans substitute for various types of shortenings. Oil binding capacity as well as the thermal, rheological and structural properties of FHSO:SO blends containing 0, 20 and 25wt.% β-sitosterol or stigmasterol were investigated in this study. Differential interference contrast (DIC) microscopy and wide angle X-ray diffraction (WAXRD) revealed that co-crystallization of FPSs with FHSO:SO blends occurred. Polymorphic forms were characterized as a mixture of β' and β for all samples. The addition of FPSs decreased oil loss (OL) of FHSO:SO samples. Melting profiles of the prepared FPS-TAG (triacylglycerol) blends were extended to higher temperatures compared to a commercial shortening. Rheological properties were comparable to those of commercial puff pastry shortening suggesting that FPS-TAG blends may be acceptable for bakery applications. FPSs co-crystallized with FHSO and SO may be a suitable trans-fat free substitute for a number of types of shortening, including puff pastry shortening. The manufacturing of co-crystallized /FPS-TAG matrices will possibly bring large economic benefits as their functionalization can potentially be achieved by using existing simple shear processing. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Detection of Lard in Cocoa Butter—Its Fatty Acid Composition, Triacylglycerol Profiles, and Thermal Characteristics

    Directory of Open Access Journals (Sweden)

    Marliana Azir

    2017-11-01

    Full Text Available The present study investigates the detection of lard in cocoa butter through changes in fatty acids composition, triacylglycerols profile, and thermal characteristics. Cocoa butter was mixed with 1% to 30% (v/v of lard and analyzed using a gas chromatography flame ionization detector, high performance liquid chromatography, and differential scanning calorimetry. The results revealed that the mixing of lard in cocoa butter showed an increased amount of oleic acid in the cocoa butter while there was a decrease in the amount of palmitic acid and stearic acids. The amount of POS, SOS, and POP also decreased with the addition of lard. A heating thermogram from the DSC analysis showed that as the concentration of lard increased from 3% to 30%, two minor peaks at −26 °C and 34.5 °C started to appear and a minor peak at 34.5 °C gradually overlapped with the neighbouring major peak. A cooling thermogram of the above adulterated cocoa butter showed a minor peak shift to a lower temperature of −36 °C to −41.5 °C. Values from this study could be used as a basis for the identification of lard from other fats in the food authentication process.

  2. Detection of Lard in Cocoa Butter—Its Fatty Acid Composition, Triacylglycerol Profiles, and Thermal Characteristics

    Science.gov (United States)

    Azir, Marliana; Abbasiliasi, Sahar; Tengku Ibrahim, Tengku Azmi; Manaf, Yanty Noorzianna Abdul; Sazili, Awis Qurni; Mustafa, Shuhaimi

    2017-01-01

    The present study investigates the detection of lard in cocoa butter through changes in fatty acids composition, triacylglycerols profile, and thermal characteristics. Cocoa butter was mixed with 1% to 30% (v/v) of lard and analyzed using a gas chromatography flame ionization detector, high performance liquid chromatography, and differential scanning calorimetry. The results revealed that the mixing of lard in cocoa butter showed an increased amount of oleic acid in the cocoa butter while there was a decrease in the amount of palmitic acid and stearic acids. The amount of POS, SOS, and POP also decreased with the addition of lard. A heating thermogram from the DSC analysis showed that as the concentration of lard increased from 3% to 30%, two minor peaks at −26 °C and 34.5 °C started to appear and a minor peak at 34.5 °C gradually overlapped with the neighbouring major peak. A cooling thermogram of the above adulterated cocoa butter showed a minor peak shift to a lower temperature of −36 °C to −41.5 °C. Values from this study could be used as a basis for the identification of lard from other fats in the food authentication process. PMID:29120362

  3. Identification of triacylglycerol using automated annotation of high resolution multistage mass spectral trees.

    Science.gov (United States)

    Wang, Xiupin; Peng, Qingzhi; Li, Peiwu; Zhang, Qi; Ding, Xiaoxia; Zhang, Wen; Zhang, Liangxiao

    2016-10-12

    High complexity of identification for non-target triacylglycerols (TAGs) is a major challenge in lipidomics analysis. To identify non-target TAGs, a powerful tool named accurate MS(n) spectrometry generating so-called ion trees is used. In this paper, we presented a technique for efficient structural elucidation of TAGs on MS(n) spectral trees produced by LTQ Orbitrap MS(n), which was implemented as an open source software package, or TIT. The TIT software was used to support automatic annotation of non-target TAGs on MS(n) ion trees from a self-built fragment ion database. This database includes 19108 simulate TAG molecules from a random combination of fatty acids and corresponding 500582 self-built multistage fragment ions (MS ≤ 3). Our software can identify TAGs using a "stage-by-stage elimination" strategy. By utilizing the MS(1) accurate mass and referenced RKMD, the TIT software can discriminate unique elemental composition candidates. The regiospecific isomers of fatty acyl chains will be distinguished using MS(2) and MS(3) fragment spectra. We applied the algorithm to the selection of 45 TAG standards and demonstrated that the molecular ions could be 100% correctly assigned. Therefore, the TIT software could be applied to TAG identification in complex biological samples such as mouse plasma extracts. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Effects of medium-chain triacylglycerols on Maillard reaction in bread baking.

    Science.gov (United States)

    Toyosaki, Toshiyuki

    2018-06-01

    To investigate the relationship between the fatty acid composition of medium-chain triacylglycerols (MCTs) and the Maillard reaction induced during bread baking, a comparison with various fatty acids was conducted. Saturated fatty acids had a remarkable inhibitory effect on the amount of advanced glycation end products (AGEs) generated from the Maillard reaction in bread baking compared to unsaturated fatty acids. The amount of AGEs produced by each fatty acid (mg kg -1 ) was as follows: C18:0, 18.7; C12:0, 35.2; C16:0, 21.4; C18:0, 38.2; C18:1, 68.7; C18:2, 80.1; C20:4, 80.8; C22:4, 89.8. Saturated fatty acids were possibly involved in the Maillard reaction and, as a result, acted to inhibit it. In the case of unsaturated fatty acids, amounts of AGEs during the Maillard reaction in baking tended to increase as the degree of unsaturation increased. In other words, there was a positive correlation between the degree of unsaturation and the amount of AGEs. It was also confirmed that the air pore distribution in baked bread was closely related to AGEs. These results led us to conclude that the fatty acid composition of the added lipids also influences properties that determine the tastiness of bread. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  5. Intramyocellular triacylglycerol accumulation across weight loss strategies; Sub-study of the CENTRAL trial.

    Science.gov (United States)

    Gepner, Yftach; Shelef, Ilan; Schwarzfuchs, Dan; Cohen, Noa; Bril, Nitzan; Rein, Michal; Tsaban, Gal; Zelicha, Hila; Yaskolka Meir, Anat; Tene, Lilac; Sarusy, Benjamin; Rosen, Philip; Hoffman, Jay R; Stout, Jeffrey R; Thiery, Joachim; Ceglarek, Uta; Stumvoll, Michael; Blüher, Matthias; Stampfer, Meir J; Shai, Iris

    2017-01-01

    Intramyocellular triacylglycerol (IMTG) is utilized as metabolic fuel during exercise and is linked to insulin resistance, but the long-term effect of weight loss strategies on IMTG among participants with abdominal fat, remain unclear. In an 18-month trial, sedentary participants with abdominal fat/dyslipidemia were randomized to either a low-fat (LF) or Mediterranean/low-carbohydrate (MED/LC) diet (including 28g·day-1 of walnuts). After 6-months, the participants were re-randomized to moderate intense physical activity (PA+) or non-physical activity (PA-). Magnetic resonance imaging (MRI) was used to quantify changes of IMTG, abdominal sub-depots, hepatic and intermuscular fats. Across the 277 participants [86% men, age = 48 years, body-mass-index (BMI) = 31kg/m2, visceral fat = 33%] 86% completed the 18-m trial. At baseline, women had higher IMTG than men (3.4% vs. 2.3%, pvs. 9.5-18.5%, p<0.05). Changes in IMTG were associated with visceral and intermuscular fat, metabolic syndrome, insulin and leptin (p<0.05 for all), however, these associations did not remain after adjustment for visceral fat changes. Lifestyle strategies differentially affect IMTG accumulation; combination of exercise with decreased carbohydrate/increased unsaturated fat proportion intake greatly increase IMTG. Our findings suggest that increased IMTG during diet-induced moderate weight loss may not be directly related to cardiometabolic risk. ClinicalTrials.gov NCT01530724.

  6. Triacylglycerol profiling of microalgae strains for biofuel feedstock by liquid chromatography-high-resolution mass spectrometry

    Energy Technology Data Exchange (ETDEWEB)

    MacDougall, Karen M.; McNichol, Jesse; McGinn, Patrick J.; O' Leary, Stephen J.B.; Melanson, Jeremy E. [Institute for Marine Biosciences, National Research Council of Canada, Halifax, NS (Canada)

    2011-11-15

    Biofuels from photosynthetic microalgae are quickly gaining interest as a viable carbon-neutral energy source. Typically, characterization of algal feedstock involves breaking down triacylglycerols (TAG) and other intact lipids, followed by derivatization of the fatty acids to fatty acid methyl esters prior to analysis by gas chromatography (GC). However, knowledge of the intact lipid profile could offer significant advantages for discovery stage biofuel research such as the selection of an algal strain or the optimization of growth and extraction conditions. Herein, lipid extracts from microalgae were directly analyzed by ultra-high pressure liquid chromatography-mass spectrometry (UHPLC-MS) using a benchtop Orbitrap mass spectrometer. Phospholipids, glycolipids, and TAGs were analyzed in the same chromatographic run, using a combination of accurate mass and diagnostic fragment ions for identification. Using this approach, greater than 100 unique TAGs were identified over the six algal strains studied and TAG profiles were obtained to assess their potential for biofuel applications. Under the growth conditions employed, Botryococcus braunii and Scenedesmus obliquus yielded the most comprehensive TAG profile with a high abundance of TAGs containing oleic acid. (orig.)

  7. White tea (Camellia sinensis extract reduces oxidative stress and triacylglycerols in obese mice

    Directory of Open Access Journals (Sweden)

    Lílian Gonçalves Teixeira

    2012-12-01

    Full Text Available White tea is an unfermented tea made from young shoots of Camellia sinensis protected from sunlight to avoid polyphenol degradation. Although its levels of catechins are higher than those of green tea (derived from the same plant, there are no studies addressing the relationship between this tea and obesity associated with oxidative stress.The objective of this study was to evaluate the effect of white tea on obesity and its complications using a diet induced obesity model. Forty male C57BL/6 mice were fed a high-fat diet to induce obesity (Obese group or the same diet supplemented with 0.5% white tea extract (Obese + WTE for 8 weeks. Adipose tissue, serum lipid profile, and oxidative stress were studied. White tea supplementation was not able to reduce food intake, body weight, or visceral adiposity. Similarly, there were no changes in cholesterol rich lipoprotein profile between the groups. A reduction in blood triacylglycerols associated with increased cecal lipids was observed in the group fed the diet supplemented with white tea. White tea supplementation also reduced oxidative stress in liver and adipose tissue. In conclusion, white tea extract supplementation (0.5% does not influence body weight or adiposity in obese mice. Its benefits are restricted to the reduction in oxidative stress associated with obesity and improvement of hypertriacylglycerolemia.

  8. Meal frequency differentially alters postprandial triacylglycerol and insulin concentrations in obese women.

    Science.gov (United States)

    Heden, Timothy D; Liu, Ying; Sims, Lauren J; Whaley-Connell, Adam T; Chockalingam, Anand; Dellsperger, Kevin C; Kanaley, Jill A

    2013-01-01

    The aim of this study was to compare postprandial lipemia, oxidative stress, antioxidant activity, and insulinemia between a three and six isocaloric high-carbohydrate meal frequency pattern in obese women. In a counterbalanced order, eight obese women completed two, 12-h conditions in which they consumed 1,500 calories (14% protein, 21% fat, and 65% carbohydrate) either as three 500 calorie liquid meals every 4-h or six 250 calorie liquid meals every 2-h. Blood samples were taken every 30 min and analyzed for triacylglycerol (TAG), total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, oxidized low-density lipoprotein cholesterol, myeloperoxidase, paraoxonase-1 activity, and insulin. The TAG incremental area under the curve (iAUC) during the three meal condition (321 ± 129 mg/dl · 12 h) was significantly lower (P = 0.04) compared with the six meal condition (481 ± 155 mg/dl · 12 h). The insulin iAUC during the three meal condition (5,549 ± 1,007 pmol/l · 12 h) was significantly higher (P = 0.05) compared with the six meal condition (4,230 ± 757 pmol/l(.) 12 h). Meal frequency had no influence on the other biochemical variables. Collectively, a three and six isocaloric high-carbohydrate meal frequency pattern differentially alters postprandial TAG and insulin concentrations but has no effect on postprandial cholesterol, oxidative stress, or antioxidant activity in obese women. Copyright © 2012 The Obesity Society.

  9. Triacylglycerol secretion in rats: validation of a tracer method employing radioactive glycerol

    International Nuclear Information System (INIS)

    Bird, M.; Williams, M.A.; Baker, N.

    1984-01-01

    A two-compartment model was developed to analyze the temporal changes in plasma triacylglycerol (TG)-specific radioactivity after injection of [2- 3 H]glycerol into rats. The analysis, which yielded fractional rate constants of TG secretion, was tested in rats fed diets either adequate or deficient in essential fatty acids (EFA) and containing either glucose, fructose or sucrose as the dietary carbohydrate. The method of analysis appeared valid, first, because of a close agreement between experimental and computer-fitted TG-specific radioactivity curves, and second, because the fractional rate constants obtained were quite similar to fractional rate constants determined previously by the Triton WR-1339 technique in rats maintained on identical diets. The results show that EFA deficiency increased the fractional rate constant of TG secretion 1.7-, 1.8- and 3.3-fold and the rate of TG secretion 1.8-, 1.6- and 1.4-fold when the dietary carbohydrate was glucose, sucrose and fructose, respectively, in comparison with control rats fed diets supplying these same carbohydrates but adequate in EFA. In the latter groups, the rates of plasma TG secretion were in the range of 0.14-0.17 mg/min per 100 g body weight, and the rate of secretion in the fructose-fed rats was only 20% higher than in the glucose-fed rats

  10. A soluble diacylglycerol acyltransferase is involved in triacylglycerol biosynthesis in the oleaginous yeast Rhodotorula glutinis.

    Science.gov (United States)

    Rani, Sapa Hima; Saha, Saikat; Rajasekharan, Ram

    2013-01-01

    The biosynthesis of triacylglycerol (TAG) occurs in the microsomal membranes of eukaryotes. Here, we report the identification and functional characterization of diacylglycerol acyltransferase (DGAT), a member of the 10 S cytosolic TAG biosynthetic complex (TBC) in Rhodotorula glutinis. Both a full-length and an N-terminally truncated cDNA clone of a single gene were isolated from R. glutinis. The DGAT activity of the protein encoded by RgDGAT was confirmed in vivo by the heterologous expression of cDNA in a Saccharomyces cerevisiae quadruple mutant (H1246) that is defective in TAG synthesis. RgDGAT overexpression in yeast was found to be capable of acylating diacylglycerol (DAG) in an acyl-CoA-dependent manner. Quadruple mutant yeast cells exhibit growth defects in the presence of oleic acid, but wild-type yeast cells do not. In an in vivo fatty acid supplementation experiment, RgDGAT expression rescued quadruple mutant growth in an oleate-containing medium. We describe a soluble acyl-CoA-dependent DAG acyltransferase from R. glutinis that belongs to the DGAT3 class of enzymes. The study highlights the importance of an alternative TAG biosynthetic pathway in oleaginous yeasts.

  11. The position of rumenic acid on triacylglycerols alters its bioavailability in rats.

    Science.gov (United States)

    Chardigny, J M; Masson, E; Sergiel, J P; Darbois, M; Loreau, O; Noël, J P; Sébédio, J-L

    2003-12-01

    The metabolic fate of rumenic acid (9cis,11trans-octadecenoic acid) related to its position on the glycerol moiety has not yet been studied. In the present work, synthetic triacylglycerols (TAG) esterified with oleic and rumenic acids were prepared. Rats were force-fed synthetic dioleyl monorumenyl glycerol with (14)C labeled rumenic acid in the internal (sn-2) or in the external position (sn-1 or sn-3). Rats were then placed in metabolic cages for 16 h. At the end of the experiment, the radioactivity in tissues, carcass and expired CO(2) was measured. Rumenic acid that was esterified at the external positions on the TAG was better absorbed and oxidized to a greater extent than when esterified at the internal position. The fatty acid from the 2-TAG form was also better incorporated into the rat carcass. In the liver, rumenic acid appeared mainly in TAG (50%) and to a lesser extent in phospholipids (33%) whatever its dietary form. Moreover, analyses of lipids from Camembert cheese and butter revealed that rumenic acid was located mainly on the sn-1 or sn-3 positions (74%). Taken together, these data suggest that rumenic acid from dairy fat may be well absorbed and used extensively for energy production.

  12. Improved fatty acid analysis of conjugated linoleic acid rich egg yolk triacylglycerols and phospholipid species.

    Science.gov (United States)

    Shinn, Sara; Liyanage, Rohana; Lay, Jack; Proctor, Andrew

    2014-07-16

    Reports from chicken conjugated linoleic acid (CLA) feeding trials are limited to yolk total fatty acid composition, which consistently described increased saturated fatty acids and decreased monounsaturated fatty acids. However, information on CLA triacylglycerol (TAG) and phospholipid (PL) species is limited. This study determined the fatty acid composition of total lipids in CLA-rich egg yolk produced with CLA-rich soy oil, relative to control yolks using gas chromatography with flame ionization detection (GC-FID), determined TAG and PL fatty acid compositions by thin-layer chromatography-GC-FID (TLC-GC-FID), identified intact PL and TAG species by TLC-matrix-assisted laser desorption/ionization mass spectrometry (TLC-MALDI-MS), and determined the composition of TAG and PL species in CLA and control yolks by direct flow infusion electrospray ionization MS (DFI ESI-MS). In total, 2 lyso-phosphatidyl choline (LPC) species, 1 sphingomyelin species, 17 phosphatidyl choline species, 19 TAG species, and 9 phosphatidyl ethanolamine species were identified. Fifty percent of CLA was found in TAG, occurring predominantly in C52:5 and C52:4 TAG species. CLA-rich yolks contained significantly more LPC than did control eggs. Comprehensive lipid profiling may provide insight on relationships between lipid composition and the functional properties of CLA-rich eggs.

  13. Reduction of serum triacylglycerol-rich lipoprotein concentrations in cows with hepatic lipidosis.

    Science.gov (United States)

    Herdt, T H; Liesman, J S; Gerloff, B J; Emery, R S

    1983-02-01

    The hepatic and serum lipid concentrations in 49 dairy cows with displaced abomasum, 7 postpartum cows fasted for 6 days, and 14 healthy postpartum cows were studied. The cows with displaced abomasums were retrospectively allotted to 2 groups: those with greater than 15% liver fat (DAHF) and those with less than 15% liver fat (DALF). Liver total lipid concentrations were high in the DAHF group, exceeding these values in the fasted cows by 30% and in the healthy and DALF cows by 63% on the average. In contrast, the liver phospholipid concentrations were low in the DAHF group, intermediate in the fasted and DALF groups and high in the healthy group. On a group basis, an inverse relationship was observed between serum and liver lipid concentrations. The serum concentrations of both total and dextran-sulfate-precipitable (DSP) lipids were high in the fasted cows and were less in the DALF and healthy cows and in the DAHF cows (lowest). The between-group differences in serum total and serum DSP concentrations of triacylglycerol, cholesterol, and phospholipid followed the same quantitative pattern as the total lipids. However, the relative difference between groups was greater for each of the DSP lipid fractions. These results support the hypothesis that severe hepatic lipidosis in cattle occurs due to impaired hepatic lipoprotein synthesis and secretion.

  14. Chemistry and liquid chromatography methods for the analyses of primary oxidation products of triacylglycerols.

    Science.gov (United States)

    Zeb, A

    2015-05-01

    Triacylglycerols (TAGs) are one of the major components of the cells in higher biological systems, which can act as an energy reservoir in the living cells. The unsaturated fatty acid moiety is the key site of oxidation and formation of oxidation compounds. The TAG free radical generates several primary oxidation compounds. These include hydroperoxides, hydroxides, epidioxides, hydroperoxy epidioxides, hydroxyl epidioxides, and epoxides. The presence of these oxidized TAGs in the cell increases the chances of several detrimental processes. For this purpose, several liquid chromatography (LC) methods were reported in their analyses. This review is therefore focused on the chemistry, oxidation, extraction, and the LC methods reported in the analyses of oxidized TAGs. The studies on thin-layer chromatography were mostly focused on the total oxidized TAGs separation and employ hexane as major solvent. High-performance LC (HPLC) methods were discussed in details along with their merits and demerits. It was found that most of the HPLC methods employed isocratic elution with methanol and acetonitrile as major solvents with an ultraviolet detector. The coupling of HPLC with mass spectrometry (MS) highly increases the efficiency of analysis as well as enables reliable structural elucidation. The use of MS was found to be helpful in studying the oxidation chemistry of TAGs and needs to be extended to the complex biological systems.

  15. Greater impairment of postprandial triacylglycerol than glucose response in metabolic syndrome subjects with fasting hyperglycaemia.

    Science.gov (United States)

    Jackson, Kim G; Walden, Charlotte M; Murray, Peter; Smith, Adrian M; Minihane, Anne M; Lovegrove, Julie A; Williams, Christine M

    2013-08-01

    Studies have started to question whether a specific component or combinations of metabolic syndrome (MetS) components may be more important in relation to cardiovascular disease risk. Our aim was to examine the impact of the presence of raised fasting glucose as a MetS component on postprandial lipaemia. Men classified with the MetS underwent a sequential test meal investigation, in which blood samples were taken at regular intervals after a test breakfast (t=0 min) and lunch (t=330 min). Lipids, glucose and insulin were measured in the fasting and postprandial samples. MetS subjects with 3 or 4 components were subdivided into those without (n=34) and with (n=23) fasting hyperglycaemia (≥5.6 mmol/l), irrespective of the combination of components. Fasting lipids and insulin were similar in the two groups, with glucose significantly higher in the men with glucose as a MetS component (Pcurve (AUC) and incremental AUC (P ≤0.016) for the postprandial triacylglycerol (TAG) response in men with fasting hyperglycaemia. Greater glucose AUC (Pglucose to be an important predictor of the postprandial TAG and glucose response. Our data analysis has revealed a greater impairment of postprandial TAG than glucose response in MetS subjects with raised fasting glucose. The worsening of postprandial lipaemic control may contribute to the greater CVD risk reported in individuals with MetS component combinations which include hyperglycaemia. Copyright © 2013 Elsevier Inc. All rights reserved.

  16. The small molecule fenpropimorph rapidly converts chloroplast membrane lipids to triacylglycerols in Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Hanul eKim

    2015-02-01

    Full Text Available Concern about global warming has prompted an intense interest in developing economical methods of producing biofuels. Microalgae provide a promising platform for biofuel production, because they accumulate high levels of lipids, and do not compete with food or feed sources. However, current methods of producing algal oil involve subjecting the microalgae to stress conditions, such as nitrogen deprivation, and are prohibitively expensive. Here, we report that the fungicide fenpropimorph rapidly causes high levels of neutral lipids to accumulate in Chlamydomonas reinhardtii cells. When treated with fenpropimorph (10 μg mL–1 for 1 h, Chlamydomonas cells accumulated at least four-fold the amount of triacylglycerols (TAGs present in the untreated control cells. Furthermore, the quantity of TAGs present after 1 h of fenpropimorph treatment was over two-fold higher than that formed after 9 days of nitrogen starvation in medium with no acetate supplement. Biochemical analysis of lipids revealed that the accumulated TAGs were derived mainly from chloroplast polar membrane lipids. Such a conversion of chloroplast polar lipids to TAGs is desirable for biodiesel production, because polar lipids are usually removed during the biodiesel production process. Thus, our data exemplified that a cost and time effective method of producing TAGs is possible using fenpropimorph or similar drugs.

  17. Tick thioester-containing proteins and phagocytosis do not affect transmission of Borrelia afzelii from the competent vector ixodes ricinus

    Czech Academy of Sciences Publication Activity Database

    Urbanová, Veronika; Hajdušek, Ondřej; Hönig Mondeková, Helena; Šíma, Radek; Kopáček, Petr

    2017-01-01

    Roč. 7, MAR (2017), č. článku 73. ISSN 2235-2988 R&D Projects: GA ČR GJ15-12006Y; GA ČR GA13-11043S; GA ČR GA17-27386S; GA ČR GA17-27393S EU Projects: European Commission(XE) 602272 - ANTIDotE Institutional support: RVO:60077344 Keywords : Borrelia * complement * Ixodes * phagocytosis * thioester-containing proteins Subject RIV: EC - Immunology OBOR OECD: Immunology Impact factor: 4.300, year: 2016

  18. Simultaneous determination of phagocytosis of Plasmodium falciparum-parasitized and non-parasitized red blood cells by flow cytometry

    Directory of Open Access Journals (Sweden)

    Gallo Valentina

    2012-12-01

    Full Text Available Abstract Background Severe falciparum malaria anaemia (SMA is a frequent cause of mortality in children and pregnant women. The most important determinant of SMA appears to be the loss of non-parasitized red blood cells (np-RBCs in excess of loss of parasitized (p- RBCs at schizogony. Based on data from acute SMA where excretion of haemoglobin in urine and increased plasma haemoglobin represented respectively less than 1% and 0.5% of total Hb loss, phagocytosis appears to be the predominant mechanism of removal of np- and p-RBC. Estimates indicate that np-RBCs are cleared in approximately 10-fold excess compared to p-RBCs. An even larger removal of np-RBCs has been described in vivax malaria anaemia. Estimates were based on two single studies both performed on neurosyphilitic patients who underwent malaria therapy. As the share of np-RBC removal is likely to vary between wide limits, it is important to assess the contribution of both np- and p-RBC populations to overall RBC loss, and disclose the mechanism of such variability. As available methods do not discriminate between the removal of np- vs p-RBCs, the purpose of this study was to set up a system allowing the simultaneous determination of phagocytosis of p- and np-RBC in the same sample. Methods and Results Phagocytosis of p- and np-RBCs was quantified in the same sample using double-labelled target cells and the human phagocytic cell-line THP-1, pre-activated by TNF and IFNγ to enhance their phagocytic activity. Target RBCs were double-labelled with fluorescent carboxyfluorescein-succinimidyl ester (CF-SE and the DNA label ethidium bromide (EB. EB, a DNA label, allowed to discriminate p-RBCs that contain parasitic DNA from the np-RBCs devoid of DNA. FACS analysis of THP-1 cells fed with double-labelled RBCs showed that p- and np-RBCs were phagocytosed in different proportions in relation to parasitaemia. Conclusions The assay allowed the analysis of phagocytosis rapidly and with low

  19. Neuro-immune relationships at patients with chronic pyelonephrite and cholecystite. Communication 2. Correlations between parameters EEG, HRV and Phagocytosis

    OpenAIRE

    Kul’chyns’kyi, Andriy B; Kovbasnyuk, Marta M; Korolyshyn, Tetyana A; Kyjenko, Valeriy M; Zukow, Walery; Popovych, Igor L

    2016-01-01

    Kul’chyns’kyi Andriy B, Kovbasnyuk Marta M, Korolyshyn Tetyana A, Kyjenko Valeriy M, Zukow Walery, Popovych Igor L. Neuro-immune relationships at patients with chronic pyelonephrite and cholecystite. Communication 2. Correlations between parameters EEG, HRV and Phagocytosis. Journal of Education, Health and Sport. 2016;6(10):377-401. eISSN 2391-8306. DOI http://dx.doi.org/10.5281/zenodo.163221 http://ojs.ukw.edu.pl/index.php/johs/article/view/3957 The journal h...

  20. Interaction between Salmonella typhimurium and phagocytic cells in pigs - Phagocytosis, oxidative burst and killing in polymorphonuclear leukocytes and monocytes

    DEFF Research Database (Denmark)

    Riber, Ulla; Lind, Peter

    1999-01-01

    Interactions between Salmonella typhimurium and peripheral blood leucocytes from healthy, Salmonella-free pigs were investigated in vitro. Both granulocytes and monocytes phagocytized FITC-labelled heat-killed Salmonella bacteria as shown by flow cytometry. Phagocytosis in whole blood and isolated...... with the exhaustion of oxidative burst in non-adherent monocytes were performed by prestimulation with PMA, heat-killed Salmonella or buffer. Prestimulation with PMA led to a strong reduction in oxidative burst induced by living opsonized Salmonella bacteria, whereas prestimulation with heat-killed bacteria gave rise...

  1. Diverse regulation of retinal pigment epithelium phagocytosis of photoreceptor outer segments by calcium-independent phospholipase A₂, group VIA and secretory phospholipase A₂, group IB

    DEFF Research Database (Denmark)

    Zhan, Chen; Wang, Jinmei; Kolko, Miriam

    2012-01-01

    PURPOSE: To investigate the roles of the phospholipases A(2) (PLA(2)) subtypes, iPLA(2)-VIA and sPLA(2)-IB in retinal pigment epithelium (RPE) phagocytosis of photoreceptor outer segments (POS) and to explore a possible interaction between sPLA(2)-IB and iPLA(2)-VIA in the RPE. METHODS: To explore...... the role of iPLA(2)-VIA in RPE phagocytosis of POS, experiments with iPLA(2)-VIA vector transfection, iPLA(2)-VIA(-/-) knockout (KO) mice, and iPLA(2)-VIA inhibition by bromoenol lactone (BEL) were done. Exogenous addition of sPLA(2)-IB was used to investigate the role of sPLA(2)-IB in RPE phagocytosis....... A Luciferase Reporter Vector containing the iPLA(2)-VIA promoter was used to study the effects of sPLA(2)-IB on the iPLA(2)-VIA promoter. RESULTS: ARPE-19 and primary mouse RPE cells transfected with iPLA(2)-VIA showed increased phagocytosis. Phagocytosis was reduced in primary mouse RPE inhibited with BEL...

  2. Decoupling Internalization, Acidification and Phagosmal-Endosomal/Iysosomal Phagocytosis of Internalin A coated Beads in epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Blanchette, C D; Woo, Y; Thomas, C; Shen, N; Sulchek, T A; Hiddessen, A L

    2008-12-22

    Phagocytosis has been extensively examined in 'professional' phagocytic cells using pH sensitive dyes. However, in many of the previous studies, a separation between the end of internalization, beginning of acidification and completion of phagosomal-endosomal/lysosomal fusion was not clearly established, and in several cases, it was treated as a one-step process. In addition, very little work has been done to systematically examine phagosomal maturation in 'non-professional' phagocytic cells, such as epithelial cells. Therefore, in this study, we developed a simple and novel method to decouple and accurately measure particle internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in Madin-Darby Canine Kidney (MDCK) and Caco-2 epithelial cells. Our method was developed using a pathogen mimetic system consisting of polystyrene beads coated with Internalin A (InlA), a membrane surface protein from Listeria monocytogenes known to trigger receptor-mediated internalization. We achieved independent measurements of the rates of internalization, phagosomal acidification and phagosomal-endosomal/lysosomal fusion in epithelial cells by combining the InlA-coated beads (InlA-beads) with antibody quenching, pH sensitive dyes and endosomal/lysosomal dyes, as follows: the rate of InlA bead internalization was measured via antibody quenching of a pH independent dye (Alexa488) conjugated to InlA-beads, the rate at which phagosomes containing internalized InlA beads became acidified was measured using a pH dependent dye (FITC) conjugated to the beads and the rate of phagosomal-endosomal/lysosomal fusion was measured using a combination of unlabeled InlA-beads and an endosomal/lysosomal dye. By performing these independent measurements under identical experimental conditions, we were able to decouple the three processes and establish time scales for each. In a separate set of experiments, we also exploited the phagosomal acidification

  3. Actin-binding protein regulation by microRNAs as a novel microbial strategy to modulate phagocytosis by host cells: the case of N-Wasp and miR-142-3p.

    Science.gov (United States)

    Bettencourt, Paulo; Marion, Sabrina; Pires, David; Santos, Leonor F; Lastrucci, Claire; Carmo, Nuno; Blake, Jonathon; Benes, Vladimir; Griffiths, Gareth; Neyrolles, Olivier; Lugo-Villarino, Geanncarlo; Anes, Elsa

    2013-01-01

    Mycobacterium tuberculosis (Mtb) is a successful intracellular pathogen that thrives in macrophages (Mφs). There is a need to better understand how Mtb alters cellular processes like phagolysosome biogenesis, a classical determinant of its pathogenesis. A central feature of this bacteria's strategy is the manipulation of Mφ actin. Here, we examined the role of microRNAs (miRNAs) as a potential mechanism in the regulation of actin-mediated events leading to phagocytosis in the context of mycobacteria infection. Given that non-virulent Mycobacterium smegmatis also controls actin filament assembly to prolong its intracellular survival inside host cells, we performed a global transcriptomic analysis to assess the modulation of miRNAs upon M. smegmatis infection of the murine Mφ cell line, J774A.1. This approach identified miR-142-3p as a key candidate to be involved in the regulation of actin dynamics required in phagocytosis. We unequivocally demonstrate that miR-142-3p targets N-Wasp, an actin-binding protein required during microbial challenge. A gain-of-function approach for miR-142-3p revealed a down-regulation of N-Wasp expression accompanied by a decrease of mycobacteria intake, while a loss-of-function approach yielded the reciprocal increase of the phagocytosis process. Equally important, we show Mtb induces the early expression of miR-142-3p and partially down-regulates N-Wasp protein levels in both the murine J774A.1 cell line and primary human Mφs. As proof of principle, the partial siRNA-mediated knock down of N-Wasp resulted in a decrease of Mtb intake by human Mφs, reflected in lower levels of colony-forming units (CFU) counts over time. We therefore propose the modulation of miRNAs as a novel strategy in mycobacterial infection to control factors involved in actin filament assembly and other early events of phagolysosome biogenesis.

  4. Inhaled corticosteroid treatment for 6 months was not sufficient to normalize phagocytosis in asthmatic children.

    Science.gov (United States)

    da Silva-Martins, Carmen Lívia Faria; Couto, Shirley Claudino; Muniz-Junqueira, Maria Imaculada

    2013-08-30

    Corticosteroids are the first-line therapy for asthma; however, the effect of corticosteroids on the innate immune system remains unclear. This study's objective was to evaluate the effect of inhaled corticosteroid therapy (ICT) on phagocytic functions. To evaluate the impact of ICT, the phagocytosis of Saccharomyces cerevisiae by blood monocytes and neutrophils and the production of superoxide anions were assessed before and after three and six months of ICT treatment in 58 children with persistent asthma and 21 healthy controls. We showed that the phagocytic capacity of monocytes and neutrophils that occurred via pattern recognition receptors or was mediated by complement and immunoglobulin receptors in asthmatic children before treatment was significantly lower than in healthy controls (pICT for 6 months was not sufficient to normalize phagocytosis by the phagocytes. Superoxide anion production was also decreased in the asthmatic children before treatment, and ICT normalized the O- production only for children with mild persistent asthma when assessed at baseline but caused this function to decrease after stimulation (p<0.05, Kruskal-Wallis test). Our data suggest that an immunodeficiency in phagocytes remained even after treatment. However, this immunodeficiency does not appear to correspond with the clinical evolution of asthma because an improvement in clinical parameters occurred.

  5. Cromolyn Reduces Levels of the Alzheimer's Disease-Associated Amyloid β-Protein by Promoting Microglial Phagocytosis.

    Science.gov (United States)

    Zhang, Can; Griciuc, Ana; Hudry, Eloise; Wan, Yu; Quinti, Luisa; Ward, Joseph; Forte, Angela M; Shen, Xunuo; Ran, ChongZhao; Elmaleh, David R; Tanzi, Rudolph E

    2018-01-18

    Amyloid-beta protein (Aβ) deposition is a pathological hallmark of Alzheimer's disease (AD). Aβ deposition triggers both pro-neuroinflammatory microglial activation and neurofibrillary tangle formation. Cromolyn sodium is an asthma therapeutic agent previously shown to reduce Aβ levels in transgenic AD mouse brains after one-week of treatment. Here, we further explored these effects as well as the mechanism of action of cromolyn, alone, and in combination with ibuprofen in APP Swedish -expressing Tg2576 mice. Mice were treated for 3 months starting at 5 months of age, when the earliest stages of β-amyloid deposition begin. Cromolyn, alone, or in combination with ibuprofen, almost completely abolished longer insoluble Aβ species, i.e. Aβ40 and Aβ42, but increased insoluble Aβ38 levels. In addition to its anti-aggregation effects on Aβ, cromolyn, alone, or plus ibuprofen, but not ibuprofen alone, increased microglial recruitment to, and phagocytosis of β-amyloid deposits in AD mice. Cromolyn also promoted Aβ42 uptake in microglial cell-based assays. Collectively, our data reveal robust effects of cromolyn, alone, or in combination with ibuprofen, in reducing aggregation-prone Aβ levels and inducing a neuroprotective microglial activation state favoring Aβ phagocytosis versus a pro-neuroinflammatory state. These findings support the use of cromolyn, alone, or with ibuprofen, as a potential AD therapeutic.

  6. Phagocytosis-induced 51Cr release from activated macrophages and blood mononuclears. Effect of colchicine and antioxidants

    International Nuclear Information System (INIS)

    McGee, M.P.; Hale, A.H.

    1981-01-01

    The chromium-release test was adapted to the measurement of the cellular injury induced when activated macrophages phagocytose particulates. Macrophages obtained from rabbit lungs undergoing BCG-induced chronic inflammation released more chromium when incubated in the presence of phagocytosable particles than when incubated under resting conditions. Blood mononuclear cells, 40-60% monocytes, procured from the same BCG-injected animals, were less susceptible to phagocytosis-induced injury than the macrophages obtained from the lungs. The amount of chromium released by the activated macrophages was proportional to the number of particles present during incubation. In the presence of catalase, the amounts of chromium released by phagocytosing and resting macrophages were similar; in the presence of superoxide dismutase and cytochrome c, the amount of chromium released by phagocytosing macrophages was 13-35% less than the amount of chromium released by macrophages incubated without the antioxidants. In addition, colchicine, an inhibitor of degranulation also exerted partial inhibition of the chromium release. These results suggest that oxygen radicals and lysosomal contents contribute to the cellular injury that results from phagocytosis

  7. Shrimp miR-12 Suppresses White Spot Syndrome Virus Infection by Synchronously Triggering Antiviral Phagocytosis and Apoptosis Pathways

    Science.gov (United States)

    Shu, Le; Zhang, Xiaobo

    2017-01-01

    Growing evidence has indicated that the innate immune system can be regulated by microRNAs (miRNAs). However, the mechanism underlying miRNA-mediated simultaneous activation of multiple immune pathways remains unknown. To address this issue, the role of host miR-12 in shrimp (Marsupenaeus japonicus) antiviral immune responses was characterized in the present study. The results indicated that miR-12 participated in virus infection, host phagocytosis, and apoptosis in defense against white spot syndrome virus invasion. miR-12 could simultaneously trigger phagocytosis, apoptosis, and antiviral immunity through the synchronous downregulation of the expression of shrimp genes [PTEN (phosphatase and tensin homolog) and BI-1(transmembrane BAX inhibitor motif containing 6)] and the viral gene (wsv024). Further analysis showed that miR-12 could synchronously mediate the 5′–3′ exonucleolytic degradation of its target mRNAs, and this degradation terminated in the vicinity of the 3′ untranslated region sequence complementary to the seed sequence of miR-12. Therefore, the present study showed novel aspects of the miRNA-mediated simultaneous regulation of multiple immune pathways. PMID:28824612

  8. Virulent and Vaccine Strains of Streptococcus equi ssp. zooepidemicus Have Different Influences on Phagocytosis and Cytokine Secretion of Macrophages.

    Science.gov (United States)

    Jie, Peng; Zhe, Ma; Chengwei, Hua; Huixing, Lin; Hui, Zhang; Chengping, Lu; Hongjie, Fan

    2017-01-06

    Swine streptococcosis is a significant threat to the Chinese pig industry, and Streptococcus equi ssp. zooepidemicus (SEZ) is one of the major pathogens. SEZ ATCC35246 is a classical virulent strain, while SEZ ST171 is a Chinese attenuated vaccine strain. In this study, we employed stable isotope labeling by amino acids in cell culture and liquid chromatography-mass spectrometry (LC-MS) to determine the differential response of macrophages to infection by these two strains. Eighty-seven upregulated proteins and 135 downregulated proteins were identified. The proteomic results were verified by real-time polymerase chain reaction for 10 chosen genes and Western blotting for three proteins. All differentially abundant proteins were analyzed for their Gene Ontology and Kyoto Encyclopedia of Genes and Genomes annotations. Certain downregulated proteins were associated with immunity functions, and the upregulated proteins were related to cytomembrane and cytoskeleton regulation. The phagocytosis rate and cytokine genes transcription in Raw264.7 cells during SEZ ATCC35246 and ST171 infection were detected to confirm the bioinformatics results. These results showed that different effects on macrophage phagocytosis and cytokine expression might explain the different phenotypes of SEZ ATCC35246 and ST171 infection. This research provided clues to the mechanisms of host immunity responses to SEZ ST171and SEZ ATCC35246, which could identify potential therapy and vaccine development targets.

  9. The cellular prion protein negatively regulates phagocytosis and cytokine expression in murine bone marrow-derived macrophages.

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    Min Wang

    Full Text Available The cellular prion protein (PrP(C is a glycosylphosphatidylinositol (GPI-anchored glycoprotein on the cell surface. Previous studies have demonstrated contradictory roles for PrP(C in connection with the phagocytic ability of macrophages. In the present work, we investigated the function of PrP(C in phagocytosis and cytokine expression in bone marrow-derived macrophages infected with Escherichia coli. E. coli infection induced an increase in the PRNP mRNA level. Knockout of PrP(C promoted bacterial uptake; upregulated Rab5, Rab7, and Eea1 mRNA expression; and increased the recruitment of lysosomal-associated membrane protein-2 to phagosomes, suggesting enhanced microbicidal activity. Remarkably, knockout of PrP(C suppressed the proliferation of internalized bacteria and increased the expression of cytokines such as interleukin-1β. Collectively, our data reveal an important role of PrP(C as a negative regulator for phagocytosis, phagosome maturation, cytokine expression, and macrophage microbicidal activity.

  10. Identification of FadAB Complexes Involved in Fatty Acid β-Oxidation in Streptomyces coelicolor and Construction of a Triacylglycerol Overproducing strain

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    Simón Menendez-Bravo

    2017-08-01

    Full Text Available Oleaginous microorganisms represent possible platforms for the sustainable production of oleochemicals and biofuels due to their metabolic robustness and the possibility to be engineered. Streptomyces coelicolor is among the narrow group of prokaryotes capable of accumulating triacylglycerol (TAG as carbon and energy reserve. Although the pathways for TAG biosynthesis in this organism have been widely addressed, the set of genes required for their breakdown have remained elusive so far. Here, we identified and characterized three gene clusters involved in the β-oxidation of fatty acids (FA. The role of each of the three different S. coelicolor FadAB proteins in FA catabolism was confirmed by complementation of an Escherichia coliΔfadBA mutant strain deficient in β-oxidation. In S. coelicolor, the expression profile of the three gene clusters showed variation related with the stage of growth and the presence of FA in media. Flux balance analyses using a corrected version of the current S. coelicolor metabolic model containing detailed TAG biosynthesis reactions suggested the relevance of the identified fadAB genes in the accumulation of TAG. Thus, through the construction and analysis of fadAB knockout mutant strains, we obtained an S. coelicolor mutant that showed a 4.3-fold increase in the TAG content compared to the wild type strain grown under the same culture conditions.

  11. Correlation Between Total Flavonoid Contents and Macrophage Phagocytosis Activity of Fractions From Faloak (Sterculia quadrifida R.Br. Barks Ethanolic Extract In Vitro

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    Rima Munawaroh

    2018-04-01

    Full Text Available On Timor island, Nusa Tenggara Timur, faloak barks (Sterculia quadrifida R.Br. has been used empirically to restore stamina. Faloak bark ethanolic extract proved to have immunomodulatory activity in vitro, which can increase macrophage phagocytosis activity. This research aimed: (i to determine the immunomodulatory active fraction of faloak bark ethanolic extract, (ii to determine the total flavonoid contents of faloak extract and fractions, and (iii to evaluate the correlation of the total flavonoid contents of those extract and fractions with their macrophage phagocytosis activity. The simplisia powder is macerated with 96% ethanol. The extract was dissolved in methanol:water (9:1v/v was then subsequently partitioned with n-hexane, ethyl acetate, and water to obtain n-hexane fraction, ethyl acetate fraction, water fraction, and insoluble fraction. Faloak extract and fractions at concentration 62,5; 125; 250; 500μg/mL were tested for their effect on the peritoneal macrophage phagocytosis of Balb/c mice in vitro by the latex beads method. Phagocytosis capacity and phagocytosis index were analyzed using one-way anova and post hoc Tukey HSD test with 95% confidence level. The results showed that ethyl acetate fraction had the highest macrophage phagocytosis capacity and the highest total flavonoid content compared to other fractions. The highest macrophage phagocytosis capacity of ethyl acetate fraction at concentration of 250 μg/mL was 51,94±4,67%, this value was significantly different from cell control (7,50±1,29%, negative controls of 0,0625% dimethylsulphoxide (6,25±0,36%, as well as positive control of 200 μg/mL echinaceae extract syrup® (9,97±0,33%. The total flavonoid content of ethyl acetate fraction determined by aluminum chloride method was 4,290±0.029 mg of quercetin equivalent/g fraction. There was a positive and strong correlation between the total flavonoid content of these extract and fractions with their macrophage

  12. Diacylglycerol acyltransferase-2 (DGAT2) and monoacylglycerol acyltransferase-2 (MGAT2) interact to promote triacylglycerol synthesis.

    Science.gov (United States)

    Jin, Youzhi; McFie, Pamela J; Banman, Shanna L; Brandt, Curtis; Stone, Scot J

    2014-10-10

    Acyl CoA:1,2-diacylglycerol acyltransferase (DGAT)-2 is an integral membrane protein that catalyzes triacylglycerol (TG) synthesis using diacylglycerol and fatty acyl CoA as substrates. DGAT2 resides in the endoplasmic reticulum (ER), but when cells are incubated with fatty acids, DGAT2 interacts with lipid droplets presumably to catalyze localized TG synthesis for lipid droplet expansion. Previous studies have shown that DGAT2 interacts with proteins that synthesize its fatty acyl CoA substrates. In this study, we provide additional evidence that DGAT2 is present in a protein complex. Using a chemical cross-linker, disuccinimidyl suberate (DSS), we demonstrated that DGAT2 formed a dimer and was also part of a protein complex of ∼ 650 kDa, both in membranes and on lipid droplets. Using co-immunoprecipitation experiments and an in situ proximity ligation assay, we found that DGAT2 interacted with monoacylglycerol acyltransferase (MGAT)-2, an enzyme that catalyzes the synthesis of diacylglycerol. Deletion mutagenesis showed that the interaction with MGAT2 was dependent on the two transmembrane domains of DGAT2. No significant interaction of DGAT2 with lipin1, another enzyme that synthesizes diacylglycerol, could be detected. When co-expressed in cells, DGAT2 and MGAT2 co-localized in the ER and on lipid droplets. Co-expression also resulted in increased TG storage compared with expression of DGAT2 or MGAT2 alone. Incubating McArdle rat hepatoma RH7777 cells with 2-monoacylglycerol caused DGAT2 to translocate to lipid droplets. This also led to the formation of large cytosolic lipid droplets, characteristic of DGAT2, but not DGAT1, and indicated that DGAT2 can utilize monoacylglycerol-derived diacylglycerol. These findings suggest that the interaction of DGAT2 and MGAT2 serves to channel lipid substrates efficiently for TG biosynthesis. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Diacylglycerol Acyltransferase-2 (DGAT2) and Monoacylglycerol Acyltransferase-2 (MGAT2) Interact to Promote Triacylglycerol Synthesis*

    Science.gov (United States)

    Jin, Youzhi; McFie, Pamela J.; Banman, Shanna L.; Brandt, Curtis; Stone, Scot J.

    2014-01-01

    Acyl CoA:1,2-diacylglycerol acyltransferase (DGAT)-2 is an integral membrane protein that catalyzes triacylglycerol (TG) synthesis using diacylglycerol and fatty acyl CoA as substrates. DGAT2 resides in the endoplasmic reticulum (ER), but when cells are incubated with fatty acids, DGAT2 interacts with lipid droplets presumably to catalyze localized TG synthesis for lipid droplet expansion. Previous studies have shown that DGAT2 interacts with proteins that synthesize its fatty acyl CoA substrates. In this study, we provide additional evidence that DGAT2 is present in a protein complex. Using a chemical cross-linker, disuccinimidyl suberate (DSS), we demonstrated that DGAT2 formed a dimer and was also part of a protein complex of ∼650 kDa, both in membranes and on lipid droplets. Using co-immunoprecipitation experiments and an in situ proximity ligation assay, we found that DGAT2 interacted with monoacylglycerol acyltransferase (MGAT)-2, an enzyme that catalyzes the synthesis of diacylglycerol. Deletion mutagenesis showed that the interaction with MGAT2 was dependent on the two transmembrane domains of DGAT2. No significant interaction of DGAT2 with lipin1, another enzyme that synthesizes diacylglycerol, could be detected. When co-expressed in cells, DGAT2 and MGAT2 co-localized in the ER and on lipid droplets. Co-expression also resulted in increased TG storage compared with expression of DGAT2 or MGAT2 alone. Incubating McArdle rat hepatoma RH7777 cells with 2-monoacylglycerol caused DGAT2 to translocate to lipid droplets. This also led to the formation of large cytosolic lipid droplets, characteristic of DGAT2, but not DGAT1, and indicated that DGAT2 can utilize monoacylglycerol-derived diacylglycerol. These findings suggest that the interaction of DGAT2 and MGAT2 serves to channel lipid substrates efficiently for TG biosynthesis. PMID:25164810

  14. Determination of the conjugated linoleic acid-containing triacylglycerols in New Zealand bovine milk fat.

    Science.gov (United States)

    Robinson, N P; MacGibbon, A K

    2000-07-01

    Reversed-phase high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection at 233 nm was used to separate, quantify, and identify the triacylglycerols (TAG) of milk fat that contain conjugated linoleic acid (CLA). The absorbance at 233 nm was substantially due to CLA-TAG (chromatography of some representative TAG devoid of CLA, such as tripalmitin and triolein, showed poor responses at 233 nm, 1/800th that of CLA-TAG). A CLA molar extinction coefficient at 233 nm of 23,360 L mol(-1) cm(-1) and an HPLC UV response factor were obtained from a commercially available cis-9,trans-11-CLA standard. This molar extinction coefficient was only 86% of reported literature values. Summation of all chromatographic peaks absorbing at 233 nm using the corrected response factor gave good agreement with independent determinations of total CLA by gas chromatography and UV spectrophotometry. This agreement allowed quantification of individual CLA-TAG peaks in the HPLC separation of a typical New Zealand bovine milk fat. Three CLA-containing TAG, CLA-dipalmitin, CLA-oleoyl-palmitin and CLA-diolein, were prepared by interesterification of tripalmitin with the respective fatty acid methyl esters and used to assign individual peaks in the reversed-phase chromatography of total milk fat, of which CLA-oleoyl-palmitin was coincident with the largest UV peak. Band fractions from argentation thin-layer chromatography of total milk fat were similarly employed to identify five predominant CLA-TAG groups in total milk fat: CLA-disaturates, CLA-oleoyl-saturates, CLA-vaccenyl-saturates, CLA-vaccenyl-olein, and CLA-diolein.

  15. Intramyocellular triacylglycerol accumulation across weight loss strategies; Sub-study of the CENTRAL trial.

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    Yftach Gepner

    Full Text Available Intramyocellular triacylglycerol (IMTG is utilized as metabolic fuel during exercise and is linked to insulin resistance, but the long-term effect of weight loss strategies on IMTG among participants with abdominal fat, remain unclear.In an 18-month trial, sedentary participants with abdominal fat/dyslipidemia were randomized to either a low-fat (LF or Mediterranean/low-carbohydrate (MED/LC diet (including 28g·day-1 of walnuts. After 6-months, the participants were re-randomized to moderate intense physical activity (PA+ or non-physical activity (PA-. Magnetic resonance imaging (MRI was used to quantify changes of IMTG, abdominal sub-depots, hepatic and intermuscular fats.Across the 277 participants [86% men, age = 48 years, body-mass-index (BMI = 31kg/m2, visceral fat = 33%] 86% completed the 18-m trial. At baseline, women had higher IMTG than men (3.4% vs. 2.3%, p<0.001 and increased IMTG was associated with aging and higher BMI, visceral and intermuscular fats, HbA1c%, HDL-c and leptin(p<0.05, but not with intra-hepatic fat. After 18 month of intervention and a -3 kg mean weight loss, participants significantly increased IMTG by 25%, with a distinct effect in the MED/LCPA+ group as compared to the other intervention groups (57% vs. 9.5-18.5%, p<0.05. Changes in IMTG were associated with visceral and intermuscular fat, metabolic syndrome, insulin and leptin (p<0.05 for all, however, these associations did not remain after adjustment for visceral fat changes.Lifestyle strategies differentially affect IMTG accumulation; combination of exercise with decreased carbohydrate/increased unsaturated fat proportion intake greatly increase IMTG. Our findings suggest that increased IMTG during diet-induced moderate weight loss may not be directly related to cardiometabolic risk.ClinicalTrials.gov NCT01530724.

  16. Lipidomic profiling of di- and tri-acylglycerol species in weight-controlled mice.

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    Brenee S King

    Full Text Available Weight control by dietary calorie restriction (DCR or exercise has been shown to prevent cancer in various models. However, the mechanisms as to how weight control is beneficial are not well understood. While previous reports have investigated the effects of weight control on total lipid levels or lipid composition within cellular membranes, there has been little work surrounding changes to individual lipids following weight control interventions. In this study, using a model of skin carcinogenesis centered on the tumor promotion stage, CD-1 mice were randomly assigned into 4 groups: ad libitum and sedentary (control, ad libitum with exercise (AL+Exe, exercise with pair feeding of a diet isocaloric with control (PF+Exe, and sedentary with 20% DCR compared to control. After ten weeks, body weight and body fat percentages significantly decreased in the PF+Exe and DCR groups but not AL+Exe when compared with sedentary controls. Murine skin and plasma samples were obtained for analysis. Lipidomics using electrospray ionization MS/MS was employed to profile triacylglycerol (TG and diacylglycerol (DG species. Both plasma and tissue TG species containing fatty acid chains with length 18:1 were significantly decreased following DCR when compared to sedentary control animals. In regards to DG, the most significant changes occurred in the plasma. DG species containing fatty acids with lengths 16:1 or 18:1 were significantly decreased in PF+Exe and DCR groups when compared to sedentary controls. Due to the significant role of TG in energy storage and DG in cellular signaling, our findings of the effects of weight control on individual TG and DG species in plasma and skin tissue following exposure to a tumor promoter, may provide insight into the mechanism of weight control on cancer prevention.

  17. Recruiting a new substrate for triacylglycerol synthesis in plants: the monoacylglycerol acyltransferase pathway.

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    James R Petrie

    Full Text Available BACKGROUND: Monoacylglycerol acyltransferases (MGATs are predominantly associated with lipid absorption and resynthesis in the animal intestine where they catalyse the first step in the monoacylglycerol (MAG pathway by acylating MAG to form diacylglycerol (DAG. Typical plant triacylglycerol (TAG biosynthesis routes such as the Kennedy pathway do not include an MGAT step. Rather, DAG and TAG are synthesised de novo from glycerol-3-phosphate (G-3-P by a series of three subsequent acylation reactions although a complex interplay with membrane lipids exists. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrate that heterologous expression of a mouse MGAT acyltransferase in Nicotiana benthamiana significantly increases TAG accumulation in vegetative tissues despite the low levels of endogenous MAG substrate available. In addition, DAG produced by this acyltransferase can serve as a substrate for both native and coexpressed diacylglycerol acyltransferases (DGAT. Finally, we show that the Arabidopsis thaliana GPAT4 acyltransferase can produce MAG in Saccharomyces cerevisiae using oleoyl-CoA as the acyl-donor. CONCLUSIONS/SIGNIFICANCE: This study demonstrates the concept of a new method of increasing oil content in vegetative tissues by using MAG as a substrate for TAG biosynthesis. Based on in vitro yeast assays and expression results in N. benthamiana, we propose that co-expression of a MAG synthesising enzyme such as A. thaliana GPAT4 and a MGAT or bifunctional M/DGAT can result in DAG and TAG synthesis from G-3-P via a route that is independent and complementary to the endogenous Kennedy pathway and other TAG synthesis routes.

  18. Arabidopsis SEIPIN Proteins Modulate Triacylglycerol Accumulation and Influence Lipid Droplet Proliferation[OPEN

    Science.gov (United States)

    2015-01-01

    The lipodystrophy protein SEIPIN is important for lipid droplet (LD) biogenesis in human and yeast cells. In contrast with the single SEIPIN genes in humans and yeast, there are three SEIPIN homologs in Arabidopsis thaliana, designated SEIPIN1, SEIPIN2, and SEIPIN3. Essentially nothing is known about the functions of SEIPIN homologs in plants. Here, a yeast (Saccharomyces cerevisiae) SEIPIN deletion mutant strain and a plant (Nicotiana benthamiana) transient expression system were used to test the ability of Arabidopsis SEIPINs to influence LD morphology. In both species, expression of SEIPIN1 promoted accumulation of large-sized lipid droplets, while expression of SEIPIN2 and especially SEIPIN3 promoted small LDs. Arabidopsis SEIPINs increased triacylglycerol levels and altered composition. In tobacco, endoplasmic reticulum (ER)-localized SEIPINs reorganized the normal, reticulated ER structure into discrete ER domains that colocalized with LDs. N-terminal deletions and swapping experiments of SEIPIN1 and 3 revealed that this region of SEIPIN determines LD size. Ectopic overexpression of SEIPIN1 in Arabidopsis resulted in increased numbers of large LDs in leaves, as well as in seeds, and increased seed oil content by up to 10% over wild-type seeds. By contrast, RNAi suppression of SEIPIN1 resulted in smaller seeds and, as a consequence, a reduction in the amount of oil per seed compared with the wild type. Overall, our results indicate that Arabidopsis SEIPINs are part of a conserved LD biogenesis machinery in eukaryotes and that in plants these proteins may have evolved specialized roles in the storage of neutral lipids by differentially modulating the number and sizes of lipid droplets. PMID:26362606

  19. Sorting cells of the microalga Chlorococcum littorale with increased triacylglycerol productivity.

    Science.gov (United States)

    Cabanelas, Iago Teles Dominguez; van der Zwart, Mathijs; Kleinegris, Dorinde M M; Wijffels, René H; Barbosa, Maria J

    2016-01-01

    Despite extensive research in the last decades, microalgae are still only economically feasible for high valued markets. Strain improvement is a strategy to increase productivities, hence reducing costs. In this work, we focus on microalgae selection: taking advantage of the natural biological variability of species to select variations based on desired characteristics. We focused on triacylglycerol (TAG), which have applications ranging from biodiesel to high-value omega-3 fatty-acids. Hence, we demonstrated a strategy to sort microalgae cells with increased TAG productivity. 1. We successfully identified sub-populations of cells with increased TAG productivity using Fluorescence assisted cell sorting (FACS). 2. We sequentially sorted cells after repeated cycles of N-starvation, resulting in five sorted populations (S1-S5). 3. The comparison between sorted and original populations showed that S5 had the highest TAG productivity [0.34 against 0.18 g l(-1) day(-1) (original), continuous light]. 4. Original and S5 were compared in lab-scale reactors under simulated summer conditions confirming the increased TAG productivity of S5 (0.4 against 0.2 g l(-1) day(-1)). Biomass composition analyses showed that S5 produced more biomass under N-starvation because of an increase only in TAG content and, flow cytometry showed that our selection removed cells with lower efficiency in producing TAGs. All combined, our results present a successful strategy to improve the TAG productivity of Chlorococcum littorale, without resourcing to genetic manipulation or random mutagenesis. Additionally, the improved TAG productivity of S5 was confirmed under simulated summer conditions, highlighting the industrial potential of S5 for microalgal TAG production.

  20. HPLC-APCI-MS analysis of triacylglycerols (TAGs) in historical pharmaceutical ointments from the eighteenth century.

    Science.gov (United States)

    Saliu, Francesco; Modugno, Francesca; Orlandi, Marco; Colombini, Maria Perla

    2011-10-01

    The lipid fractions of residues from historical pharmaceutical ointments were analysed by reversed-phase liquid chromatography coupled with atmospheric pressure chemical ionization and mass spectrometer detection. The residues were contained in a series of historical apothecary jars, dating from the eighteenth century and conserved at the "Aboca Museum" in Sansepolcro (Arezzo, Italy) and at the pharmacy of the "Real Cartuja de Valldemossa" in Palma de Majorca (Spain). The analytical protocol was set up using a comparative study based on the evaluation of triacylglycerol (TAG) compositions in raw natural lipid materials and in laboratory-reproduced ointments. These ointments were prepared following pharmaceutical recipes reported in historical treatises and used as reference materials. The reference materials were also subjected to stress treatments in order to evaluate the modification occurring in the TAG profiles as an effect of ageing. TAGs were successfully detected in the reproduced formulations even in mixtures of up to ten ingredients and after harsh degradative treatments, and also in real historical samples. No particular interferences were detected from other non-lipid ingredients of the formulations. The TAG compositions detected in the historical ointments indicated a predominant use of olive oil and pig adipose material as lipid ingredients. The detection of a high level of tristearine and myristyl-palmitoyl-stearyl glycerol in two of the samples suggested the presence of a fatty material of a different origin (maybe a ruminant). On the basis of the positional isomer ratio, sn-PPO/sn-POP, it was possible to hypothesize an exclusive use of pig fat in one sample. We also evaluated the application of principal component analysis of TAG profiles as an approach for the multivariate statistical comparison of the reference and historical ointments.

  1. Efficient production of triacylglycerols rich in docosahexaenoic acid (DHA) by osmo-heterotrophic marine protists.

    Science.gov (United States)

    Liu, Ying; Tang, Jie; Li, Jingjing; Daroch, Maurycy; Cheng, Jay J

    2014-12-01

    Thraustochytrids have recently emerged as a promising source for docosahexaenoic acid (DHA) production due to their high growth rate and oil content. In this study, two thraustochytrid isolates, Aurantiochytrium sp. PKU#SW7 and Thraustochytriidae sp. PKU#Mn16 were used for DHA production. Following growth parameters were optimized to maximize DHA production: temperature, pH, salinity, and glucose concentration. Both isolates achieved the highest DHA yield at the cultivation temperature of 28 °C, pH 6, 100 % seawater, and 2 % glucose. A DHA yield of 1.395 g/l and 1.426 g/l was achieved under the optimized culture conditions. Further investigation revealed that both isolates possess simple fatty acids profiles with palmitic acid and DHA as their dominant constituents, accounting for ∼79 % of total fatty acids. To date, very few studies have focused on the DHA distribution in various lipid fractions which is an important factor for identifying strains with a potential for industrial DHA production. In the present study, the lipids profiles of each strain both revealed that the majority of DHA was distributed in neutral lipids (NLs), and the DHA distribution in NLs of PKU#SW7 was exclusively in the form of triacylglycerols (TAGs) which suggest that PKU#SW7 could be utilized as an alternative source of DHA for dietary supplements. The fermentation process established for both strains also indicating that Aurantiochytrium sp. PKU#SW7 was more suitable for cultivation in fermenter. In addition, the high percentage of saturated fatty acids produced by the two thraustochytrids indicates their potential application in biodiesel production. Overall, our findings suggest that two thraustochytrid isolates are suitable candidates for biotechnological applications.

  2. HPLC separation of triacylglycerol positional isomers on a polymeric ODS column.

    Science.gov (United States)

    Kuroda, Ikuma; Nagai, Toshiharu; Mizobe, Hoyo; Yoshimura, Nobuhito; Gotoh, Naohiro; Wada, Shun

    2008-07-01

    A polymeric ODS column was applied to the resolution of triacylglycerol positional isomers (TAG-PI), i.e. 1,3-dioleoyl-2-palmitoyl-glycerol (OPO) and 1,2-dioleoyl-3-palmitoyl-rac-glycerol (OOP), with a recycle HPLC system. To investigate the ODS column species and the column temperatures for the resolution of a TAG-PI pair, a mixture of OPO and OOP was subjected to an HPLC system equipped with a non-endcapped polymeric, endcapped monomeric, endcapped intermediate, or non-endcapped monomeric ODS column at three different column temperatures (40, 25, or 10 degrees C). Only the non-endcapped polymeric ODS column achieved the separation of OPO and OOP, and the lowest column temperature (10 degrees C) showed the best resolution for them. The other pair of TAG-PI, a mixture of 1,3-dipalmitoyl-2-oleoyl-glycerol (POP) and 1,2-dipalmitoyl-3-oleoyl-rac-glycerol (PPO) was also subjected to the system equipped with a non-endcapped polymeric or monomeric ODS column at five different column temperatures (40, 32, 25, 17, and 10 degrees C). Thus, POP and PPO were also separated on only the non-endcapped polymeric ODS column at 25 degrees C. However, no clear peak appeared at 10 degrees C. These results would indicate that the polymeric ODS stationary phase has an ability to recognize the structural differences between TAG-PI pairs. Also, the column temperature is a very important factor for separating the TAG-PI pair, and the optimal temperature would relate to the solubility of TAG-PI in the mobile phase. Furthermore, the recycle HPLC system provided measurements for the separation and analysis of TAG-PI pairs.

  3. Impact of temperature and light intensity on triacylglycerol accumulation in marine microalgae

    International Nuclear Information System (INIS)

    Kurpan Nogueira, Daniel P.; Silva, Anita F.; Araújo, Ofélia Q.F.; Chaloub, Ricardo M.

    2015-01-01

    Triacylglycerol (TAG) productivity of Isochrysis galbana, Nannochloropsis oceanica and Phaeodactylum tricornutum was compared to study their suitability for biotechnological applications. Photoautotrophic batch cultures grown at 20 °C and 50 μmol photons m −2  s −1 showed that N. oceanica had the least TAG content and TAG productivity of the three microalgae. Hence, effects of temperature and light intensity on growth rate and accumulation of TAG were subsequently assessed only in I. galbana and P. tricornutum by cultivation at 20 and 30 °C under 50, 300 and 600 μmol photons m −2  s −1 . Although P. tricornutum did not grow at temperatures higher than 20 °C, an increase in both TAG content (from 28.37 to 39.53%) and productivity (from 15.58 to 31.39 mg L −1  d −1 ) was observed at the highest irradiance values. We also found that combined effects of temperature and light intensity enhanced TAG content (from 18.59 to 31.71%) and productivity (from 11.76 to 21.67 mg L −1  d −1 ) in I. galbana. - Highlights: • Productivity of oil and biomass in batch-cultured marine microalgae was compared. • Increase in temperature and irradiance rose oil productivity in Isochrysis galbana. • An increase in light intensity rose oil productivity in Phaeodactylum tricornutum. • Phaeodactylum tricornutum showed the highest productivity in biomass and neutral lipids

  4. Conjugated Linoleic Triacylglycerols Exhibit Superior Lymphatic Absorption Than Free Conjugate Linoleic Acids and Have Antiobesity Properties.

    Science.gov (United States)

    Woo, Hyunjoon; Chung, Min-Yu; Kim, Juyeon; Kong, Daecheol; Min, Jinyoung; Choi, Hee-Don; Choi, In-Wook; Kim, In-Hwan; Noh, Sang K; Kim, Byung Hee

    2016-05-01

    This study aimed to compare lymphatic absorption of conjugated linoleic acids (CLAs) in the triacylglycerol (TAG) or free fatty acid (FFA) form and to examine the antiobesity effects of different doses of CLAs in the TAG form in animals. Conjugated linoleic TAGs (containing 70.3 wt% CLAs; CLA-TAG) were prepared through lipase-catalyzed esterification of glycerol with commercial CLA mixtures (CLA-FFA). Lymphatic absorption of CLA-TAG and CLA-FFA was compared in a rat model of lymphatic cannulation. Greater amounts of cis-9,trans-11 and trans-10,cis-12 CLAs were detected in the collected lymph from a lipid emulsion containing CLA-TAG. This result suggests that CLA-TAG has greater capacity for lymphatic absorption than does CLA-FFA. The antiobesity efficacy of CLA-TAG at different doses was examined in mice with diet-induced obesity. A high-fat diet (HFD) for 12 weeks caused a significant increase in body weight and epididymal and retroperitoneal fat weights, which were significantly decreased by 2% dietary supplementation (w/w) with CLA-TAG. CLA-TAG at 2% significantly attenuated the HFD-induced upregulation of serum TAG, but led to hepatomegaly and exacerbated HFD-induced hypercholesterolemia. CLA-TAG at 1% significantly attenuated upregulation of retroperitoneal fat weight and significantly increased liver weight, which was decreased by the HFD. Nonetheless, the liver weight in group "HFD +1% CLA-TAG" was not significantly different from that of normal diet controls. CLA-TAG at 1% significantly reduced serum TAG levels and did not exacerbate HFD-induced hypercholesterolemia. Thus, 1% dietary supplementation with CLA-TAG reduces retroperitoneal fat weight without apparent hepatomegaly, a known side-effect of CLAs in mouse models of obesity.

  5. The Antiphagocytic Activity of SeM of Streptococcus equi Requires Capsule.

    Science.gov (United States)

    Timoney, John F; Suther, Pranav; Velineni, Sridhar; Artiushin, Sergey C

    2014-01-01

    Resistance to phagocytosis is a crucial virulence property of Streptococcus equi (Streptococcus equi subsp. equi; Se), the cause of equine strangles. The contribution and interdependence of capsule and SeM to killing in equine blood and neutrophils were investigated in naturally occurring strains of Se. Strains CF32, SF463 were capsule and SeM positive, strains Lex90, Lex93 were capsule negative and SeM positive and strains Se19, Se1-8 were capsule positive and SeM deficient. Phagocytosis and killing of Se19, Se1-8, Lex90 and Lex93 in equine blood and by neutrophils suspended in serum were significantly (P ≤ 0.02) greater compared to CF32 and SF463. The results indicate capsule and SeM are both required for resistance to phagocytosis and killing and that the anti-phagocytic property of SeM is greatly reduced in the absence of capsule.

  6. Replacement of Retinyl Esters by Polyunsaturated Triacylglycerol Species in Lipid Droplets of Hepatic Stellate Cells during Activation

    Science.gov (United States)

    Testerink, Nicole; Ajat, Mokrish; Houweling, Martin; Brouwers, Jos F.; Pully, Vishnu V.; van Manen, Henk-Jan; Otto, Cees; Helms, J. Bernd; Vaandrager, Arie B.

    2012-01-01

    Activation of hepatic stellate cells has been recognized as one of the first steps in liver injury and repair. During activation, hepatic stellate cells transform into myofibroblasts with concomitant loss of their lipid droplets (LDs) and production of excessive extracellular matrix. Here we aimed to obtain more insight in the dynamics and mechanism of LD loss. We have investigated the LD degradation processes in rat hepatic stellate cells in vitro with a combined approach of confocal Raman microspectroscopy and mass spectrometric analysis of lipids (lipidomics). Upon activation of the hepatic stellate cells, LDs reduce in size, but increase in number during the first 7 days, but the total volume of neutral lipids did not decrease. The LDs also migrate to cellular extensions in the first 7 days, before they disappear. In individual hepatic stellate cells. all LDs have a similar Raman spectrum, suggesting a similar lipid profile. However, Raman studies also showed that the retinyl esters are degraded more rapidly than the triacylglycerols upon activation. Lipidomic analyses confirmed that after 7 days in culture hepatic stellate cells have lost most of their retinyl esters, but not their triacylglycerols and cholesterol esters. Furthermore, we specifically observed a large increase in triacylglycerol-species containing polyunsaturated fatty acids, partly caused by an enhanced incorporation of exogenous arachidonic acid. These results reveal that lipid droplet degradation in activated hepatic stellate cells is a highly dynamic and regulated process. The rapid replacement of retinyl esters by polyunsaturated fatty acids in LDs suggests a role for both lipids or their derivatives like eicosanoids during hepatic stellate cell activation. PMID:22536341

  7. Replacement of retinyl esters by polyunsaturated triacylglycerol species in lipid droplets of hepatic stellate cells during activation.

    Directory of Open Access Journals (Sweden)

    Nicole Testerink

    Full Text Available Activation of hepatic stellate cells has been recognized as one of the first steps in liver injury and repair. During activation, hepatic stellate cells transform into myofibroblasts with concomitant loss of their lipid droplets (LDs and production of excessive extracellular matrix. Here we aimed to obtain more insight in the dynamics and mechanism of LD loss. We have investigated the LD degradation processes in rat hepatic stellate cells in vitro with a combined approach of confocal Raman microspectroscopy and mass spectrometric analysis of lipids (lipidomics. Upon activation of the hepatic stellate cells, LDs reduce in size, but increase in number during the first 7 days, but the total volume of neutral lipids did not decrease. The LDs also migrate to cellular extensions in the first 7 days, before they disappear. In individual hepatic stellate cells. all LDs have a similar Raman spectrum, suggesting a similar lipid profile. However, Raman studies also showed that the retinyl esters are degraded more rapidly than the triacylglycerols upon activation. Lipidomic analyses confirmed that after 7 days in culture hepatic stellate cells have lost most of their retinyl esters, but not their triacylglycerols and cholesterol esters. Furthermore, we specifically observed a large increase in triacylglycerol-species containing polyunsaturated fatty acids, partly caused by an enhanced incorporation of exogenous arachidonic acid. These results reveal that lipid droplet degradation in activated hepatic stellate cells is a highly dynamic and regulated process. The rapid replacement of retinyl esters by polyunsaturated fatty acids in LDs suggests a role for both lipids or their derivatives like eicosanoids during hepatic stellate cell activation.

  8. Emergence of anti-red blood cell antibodies triggers red cell phagocytosis by activated macrophages in a rabbit model of Epstein-Barr virus-associated hemophagocytic syndrome.

    Science.gov (United States)

    Hsieh, Wen-Chuan; Chang, Yao; Hsu, Mei-Chi; Lan, Bau-Shin; Hsiao, Guan-Chung; Chuang, Huai-Chia; Su, Ih-Jen

    2007-05-01

    Hemophagocytic syndrome (HPS) is a fatal complication frequently associated with viral infections. In childhood HPS, Epstein-Barr virus (EBV) is the major causative agent, and red blood cells (RBCs) are predominantly phagocytosed by macrophages. To investigate the mechanism of RBC phagocytosis triggered by EBV infection, we adopted a rabbit model of EBV-associated HPS previously established by using Herpesvirus papio (HVP). The kinetics of virus-host interaction was studied. Using flow cytometry, we detected the emergence of antibody-coated RBCs, as well as anti-platelet antibodies, at peak virus load period at weeks 3 to 4 after HVP injection, and the titers increased thereafter. The presence of anti-RBCs preceded RBC phagocytosis in tissues and predicted the full-blown development of HPS. The anti-RBC antibodies showed cross-reactivity with Paul-Bunnell heterophile antibodies. Preabsorption of the HVP-infected serum with control RBCs removed the majority of anti-RBC activities and remarkably reduced RBC phagocytosis. The RBC phagocytosis was specifically mediated via an Fc fragment of antibodies in the presence of macrophage activation. Therefore, the emergence of anti-RBC antibodies and the presence of macrophage activation are both essential in the development of HPS. Our observations in this animal model provide a potential mechanism for hemophagocytosis in EBV infection.

  9. IrAM—An .alpha..sub.2./sub.-macroglobulin from the hard tick Ixodes ricinus: Characterization and function in phagocytosis of a potential pathogen Chryseobacterium indologenes

    Czech Academy of Sciences Publication Activity Database

    Burešová, Veronika; Hajdušek, Ondřej; Franta, Zdeněk; Sojka, Daniel; Kopáček, Petr

    2009-01-01

    Roč. 33, č. 4 (2009), s. 489-198 ISSN 0145-305X R&D Projects: GA AV ČR IAA600220603; GA MŠk(CZ) LC06009 Institutional research plan: CEZ:AV0Z60220518 Keywords : α2-macroglobulin * tick * phagocytosis Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.290, year: 2009

  10. Apparent and quantitative loss of fatty acids and triacylglycerols at frying temperatures

    Directory of Open Access Journals (Sweden)

    Berdeaux, O.

    2012-09-01

    Full Text Available Olive and sunflower oils were heated at 180 °C for 5, 10 and 15 hours with the aim of defining the changes in the contents of fatty acid methyl esters (FAME and triacylglycerols (TAG under frying conditions. Differences between apparent and real loss of FAME or TAG are defined for an adequate interpretation of the changes taking place at high temperature. Such differences depend on the expression of the results and frequently result in erroneous conclusions. Results showed that from the normalized composition it could be deduced that only the most unsaturated FAME (C18:2 or TAG containing it was significantly altered. However, quantitative data indicated that all the unsaturated FAME were degraded in samples of olive oil and sunflower oil whose levels of alteration ranged from 14.6 to 35.4% polar compounds. With regard to TAG, their loss depended on both the number of double bonds and the concentration of the linoleyl group, which is the most unsaturated fatty acyl group in both oils.

    Aceites de oliva y girasol se calentaron a 180 °C durante 5, 10 y 15 horas con el objetivo de definir los cambios en la concentración de ácidos grasos y especies de triglicéridos en condiciones de fritura. Se definen las diferencias entre la pérdida aparente y cuantitativa de ácidos grasos y triglicéridos que dependen de la forma de expresión de los resultados dando lugar en muchas ocasiones a interpretaciones erróneas. Así, a partir de la composición normalizada, podría deducirse que sólo el ácido graso más insaturado en los aceites (C18:2 se altera. Sin embargo, los datos cuantitativos muestran claramente que en muestras con niveles de compuestos polares entre 14.6 y 35.4%, todos los ácidos grasos insaturados sufren degradación significativa en ambos aceites. Con respecto a los cambios en los distintos triglicéridos, su pérdida cuantitativa dependió no sólo de su número de dobles enlaces sino también del contenido del ácido m

  11. Synthesis of 2-monoacylglycerols and structured triacylglycerols rich in polyunsaturated fatty acids by enzyme catalyzed reactions.

    Science.gov (United States)

    Rodríguez, Alicia; Esteban, Luis; Martín, Lorena; Jiménez, María José; Hita, Estrella; Castillo, Beatriz; González, Pedro A; Robles, Alfonso

    2012-08-10

    This paper studies the synthesis of structured triacylglycerols (STAGs) by a four-step process: (i) obtaining 2-monoacylglycerols (2-MAGs) by alcoholysis of cod liver oil with several alcohols, catalyzed by lipases Novozym 435, from Candida antartica and DF, from Rhizopus oryzae, (ii) purification of 2-MAGs, (iii) formation of STAGs by esterification of 2-MAGs with caprylic acid catalyzed by lipase DF, from R. oryzae, and (iv) purification of these STAGs. For the alcoholysis of cod liver oil, absolute ethanol, ethanol 96% (v/v) and 1-butanol were compared; the conditions with ethanol 96% were then optimized and 2-MAG yields of around 54-57% were attained using Novozym 435. In these 2-MAGs, DHA accounted for 24-31% of total fatty acids. In the operational conditions this lipase maintained a stable level of activity over at least 11 uses. These results were compared with those obtained with lipase DF, which deactivated after only three uses. The alcoholysis of cod liver oil and ethanol 96% catalyzed by Novozym 435 was scaled up by multiplying the reactant amounts 100-fold and maintaining the intensity of treatment constant (IOT=3g lipase h/g oil). In these conditions, the 2-MAG yield attained was about 67%; these 2-MAGs contained 36.6% DHA. The synthesized 2-MAGs were separated and purified from the alcoholysis reaction products by solvent extraction using solvents of low toxicity (ethanol and hexane); 2-MAG recovery yield and purity of the target product were approximately 96.4% and 83.9%, respectively. These 2-MAGs were transformed to STAGs using the optimal conditions obtained in a previous work. After synthesis and purification, 93% pure STAGs were obtained, containing 38% DHA at sn-2 position and 60% caprylic acid (CA) at sn-1,3 positions (of total fatty acids at these positions), i.e. the major TAG is the STAG with the structure CA-DHA-CA. Copyright © 2012 Elsevier Inc. All rights reserved.

  12. Metabolic link between phosphatidylethanolamine and triacylglycerol metabolism in the yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Horvath, Susanne E; Wagner, Andrea; Steyrer, Ernst; Daum, Günther

    2011-12-01

    In the yeast Saccharomyces cerevisiae triacylglycerols (TAG) are synthesized by the acyl-CoA dependent acyltransferases Dga1p, Are1p, Are2p and the acyl-CoA independent phospholipid:diacylglycerol acyltransferase (PDAT) Lro1p which uses phosphatidylethanolamine (PE) as a preferred acyl donor. In the present study we investigated a possible link between TAG and PE metabolism by analyzing the contribution of the four different PE biosynthetic pathways to TAG formation, namely de novo PE synthesis via Psd1p and Psd2p, the CDP-ethanolamine (CDP-Etn) pathway and lyso-PE acylation by Ale1p. In cells grown on the non-fermentable carbon source lactate supplemented with 5mM ethanolamine (Etn) the CDP-Etn pathway contributed most to the cellular TAG level, whereas mutations in the other pathways displayed only minor effects. In cki1∆dpl1∆eki1∆ mutants bearing defects in the CDP-Etn pathway both the cellular and the microsomal levels of PE were markedly decreased, whereas in other mutants of PE biosynthetic routes depletion of this aminoglycerophospholipid was less pronounced in microsomes. This observation is important because Lro1p similar to the enzymes of the CDP-Etn pathway is a component of the ER. We conclude from these results that in cki1∆dpl1∆eki1∆ insufficient supply of PE to the PDAT Lro1p was a major reason for the strongly reduced TAG level. Moreover, we found that Lro1p activity was markedly decreased in cki1∆dpl1∆eki1∆, although transcription of LRO1 was not affected. Our findings imply that (i) TAG and PE syntheses in the yeast are tightly linked; and (ii) TAG formation by the PDAT Lro1p strongly depends on PE synthesis through the CDP-Etn pathway. Moreover, it is very likely that local availability of PE in microsomes is crucial for TAG synthesis through the Lro1p reaction. Copyright © 2011 Elsevier B.V. All rights reserved.

  13. Effect of UVB irradiation of the blood on cellular volume adherence and phagocytosis in probands and patients with multiple sclerosis

    Energy Technology Data Exchange (ETDEWEB)

    Mix, E; Jenssen, H L; Lehmitz, R; Buddenhagen, F; Hitzschke, B; Richter, M

    1988-01-01

    UVB-induced changes of blood cell properties were investigated in 12 multiple sclerosis (MS) patients and in 10 healthy volunteers. The mean cell volume (MCV) was determined by electronic sizing, the granulocyte and lymphocyte adherence was estimated in a capillary assay, and the phagocytic activity of granulocytes was measured in a test system based on the incorporation of opsonized baker's yeast. In MS patients the MCV of red cells and lymphocytes decreased rapidly within 6 UVB treatments. In contrast, the reduction of the granulocyte volume was delayed (between the 6th and 12th UVB). In the control group the mean value of the red cell and lymphocyte MCV remained rather unaffected. There was a slight rise of the granulocyte volume after the 6th UVB. The only significant change of adherence was an increase of granulocyte adherence in MS patients. Untreated patients had a significantly enhanced phagocytic activity in comparison to the control group. 6 UVB treatments induced a singificant reduction of the phagocytic activity in MS patients. However, subsequently the percentage of phagocytizing cells increased again, whereas the particle uptake per cell continued to decrease. In the control group only minor UVB-induced changes of phagocytosis were observed. The in vitro UV irradiation caused an enhanced phagocytosis in the majority of cases in both controls and MS patients. In general, under the UVB treatment all parameters examined changed in the sense of a normalisation, in that the measured values reached a new level lying between the extreme pretreatment values accompanied by a reduced standard deviation. The effect of UVB was more pronounced in MS patients when compared with normal control. This could result from an enhanced sensitivity to the influence of UVB of pathologically altered cells in MS patients. (Abstract Truncated)

  14. Effect of trastuzumab interchain disulfide bond cleavage on Fcγ receptor binding and antibody-dependent tumour cell phagocytosis.

    Science.gov (United States)

    Suzuki, Mami; Yamanoi, Ayaka; Machino, Yusuke; Ootsubo, Michiko; Izawa, Ken-ichi; Kohroki, Junya; Masuho, Yasuhiko

    2016-01-01

    The Fc domain of human IgG1 binds to Fcγ receptors (FcγRs) to induce effector functions such as phagocytosis. There are four interchain disulfide bonds between the H and L chains. In this study, the disulfide bonds within the IgG1 trastuzumab (TRA), which is specific for HER2, were cleaved by mild S-sulfonation or by mild reduction followed by S-alkylation with three different reagents. The cleavage did not change the binding activities of TRA to HER2-bearing SK-BR-3 cells. The binding activities of TRA to FcγRIIA and FcγRIIB were greatly enhanced by modification with mild reduction and S-alkylation with ICH2CONH2 or N-(4-aminophenyl) maleimide, while the binding activities of TRA to FcγRI and FcγRIIIA were decreased by any of the four modifications. However, the interchain disulfide bond cleavage by the different modifications did not change the antibody-dependent cell-mediated phagocytosis (ADCP) of SK-BR-3 cells by activated THP-1 cells. The order of FcγR expression levels on the THP-1 cells was FcγRII > FcγRI > FcγRIII and ADCP was inhibited by blocking antibodies against FcγRI and FcγRII. These results imply that the effect of the interchain disulfide bond cleavage on FcγRs binding and ADCP is dependent on modifications of the cysteine residues and the FcγR isotypes. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  15. Distribution of fatty acids from dietary oils into phospholipid classes of triacylglycerol-rich lipoproteins in healthy subjects.

    Science.gov (United States)

    Abia, Rocio; Pacheco, Yolanda M; Montero, Emilio; Ruiz-Gutierrez, Valentina; Muriana, Francisco J G

    2003-02-21

    Several studies have suggested that lipoprotein metabolism can be affected by lipoprotein phospholipid composition. We investigated the effect of virgin olive oil (VOO) and high-oleic sunflower oil (HOSO) intake on the distribution of fatty acids in triacylglycerols (TG), cholesteryl esters (CE) and phospholipid (PL) classes of triacylglycerol-rich lipoproteins (TRL) from normolipidemic males throughout a 7 h postprandial metabolism. Particularly, changes in oleic acid (18:1n-9) concentration of PL were used as a marker of in vivo hydrolysis of TRL external monolayer. Both oils equally promoted the incorporation of oleic acid into the TG and CE of postprandial TRL. However, PL was enriched in oleic acid (18:1n-9) and n-3 polyunsaturated fatty acids (PUFA) after VOO meal, whereas in stearic (18:0) and linoleic (18:2n-6) acids after HOSO meal. We also found that VOO produced TRL which PL 18:1n-9 content was dramatically reduced along the postprandial period. We conclude that the fatty acid composition of PL can be a crucial determinant for the clearance of TRL during the postprandial metabolism of fats.

  16. Analysis and optimization of triacylglycerol synthesis in novel oleaginous Rhodococcus and Streptomyces strains isolated from desert soil.

    Science.gov (United States)

    Röttig, Annika; Hauschild, Philippa; Madkour, Mohamed H; Al-Ansari, Ahmed M; Almakishah, Naief H; Steinbüchel, Alexander

    2016-05-10

    As oleaginous microorganisms represent an upcoming novel feedstock for the biotechnological production of lipids or lipid-derived biofuels, we searched for novel, lipid-producing strains in desert soil. This was encouraged by the hypothesis that neutral lipids represent an ideal storage compound, especially under arid conditions, as several animals are known to outlast long periods in absence of drinking water by metabolizing their body fat. Ten lipid-accumulating bacterial strains, affiliated to the genera Bacillus, Cupriavidus, Nocardia, Rhodococcus and Streptomyces, were isolated from arid desert soil due to their ability to synthesize poly(β-hydroxybutyrate), triacylglycerols or wax esters. Particularly two Streptomyces sp. strains and one Rhodococcus sp. strain accumulate significant amounts of TAG under storage conditions under optimized cultivation conditions. Rhodococcus sp. A27 and Streptomyces sp. G49 synthesized approx. 30% (w/w) fatty acids from fructose or cellobiose, respectively, while Streptomyces isolate G25 reached a cellular fatty acid content of nearly 50% (w/w) when cultivated with cellobiose. The stored triacylglycerols were composed of 30-40% branched fatty acids, such as anteiso-pentadecanoic or iso-hexadecanoic acid. To date, this represents by far the highest lipid content described for streptomycetes. A biotechnological production of such lipids using (hemi)cellulose-derived raw material could be used to obtain sustainable biodiesel with a high proportion of branched-chain fatty acids to improve its cold-flow properties and oxidative stability. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. The Use of Atmospheric Pressure Chemical Ionization Mass Spectrometry with High Performance Liquid Chromatography and Other Separation Techniques for Identification of Triacylglycerols

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Sigler, Karel

    2007-01-01

    Roč. 3, - (2007), s. 252-271 ISSN 1573-4110 R&D Projects: GA ČR GA203/06/0219 Institutional research plan: CEZ:AV0Z50200510 Keywords : triacylglycerols * atmospheric presssure chemical ionization * mass spectrometry Subject RIV: EE - Microbiology, Virology Impact factor: 1.815, year: 2007

  18. A plant stanol yogurt drink alone or combined with a low-dose statin lowers serum triacylglycerol and non-HDL cholesterol in metabolic syndrome patients

    NARCIS (Netherlands)

    Plat, Jogchum; Brufau, Gemma; Dallinga-Thie, Geesje M.; Dasselaar, Margreet; Mensink, Ronald P.

    2009-01-01

    We evaluated the effects of 2 commonly available strategies (plant stanol ester drink and 10 mg simvastatin) on coronary heart disease (CHD) risk variables in participants with metabolic syndrome. Metabolic syndrome patients are at increased risk to develop CHD, partly due to high triacylglycerol

  19. Regioisomeric analysis of triacylglycerols using silver-ion liquid chromatography-atmospheric pressure chemical ionization mass spectrometry: Comparison of five different mass analyzers

    Czech Academy of Sciences Publication Activity Database

    Holčapek, M.; Dvořáková, H.; Lísa, M.; Girón, A. J.; Sandra, P.; Cvačka, Josef

    2010-01-01

    Roč. 1217, č. 52 (2010), s. 8186-8194 ISSN 0021-9673 R&D Projects: GA ČR GA203/09/0139 Institutional research plan: CEZ:AV0Z40550506 Keywords : triacylglycerols * silver-ion liquid chromatography * regioisomer * HPLC/MS * APCI Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.194, year: 2010

  20. Consumption of diets with different type of fat influences triacylglycerols-rich lipoproteins particle number and size during the postprandial state

    Science.gov (United States)

    Background and aims: Previous evidence suggests that dietary fat could influence the composition and size of triacylglycerols-rich lipoproteins (TRL). In a controlled intervention study on healthy subjects, we evaluated the influence of 3 dietary interventions, with different types of fat on postpra...

  1. HIV-1-Specific IgA Monoclonal Antibodies from an HIV-1 Vaccinee Mediate Galactosylceramide Blocking and Phagocytosis

    Science.gov (United States)

    2018-01-01

    ABSTRACT Vaccine-elicited humoral immune responses comprise an array of antibody forms and specificities, with only a fraction contributing to protective host immunity. Elucidation of antibody effector functions responsible for protective immunity against human immunodeficiency virus type 1 (HIV-1) acquisition is a major goal for the HIV-1 vaccine field. Immunoglobulin A (IgA) is an important part of the host defense against pathogens; however, little is known about the role of vaccine-elicited IgA and its capacity to mediate antiviral functions. To identify the antiviral functions of HIV-1-specific IgA elicited by vaccination, we cloned HIV-1 envelope-specific IgA monoclonal antibodies (MAbs) by memory B cell cultures from peripheral blood mononuclear cells from an RV144 vaccinee and produced two IgA clonal cell lines (HG129 and HG130) producing native, nonrecombinant IgA MAbs. The HG129 and HG130 MAbs mediated phagocytosis by monocytes, and HG129 blocked HIV-1 Env glycoprotein binding to galactosylceramide, an alternative HIV-1 receptor. These findings elucidate potential antiviral functions of vaccine-elicited HIV-1 envelope-specific IgA that may act to block HIV-1 acquisition at the portal of entry by preventing HIV-1 binding to galactosylceramide and mediating antibody Fc receptor-mediated virion phagocytosis. Furthermore, these findings highlight the complex and diverse interactions of vaccine-elicited IgA with pathogens that depend on IgA fine specificity and form (e.g., multimeric or monomeric) in the systemic circulation and mucosal compartments. IMPORTANCE Host-pathogen interactions in vivo involve numerous immune mechanisms that can lead to pathogen clearance. Understanding the nature of antiviral immune mechanisms can inform the design of efficacious HIV-1 vaccine strategies. Evidence suggests that both neutralizing and nonneutralizing antibodies can mediate some protection against HIV in animal models. Although numerous studies have characterized the

  2. Determination and Comparison of Seed Oil Triacylglycerol Composition of Various Soybeans (Glycine max (L. Using 1H-NMR Spectroscopy

    Directory of Open Access Journals (Sweden)

    Won Woo Kim

    2013-11-01

    Full Text Available Seed oil triacylglycerol (TAG composition of 32 soybean varieties were determined and compared using 1H-NMR. The contents of linolenic (Ln, linoleic (L, and oleic (O ranged from 10.7% to 19.3%, 37.4%–50.1%, and 15.7%–34.1%, respectively. As is evident, linoleic acid was the major fatty acid of soybean oil. Compositional differences among the varieties were observed. Natural oils containing unsaturated groups have been regarded as important nutrient and cosmetic ingredients because of their various biological activities. The TAG profiles of the soy bean oils could be useful for distinguishing the origin of seeds and controlling the quality of soybean oils. To the best of our knowledge, this is the first study in which the TAG composition of various soybean oils has been analyzed using the 1H-NMR method.

  3. Impact of alpha-, gamma-, and delta-tocopherol on the radiation induced oxidation of rapeseed oil triacylglycerols

    Energy Technology Data Exchange (ETDEWEB)

    Braunrath, Robert; Isnardy, Bettina; Solar, Sonja, E-mail: sonja.solar@univie.ac.at; Elmadfa, Ibrahim

    2010-07-15

    Gamma-irradiation (doses: 2, 4, 7, and 10 kGy) was used as oxidation tool to study the antioxidant effects of alpha-, gamma-, and delta-tocopherol (enrichments 500-5000 ppm) in purified rapeseed oil triacylglycerols (RSOTG). Fatty acid composition, tocopherol degradation, primary (conjugated dienes (CD) and peroxide value (POV)) and secondary (p-anisidine value) oxidation products were chosen as test parameters. Fatty acid composition did not change. While secondary oxidation products could not be found in the irradiated samples, the POVs and CDs showed a significant, dose-dependent increase. alpha-Tocopherol did not inhibit the formation of peroxides, whereas gamma- and delta-tocopherol reduced the POVs by more than 30%. No uniform effect of the different tocopherol concentrations at the particular doses could be established. The influence of the individual tocopherols on the CD formation was not pronounced. The degradation of the tocopherols decreased with increasing concentration. None of the tocopherols showed a prooxidant effect.

  4. Quantitation of acyl migration during lipase-catalyzed acidolysis, and of the regioisomers of structured triacylglycerols formed

    DEFF Research Database (Denmark)

    Mu, Huiling; Kurvinen, J.P.; Kallio, H.

    2001-01-01

    degradation, and ranged from 39.0 to 48.7% and 0.6 to 9.3%, respectively. Quantitation of triacylglycerol molecular species was performed by ammonia negative ion chemical ionization (NICI) mass spectrometry (MS). The proportion of ACN (acyl carbon number) 34 species that contained one C-18 fatty acid and two...... C-8:0, in samples analyzed, varied from 12.5 to 23.2%. The selected regioisomers MLM and MML within the ACN 34 species group were quantified by NICI tandem MS (MS/MS) and were in the range of 97.1 to 98.4% and 1.6 to 2.9%, respectively. There was no correlation between the level of acyl migration...

  5. Impact of α-, γ-, and δ-tocopherol on the radiation induced oxidation of rapeseed oil triacylglycerols

    International Nuclear Information System (INIS)

    Braunrath, Robert; Isnardy, Bettina; Solar, Sonja; Elmadfa, Ibrahim

    2010-01-01

    Gamma-irradiation (doses: 2, 4, 7, and 10 kGy) was used as oxidation tool to study the antioxidant effects of α-, γ-, and δ-tocopherol (enrichments 500-5000 ppm) in purified rapeseed oil triacylglycerols (RSOTG). Fatty acid composition, tocopherol degradation, primary (conjugated dienes (CD) and peroxide value (POV)) and secondary (p-anisidine value) oxidation products were chosen as test parameters. Fatty acid composition did not change. While secondary oxidation products could not be found in the irradiated samples, the POVs and CDs showed a significant, dose-dependent increase. α-Tocopherol did not inhibit the formation of peroxides, whereas γ- and δ-tocopherol reduced the POVs by more than 30%. No uniform effect of the different tocopherol concentrations at the particular doses could be established. The influence of the individual tocopherols on the CD formation was not pronounced. The degradation of the tocopherols decreased with increasing concentration. None of the tocopherols showed a prooxidant effect.

  6. Effect of dietary fatty acids on the postprandial fatty acid composition of triacylglycerol-rich lipoproteins in healthy male subjects

    DEFF Research Database (Denmark)

    Bysted, Anette; Holmer, G.; Lund, Pia

    2005-01-01

    interesterified test fats with equal amounts of palmitic acid ( P fat), stearic acid (S fat), trans-18: 1 isomers (T fat), oleic acid (O fat), or linoleic acid (L fat) were tested. Subjects: A total of 16 healthy, normolipidaemic males ( age 23 +/- 2 y) were recruited. Interventions: The participants ingested fat......Objective: The aim of the present study was to investigate the effect of trans-18: 1 isomers compared to other fatty acids, especially saturates, on the postprandial fatty acid composition of triacylglycerols ( TAG) in chylomicrons and VLDL. Design: A randomised crossover experiment where five......-rich test meals ( 1 g fat per kg body weight) and the fatty acid profiles of chylomicron and VLDL TAG were followed for 8 h. Results: The postprandial fatty acid composition of chylomicron TAG resembled that of the ingested fats. The fatty acids in chylomicron TAG were randomly distributed among the three...

  7. A simple liquid extraction protocol for overcoming the ion suppression of triacylglycerols by phospholipids in liquid chromatography mass spectrometry studies.

    Science.gov (United States)

    Araujo, Pedro; Tilahun, Ephrem; Breivik, Joar Fjørtoft; Abdulkader, Bashir M; Frøyland, Livar; Zeng, Yingxu

    2016-02-01

    It is well-known that triacylglycerol (TAG) ions are suppressed by phospholipid (PL) ions in regiospecific analysis of TAG by mass spectrometry (MS). Hence, it is essential to remove the PL during sample preparation prior to MS analysis. The present article proposes a cost-effective liquid-liquid extraction (LLE) method to remove PL from TAG in different kinds of biological samples by using methanol, hexane and water. High performance thin layer chromatography confirmed the lack of PL in krill oil and salmon liver samples, submitted to the proposed LLE protocol, and liquid chromatography tandem MS confirmed that the identified TAG ions were highly enhanced after implementing the LLE procedure. Copyright © 2015 Elsevier B.V. All rights reserved.

  8. Simultaneous Production of Triacylglycerol and High-Value Carotenoids by the Astaxanthin-Producing Oleaginous Green Microalga Chlorella zofingiensis

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jin; Mao, Xuemei; Zhou, Wenguang; Guarnieri, Michael T.

    2016-08-01

    The production of lipids and astaxanthin, a high-value carotenoid, by Chlorella zofingiensis was investigated under different culture conditions. Comparative analysis revealed a good correlation between triacylglycerol (TAG) and astaxanthin accumulation in C. zofingiensis. Stress conditions promoted cell size and weight and induced the accumulation of neutral lipids, especially TAG and astaxanthin, with a concomitant decrease in membrane lipids. The highest contents of TAG and astaxanthin achieved were 387 and 4.89 mg g-1 dry weight, respectively. A semi-continuous culture strategy was developed to optimize the TAG and astaxanthin productivities, which reached 297 and 3.3 mg L-1 day-1, respectively. Additionally, astaxanthin accumulation was enhanced by inhibiting de novo fatty acid biosynthesis. In summary, our study represents a pioneering work of utilizing Chlorella for the integrated production of lipids and high-value products and C. zofingiensis has great potential to be a promising production strain and serve as an emerging oleaginous model alga.

  9. Synergistic antioxidant activity of milk sphingomyeline and its sphingoid base with α-tocopherol on fish oil triacylglycerol.

    Science.gov (United States)

    Shimajiri, Junki; Shiota, Makoto; Hosokawa, Masashi; Miyashita, Kazuo

    2013-08-21

    The effects of milk phospholipids (PLs), sphingolipids (SLs), and their sphingoid backbone on the oxidation of fish oil triacylglycerol (TAG) were examined with or without α-tocopherol. All compounds had little effect on the TAG oxidation in the absence of α-tocopherol. On the other hand, they could act synergistically with α-tocopherol. The highest synergistic activity was shown by sphingoid bases, followed by sphingomyelin (SPM) and other amine-containing PLs and SLs. This result showed that the synergistic activity increased with an increasing concentration of amine group of PLs, SLs, or sphingoid bases in the reaction mixture. The comparison of changes in α-tocopherol content in fish oil TAG and tricaprylin suggested that antioxidant compounds would be formed from the amine group and the lipid oxidation products in a mild oxidation condition controlled by α-tocopherol.

  10. Characterization of blood lipoproteins and validation of cholesterol and triacylglycerol assays for free-ranging polar bears (Ursus maritimus).

    Science.gov (United States)

    Whiteman, John P; Frank, Nicholas; Greller, Katie A; Harlow, Henry J; Ben-David, Merav

    2013-05-01

    Blood triacylglycerol (TG) and lipoproteins are important variables for evaluating nutritional status of wildlife, but measurements are often expensive and difficult. Performance of a small, portable blood analyzer intended for human medical diagnostics was evaluated in measuring these variables in plasma and serum from free-ranging polar bears (Ursus maritimus), which are experiencing nutritional stress related to sea ice loss. The analyzer accurately tracked changes in concentration of total cholesterol (Ctotal), cholesterol associated with high-density lipoprotein (CHDL), and TG during a validation protocol of diluting samples and spiking them with exogenous cholesterol and glycerol. Values of Ctotal and TG agreed well with values obtained by other methods (ultracentrifugation followed by colorimetric assays); agreement was variable for values of cholesterol associated with specific lipoproteins. Similar to a study of captive polar bears, ultracentrifugation methods revealed greater TG in very low-density lipoproteins than in low-density lipoprotein, which is unusual and merits additional study.

  11. Determination and comparison of seed oil triacylglycerol composition of various soybeans (Glycine max (L.)) using ¹H-NMR spectroscopy.

    Science.gov (United States)

    Kim, Won Woo; Rho, Ho Sik; Hong, Yong Deog; Yeom, Myung Hun; Shin, Song Seok; Yi, Jun Gon; Lee, Min-Seuk; Park, Hye Yoon; Cho, Dong Ha

    2013-11-21

    Seed oil triacylglycerol (TAG) composition of 32 soybean varieties were determined and compared using ¹H-NMR. The contents of linolenic (Ln), linoleic (L), and oleic (O) ranged from 10.7% to 19.3%, 37.4%-50.1%, and 15.7%-34.1%, respectively. As is evident, linoleic acid was the major fatty acid of soybean oil. Compositional differences among the varieties were observed. Natural oils containing unsaturated groups have been regarded as important nutrient and cosmetic ingredients because of their various biological activities. The TAG profiles of the soy bean oils could be useful for distinguishing the origin of seeds and controlling the quality of soybean oils. To the best of our knowledge, this is the first study in which the TAG composition of various soybean oils has been analyzed using the ¹H-NMR method.

  12. An interspecies comparison of the phagocytosis and dissolution of 241AmO2 particles by rat, dog and monkey alveolar macrophages in vitro

    International Nuclear Information System (INIS)

    Taya, A.; Carmack, D.B.; Muggenburg, B.A.; Mewhinney, J.A.

    1992-01-01

    Experiments were conducted to study the phagocytosis and dissolution of 241 AmO 2 particles by rat, dog and monkey alveolar macrophages (PAM) in vitro. The phagocytosis and dissolution of 241 AmO 2 particles were followed up to 20 and 72 h, respectively. Dog and monkey PAM took up 241 AmO 2 particles at similar rates, whereas rat PAM phagocytosed only 60% of the amount phagocytosed by dog and monkey PAM at 20h. The PAM of the three species dissolved 241 AmO 2 particles at similar rates; 8-10% was dissolved by 72h. The results of the 241 AmO 2 uptake in vitro may reflect in vivo situations, where the differences in uptake seen in vitro would probably diminish at later times after exposure. The dissolution results imply that the dissolution of 241 AmO 2 particles by alveolar macrophages of the three species might be species-independent. (author)

  13. The role of phagocytosis, oxidative burst and neutrophil extracellular traps in the interaction between neutrophils and the periodontal pathogen Porphyromonas gingivalis.

    Science.gov (United States)

    Jayaprakash, K; Demirel, I; Khalaf, H; Bengtsson, T

    2015-10-01

    Neutrophils are regarded as the sentinel cells of innate immunity and are found in abundance within the gingival crevice. Discovery of neutrophil extracellular traps (NETs) within the gingival pockets prompted us to probe the nature of the interactions of neutrophils with the prominent periopathogen Porphyromonas gingivalis. Some of the noted virulence factors of this Gram-negative anaerobe are gingipains: arginine gingipains (RgpA/B) and lysine gingipain (Kgp). The aim of this study was to evaluate the role of gingipains in phagocytosis, formation of reactive oxygen species, NETs and CXCL8 modulation by using wild-type strains and isogenic gingipain mutants. Confocal imaging showed that gingipain mutants K1A (Kgp) and E8 (RgpA/B) induced extracellular traps in neutrophils, whereas ATCC33277 and W50 were phagocytosed. The viability of both ATCC33277 and W50 dwindled as the result of phagocytosis and could be salvaged by cytochalasin D, and the bacteria released high levels of lipopolysaccharide in the culture supernatant. Porphyromonas gingivalis induced reactive oxygen species and CXCL8 with the most prominent effect being that of the wild-type strain ATCC33277, whereas the other wild-type strain W50 was less effective. Quantitative real-time polymerase chain reaction revealed a significant CXCL8 expression by E8. All the tested P. gingivalis strains increased cytosolic free calcium. In conclusion, phagocytosis is the primary neutrophil response to P. gingivalis, although NETs could play an accessory role in infection control. Although gingipains do not seem to directly regulate phagocytosis, NETs or oxidative burst in neutrophils, their proteolytic properties could modulate the subsequent outcomes such as nutrition acquisition and survival by the bacteria. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Ocean Acidification Affects the Cytoskeleton, Lysozymes, and Nitric Oxide of Hemocytes: A Possible Explanation for the Hampered Phagocytosis in Blood Clams, Tegillarca granosa.

    Science.gov (United States)

    Su, Wenhao; Rong, Jiahuan; Zha, Shanjie; Yan, Maocang; Fang, Jun; Liu, Guangxu

    2018-01-01

    An enormous amount of anthropogenic carbon dioxide (CO 2 ) has been dissolved into the ocean, leading to a lower pH and changes in the chemical properties of seawater, which has been termed ocean acidification (OA). The impacts of p CO 2 -driven acidification on immunity have been revealed recently in various marine organisms. However, the mechanism causing the reduction in phagocytosis still remains unclear. Therefore, the impacts of p CO 2 -driven OA at present and near-future levels (pH values of 8.1, 7.8, and 7.4) on the rate of phagocytosis, the abundance of cytoskeleton components, the levels of nitric oxide (NO), and the concentration and activity of lysozymes (LZM) of hemocytes were investigated in a commercial bivalve species, the blood clam ( Tegillarca granosa ). In addition, the effects of OA on the expression of genes regulating actin skeleton and nitric oxide synthesis 2 ( NOS2 ) were also analyzed. The results obtained showed that the phagocytic rate, cytoskeleton component abundance, concentration and activity of LZM of hemocytes were all significantly reduced after a 2-week exposure to the future OA scenario of a pH of 7.4. On the contrary, a remarkable increase in the concentration of NO compared to that of the control was detected in clams exposed to OA. Furthermore, the expression of genes regulating the actin cytoskeleton and NOS were significantly up-regulated after OA exposure. Though the mechanism causing phagocytosis seemed to be complicated based on the results obtained in the present study and those reported previously, our results suggested that OA may reduce the phagocytosis of hemocytes by (1) decreasing the abundance of cytoskeleton components and therefore hampering the cytoskeleton-mediated process of engulfment, (2) reducing the concentration and activity of LZM and therefore constraining the degradation of the engulfed pathogen through an oxygen-independent pathway, and (3) inducing the production of NO, which may negatively

  15. In vivo and in vitro phagocytosis of Leishmania (Leishmania) amazonensis promastigotes by B-1 cells.

    Science.gov (United States)

    Geraldo, M M; Costa, C R; Barbosa, F M C; Vivanco, B C; Gonzaga, W F K M; Novaes E Brito, R R; Popi, A F; Lopes, J D; Xander, P

    2016-06-01

    Leishmaniasis is caused by Leishmania parasites that infect several cell types. The promastigote stage of Leishmania is internalized by phagocytic cells and transformed into the obligate intracellular amastigote form. B-1 cells are a subpopulation of B cells that are able to differentiate in vitro and in vivo into mononuclear phagocyte-like cells with phagocytic properties. B-1 cells use several receptors for phagocytosis, such as the mannose receptor and third complement receptor. Leishmania binds to the same receptors on macrophages. In this study, we demonstrated that phagocytes derived from B-1 cells (B-1 CDP) were able to internalize promastigotes of L. (L.) amazonensis in vitro. The internalized promastigotes differentiated into amastigotes. Our results showed that the phagocytic index was higher in B-1 CDP compared to peritoneal macrophages and bone marrow-derived macrophages. The in vivo phagocytic ability of B-1 cells was also demonstrated. Parasites were detected inside purified B-1 cells after intraperitoneal infection with L. (L.) amazonensis promastigotes. Intraperitoneal stimulation with the parasites led to an increase in both IL-10 and TNF-α. These results highlight the importance of studying B-1 CDP cells as phagocytic cells that can participate and contribute to immunity to parasites. © 2016 John Wiley & Sons Ltd.

  16. Sialoglycoproteins in morphological distinct stages of Mucor polymorphosporus and their influence on phagocytosis by human blood phagocytes.

    Science.gov (United States)

    Almeida, Catia Amancio; de Campos-Takaki, Galba Maria; Portela, Maristela Barbosa; Travassos, Luiz R; Alviano, Celuta Sales; Alviano, Daniela Sales

    2013-10-01

    The possible role of sialic acids in host cells-fungi interaction and their association with glycoproteins were evaluated using a clinical isolate of the dimorphic fungus Mucor polymorphosporus. Lectin-binding assays with spores and yeast cells denoted the presence of surface sialoglycoconjugates containing 2,3- and 2,6-linked sialylglycosyl groups. Western blotting with peroxidase-labeled Limulus polyphemus agglutinin revealed the occurrence of different sialoglycoprotein types in both cell lysates and cell wall protein extracts of mycelia, spores, and yeasts of M. polymorphosporus. Sialic acids contributed to the surface negative charge of spores and yeast forms as evaluated by adherence to a cationic substrate. Sialidase-treated spores were less resistant to phagocytosis by human neutrophils and monocytes from healthy individuals than control (untreated) fungal suspensions. The results suggest that sialic acids are terminal units of various glycoproteins of M. polymorphosporus, contributing to negative charge of yeasts and spore cells and protecting infectious propagules from destruction by host cells.

  17. Cross-talk between miR-471-5p and autophagy component proteins regulates LC3-associated phagocytosis (LAP) of apoptotic germ cells.

    Science.gov (United States)

    Panneerdoss, Subbarayalu; Viswanadhapalli, Suryavathi; Abdelfattah, Nourhan; Onyeagucha, Benjamin C; Timilsina, Santosh; Mohammad, Tabrez A; Chen, Yidong; Drake, Michael; Vuori, Kristiina; Kumar, T Rajendra; Rao, Manjeet K

    2017-09-19

    Phagocytic clearance of apoptotic germ cells by Sertoli cells is vital for germ cell development and differentiation. Here, using a tissue-specific miRNA transgenic mouse model, we show that interaction between miR-471-5p and autophagy member proteins regulates clearance of apoptotic germ cells via LC3-associated phagocytosis (LAP). Transgenic mice expressing miR-471-5p in Sertoli cells show increased germ cell apoptosis and compromised male fertility. Those effects are due to defective engulfment and impaired LAP-mediated clearance of apoptotic germ cells as miR-471-5p transgenic mice show lower levels of Dock180, LC3, Atg12, Becn1, Rab5 and Rubicon in Sertoli cells. Our results reveal that Dock180 interacts with autophagy member proteins to constitute a functional LC3-dependent phagocytic complex. We find that androgen regulates Sertoli cell phagocytosis by controlling expression of miR-471-5p and its target proteins. These findings suggest that recruitment of autophagy machinery is essential for efficient clearance of apoptotic germ cells by Sertoli cells using LAP.Although phagocytic clearance of apoptotic germ cells by Sertoli cells is essential for spermatogenesis, little of the mechanism is known. Here the authors show that Sertoli cells employ LC3-associated phagocytosis (LAP) by recruiting autophagy member proteins to clear apoptotic germ cells.

  18. Effect of a 2.45-GHz radiofrequency electromagnetic field on neutrophil chemotaxis and phagocytosis in differentiated human HL-60 cells.

    Science.gov (United States)

    Koyama, Shin; Narita, Eijiro; Suzuki, Yoshihisa; Taki, Masao; Shinohara, Naoki; Miyakoshi, Junji

    2015-01-01

    The potential public health risks of radiofrequency (RF) fields have been discussed at length, especially with the use of mobile phones spreading extensively throughout the world. In order to investigate the properties of RF fields, we examined the effect of 2.45-GHz RF fields at the specific absorption rate (SAR) of 2 and 10 W/kg for 4 and 24 h on neutrophil chemotaxis and phagocytosis in differentiated human HL-60 cells. Neutrophil chemotaxis was not affected by RF-field exposure, and subsequent phagocytosis was not affected either compared with that under sham exposure conditions. These studies demonstrated an initial immune response in the human body exposed to 2.45-GHz RF fields at the SAR of 2 W/kg, which is the maximum value recommended by the International Commission for Non-Ionizing Radiation Protection (ICNIRP) guidelines. The results of our experiments for RF-field exposure at an SAR under 10 W/kg showed very little or no effects on either chemotaxis or phagocytosis in neutrophil-like human HL-60 cells. © The Author 2014. Published by Oxford University Press on behalf of The Japan Radiation Research Society and Japanese Society for Radiation Oncology.

  19. Antibody-mediated platelet phagocytosis by human macrophages is inhibited by siRNA specific for sequences in the SH2 tyrosine kinase, Syk.

    Science.gov (United States)

    Lu, Ying; Wang, Weiming; Mao, Huiming; Hu, Hai; Wu, Yanling; Chen, Bing-Guan; Liu, Zhongmin

    2011-01-01

    Immune thrombocytopenia depends upon Fc receptor-mediated phagocytosis that involves signaling through the SH2 tyrosine kinase, Syk. We designed small interfering (siRNA) sequences complementary to Syk coding regions to decrease the expression of Syk in the human macrophage cell line, THP-1. To evaluate the functional effect of siRNA on phagocytosis, we developed a new in vitro assay for antibody-mediated platelet ingestion by THP-1 cells. Incubation of THP-1 cells at 37°C with fluorescence-labeled platelets and anti-platelet antibody promoted ingestion of platelets that could be quantitated by flow cytometry. Transfection of THP-1 cells with Syk-specific siRNA resulted in a reduction in the amount of FcγRII-associated Syk protein. Coincident with decreased Syk expression, we observed inhibition of antibody-mediated platelet ingestion. These results confirm a key role for Syk in antibody-mediated phagocytosis and suggest Syk-specific siRNA as a possible therapeutic candidate for immune thrombocytopenia. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Effects of dexamethasone and insulin on the synthesis of triacylglycerols and phosphatidylcholine and the secretion of very-low-density lipoproteins and lysophosphatidylcholine by monolayer cultures of rat hepatocytes.

    OpenAIRE

    Mangiapane, E H; Brindley, D N

    1986-01-01

    Rat hepatocytes in monolayer culture were preincubated for 19 h with 1 microM-dexamethasone, and the incubation was continued for a further 23 h with [14C]oleate, [3H]glycerol and 1 microM-dexamethasone. Dexamethasone increased the secretion of triacylglycerol into the medium in particles that had the properties of very-low-density lipoproteins. The increased secretion was matched by a decrease in the triacylglycerol and phosphatidylcholine that remained in the hepatocytes. Preincubating the ...

  1. Entamoeba histolytica phagocytosis of human erythrocytes involves PATMK, a member of the transmembrane kinase family.

    Directory of Open Access Journals (Sweden)

    Douglas R Boettner

    2008-01-01

    Full Text Available Entamoeba histolytica is the cause of amebic colitis and liver abscess. This parasite induces apoptosis in host cells and utilizes exposed ligands such as phosphatidylserine to ingest the apoptotic corpses and invade deeper into host tissue. The purpose of this work was to identify amebic proteins involved in the recognition and ingestion of dead cells. A member of the transmembrane kinase family, phagosome-associated TMK96 (PATMK, was identified in a proteomic screen for early phagosomal proteins. Anti-peptide affinity-purified antibody produced against PATMK demonstrated that it was a type I integral membrane protein that was expressed on the trophozoite surface, and that co-localized with human erythrocytes at the site of contact. The role of PATMK in erythrophagocytosis in vitro was demonstrated by: (i incubation of ameba with anti-PATMK antibodies; (ii PATMK mRNA knock-down using a novel shRNA expression system; and (iii expression of a carboxy-truncation of PATMK (PATMK(delta932. Expression of the carboxy-truncation of PATMK(delta932 also caused a specific reduction in the ability of E. histolytica to establish infection in the intestinal model of amebiasis, however these amebae retained the ability to cause hepatic abscesses when directly injected in the liver. In conclusion, PATMK was identified as a member of the TMK family that participates in erythrophagocytosis and is uniquely required for intestinal infection.

  2. Entamoeba histolytica phagocytosis of human erythrocytes involves PATMK, a member of the transmembrane kinase family.

    Science.gov (United States)

    Boettner, Douglas R; Huston, Christopher D; Linford, Alicia S; Buss, Sarah N; Houpt, Eric; Sherman, Nicholas E; Petri, William A

    2008-01-01

    Entamoeba histolytica is the cause of amebic colitis and liver abscess. This parasite induces apoptosis in host cells and utilizes exposed ligands such as phosphatidylserine to ingest the apoptotic corpses and invade deeper into host tissue. The purpose of this work was to identify amebic proteins involved in the recognition and ingestion of dead cells. A member of the transmembrane kinase family, phagosome-associated TMK96 (PATMK), was identified in a proteomic screen for early phagosomal proteins. Anti-peptide affinity-purified antibody produced against PATMK demonstrated that it was a type I integral membrane protein that was expressed on the trophozoite surface, and that co-localized with human erythrocytes at the site of contact. The role of PATMK in erythrophagocytosis in vitro was demonstrated by: (i) incubation of ameba with anti-PATMK antibodies; (ii) PATMK mRNA knock-down using a novel shRNA expression system; and (iii) expression of a carboxy-truncation of PATMK (PATMK(delta932)). Expression of the carboxy-truncation of PATMK(delta932) also caused a specific reduction in the ability of E. histolytica to establish infection in the intestinal model of amebiasis, however these amebae retained the ability to cause hepatic abscesses when directly injected in the liver. In conclusion, PATMK was identified as a member of the TMK family that participates in erythrophagocytosis and is uniquely required for intestinal infection.

  3. Phagocytosis of Giardia muris by macrophages in Peyer's patch epithelium in mice.

    Science.gov (United States)

    Owen, R L; Allen, C L; Stevens, D P

    1981-08-01

    No mechanism for the initiation of immunological clearance of Giardia from the mammalian intestinal tract has been identified. In normal and nude mice experimentally infected with G. muris, we examined antigen-sampling epithelium over Peyer's patch follicles by electron microscopy for evidence of interaction between G. muris and lymphoid cells. Invading G. muris were found in the epithelium near dying or desquamating columnar cells. Macrophages beneath the basal lamina extended pseudopods into the epithelium, trapping invading G. muris and enclosing them in phagolysosomes. In normal mice, which clear G. muris in 4 to 6 weeks, macrophages containing digested G. muris were surrounded by rosettes of lymphoblasts in the epithelium. In nude mice deficient in lymphocytes, there was apparent hyperplasia of macrophages, which filled the follicle domes, resulting in more frequent entrapment of G. muris but no contact between macrophages and lymphoblasts in the epithelium. In nude mice, which require 6 months to control G. muris infection, lymphoblast contact with macrophages containing distinctive microtubular remnants of G. muris was only identified in the follicle dome. This close physical association of lymphoblasts and macrophages containing G. muris remnants suggests that this macrophage activity represents intraepithelial antigen processing as well as a defense against the effects of the uncontrolled entrance of microorganisms and other antigenic particles into Peyer's patch lymphoid follicles.

  4. Phagocytosis and immune response studies of Macrophage-Nanodiamond Interactions in vitro and in vivo.

    Science.gov (United States)

    Huang, K-J; Lee, C-Y; Lin, Y-C; Lin, C-Y; Perevedentseva, E; Hung, S-F; Cheng, C-L

    2017-10-01

    The applications of nanodiamond as drug delivery and bio-imaging can require the relinquishing ND-drug conjugate via blood flow, where interaction with immune cells may occur. In this work, we investigated the ND penetration in macrophage and the immune response using the tissue-resident murine macrophages (RAW 264.7). Confocal fluorescence imaging, immunofluorescence analysis of nuclear translocation of interferon regulatory factor IRF-3 and transcriptional factor NF-κΒ, analysis of pro-inflammatory cytokines production IL-1β, IL-6 IL-10 with a reverse transcription-polymerase chain reaction technique were applied. The TNF-α factor production has been studied both in vitro at ND interaction with the macrophage and in vivo after ND injection in the mice blood system using immunoassay. The macrophage antibacterial function was estimated through E. coli bacterial colony formation. ND didn't stimulate the immune response and functionality of the macrophage was not altered. Using MTT test, ND was found negligibly cytotoxic to macrophages. Thus, ND can serve as a biocompatible platform for bio-medical applications. Left: Graphic representation of Nanodiamond internalization in macrophage. Right: (a) Fluorescence images of lysosomes, (b) nanodiamond and (c) merged image of nanodiamond internalization in macrophage. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Stochastic cellular automata model of neurosphere growth: Roles of proliferative potential, contact inhibition, cell death, and phagocytosis.

    Science.gov (United States)

    Sipahi, Rifat; Zupanc, Günther K H

    2018-05-14

    Neural stem and progenitor cells isolated from the central nervous system form, under specific culture conditions, clonal cell clusters known as neurospheres. The neurosphere assay has proven to be a powerful in vitro system to study the behavior of such cells and the development of their progeny. However, the theory of neurosphere growth has remained poorly understood. To overcome this limitation, we have, in the present paper, developed a cellular automata model, with which we examined the effects of proliferative potential, contact inhibition, cell death, and clearance of dead cells on growth rate, final size, and composition of neurospheres. Simulations based on this model indicated that the proliferative potential of the founder cell and its progenitors has a major influence on neurosphere size. On the other hand, contact inhibition of proliferation limits the final size, and reduces the growth rate, of neurospheres. The effect of this inhibition is particularly dramatic when a stem cell becomes encapsulated by differentiated or other non-proliferating cells, thereby suppressing any further mitotic division - despite the existing proliferative potential of the stem cell. Conversely, clearance of dead cells through phagocytosis is predicted to accelerate growth by reducing contact inhibition. A surprising prediction derived from our model is that cell death, while resulting in a decrease in growth rate and final size of neurospheres, increases the degree of differentiation of neurosphere cells. It is likely that the cellular automata model developed as part of the present investigation is applicable to the study of tissue growth in a wide range of systems. Copyright © 2018 Elsevier Ltd. All rights reserved.

  6. Comparison of HPLC/MS and MALDI-MS for characterizing triacylglycerols in insects: Specie-specific composition of lipids in fat bodies of bumblebee males

    Czech Academy of Sciences Publication Activity Database

    Kofroňová, Edita; Cvačka, Josef; Vrkoslav, Vladimír; Hanus, Robert; Jiroš, Pavel; Kindl, Jiří; Hovorka, Oldřich; Valterová, Irena

    2009-01-01

    Roč. 877, č. 30 (2009), s. 3878-3884 ISSN 1570-0232 R&D Projects: GA ČR GA203/09/0139; GA ČR GA203/09/1446; GA MŠk 2B06007 Institutional research plan: CEZ:AV0Z40550506 Keywords : Bumblebee * fat body * triacylglycerols * HPLC/APCI-MS * MALDI-MS Subject RIV: CC - Organic Chemistry Impact factor: 2.777, year: 2009

  7. A Phospholipid-Protein Complex from Krill with Antioxidative and Immunomodulating Properties Reduced Plasma Triacylglycerol and Hepatic Lipogenesis in Rats

    Directory of Open Access Journals (Sweden)

    Marie S. Ramsvik

    2015-07-01

    Full Text Available Dietary intake of marine omega-3 polyunsaturated fatty acids (n-3 PUFAs can change the plasma profile from atherogenic to cardioprotective. In addition, there is growing evidence that proteins of marine origin may have health benefits. We investigated a phospholipid-protein complex (PPC from krill that is hypothesized to influence lipid metabolism, inflammation, and redox status. Male Wistar rats were fed a control diet (2% soy oil, 8% lard, 20% casein, or diets where corresponding amounts of casein and lard were replaced with PPC at 3%, 6%, or 11% (wt %, for four weeks. Dietary supplementation with PPC resulted in significantly lower levels of plasma triacylglycerols in the 11% PPC-fed group, probably due to reduced hepatic lipogenesis. Plasma cholesterol levels were also reduced at the highest dose of PPC. In addition, the plasma and liver content of n-3 PUFAs increased while n-6 PUFAs decreased. This was associated with increased total antioxidant capacity in plasma and increased liver gene expression of mitochondrial superoxide dismutase (Sod2. Finally, a reduced plasma level of the inflammatory mediator interleukin-2 (IL-2 was detected in the PPC-fed animals. The present data show that PPC has lipid-lowering effects in rats, and may modulate risk factors related to cardiovascular disease progression.

  8. Viral infection of the marine alga Emiliania huxleyi triggers lipidome remodeling and induces the production of highly saturated triacylglycerol.

    Science.gov (United States)

    Malitsky, Sergey; Ziv, Carmit; Rosenwasser, Shilo; Zheng, Shuning; Schatz, Daniella; Porat, Ziv; Ben-Dor, Shifra; Aharoni, Asaph; Vardi, Assaf

    2016-04-01

    Viruses that infect marine photosynthetic microorganisms are major ecological and evolutionary drivers of microbial food webs, estimated to turn over more than a quarter of the total photosynthetically fixed carbon. Viral infection of the bloom-forming microalga Emiliania huxleyi induces the rapid remodeling of host primary metabolism, targeted towards fatty acid metabolism. We applied a liquid chromatography-mass spectrometry (LC-MS)-based lipidomics approach combined with imaging flow cytometry and gene expression profiling to explore the impact of viral-induced metabolic reprogramming on lipid composition. Lytic viral infection led to remodeling of the cellular lipidome, by predominantly inducing the biosynthesis of highly saturated triacylglycerols (TAGs), coupled with a significant accumulation of neutral lipids within lipid droplets. Furthermore, TAGs were found to be a major component (77%) of the lipidome of isolated virions. Interestingly, viral-induced TAGs were significantly more saturated than TAGs produced under nitrogen starvation. This study highlights TAGs as major products of the viral-induced metabolic reprogramming during the host-virus interaction and indicates a selective mode of membrane recruitment during viral assembly, possibly by budding of the virus from specialized subcellular compartments. These findings provide novel insights into the role of viruses infecting microalgae in regulating metabolism and energy transfer in the marine environment and suggest their possible biotechnological application in biofuel production. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

  9. Fostering triacylglycerol accumulation in novel oleaginous yeast Cryptococcus psychrotolerans IITRFD utilizing groundnut shell for improved biodiesel production.

    Science.gov (United States)

    Deeba, Farha; Pruthi, Vikas; Negi, Yuvraj S

    2017-10-01

    The investigation was carried out to examine the potential of triacylglycerol (TAG) accumulation by novel oleaginous yeast isolate Cryptococcus psychrotolerans IITRFD on utilizing groundnut shell acid hydrolysate (GSH) as cost-effective medium. The maximum biomass productivity and lipid productivity of 0.095±0.008g/L/h and 0.044±0.005g/L/h, respectively with lipid content 46% was recorded on GSH. Fatty acid methyl ester (FAME) profile obtained by GC-MS analysis revealed oleic acid (37.8%), palmitic (29.4%) and linoleic (32.8%) as major fatty acids representing balance between oxidative stability (OS) and cold flow filter properties (CFFP) for improved biodiesel quality. The biodiesel property calculated were correlated well with the fuel standards limits of ASTM D6751, EN 14214 and IS 15607. The present findings raise the possibility of using agricultural waste groundnut shell as a substrate for production of biodiesel by novel oleaginous yeast isolate C. psychrotolerans IITRFD. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Functional analysis of three type-2 DGAT homologue genes for triacylglycerol production in the green microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    La Russa, M; Bogen, C; Uhmeyer, A; Doebbe, A; Filippone, E; Kruse, O; Mussgnug, J H

    2012-11-30

    Photosynthetic organisms like plants and algae can use sunlight to produce lipids as important metabolic compounds. Plant-derived triacylglycerols (TAGs) are valuable for human and animal nutrition because of their high energy content and are becoming increasingly important for the production of renewable biofuels. Acyl-CoA:diacylglycerol acyltransferases (DGATs) have been demonstrated to play an important role in the accumulation of TAG compounds in higher plants. DGAT homologue genes have been identified in the genome of the green alga Chlamydomonas reinhardtii, however their function in vivo is still unknown. In this work, the three most promising type-2 DGAT candidate genes potentially involved in TAG lipid accumulation (CrDGAT2a, b and c) were investigated by constructing overexpression strains. For each of the genes, three strains were identified which showed enhanced mRNA levels of between 1.7 and 29.1 times that of the wild type (wt). Total lipid contents, neutral lipids and fatty acid profiles were determined and showed that an enhanced mRNA expression level of the investigated DGAT genes did not boost the intracellular TAG accumulation or resulted in alterations of the fatty acid profiles compared to wild type during standard growth condition or during nitrogen or sulfur stress conditions. We conclude that biotechnological efforts to enhance cellular TAG amount in microalgae need further insights into the complex network of lipid biosynthesis to identify potential bottlenecks of neutral lipid production. Copyright © 2012 Elsevier B.V. All rights reserved.

  11. Comparison of CO(2) and bicarbonate as inorganic carbon sources for triacylglycerol and starch accumulation in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gardner, Robert D; Lohman, Egan; Gerlach, Robin; Cooksey, Keith E; Peyton, Brent M

    2013-01-01

    Microalgae are capable of accumulating high levels of lipids and starch as carbon storage compounds. Investigation into the metabolic activities involved in the synthesis of these compounds has escalated since these compounds can be used as precursors for food and fuel. Here, we detail the results of a comprehensive analysis of Chlamydomonas reinhardtii using high or low inorganic carbon concentrations and speciation between carbon dioxide and bicarbonate, and the effects these have on inducing lipid and starch accumulation during nitrogen depletion. High concentrations of CO(2) (5%; v/v) produced the highest amount of biofuel precursors, transesterified to fatty acid methyl esters, but exhibited rapid accumulation and degradation characteristics. Low CO(2) (0.04%; v/v) caused carbon limitation and minimized triacylglycerol (TAG) and starch accumulation. High bicarbonate caused a cessation of cell cycling and accumulation of both TAG and starch that was more stable than the other experimental conditions. Starch accumulated prior to TAG and then degraded as maximum TAG was reached. This suggests carbon reallocation from starch-based to TAG-based carbon storage. Copyright © 2012 Wiley Periodicals, Inc.

  12. Determination of triacylglycerol regioisomers using electrospray ionization-quadrupole ion trap mass spectrometry with a kinetic method.

    Science.gov (United States)

    Leveque, Nathalie L; Acheampong, Akwasi; Heron, Sylvie; Tchapla, Alain

    2012-04-13

    The kinetic method was applied to differentiate and quantify mixtures of regioisomeric triacylglycerols (TAGs) by generating and mass selecting alkali ion bound metal dimeric clusters with a TAG chosen as reference (ref) and examining their competitive dissociations in a quadrupole ion trap mass spectrometer. This methodology readily distinguished pairs of regioisomers (AAB/ABA) such as LLO/LOL, OOP/OPO and SSP/SPS and consequently distinguished sn-1/sn-3, sn-2 substituents on the glycerol backbone. The dimeric complex ions [ref, Li, TAG((AAB and/or ABA))](+) generated by electrospray ionization mass spectrometry were subjected to collision induced dissociation causing competitive loss of either the neutral TAG reference (ref) leading to [Li(AAB and/or ABA)](+) or the neutral TAG molecule (TAG((AAB and/or ABA))) leading to [ref, Li](+). The ratio of the two competitive dissociation rates, defined by the product ion branching ratio (R(iso)), was related via the kinetic method to the regioisomeric composition of the investigated TAG mixture. In this work, a linear correlation was established between composition of the mixture of each TAG regioisomer and the logarithm of the branching ratio for competitive fragmentation. Depending on the availability of at least one TAG regioisomer as standard, the kinetic method and the standard additions method led to the quantitative analysis of natural TAG mixtures. Copyright © 2012 Elsevier B.V. All rights reserved.

  13. Simultaneous production of intracellular triacylglycerols and extracellular polyol esters of fatty acids by Rhodotorula babjevae and Rhodotorula aff. paludigena.

    Science.gov (United States)

    Garay, Luis A; Sitepu, Irnayuli R; Cajka, Tomas; Cathcart, Erin; Fiehn, Oliver; German, J Bruce; Block, David E; Boundy-Mills, Kyria L

    2017-10-01

    Microbial oils have been analyzed as alternatives to petroleum. However, just a handful of microbes have been successfully adapted to produce chemicals that can compete with their petroleum counterparts. One of the reasons behind the low success rate is the overall economic inefficiency of valorizing a single product. This study presents a lab-scale analysis of two yeast species that simultaneously produce multiple high-value bioproducts: intracellular triacylglycerols (TG) and extracellular polyol esters of fatty acids (PEFA), two lipid classes with immediate applications in the biofuels and surfactant industries. At harvest, the yeast strain Rhodotorula aff. paludigena UCDFST 81-84 secreted 20.9 ± 0.2 g L -1 PEFA and produced 8.8 ± 1.0 g L -1 TG, while the yeast strain Rhodotorula babjevae UCDFST 04-877 secreted 11.2 ± 1.6 g L -1 PEFA and 18.5 ± 1.7 g L -1 TG. The overall glucose conversion was 0.24 and 0.22 g (total lipid) g (glucose) -1 , respectively. The results present a stable and scalable microbial growth platform yielding multiple co-products.

  14. Reduced triacylglycerol mobilization during seed germination and early seedling growth in Arabidopsis containing nutritionally important polyunsaturated fatty acids

    Directory of Open Access Journals (Sweden)

    Pushkar Shrestha

    2016-09-01

    Full Text Available There are now several examples of plant species engineered to synthesise and accumulate nutritionally important polyunsaturated fatty acids in their seed triacylglycerols (TAG. The utilization of such TAG in germinating seeds of such transgenic plants was unknown. In this study, we examined the TAG utilization efficiency during seed germination in transgenic Arabidopsis seeds containing several examples of these fatty acids. Seed TAG species with native fatty acids had higher utilization rate than the TAG species containing transgenically produced polyunsaturated fatty acids. Conversely, quantification of the fatty acid components remaining in the total TAG after early stages of seed germination revealed that the undigested TAGs tended to contain an elevated level of the engineered polyunsaturated fatty acids (PUFA. LC-MS analysis further revealed asymmetrical mobilization rates for the individual TAG species. TAGs which contained multiple PUFA fatty acids were mobilized slower than the species containing single PUFA. The mobilised engineered fatty acids were used in de novo membrane lipid synthesis during seedling development.

  15. The effects of epidermal fatty acid profiles, 1-oleoglycerol, and triacylglycerols on the susceptibility of hibernating bats to Pseudogymnoascus destructans.

    Directory of Open Access Journals (Sweden)

    Melissa R Ingala

    Full Text Available White Nose Syndrome (WNS greatly increases the over-winter mortality of little brown (Myotis lucifugus, Indiana (M. sodalis, northern (M. septentrionalis, and tricolored (Perimyotis subflavus bats, and is caused by cutaneous infections with Pseudogymnoascus destructans (Pd. Big brown bats (Eptesicus fuscus are highly resistant to Pd infections. Seven different fatty acids (myristic, pentadecanoic, palmitic, palmitoleic, oleic, and, linoleic acids occur in the wing epidermis of both M. lucifugus and E. fuscus, 4 of which (myristic, palmitoleic, oleic, and, linoleic acids inhibit Pd growth. The amounts of myristic and linoleic acids in the epidermis of M. lucifugus decrease during hibernation, thus we predicted that the epidermal fatty acid profile of M. lucifugus during hibernation has a reduced ability to inhibit Pd growth. Laboratory Pd growth experiments were conducted to test this hypothesis. The results demonstrated that the fatty acid profile of M. lucifugus wing epidermis during hibernation has a reduced ability to inhibit the growth of Pd. Additional Pd growth experiments revealed that: a triacylglycerols composed of known anti-Pd fatty acids do not significantly affect growth, b pentadecanoic acid inhibits Pd growth, and c 1-oleoglycerol, which is found in the wing epidermis of E. fuscus, also inhibits the growth of this fungus. Analyses of white adipose from M. lucifugus also revealed the selective retention of oleic and linoleic acids in this tissue during hibernation.

  16. The effects of epidermal fatty acid profiles, 1-oleoglycerol, and triacylglycerols on the susceptibility of hibernating bats to Pseudogymnoascus destructans.

    Science.gov (United States)

    Ingala, Melissa R; Ravenelle, Rebecca E; Monro, Johanna J; Frank, Craig L

    2017-01-01

    White Nose Syndrome (WNS) greatly increases the over-winter mortality of little brown (Myotis lucifugus), Indiana (M. sodalis), northern (M. septentrionalis), and tricolored (Perimyotis subflavus) bats, and is caused by cutaneous infections with Pseudogymnoascus destructans (Pd). Big brown bats (Eptesicus fuscus) are highly resistant to Pd infections. Seven different fatty acids (myristic, pentadecanoic, palmitic, palmitoleic, oleic, and, linoleic acids) occur in the wing epidermis of both M. lucifugus and E. fuscus, 4 of which (myristic, palmitoleic, oleic, and, linoleic acids) inhibit Pd growth. The amounts of myristic and linoleic acids in the epidermis of M. lucifugus decrease during hibernation, thus we predicted that the epidermal fatty acid profile of M. lucifugus during hibernation has a reduced ability to inhibit Pd growth. Laboratory Pd growth experiments were conducted to test this hypothesis. The results demonstrated that the fatty acid profile of M. lucifugus wing epidermis during hibernation has a reduced ability to inhibit the growth of Pd. Additional Pd growth experiments revealed that: a) triacylglycerols composed of known anti-Pd fatty acids do not significantly affect growth, b) pentadecanoic acid inhibits Pd growth, and c) 1-oleoglycerol, which is found in the wing epidermis of E. fuscus, also inhibits the growth of this fungus. Analyses of white adipose from M. lucifugus also revealed the selective retention of oleic and linoleic acids in this tissue during hibernation.

  17. Korean Pine Nut Oil Attenuated Hepatic Triacylglycerol Accumulation in High-Fat Diet-Induced Obese Mice

    Directory of Open Access Journals (Sweden)

    Soyoung Park

    2016-01-01

    Full Text Available Korean pine nut oil (PNO has been reported to influence weight gain and lipid metabolism. We examined whether PNO replacement in a high-fat diet (HFD can ameliorate HFD-induced hepatic steatosis. Five-week-old male C57BL mice were fed control diets containing 10% of the energy from fat from PNO or soybean oil (SBO (PC, SC or HFDs with 45% of the energy from fat, with 10% from PNO or SBO and 35% from lard (PHFD, SHFD, for 12 weeks. Body weight gain and amount of white adipose tissue were lower in PHFD (10% and 18% lower, respectively compared with SHFD. Hepatic triacylglycerol (TG level was significantly lower in PHFD than the SHFD (26% lower. PNO consumption upregulated hepatic ACADL mRNA levels. The hepatic PPARG mRNA level was lower in the PC than in the SC. Expression of the sirtuin (SIRT 3 protein in white adipose tissue was down-regulated in the SHFD and restored in the PHFD to the level in the lean control mice. SIRT 3 was reported to be upregulated under conditions of caloric restriction (CR and plays a role in regulating mitochondrial function. PNO consumption resulted in lower body fat and hepatic TG accumulation in HFD-induced obesity, which seemed to be associated with the CR-mimetic response.

  18. Impact of liver fat on the differential partitioning of hepatic triacylglycerol into VLDL subclasses on high and low sugar diets.

    Science.gov (United States)

    Umpleby, A Margot; Shojaee-Moradie, Fariba; Fielding, Barbara; Li, Xuefei; Marino, Andrea; Alsini, Najlaa; Isherwood, Cheryl; Jackson, Nicola; Ahmad, Aryati; Stolinski, Michael; Lovegrove, Julie A; Johnsen, Sigurd; Jeewaka R Mendis, A S; Wright, John; Wilinska, Malgorzata E; Hovorka, Roman; Bell, Jimmy D; Thomas, E Louise; Frost, Gary S; Griffin, Bruce A

    2017-11-01

    Dietary sugars are linked to the development of non-alcoholic fatty liver disease (NAFLD) and dyslipidaemia, but it is unknown if NAFLD itself influences the effects of sugars on plasma lipoproteins. To study this further, men with NAFLD ( n = 11) and low liver fat 'controls' ( n = 14) were fed two iso-energetic diets, high or low in sugars (26% or 6% total energy) for 12 weeks, in a randomised, cross-over design. Fasting plasma lipid and lipoprotein kinetics were measured after each diet by stable isotope trace-labelling.There were significant differences in the production and catabolic rates of VLDL subclasses between men with NAFLD and controls, in response to the high and low sugar diets. Men with NAFLD had higher plasma concentrations of VLDL 1 -triacylglycerol (TAG) after the high ( P sugar ( P diets, a lower VLDL 1 -TAG fractional catabolic rate after the high sugar diet ( P sugar diet ( P sugar diet, was to channel hepatic TAG into a higher production of VLDL 1 -TAG ( P sugars. © 2017 The Author(s).

  19. Laser desorption/ionization time-of-flight mass spectrometry of triacylglycerols and other components in fingermark samples.

    Science.gov (United States)

    Emerson, Beth; Gidden, Jennifer; Lay, Jackson O; Durham, Bill

    2011-03-01

    The chemical composition of fingermarks could potentially be important for determining investigative leads, placing individuals at the time of a crime, and has applications as biomarkers of disease. Fingermark samples containing triacylglycerols (TAGs) and other components were analyzed using laser desorption/ionization (LDI) time-of-flight mass spectrometry (TOF MS). Only LDI appeared to be useful for this application while conventional matrix-assisted LDI-TOF MS was not. Tandem MS was used to identify/confirm selected TAGs. A limited gender comparison, based on a simple t-distribution and peaks intensities, indicated that two TAGs showed gender specificity at the 95% confidence level and two others at 97.5% confidence. Because gender-related TAGs differences were most often close to the standard deviation of the measurements, the majority of the TAGs showed no gender specificity. Thus, LDI-TOF MS is not a reliable indicator of gender based on fingermark analysis. Cosmetic ingredients present in some samples were identified. © 2011 American Academy of Forensic Sciences.

  20. Participation of 14-3-3ε and 14-3-3ζ proteins in the phagocytosis, component of cellular immune response, in Aedes mosquito cell lines.

    Science.gov (United States)

    Trujillo-Ocampo, Abel; Cázares-Raga, Febe Elena; Del Angel, Rosa María; Medina-Ramírez, Fernando; Santos-Argumedo, Leopoldo; Rodríguez, Mario H; Hernández-Hernández, Fidel de la Cruz

    2017-08-01

    Better knowledge of the innate immune system of insects will improve our understanding of mosquitoes as potential vectors of diverse pathogens. The ubiquitously expressed 14-3-3 protein family is evolutionarily conserved from yeast to mammals, and at least two isoforms of 14-3-3, the ε and ζ, have been identified in insects. These proteins have been shown to participate in both humoral and cellular immune responses in Drosophila. As mosquitoes of the genus Aedes are the primary vectors for arboviruses, causing several diseases such as dengue fever, yellow fever, Zika and chikungunya fevers, cell lines derived from these mosquitoes, Aag-2 from Aedes aegypti and C6/36 HT from Aedes albopictus, are currently used to study the insect immune system. Here, we investigated the role of 14-3-3 proteins (ε and ζ isoform) in phagocytosis, the main cellular immune responses executed by the insects, using Aedes spp. cell lines. We evaluated the mRNA and protein expression of 14-3-3ε and 14-3-3ζ in C6/36 HT and Aag-2 cells, and demonstrated that both proteins were localised in the cytoplasm. Further, in C6/36 HT cells treated with a 14-3-3 specific inhibitor we observed a notable modification of cell morphology with filopodia-like structure caused through cytoskeleton reorganisation (co-localization of 14-3-3 proteins with F-actin), more importantly the decrease in Salmonella typhimurium, Staphylococcus aureus and E. coli phagocytosis and reduction in phagolysosome formation. Additionally, silencing of 14-3-3ε and 14-3-3ζ expression by mean of specific DsiRNA confirmed the decreased phagocytosis and phagolysosome formation of pHrodo labelled E. coli and S. aureus bacteria by Aag-2 cells. The 14-3-3ε and 14-3-3ζ proteins modulate cytoskeletal remodelling, and are essential for phagocytosis of Gram-positive and Gram-negative bacteria in Aedes spp. cell lines.

  1. Thioester-containing proteins of the tick Ixodes ricinus: Gene expression, response to microbial challenge and their role in phagocytosis of the yeast Candida albicans

    Czech Academy of Sciences Publication Activity Database

    Urbanová, Veronika; Šíma, Radek; Šauman, Ivo; Hajdušek, Ondřej; Kopáček, Petr

    2015-01-01

    Roč. 48, č. 1 (2015), s. 55-64 ISSN 0145-305X R&D Projects: GA ČR GAP506/10/2136; GA ČR GA13-11043S; GA ČR GP13-27630P; GA ČR GP13-12816P; GA MŠk(CZ) EE2.3.30.0032 EU Projects: European Commission(XE) 316304 - MODBIOLIN Institutional support: RVO:60077344 Keywords : Candida albicans * Complement * Innate immunity * Phagocytosis * Thioester-containing proteins * Tick Ixodes ricinus Subject RIV: EC - Immunology Impact factor: 3.620, year: 2015

  2. Antibody dependent cellular phagocytosis (ADCP) and antibody dependent cellular cytotoxicity (ADCC) of breast cancer cells mediated by bispecific antibody, MDX-210.

    Science.gov (United States)

    Watanabe, M; Wallace, P K; Keler, T; Deo, Y M; Akewanlop, C; Hayes, D F

    1999-02-01

    MDX-210 is a bispecific antibody (BsAb) with specificity for both the proto-oncogene product of HER-2/neu (c-erbB-2) and FcgammaRI (CD64). HER-2/neu is overexpressed in malignant tissue of approximately 30% of patients with breast cancer, and FcgammaRI is expressed on human monocytes, macrophages, and IFN-gamma activated granulocytes. We investigated phagocytosis and cytolysis of cultured human breast cancer cells by human monocyte-derived macrophages (MDM) mediated by BsAb MDX-210, its partially humanized derivative (MDX-H210), and its parent MoAb 520C9 (anti-HER-2/neu) under various conditions. Purified monocytes were cultured with GM-CSF, M-CSF, or no cytokine for five or six days. Antibody dependent cellular phagocytosis (ADCP) and cytolysis (ADCC) assays were performed with the MDM and HER-2/neu positive target cells (SK-BR-3). ADCP was measured by two-color fluorescence flow cytometry using PKH2 (green fluorescent dye) and phycoerythrin-conjugated (red) monoclonal antibodies (MoAb) against human CD14 and CD11b. ADCC was measured with a non-radioactive LDH detection kit. Both BsAb MDX-210 (via FcgammaRI) and MoAb 520C9 (mouse IgG1, via FcgammaRII) mediated similar levels of ADCP and ADCC. ADCP mediated by BsAb MDX-H210 was identical to that mediated by BsAb MDX-210. Confocal microscopy demonstrated that dual-labeled cells represented true phagocytosis. Both ADCP and ADCC were higher when MDM were pre-incubated with GM-CSF than when incubated with M-CSF. BsAb MDX-210 is as active in vitro as the parent MoAb 520C9 in inducing both phagocytosis and cytolysis of MDM. MDX-210 and its partially humanized derivative, MDX-H210, mediated similar levels of ADCP. GM-CSF appears to superior to M-CSF in inducing MDM-mediated ADCC and ADCP. These studies support the ongoing clinical investigations of BsAb MDX-210 and its partially humanized derivative.

  3. Trivalent pneumococcal protein recombinant vaccine protects against lethal Streptococcus pneumoniae pneumonia and correlates with phagocytosis by neutrophils during early pathogenesis.

    Science.gov (United States)

    Xu, Qingfu; Surendran, Naveen; Verhoeven, David; Klapa, Jessica; Ochs, Martina; Pichichero, Michael E

    2015-02-18

    Due to the fact that current polysaccharide-based pneumococcal vaccines have limited serotype coverage, protein-based vaccine candidates have been sought for over a decade to replace or complement current vaccines. We previously reported that a trivalent Pneumococcal Protein recombinant Vaccine (PPrV), showed protection against pneumonia and sepsis in an infant murine model. Here we investigated immunological correlates of protection of PPrV in the same model. C57BL/6J infant mice were intramuscularly vaccinated at age 1-3 weeks with 3 doses of PPrV, containing pneumococcal histidine triad protein D (PhtD), pneumococcal choline binding protein A (PcpA), and detoxified pneumolysin mutant PlyD1. 3-4 weeks after last vaccination, serum and lung antibody levels to PPrV components were measured, and mice were intranasally challenged with a lethal dose of Streptococcus pneumoniae (Spn) serotype 6A. Lung Spn bacterial burden, number of neutrophils and alveolar macrophages, phagocytosed Spn by granulocytes, and levels of cytokines and chemokines were determined at 6, 12, 24, and 48h after challenge. PPrV vaccination conferred 83% protection against Spn challenge. Vaccinated mice had significantly elevated serum and lung antibody levels to three PPrV components. In the first stage of pathogenesis of Spn induced pneumonia (6-24h after challenge), vaccinated mice had lower Spn bacterial lung burdens and more phagocytosed Spn in the granulocytes. PPrV vaccination led to lower levels of pro-inflammatory cytokines IL-6, IL-1β, and TFN-α, and other cytokines and chemokines (IL-12, IL-17, IFN-γ, MIP-1b, MIP-2 and KC, and G-CSF), presumably due to a lower lung bacterial burden. Trivalent PPrV vaccination results in increased serum and lung antibody levels to the vaccine components, a reduction in Spn induced lethality, enhanced early clearance of Spn in lungs due to more rapid and thorough phagocytosis of Spn by neutrophils, and correspondingly a reduction in lung inflammation

  4. Anthocyanins from black rice (Oryza sativa) promote immune responses in leukemia through enhancing phagocytosis of macrophages in vivo.

    Science.gov (United States)

    Fan, Ming-Jen; Yeh, Ping-Hsuan; Lin, Jing-Pin; Huang, An-Cheng; Lien, Jin-Cherng; Lin, Hui-Yi; Chung, Jing-Gung

    2017-07-01

    Rice is a staple food in numerous countries around the world. Anthocyanins found in black rice have been reported to reduce the risk of certain diseases, but the effects of crude extract of anthocyanins from Asia University-selected purple glutinous indica rice (AUPGA) on immune responses have not yet been demonstrated. The current study aimed to investigate whether AUPGA treatment could affect immune responses in murine leukemia cells in vivo . Murine acute myelomonocytic leukemia WEHI-3 cells were intraperitoneally injected into normal BALB/c mice to generate leukemia mice. A total of 50 mice were randomly divided into five groups (n=10 in each group) and were fed a diet supplemented with AUPGA at 0, 20, 50 or 100 mg/kg for three weeks. All mice were weighed and the blood, liver and spleen were collected for further experiments. The results indicated that AUPGA did not significantly affect animal body weight, but significantly increased spleen weight (P<0.05) and decreased liver weight (P<0.05) when compared with the control group. AUPGA significantly increased the T cell (CD3) population at treatments of 20 and 100 mg/kg (P<0.05). However, it only significantly increased the B cell (CD19) population at a treatment of 20 mg/kg (P<0.05). Furthermore, AUPGA at 50 and 100 mg/kg significantly increased the monocyte (CD11b) population and the level of macrophages (Mac-3; P<0.05 for both). AUPGA at 50 and 100 mg/kg significantly promoted macrophage phagocytosis in peripheral blood mononuclear cells (P<0.05), and all doses of AUPGA treatment significantly promoted macrophage phagocytotic activity in the peritoneum (P<0.05). AUPGA treatment significantly decreased natural killer cell activity from splenocytes (P<0.05). Finally, AUPGA treatment at 20 mg/kg treatment significantly promoted T cell proliferation (P<0.05), and treatment at 50 and 100 mg/kg significantly decreased B cell proliferation compared with the control group (P<0.05).

  5. Phagocytosis: history's lessons.

    Science.gov (United States)

    Garg, Manish; Chandawarkar, Rajiv Y

    2013-01-01

    The assimilation of lessons from the past is an essential component of education for scientists of tomorrow. These lessons are not easy to find. History books on science are few and usually highly dramatized and biographies of scientists tend to exaggerate the pomp of scientific discovery. Both underplay the hard and laborious work that is integral to any scientific pursuit. Here we illustrate one such example. A century ago, the Nobel Prize in Medicine was awarded to two scientists: Ilya Metchnikoff, a Russian zoologist, for the discovery ofphagocytosis-a cell-mediated ingestion ofmicrobes; and Paul Ehrlich, a distinguished physician-scientist, for discovering a highly antigen-specific serum-derived antibody-based immune defense. These two diametrically opposing views of the host-pathogen interaction set the stage for a strife that led to seminal advancements in immunology. Mirrored in this journey are important lessons for scientists today--ubiquitously as applicable to modern scientific life as they were a century ago. This commentaryhighlights these lessons--a fitting centenary to a well-deserved recognition.

  6. Nanoscale characteristics of triacylglycerol oils: phase separation and binding energies of two-component oils to crystalline nanoplatelets.

    Science.gov (United States)

    MacDougall, Colin J; Razul, M Shajahan; Papp-Szabo, Erzsebet; Peyronel, Fernanda; Hanna, Charles B; Marangoni, Alejandro G; Pink, David A

    2012-01-01

    Fats are elastoplastic materials with a defined yield stress and flow behavior and the plasticity of a fat is central to its functionality. This plasticity is given by a complex tribological interplay between a crystalline phase structured as crystalline nanoplatelets (CNPs) and nanoplatelet aggregates and the liquid oil phase. Oil can be trapped within microscopic pores within the fat crystal network by capillary action, but it is believed that a significant amount of oil can be trapped by adsorption onto crystalline surfaces. This, however, remains to be proven. Further, the structural basis for the solid-liquid interaction remains a mystery. In this work, we demonstrate that the triglyceride liquid structure plays a key role in oil binding and that this binding could potentially be modulated by judicious engineering of liquid triglyceride structure. The enhancement of oil binding is central to many current developments in this area since an improvement in the health characteristics of fat and fat-structured food products entails a reduction in the amount of crystalline triacylglycerols (TAGs) and a relative increase in the amount of liquid TAGs. Excessive amounts of unbound, free oil, will lead to losses in functionality of this important food component. Engineering fats for enhanced oil binding capacity is thus central to the design of more healthy food products. To begin to address this, we modelled the interaction of triacylglycerol oils, triolein (OOO), 1,2-olein elaidin (OOE) and 1,2-elaidin olein (EEO) with a model crystalline nanoplatelet composed of tristearin in an undefined polymorphic form. The surface of the CNP in contact with the oil was assumed to be planar. We considered pure OOO and mixtures of OOO + OOE and OOO + EEO with 80% OOO. The last two cases were taken as approximations to high oleic sunflower oil (HOSO). The intent was to investigate whether phase separation on a nanoscale took place. We defined an "oil binding capacity" parameter, B

  7. Application of artificial neural networks to determine the authentication of fattening diets of Iberian pigs according to their triacylglycerol profiles

    Directory of Open Access Journals (Sweden)

    León-Camacho, M.

    2013-04-01

    Full Text Available The triacylglycerols in the subcutaneous fat from Iberian pigs reared on four different feeding types, Montanera, Recebo, extensive Cebo and intensive Cebo, have been determined by gas chromatography with a flame ionization detector. Analyses were performed in a column coated with a bonded stationary phase (50% phenyl-50% methylpolysiloxane with hydrogen as the carrier gas. Lipids were extracted by melting the subcutaneous fat in a microwave oven and then filtering and dissolving it in hexane. A total amount of 2783 samples from several campaigns were considered. Using the triacylglycerols as chemical descriptors, a study on the discriminating power to differentiate samples according to the pig feeding type and system was performed. With this aim, pattern recognition techniques, such as linear discriminant analysis (LDA and multilayer perceptron artificial neural networks (MLPANN, have been used. ANN performed better than LDA, with a mean prediction ability of approximately 97% in the differentiation of fattening diets such as Montanera, extensive Cebo and intensive Cebo. In the case of including the recebo fattening diet, the model presents a mean performance of 82%. The differentiation of fattening systems has also been achieved by means of ANN, with a mean performance of 96%.Se ha determinado mediante cromatografía de gases con detector de ionización de llama los triglicéridos de la grasa subcutánea de cerdos ibéricos, cebados con cuatro tipos de alimentación: montanera, recebo, cebo extensivo y cebo intensivo. Los análisis se realizaron en una columna con una fase estacionaria ligada químicamente (50% fenil-50% metilpolisiloxano usando hidrógeno como gas portador. La grasa subcutánea se extrajo por fusión en horno de microondas, posteriormente se filtró y se disolvió en hexano. Un total de 2.783 muestras de varias campañas fueron analizadas. Usando los triglicéridos como descriptores químicos se ha llevado a cabo un estudio

  8. Over-expression of the mycobacterial trehalose-phosphate phosphatase OtsB2 results in a defect in macrophage phagocytosis associated with increased mycobacterial-macrophage adhesion

    Directory of Open Access Journals (Sweden)

    Hao Li

    2016-11-01

    Full Text Available Trehalose-6-phosphate phosphatase (OtsB2 is involved in the OtsAB trehalose synthesis pathway to produce free trehalose and is strictly essential for mycobacterial growth. We wished to determine the effects of OtsB2 expression on mycobacterial phenotypes such as growth, phagocytosis and survival in macrophages. Mycobacterium bovis-BCG (BCG over-expressing OtsB2 were able to better survive in stationary phase. Over-expression of OtsB2 led to a decrease in phagocytosis but not survival in THP-1 macrophage-like cells, and this was not due to a decrease in general macrophage phagocytic activity. Surprisingly, when we investigated macrophage-mycobacterial interactions by flow cytometry and atomic force microscopy, we discovered that BCG over-expressing OtsB2 have stronger binding to THP-1 cells than wild-type BCG. These results suggest that altering OtsB2 expression has implications for mycobacterial host-pathogen interactions. Macrophage-mycobacteria phagocytic interactions are complex and merit further study.

  9. Deficiencies of the lipid-signaling enzymes phospholipase D1 and D2 alter cytoskeletal organization, macrophage phagocytosis, and cytokine-stimulated neutrophil recruitment.

    Directory of Open Access Journals (Sweden)

    Wahida H Ali

    Full Text Available Cell migration and phagocytosis ensue from extracellular-initiated signaling cascades that orchestrate dynamic reorganization of the actin cytoskeleton. The reorganization is mediated by effector proteins recruited to the site of activity by locally-generated lipid second messengers. Phosphatidic acid (PA, a membrane phospholipid generated by multiple enzyme families including Phospholipase D (PLD, has been proposed to function in this role. Here, we show that macrophages prepared from mice lacking either of the classical PLD isoforms PLD1 or PLD2, or wild-type macrophages whose PLD activity has been pharmacologically inhibited, display isoform-specific actin cytoskeleton abnormalities that likely underlie decreases observed in phagocytic capacity. Unexpectedly, PA continued to be detected on the phagosome in the absence of either isoform and even when all PLD activity was eliminated. However, a disorganized phagocytic cup was observed as visualized by imaging PA, F-actin, Rac1, an organizer of the F-actin network, and DOCK2, a Rac1 activator, suggesting that PLD-mediated PA production during phagocytosis is specifically critical for the integrity of the process. The abnormal F-actin reorganization additionally impacted neutrophil migration and extravasation from the vasculature into interstitial tissues. Although both PLD1 and PLD2 were important in these processes, we also observed isoform-specific functions. PLD1-driven processes in particular were observed to be critical in transmigration of macrophages exiting the vasculature during immune responses such as those seen in acute pancreatitis or irritant-induced skin vascularization.

  10. Immunomodulatory effect of exo-polysaccharides from submerged cultured Cordyceps sinensis: enhancement of cytokine synthesis, CD11b expression, and phagocytosis.

    Science.gov (United States)

    Kuo, Mei-Chun; Chang, Chien-Yu; Cheng, Tso-Lin; Wu, Ming-Jiuan

    2007-06-01

    Cordyceps sinensis is widely used as a traditional medicine for treatment of a wide variety of diseases or to maintain health. The immunomodulatory activity of polysaccharides prepared from submerged cultured C. sinensis BCRC36421 was investigated in human peripheral blood. Results demonstrated that Fr. A (exo-polysaccharides, 0.025 approximately 0.1 mg/ml) induced the production of tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-6, and IL-10 dose-dependently. Fr. A, as low as 0.025 mg/ml, could significantly augment surface expression of CD11b in monocytes and polymorphonuclear neutrophils. Functional assay revealed that Fr. A (0.05 mg/ml) also elevated phagocytosis in monocytes and PMN. On the other hand, Fr. B (intracellular polysaccharides) only moderately induced TNF-alpha release, CD11b expression, and phagocytosis at the same concentrations. Our results indicate that the immunomodulatory components of submerged cultured C. sinensis mainly reside in the culture filtrate.

  11. PLD$ is involved in phagocytosis of microglia: expression and localization changes of PLD4 are correlated with activation state of microglia.

    Directory of Open Access Journals (Sweden)

    Yoshinori Otani

    Full Text Available Phospholipase D4 (PLD4 is a recently identified protein that is mainly expressed in the ionized calcium binding adapter molecule 1 (Iba1-positive microglia in the early postnatal mouse cerebellar white matter. Unlike PLD1 and PLD2, PLD4 exhibits no enzymatic activity for conversion of phosphatidylcholine into choline and phosphatidic acid, and its function is completely unknown. In the present study, we examined the distribution of PLD4 in mouse cerebellar white matter during development and under pathological conditions. Immunohistochemical analysis revealed that PLD4 expression was associated with microglial activation under such two different circumstances. A primary cultured microglia and microglial cell line (MG6 showed that PLD4 was mainly present in the nucleus, except the nucleolus, and expression of PLD4 was upregulated by lipopolysaccharide (LPS stimulation. In the analysis of phagocytosis of LPS-stimulated microglia, PLD4 was co-localized with phagosomes that contained BioParticles. Inhibition of PLD4 expression using PLD4 specific small interfering RNA (siRNA in MG6 cells significantly reduced the ratio of phagocytotic cell numbers. These results suggest that the increased PLD4 in the activation process is involved in phagocytosis of activated microglia in the developmental stages and pathological conditions of white matter.

  12. Tamm-Horsfall Glycoprotein Enhances PMN Phagocytosis by Binding to Cell Surface-Expressed Lactoferrin and Cathepsin G That Activates MAP Kinase Pathway

    Directory of Open Access Journals (Sweden)

    Chia-Li Yu

    2011-03-01

    Full Text Available The molecular basis of polymorphonuclear neutrophil (PMN phagocytosis-enhancing activity (PEA by human purified urinary Tamm-Horsfall glyco- protein (THP has not been elucidated. In this study, we found human THP bound to lactoferrin (LF and cathepsin G (CG expressed on the surface of PMN, identified by a proteomic study with MALDI-TOF- LC/LC/mass spectrometric analysis. Pre-incubation of 10% SDS-PAGE electrophoresed PMN lysates with monoclonal anti-LF or anti-CG antibody reduced the binding with THP. To elucidate the signaling pathway of THP on PMN activation, we found THP enhanced ERK1/2 phosphorylation, reduced p38 MAP kinase phosphorylation, but had no effect on DNA binding of the five NF-kB family members in PMN. To further clarify whether the carbohydrate-side chains or protein-core structure in THP molecule is responsible for THP-PEA, THP was cleaved by different degrading enzymes with carbohydrate specificity (neuraminidase and β-galactosidase, protein specificity (V8 protease and proteinase K or glycoconjugate specificity (carboxylpeptidase Y and O-sialoglycoprotein endopeptidase. We clearly demonstrated that the intact protein-core structure in THP molecule was more important for THP-PEA than carbohydrate-side chains. Putting these results together, we conclude that THP adheres to surface-expressed LF and CG on PMN and transduces signaling via the MAP kinase pathway to enhance PMN phagocytosis.

  13. Development of a high-throughput assay for measuring lipase activity using natural triacylglycerols coated on microtiter plates.

    Science.gov (United States)

    Serveau-Avesque, Carole; Verger, Robert; Rodriguez, Jorge A; Abousalham, Abdelkarim

    2013-09-21

    We have designed a convenient, specific, sensitive and continuous lipase assay based on the use of natural triacylglycerols (TAGs) from the Aleurites fordii seed oil which contains α-eleostearic acid (9,11,13,cis,trans,trans-octadecatrienoic acid) and which was coated in the wells of microtiter plates. The coated TAG film cannot be desorbed by the various buffers used during the lipase assay. Upon lipase action, α-eleostearic acid is liberated and desorbed from the interface and then solubilized into the micellar phase. Consequently, the UV absorbance of the α-eleostearic acid is considerably enhanced due to the transformation from an adsorbed to a water soluble state. The lipase activity can be measured continuously by recording the variations with time of the UV absorption spectra. The rate of lipolysis was monitored by measuring the increase of OD at 272 nm, which was found to be linear with time and directly proportional to the amount of added lipase. This microtiter plate lipase assay, based on coated TAGs, presents various advantages as compared to the classical systems: (i) coated TAGs on the microtiter plates could be stored for a long-time at 4 °C, (ii) higher sensitivity in lipase detection, (iii) good reproducibility, and (iv) increase of signal to noise ratio due to high UV absorption after transfer of α-eleostearic acid from an adsorbed to a soluble state. Low concentrations, down to 1 pg mL(-1) of pure Thermomyces lanuginosus or human pancreatic lipase, could be detected under standard assay conditions. The detection sensitivity of this coated method is around 1000 times higher as compared to those obtained with the classical emulsified systems. This continuous high throughput lipase assay could be used to screen new lipases and/or lipase inhibitors present in various biological samples.

  14. Formation of highly structured cubic micellar lipid nanoparticles of soy phosphatidylcholine and glycerol dioleate and their degradation by triacylglycerol lipase.

    Science.gov (United States)

    Wadsäter, Maria; Barauskas, Justas; Nylander, Tommy; Tiberg, Fredrik

    2014-05-28

    Lipid nanoparticles of reversed internal phase structures, such as cubic micellar (I2) structure show good drug loading ability of peptides and proteins as well as some small molecules. Due to their controllable small size and inner morphology, such nanoparticles are suitable for drug delivery using several different administration routes, including intravenous, intramuscular, and subcutaneous injection. A very interesting system in this regard, is the two component soy phosphatidylcholine (SPC)/glycerol dioleate (GDO) system, which depending on the ratio of the lipid components form a range of reversed liquid crystalline phases. For a 50/50 (w/w) ratio in excess water, these lipids have been shown to form a reversed cubic micellar (I2) phase of the Fd3m structure. Here, we demonstrate that this SPC/GDO phase, in the presence of small quantities (5-10 wt %) of Polysorbate 80 (P80), can be dispersed into nanoparticles, still with well-defined Fd3m structure. The resulting nanoparticle dispersion has a narrow size distribution and exhibit good long-term stability. In pharmaceutical applications, biodegradation pathways of the drug delivery vehicles and their components are important considerations. In the second part of the study we show how the structure of the particles evolves during exposure to a triacylglycerol lipase (TGL) under physiological-like temperature and pH. TGL catalyzes the lipolytic degradation of acylglycerides, such as GDO, to monoglycerides, glycerol, and free fatty acids. During the degradation, the interior phase of the particles is shown to undergo continuous phase transitions from the reversed I2 structure to structures of less negative curvature (2D hexagonal, bicontinuous cubic, and sponge), ultimately resulting in the formation of multilamellar vesicles.

  15. Extending food deprivation reverses the short-term lipolytic response to fasting: role of the triacylglycerol/fatty acid cycle.

    Science.gov (United States)

    Weber, Jean-Michel; Reidy, Shannon P

    2012-05-01

    The effects of short-term food deprivation on lipid metabolism are well documented, but little is known about prolonged fasting. This study monitored the kinetics of glycerol (rate of appearance, R(a) glycerol) and non-esterified fatty acids (R(a) NEFA) in fasting rabbits. Our goals were to determine whether lipolysis is stimulated beyond values seen for short-term fasting, and to characterize the roles of primary (intracellular) and secondary (with transit through the circulation) triacylglycerol/fatty acid cycling (TAG/FA cycling) in regulating fatty acid allocation to oxidation or re-esterification. R(a) glycerol (9.62±0.72 to 15.29±0.96 μmol kg(-1) min(-1)) and R(a) NEFA (18.05±2.55 to 31.25±1.93 μmol kg(-1) min(-1)) were stimulated during the first 2 days of fasting, but returned to baseline after 4 days. An initial increase in TAG/FA cycling was followed by a reduction below baseline after 6 days without food, with primary and secondary cycling contributing to these responses. We conclude that the classic activation of lipolysis caused by short-term fasting is abolished when food deprivation is prolonged. High rates of re-esterification may become impossible to sustain, and TAG/FA cycling could decrease to reduce its cost to 3% of total energy expenditure. Throughout prolonged fasting, fatty acid metabolism gradually shifts towards increased oxidation and reduced re-esterification. Survival is achieved by pressing fuel selection towards the fatty acid dominance of energy metabolism and by slowing substrate cycles to assist metabolic suppression. However, TAG/FA cycling remains active even after prolonged fasting, suggesting that re-esterification is a crucial mechanism that cannot be stopped without harmful consequences.

  16. Rheological profiling of organogels prepared at critical gelling concentrations of natural waxes in a triacylglycerol solvent.

    Science.gov (United States)

    Patel, Ashok R; Babaahmadi, Mehrnoosh; Lesaffer, Ans; Dewettinck, Koen

    2015-05-20

    The aim of this study was to use a detailed rheological characterization to gain new insights into the gelation behavior of natural waxes. To make a comprehensive case, six natural waxes (differing in the relative proportion of chemical components: hydrocarbons, fatty alcohols, fatty acids, and wax esters) were selected as organogelators to gel high-oleic sunflower oil. Flow and dynamic rheological properties of organogels prepared at critical gelling concentrations (Cg) of waxes were studied and compared using drag (stress ramp and steady flow) and oscillatory shear (stress and frequency sweeps) tests. Although, none of the organogels satisfied the rheological definition of a "strong gel" (G″/G' (ω) ≤ 0.1), on comparing the samples, the strongest gel (highest critical stress and dynamic, apparent, and static yield stresses) was obtained not with wax containing the highest proportion of wax esters alone (sunflower wax, SFW) but with wax containing wax esters along with a higher proportion of fatty alcohols (carnauba wax, CRW) although at a comparatively higher Cg (4%wt for latter compared to 0.5%wt for former). As expected, gel formation by waxes containing a high proportion of lower melting fatty acids (berry, BW, and fruit wax, FW) required a comparatively higher Cg (6 and 7%wt, respectively), and in addition, these gels showed the lowest values for plateau elastic modulus (G'LVR) and a prominent crossover point at higher frequency. The gelation temperatures (TG'=G″) for all the studied gels were lower than room temperature, except for SFW and CRW. The yielding-type behavior of gels was evident, with most gels showing strong shear sensitivity and a weak thixotropic recovery. The rheological behavior was combined with the results of thermal analysis and microstructure studies (optical, polarized, and cryo-scanning electron microscopy) to explain the gelation properties of these waxes.

  17. Dietary structured triacylglycerols containing docosahexaenoic acid given from birth affect visual and auditory performance and tissue fatty acid profiles of rats

    DEFF Research Database (Denmark)

    Christensen, M. M.; Lund, S. P.; Simonsen, L.

    1998-01-01

    To examine whether it is possible to enhance the level of 22:6(n-3) in the central nervous system, newborn rats were fed dietary supplements containing oils with either specific or random triacylglycerol structure, but similar concentrations of polyunsaturated fatty acids. In the specific structu...... in differences in learning ability, but caused changes in visual function, evidenced by higher latency of the b-wave and lower oscillatory potential, and in auditory brainstem response, evidenced by generally greater amplitude of wave la in the group fed specific structured oil....

  18. New isochaetochromin, an inhibitor of triacylglycerol synthesis in mammalian cells, produced by Penicillium sp. FKI-4942: I. Taxonomy, fermentation, isolation and biological properties.

    Science.gov (United States)

    Ugaki, Narihiro; Matsuda, Daisuke; Yamazaki, Hiroyuki; Nonaka, Kenichi; Masuma, Rokuro; Omura, Satoshi; Tomoda, Hiroshi

    2012-01-01

    A new bis-naphtho-γ-pyrone isomer named isochaetochromin A(1) was isolated along with known isochaetochromins B(1) and B(2) from the culture broth of Penicillium sp. FKI-4942 by solvent extraction, silica gel column chromatography and HPLC. Among them, isochaetochromin B(1) showed the most potent inhibitory activity of triacylglycerol synthesis with an IC(50) value of 5.6 μM, followed by isochaetochromins B(2) (IC(50), 11 μM) and A(1) (33 μM).

  19. Analysis of molecular species of triacylglycerols from vegetable oils containing fatty acids with non-methylene-interrupted double bonds, by HPLC in the silver-ion mode

    Energy Technology Data Exchange (ETDEWEB)

    Joh, Y.; Kim, S. [Dong A Univ., Pusan (Korea, Republic of)

    1998-10-20

    The possibilities for application of silver ion HPLC to analysis of the triacylglycerols containing conjugate trienoic acids and {Delta}{sup 5}-polymethylene-interrupted acids and proportions of triacylglycerol fractions obtained by silver-ion HPLC from the seed oil of Momordica charantia double bonds were examined, respectively. The triacylglycerols of seed oils containing conjugate trienoic acids such as {alpha}-eleostearic acid (C{sub 18:3 9c,11t,13t}) and punicic acid (C{sub 18:3} {sub 9c,11t,13c}) were resolved by silver-ion HPLC. Fractions were fractionated on the basis of the number and configuration of double bonds in the species, and the elution profile is quite different from that of the species comprising exclusively saturated and unsaturated fatty acids with methylene-interrupted double bonds ; for instance, the species (DT(c2)) composed of one dienoic acid and two conjugate trienoic acids eluted much earlier than the species (D{sub 2}T{sub c}) composed of two dienoic acids and one conjugate trienoic acid, in spite of having larger number of double bonds. This means that the interaction of conjugate double bonds with silver ions is weaker than that of methylene-interrupted double bonds, presumably because of the delocalization of {pi}-electrons in conjugate double bonds. In this instance, the strength of interaction of a conjugate trienoic double bond system with silver ions seemed to be between that of methylene-interrupted dienoic and monoenoic double bond systems. Triacylglycerols of the seeds of Ginkgo biloba have been resolved by HPLC in the silver-ion mode according to the number and position of double bonds. In this instance, the strength of interaction between the {pi}-electrons of double bonds in the fatty acyl residues and silver ions is in the order; C{sub 18:3{omega}3}>C(20:3){Delta}{sup 5,11,14}C{sub 18:3}{Delta}{sup 5,9,12}>= C{sub 18:2{omega}6}>C{sub 18:2}{Delta}{sup 5,9}>C{sub 18:1{omega}9}>C{sub 18:1ome= ga7}. 49 refs., 2 figs., 2 tabs.

  20. C-type lectin B (SpCTL-B) regulates the expression of antimicrobial peptides and promotes phagocytosis in mud crab Scylla paramamosain.

    Science.gov (United States)

    Wei, Xiaoyuan; Wang, Limin; Sun, Wanwei; Zhang, Ming; Ma, Hongyu; Zhang, Yueling; Zhang, Xinxu; Li, Shengkang

    2018-07-01

    RNA transcriptional level and protein translational level in mud crab. Meantime, the phagocytosis rate and the expression of three phagocytosis related genes were declined after RNAi of SpCTL-B in hemocytes in mud crab. Collectively, our results suggest that SpCTL-B might play its roles as a pattern recognition receptor (PRR) in immune response towards pathogens infection through influencing the expression of AMPs and the phagocytosis of hemocytes in mud crab S. paramamosain. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Effects of dexamethasone and insulin on the synthesis of triacylglycerols and phosphatidylcholine and the secretion of very-low-density lipoproteins and lysophosphatidylcholine by monolayer cultures of rat hepatocytes.

    Science.gov (United States)

    Mangiapane, E H; Brindley, D N

    1986-01-01

    Rat hepatocytes in monolayer culture were preincubated for 19 h with 1 microM-dexamethasone, and the incubation was continued for a further 23 h with [14C]oleate, [3H]glycerol and 1 microM-dexamethasone. Dexamethasone increased the secretion of triacylglycerol into the medium in particles that had the properties of very-low-density lipoproteins. The increased secretion was matched by a decrease in the triacylglycerol and phosphatidylcholine that remained in the hepatocytes. Preincubating the hepatocytes for the total 42 h period with 36 nM-insulin decreased the amount of triacylglycerol in the medium and in the cells after the final incubation for 23 h with radioactive substrates. However, insulin had no significant effect on the triacylglycerol content of the cell and medium when it was present only in the final 23 h incubation. Insulin antagonized the effects of dexamethasone in stimulating the secretion of triacylglycerol from the hepatocytes, especially when it was present throughout the total 42 h period. The labelling of lysophosphatidylcholine in the medium when hepatocytes were incubated with [14C]oleate and [3H]glycerol was greater than that of phosphatidylcholine. The appearance of this lipid in the medium, unlike that of triacylglycerol and phosphatidylcholine, was not stimulated by dexamethasone, or inhibited by colchicine. However, the presence of lysophosphatidylcholine in the medium was decreased when the hepatocytes were incubated with both dexamethasone and insulin. These findings are discussed in relation to the control of the synthesis of glycerolipids and the secretion of very-low-density lipoproteins and lysophosphatidylcholine by the liver, particularly in relation to the interactions of glucocorticoids and insulin. PMID:3513755

  2. Spirulina platensis effects on the levels of total cholesterol, HDL and triacylglycerols in rabbits fed with a hypercholesterolemic diet

    Directory of Open Access Journals (Sweden)

    Luciane Maria Colla

    2008-04-01

    Full Text Available In this work, hypercholesterolemia was induced in rabbits by feeding them a high cholesterol diet (CD, 350 mg/d and the effects of supplementing this diet with 0.5 g/d Spirulina platensis was evaluated by measuring the levels of serum total-cholesterol (TC, triacylglycerols (TAG and high-density lipoprotein (HDL-cholesterol at the start of the experiment and after 30 d and 60 d. It was found that the levels of serum cholesterol decreased from 1,054±101 mg.dL-1 in the rabbits fed a CD without S. platensis to 516±163 mg.dL-1 to those fed with a high cholesterol diet supplemented with S. platensis (significant at p A microalga Spirulina é cultivada e comercializada no mundo devido a suas características nutricionais (elevada concentração de proteínas, em torno de 65%, vitaminas e sais minerais e ao seu potencial terapêutico no tratamento de inúmeras doenças, inclusive a hipercolesterolemia. Neste trabalho foi avaliada a inibição da hipercolesterolemia induzida em coelhos por uma dieta adicionada de colesterol (350 mg.d-1, pela suplementação de 0,5 g.dia-1 de biomassa de Spirulina platensis, sendo avaliados os níveis de colesterol total, triglicerídeos e HDL nos tempos de 0 d, 30 d e 60 d de tratamento. Os resultados indicaram que a adição de Spirulina platensis na dieta ocasionou decréscimo nos níveis de colesterol total de 1054±101 mg.dL-1 para 516±163 mg.dL-1 (p<0,0001, para os coelhos alimentados com a dieta colesterolêmica em comparação com os que receberam a dieta adicionada de Spirulina platensis. A adição de Spirulina a dieta colesterolêmica não ocasionou decréscimo significativo nos níveis de triglicerídios dos coelhos. Os valores de HDL aumentaram de 73±31 mg.dL-1 para 91,0±15,7 mg.dL-1, comparando-se os coelhos alimentados com a dieta colesterolêmica e os alimentados com a dieta adicionada de Spirulina, estatisticamente diferentes a um nível de significância maior que 0,1533.

  3. Short term and dosage influences of palm based medium- and long-chain triacylglycerols on body fat and blood parameters in C57BL/6J mice.

    Science.gov (United States)

    Lee, Yee-Ying; Tang, Teck-Kim; Ab Karim, Nur Azwani; Alitheen, Noorjahan Banu Mohamed; Lai, Oi-Ming

    2014-01-01

    Structured lipid medium- and long-chain triacylglycerols (MLCT) are claimed to be able to manage obesity. The present study investigated the body fat influence of enzymatically interesterifed palm-based medium- and long-chain triacylglycerols (P-MLCT) on diet-induced obesity (DIO) C57BL/6J mice compared with commercial MLCT oil (C-MLCT) and a control, which was the non enzymatically modified palm kernel and palm oil blend (PKO-PO blend). It also investigated the low fat and high fat effects of P-MLCT. DIO C57BL/6J mice were fed ad libitum with low fat (7%) and high fat (30%) experimental diets for 8 weeks before being sacrificed to obtain blood serum for analysis. From the results, there is a trend that P-MLCT fed mice were found to have the lowest body weight, body weight gain, total fat pad accumulation (perirenal, retroperitoneal, epididymal and mesenteric), total triglyceride levels and efficiency in controlling blood glucose level, compared with C-MLCT and the PKO-PO blend in both low fat and high fat diets. Nevertheless, the PKO-PO blend and P-MLCT caused significantly (P < 0.05) higher total cholesterol levels compared to C-MLCT. P-MLCT present in low fat and high fat dosage were shown to be able to suppress body fat accumulation. This effect is more prominent with the low fat dosage.

  4. The intramolecular position of docosahexaenoic acid in the triacylglycerol sources used for pediatric nutrition has a minimal effect on its metabolic use.

    Science.gov (United States)

    Sala-Vila, Aleix; Castellote, Ana I; López-Sabater, M Carmen

    2008-03-01

    Docosahexaenoic acid (DHA) plays an important role in normal development of the brain and retina in the human. In utero, DHA is incorporated in the fetus, and its accretion continues throughout early postnatal life. Although human breast milk contains this fatty acid, several organizations recommend supplementing infant formulas with DHA for infants and premature infants. Traditionally, certain types of fish oil have been used for fortifying some infant formulas, but with the decline in world fisheries, the search for alternative sources of DHA continues. Among the viable ingredient sources of DHA is oil derived from single-cell organisms (marine microorganisms); however, these oil sources display different positional specificity of DHA in the glycerol lipids compared with that found in human breast milk lipids. In the latter, the DHA is mainly esterified in the central position of the glycerol backbone. Because of these differences in human milk and oils derived from single-cell organisms, recent research in biotechnology has focused on developing new structured triacylglycerols with an intramolecular structure resembling that found in human milk lipids. This research is justified by the potential differences in metabolism of DHA based on the hypothetical bioavailability and benefits in DHA found in human milk lipids. Presented herein is a review of the published research on the metabolism of DHA from different triacylglycerol sources including in vitro studies and animal studies. Despite small differences observed in digestion, the current data reveal a minimal effect on the parameters of development studied for the intramolecular position in which DHA is esterified.

  5. Triacylglycerols profiling in plant oils important in food industry, dietetics and cosmetics using high-performance liquid chromatography-atmospheric pressure chemical ionization mass spectrometry.

    Science.gov (United States)

    Lísa, Miroslav; Holcapek, Michal

    2008-07-11

    Optimized non-aqueous reversed-phase high-performance liquid chromatography method using acetonitrile-2-propanol gradient elution and the column coupling in the total length of 45 cm has been applied for the high resolution separation of plant oils important in food industry, dietetics and cosmetics. Positive-ion atmospheric pressure chemical ionization mass spectrometry is used for the unambiguous identification and also the reliable quantitation with the response factors approach. Based on the precise determination of individual triacyglycerol concentrations, the calculation of average parameters important in the nutrition is performed, i.e. average carbon number, average double bond number, relative concentrations of essential, saturated, monounsaturated and polyunsaturated fatty acids. Results are reported in the form of both chromatographic fingerprints and tables containing relative concentrations for all triacylglycerols and fatty acids in individual samples. In total, 264 triacylglycerols consisting of 28 fatty acids with the alkyl chain length from 6 to 26 carbon atoms and 0 to 4 double bonds have been identified in 26 industrial important plant oils.

  6. Factor H binds to the hypervariable region of many Streptococcus pyogenes M proteins but does not promote phagocytosis resistance or acute virulence

    DEFF Research Database (Denmark)

    Gustafsson, Caj Ulrik Mattias; Lannergård, Jonas; Nilsson, Olof Rickard

    2013-01-01

    Many pathogens express a surface protein that binds the human complement regulator factor H (FH), as first described for Streptococcus pyogenes and the antiphagocytic M6 protein. It is commonly assumed that FH recruited to an M protein enhances virulence by protecting the bacteria against...... represents a distinct ligand-binding domain. The isolated HVRs specifically interacted with FH among all human serum proteins, interacted with the same region in FH and showed species specificity, but exhibited little or no antigenic cross-reactivity. Although these findings suggested that FH recruited...... to an M protein promotes virulence, studies in transgenic mice did not demonstrate a role for bound FH during acute infection. Moreover, phagocytosis tests indicated that ability to bind FH is neither sufficient nor necessary for S. pyogenes to resist killing in whole human blood. While these data shed...

  7. Factor H Binds to the Hypervariable Region of Many Streptococcus pyogenes M Proteins but Does Not Promote Phagocytosis Resistance or Acute Virulence

    Science.gov (United States)

    Kristensen, Bodil M.; Olsen, John E.; Harris, Claire L.; Ufret-Vincenty, Rafael L.; Stålhammar-Carlemalm, Margaretha; Lindahl, Gunnar

    2013-01-01

    Many pathogens express a surface protein that binds the human complement regulator factor H (FH), as first described for Streptococcus pyogenes and the antiphagocytic M6 protein. It is commonly assumed that FH recruited to an M protein enhances virulence by protecting the bacteria against complement deposition and phagocytosis, but the role of FH-binding in S. pyogenes pathogenesis has remained unclear and controversial. Here, we studied seven purified M proteins for ability to bind FH and found that FH binds to the M5, M6 and M18 proteins but not the M1, M3, M4 and M22 proteins. Extensive immunochemical analysis indicated that FH binds solely to the hypervariable region (HVR) of an M protein, suggesting that selection has favored the ability of certain HVRs to bind FH. These FH-binding HVRs could be studied as isolated polypeptides that retain ability to bind FH, implying that an FH-binding HVR represents a distinct ligand-binding domain. The isolated HVRs specifically interacted with FH among all human serum proteins, interacted with the same region in FH and showed species specificity, but exhibited little or no antigenic cross-reactivity. Although these findings suggested that FH recruited to an M protein promotes virulence, studies in transgenic mice did not demonstrate a role for bound FH during acute infection. Moreover, phagocytosis tests indicated that ability to bind FH is neither sufficient nor necessary for S. pyogenes to resist killing in whole human blood. While these data shed new light on the HVR of M proteins, they suggest that FH-binding may affect S. pyogenes virulence by mechanisms not assessed in currently used model systems. PMID:23637608

  8. Factor H binds to the hypervariable region of many Streptococcus pyogenes M proteins but does not promote phagocytosis resistance or acute virulence.

    Directory of Open Access Journals (Sweden)

    Mattias C U Gustafsson

    Full Text Available Many pathogens express a surface protein that binds the human complement regulator factor H (FH, as first described for Streptococcus pyogenes and the antiphagocytic M6 protein. It is commonly assumed that FH recruited to an M protein enhances virulence by protecting the bacteria against complement deposition and phagocytosis, but the role of FH-binding in S. pyogenes pathogenesis has remained unclear and controversial. Here, we studied seven purified M proteins for ability to bind FH and found that FH binds to the M5, M6 and M18 proteins but not the M1, M3, M4 and M22 proteins. Extensive immunochemical analysis indicated that FH binds solely to the hypervariable region (HVR of an M protein, suggesting that selection has favored the ability of certain HVRs to bind FH. These FH-binding HVRs could be studied as isolated polypeptides that retain ability to bind FH, implying that an FH-binding HVR represents a distinct ligand-binding domain. The isolated HVRs specifically interacted with FH among all human serum proteins, interacted with the same region in FH and showed species specificity, but exhibited little or no antigenic cross-reactivity. Although these findings suggested that FH recruited to an M protein promotes virulence, studies in transgenic mice did not demonstrate a role for bound FH during acute infection. Moreover, phagocytosis tests indicated that ability to bind FH is neither sufficient nor necessary for S. pyogenes to resist killing in whole human blood. While these data shed new light on the HVR of M proteins, they suggest that FH-binding may affect S. pyogenes virulence by mechanisms not assessed in currently used model systems.

  9. Preventive effects of a fermented dairy product against Alzheimer's disease and identification of a novel oleamide with enhanced microglial phagocytosis and anti-inflammatory activity.

    Directory of Open Access Journals (Sweden)

    Yasuhisa Ano

    Full Text Available Despite the ever-increasing number of patients with dementia worldwide, fundamental therapeutic approaches to this condition have not been established. Epidemiological studies suggest that intake of fermented dairy products prevents cognitive decline in the elderly. However, the active compounds responsible for the effect remain to be elucidated. The present study aims to elucidate the preventive effects of dairy products on Alzheimer's disease and to identify the responsible component. Here, in a mouse model of Alzheimer's disease (5xFAD, intake of a dairy product fermented with Penicillium candidum had preventive effects on the disease by reducing the accumulation of amyloid β (Aβ and hippocampal inflammation (TNF-α and MIP-1α production, and enhancing hippocampal neurotrophic factors (BDNF and GDNF. A search for preventive substances in the fermented dairy product identified oleamide as a novel dual-active component that enhanced microglial Aβ phagocytosis and anti-inflammatory activity towards LPS stimulation in vitro and in vivo. During the fermentation, oleamide was synthesized from oleic acid, which is an abundant component of general dairy products owing to lipase enzymatic amidation. The present study has demonstrated the preventive effect of dairy products on Alzheimer's disease, which was previously reported only epidemiologically. Moreover, oleamide has been identified as an active component of dairy products that is considered to reduce Aβ accumulation via enhanced microglial phagocytosis, and to suppress microglial inflammation after Aβ deposition. Because fermented dairy products such as camembert cheese are easy to ingest safely as a daily meal, their consumption might represent a preventive strategy for dementia.

  10. Glycogen synthase kinase-3 (GSK3) regulates TNF production and haemocyte phagocytosis in the immune response of Chinese mitten crab Eriocheir sinensis.

    Science.gov (United States)

    Li, Xiaowei; Jia, Zhihao; Wang, Weilin; Wang, Lingling; Liu, Zhaoqun; Yang, Bin; Jia, Yunke; Song, Xiaorui; Yi, Qilin; Qiu, Limei; Song, Linsheng

    2017-08-01

    Glycogen synthase kinase-3 (GSK3) is a serine/threonine protein kinase firstly identified as a regulator of glycogen synthesis. Recently, it has been proved to be a key regulator of the immune reaction. In the present study, a GSK3 homolog gene (designated as EsGSK3) was cloned from Chinese mitten crab, Eriocheir sinensis. The open reading frame (ORF) was 1824 bp, which encoded a predicted polypeptide of 607 amino acids. There was a conserved Serine/Threonine Kinase domain and a DNA binding domain found in EsGSK3. Phylogenetic analysis showed that EsGSK3 was firstly clustered with GSK3-β from oriental river prawn Macrobrachium nipponense in the invertebrate branch, while GSK3s from vertebrates formed the other distinct branch. EsGSK3 mRNA transcripts could be detected in all tested tissues of the crab including haepatopancreas, eyestalk, muscle, gonad, haemocytes and haematopoietic tissue with the highest expression level in haepatopancreas. And EsGSK3 protein was mostly detected in the cytoplasm of haemocyte by immunofluorescence analysis. The expression levels of EsGSK3 mRNA increased significantly at 6 h after Aeromonas hydrophila challenge (p level at 48 h (p > 0.05). The mRNA expression of lipopolysaccharide-induced tumor necrosis factor (TNF)-α factor (EsLITAF) was also induced by A. hydrophila challenge. However, the mRNA expression of EsLITAF and TNF-α production was significantly suppressed after EsGSK3 was blocked in vivo with specific inhibitor lithium, while the phagocytosis of crab haemocytes was significantly promoted. These results collectively demonstrated that EsGSK3 could regulate the innate immune responses of E. sinensis by promoting TNF-α production and inhibiting haemocyte phagocytosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Preventive effects of a fermented dairy product against Alzheimer's disease and identification of a novel oleamide with enhanced microglial phagocytosis and anti-inflammatory activity.

    Science.gov (United States)

    Ano, Yasuhisa; Ozawa, Makiko; Kutsukake, Toshiko; Sugiyama, Shinya; Uchida, Kazuyuki; Yoshida, Aruto; Nakayama, Hiroyuki

    2015-01-01

    Despite the ever-increasing number of patients with dementia worldwide, fundamental therapeutic approaches to this condition have not been established. Epidemiological studies suggest that intake of fermented dairy products prevents cognitive decline in the elderly. However, the active compounds responsible for the effect remain to be elucidated. The present study aims to elucidate the preventive effects of dairy products on Alzheimer's disease and to identify the responsible component. Here, in a mouse model of Alzheimer's disease (5xFAD), intake of a dairy product fermented with Penicillium candidum had preventive effects on the disease by reducing the accumulation of amyloid β (Aβ) and hippocampal inflammation (TNF-α and MIP-1α production), and enhancing hippocampal neurotrophic factors (BDNF and GDNF). A search for preventive substances in the fermented dairy product identified oleamide as a novel dual-active component that enhanced microglial Aβ phagocytosis and anti-inflammatory activity towards LPS stimulation in vitro and in vivo. During the fermentation, oleamide was synthesized from oleic acid, which is an abundant component of general dairy products owing to lipase enzymatic amidation. The present study has demonstrated the preventive effect of dairy products on Alzheimer's disease, which was previously reported only epidemiologically. Moreover, oleamide has been identified as an active component of dairy products that is considered to reduce Aβ accumulation via enhanced microglial phagocytosis, and to suppress microglial inflammation after Aβ deposition. Because fermented dairy products such as camembert cheese are easy to ingest safely as a daily meal, their consumption might represent a preventive strategy for dementia.

  12. Preventive Effects of a Fermented Dairy Product against Alzheimer’s Disease and Identification of a Novel Oleamide with Enhanced Microglial Phagocytosis and Anti-Inflammatory Activity

    Science.gov (United States)

    Ano, Yasuhisa; Ozawa, Makiko; Kutsukake, Toshiko; Sugiyama, Shinya; Uchida, Kazuyuki; Yoshida, Aruto; Nakayama, Hiroyuki

    2015-01-01

    Despite the ever-increasing number of patients with dementia worldwide, fundamental therapeutic approaches to this condition have not been established. Epidemiological studies suggest that intake of fermented dairy products prevents cognitive decline in the elderly. However, the active compounds responsible for the effect remain to be elucidated. The present study aims to elucidate the preventive effects of dairy products on Alzheimer’s disease and to identify the responsible component. Here, in a mouse model of Alzheimer’s disease (5xFAD), intake of a dairy product fermented with Penicillium candidum had preventive effects on the disease by reducing the accumulation of amyloid β (Aβ) and hippocampal inflammation (TNF-α and MIP-1α production), and enhancing hippocampal neurotrophic factors (BDNF and GDNF). A search for preventive substances in the fermented dairy product identified oleamide as a novel dual-active component that enhanced microglial Aβ phagocytosis and anti-inflammatory activity towards LPS stimulation in vitro and in vivo. During the fermentation, oleamide was synthesized from oleic acid, which is an abundant component of general dairy products owing to lipase enzymatic amidation. The present study has demonstrated the preventive effect of dairy products on Alzheimer’s disease, which was previously reported only epidemiologically. Moreover, oleamide has been identified as an active component of dairy products that is considered to reduce Aβ accumulation via enhanced microglial phagocytosis, and to suppress microglial inflammation after Aβ deposition. Because fermented dairy products such as camembert cheese are easy to ingest safely as a daily meal, their consumption might represent a preventive strategy for dementia. PMID:25760987

  13. Triacylglycerol composition, physico-chemical characteristics and oxidative stability of interesterified canola oil and fully hydrogenated cottonseed oil blends.

    Science.gov (United States)

    Imran, Muhammad; Nadeem, Muhammad

    2015-10-29

    Partial hydrogenation process is used worldwide to produce shortening, baking, and pastry margarines for food applications. However, demand for such products is decreased during last decade due to their possible links to consumer health and disease. This has raised the need to replace hydrogenation with alternative acceptable interesterification process which has advantage in context of modifying the physico-chemical properties of edible fat-based products. Therefore, the main mandate of research was the development of functional fat through chemical interesterification of canola oil (CaO) and fully hydrogenated cottonseed oil (FHCSO) mixtures. Blends were prepared in the proportions of 75:25 (T1), 50:50 (T2) and 25:75 (T3) of CaO:FHCSO (w/w). Interesterification was performed using sodium methoxide (0.2 %) as catalyst at 120 °C, under reduced pressure and constant agitation for 60 minutes. The non-interesterified and interesterified CaO:FHCSO blends were evaluated for triacylglycerol (TAG) composition, physico-chemical characteristics, oxidative stability and consumer acceptability at 0, 30 and 60 days of storage interval. The oleic acid (58.3 ± 0.6 %) was predominantly present in CaO while the contents of stearic acid (72 ± 0.8 %) were significantly higher in FHCSO. Maximum trisaturated (S3) contents (63.9 ± 0.5 %) were found in T3 while monounsaturated (S2U), diunsaturated (U2S) and triunsaturated (U3) contents were quite low in T2 and T3 before interesterification. A marked reduction in S3 and U3 contents with concomitant increase in S2U and U2S contents was observed for all CaO:FHCSO blends on interesterification. During storage, the changes in S3, S2U and U2S contents were not found significant (p ≥ 0.05). However, maximum decrease 13 %, 7.5 and 5.6 % in U3 contents for T1, T2 and T3 was noted after 60-days of interesterification, respectively. The Lovibond color R, melting point, refractive index, specific gravity, peroxide and free

  14. No difference between high-fructose and high-glucose diets on liver triacylglycerol or biochemistry in healthy overweight men.

    Science.gov (United States)

    Johnston, Richard D; Stephenson, Mary C; Crossland, Hannah; Cordon, Sally M; Palcidi, Elisa; Cox, Eleanor F; Taylor, Moira A; Aithal, Guruprasad P; Macdonald, Ian A

    2013-11-01

    Diets high in fructose have been proposed to contribute to nonalcoholic fatty liver disease. We compared the effects of high-fructose and matched glucose intake on hepatic triacylglycerol (TAG) concentration and other liver parameters. In a double-blind study, we randomly assigned 32 healthy but centrally overweight men to groups that received either a high-fructose or high-glucose diet (25% energy). These diets were provided during an initial isocaloric period of 2 weeks, followed by a 6-week washout period, and then again during a hypercaloric 2-week period. The primary outcome measure was hepatic level of TAG, with additional assessments of TAG levels in serum and soleus muscle, hepatic levels of adenosine triphosphate, and systemic and hepatic insulin resistance. During the isocaloric period of the study, both groups had stable body weights and concentrations of TAG in liver, serum, and soleus muscle. The high-fructose diet produced an increase of 22 ± 52 μmol/L in the serum level of uric acid, whereas the high-glucose diet led to a reduction of 23 ± 25 μmol/L (P fructose diet also produced an increase of 0.8 ± 0.9 in the homeostasis model assessment of insulin resistance, whereas the high-glucose diet produced an increase of only 0.1 ± 0.7 (P = .03). During the hypercaloric period, participants in the high-fructose and high-glucose groups had similar increases in weight (1.0 ± 1.4 vs 0.6 ± 1.0 kg; P = .29) and absolute concentration of TAG in liver (1.70% ± 2.6% vs 2.05% ± 2.9%; P = .73) and serum (0.36 ± 0.75 vs 0.33 ± 0.38 mmol/L; P = .91), and similar results in biochemical assays of liver function. Body weight changes were associated with changes in liver biochemistry and concentration of TAGs. In the isocaloric period, overweight men who were on a high-fructose or a high-glucose diet did not develop any significant changes in hepatic concentration of TAGs or serum levels of liver enzymes. However, in the hypercaloric period

  15. Characterization of fatty acid and triacylglycerol composition in animal fats using silver-ion and non-aqueous reversed-phase high-performance liquid chromatography/mass spectrometry and gas chromatography/flame ionization detection

    Czech Academy of Sciences Publication Activity Database

    Lísa, M.; Netušilová, K.; Franěk, L.; Dvořáková, H.; Vrkoslav, Vladimír; Holčapek, M.

    2011-01-01

    Roč. 1218, č. 42 (2011), s. 7499-7510 ISSN 0021-9673 R&D Projects: GA ČR GA203/09/0139 Institutional research plan: CEZ:AV0Z40550506 Keywords : silver-ion HPLC * non-aqueous reversed-phase HPLC * triacylglycerol Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 4.531, year: 2011

  16. Effect of simultaneous, single oral doses of beta-carotene with lutein or lycopene on the beta-carotene and retinyl ester responses in the triacylglycerol-rich lipoprotein fraction of men

    NARCIS (Netherlands)

    Berg, H. van den; Vliet, T. van

    1998-01-01

    The effects of lutein and lycopene on β-carotene absorption and cleavage were investigated in 12 male subjects. Responses of carotenoids and retinyl palmitate in the triacylglycerol-rich lipoprotein (TRL) fraction after a separate 15-mg β-carotene dose were compared with those after a dose of 15 mg

  17. The GCKR rs780094 polymorphism is associated with elevated fasting serum triacylglycerol, reduced fasting and OGTT-related insulinaemia, and reduced risk of type 2 diabetes

    DEFF Research Database (Denmark)

    Sparsø, T; Andersen, G; Nielsen, T

    2007-01-01

    to metabolic traits (mainly fasting hypertriacylglycerolaemia) and traits related to pancreatic beta cell function. METHODS: The polymorphism was genotyped in 16,853 Danes using Taqman allelic discrimination. Association was analysed in case-control studies and quantitative trait analyses. We also analysed...... the possible interactive effect between the GCK -30G>A polymorphism and the GCKR rs780094 variant on metabolic traits. RESULTS: The minor GCKR A-allele of rs780094 is associated with an increased level of fasting serum triacylglycerol (p = 6 x 10(-14)), impaired fasting (p = 0.001) and OGTT-related insulin...... release (p = 3 x 10(-6)), reduced homeostasis model assessment of insulin resistance (p = 0.0004), WHO-defined dyslipidaemia (p = 6 x 10(-9)) and a modestly decreased risk of type 2 diabetes (p = 0.01). Significantly increased fasting serum insulin concentrations were demonstrated when analysing the GCK...

  18. Triacylglycerol Analysis in Human Milk and Other Mammalian Species: Small-Scale Sample Preparation, Characterization, and Statistical Classification Using HPLC-ELSD Profiles.

    Science.gov (United States)

    Ten-Doménech, Isabel; Beltrán-Iturat, Eduardo; Herrero-Martínez, José Manuel; Sancho-Llopis, Juan Vicente; Simó-Alfonso, Ernesto Francisco

    2015-06-24

    In this work, a method for the separation of triacylglycerols (TAGs) present in human milk and from other mammalian species by reversed-phase high-performance liquid chromatography using a core-shell particle packed column with UV and evaporative light-scattering detectors is described. Under optimal conditions, a mobile phase containing acetonitrile/n-pentanol at 10 °C gave an excellent resolution among more than 50 TAG peaks. A small-scale method for fat extraction in these milks (particularly of interest for human milk samples) using minimal amounts of sample and reagents was also developed. The proposed extraction protocol and the traditional method were compared, giving similar results, with respect to the total fat and relative TAG contents. Finally, a statistical study based on linear discriminant analysis on the TAG composition of different types of milks (human, cow, sheep, and goat) was carried out to differentiate the samples according to their mammalian origin.

  19. The effect of Lactobacillus paracasei subsp. paracasei L. casei W8® on blood levels of triacylglycerol is independent of colonisation

    DEFF Research Database (Denmark)

    Bjerg, Anne Toksvig; Sørensen, M.B.; Krych, Lukasz

    2015-01-01

    Gut microbiota (GM) dysbiosis has been linked to obesity and its metabolic complications such as cardiovascular disease (CVD). The risk of developing CVD increases with elevated concentration of serum triacylglycerol (TAG). In a blinded, randomised two-arm parallel human intervention study we have...... study we analysed faecal samples obtained during the intervention study to investigate whether this effect was related to the ability of L. casei W8 to colonise the human gut after supplementation of L. casei W8 (1010 cfu daily) as determined by qPCR specific for L. paracasei and L. casei (L. casei...... and two weeks after the supplementation was ended, and fasting blood samples were collected at baseline and after 4 weeks. Four weeks supplementation with L. casei W8 did not affect the overall composition of the GM; however, an increase in the relative abundance of the L. casei group from 8...

  20. The utilization of the acyl-CoA and the involvement PDAT and DGAT in the biosynthesis of erucic acid-rich triacylglycerols in Crambe seed oil.

    Science.gov (United States)

    Furmanek, Tomasz; Demski, Kamil; Banaś, Walentyna; Haslam, Richard; Napier, Jonathan; Stymne, Sten; Banaś, Antoni

    2014-04-01

    The triacylglycerol of Crambe abyssinica seeds consist of 95% very long chain (>18 carbon) fatty acids (86% erucic acid; 22:1∆13) in the sn-1 and sn-3 positions. This would suggest that C. abyssinica triacylglycerols are not formed by the action of the phospholipid:diacylglycerol acyltransferase (PDAT), but are rather the results of acyl-CoA:diacylglycerol acyltransferase (DGAT) activity. However, measurements of PDAT and DGAT activities in microsomal membranes showed that C. abyssinica has significant PDAT activity, corresponding to about 10% of the DGAT activity during periods of rapid seed oil accumulation. The specific activity of DGAT for erucoyl-CoA had doubled at 19 days after flowering compared to earlier developmental stages, and was, at that stage, the preferred acyl donor, whereas the activities for 16:0-CoA and 18:1-CoA remained constant. This indicates that an expression of an isoform of DGAT with high specificity for erucoyl-CoA is induced at the onset of rapid erucic acid and oil accumulation in the C. abyssinica seeds. Analysis of the composition of the acyl-CoA pool during different stages of seed development showed that the percentage of erucoyl groups in acyl-CoA was much higher than in complex lipids at all stages of seed development except in the desiccation phase. These results are in accordance with published results showing that the rate limiting step in erucic acid accumulation in C. abyssinica oil is the utilization of erucoyl-CoA by the acyltransferases in the glycerol-3-phosphate pathway.

  1. Fatty acid and triacylglycerol composition of the subcutaneous fat from iberian pigs fattened on the traditional feed: “Montanera”. effect of anatomical location and length of feeding

    Directory of Open Access Journals (Sweden)

    Narváez-Rivas, Mónica

    2009-07-01

    Full Text Available Fatty acid and triacylglycerol compositions of 200 samples of subcutaneous fat from two different anatomical locations (rump and adipose tissue covering the Biceps femoris muscle of Iberian purebred pigs reared on “Montanera” were determined. Significant differences were found for the majority fatty acids and for some triacylglycerol species (PPS, PLPo + MLO, PLO, PLL + PoLO, SOS, SOL, OLL among the two anatomical locations, being the rump location less saturated. The activity level of the key enzyme involved in lipogenesis differed (p Biceps femoris, increases faster than that of the subcutaneous fat covering a muscle with low oxidative metabolism, as Longissimus dorsi.Se ha determinado la composición de ácidos grasos y de triglicéridos en 200 muestras de grasa subcutánea procedentes de dos localizaciones anatómicas (rabadilla y tejido adiposo que recubre el músculo Biceps femoris de cerdos ibéricos puros alimentados en “Montanera”. Se encontraron diferencias significativas para la mayoría de ácidos grasos y para algunos triglicéridos PPS, PLPo + MLO, PLO, PLL + PoLO, SOS, SOL, OLL entre las dos localizaciones anatómicas, siendo la rabadilla la menos saturada. El nivel de actividad de la enzima involucrada en la lipogénesis defirió significativamente (p B. femoris, aumenta más rápidamente que la de la grasa subcutánea que recubre un músculo con bajo metabolismo oxidativo, como el Longissimus dorsi.

  2. A rich medium-chain triacylglycerol diet benefits adiposity but has adverse effects on the markers of hepatic lipogenesis and beta-oxidation.

    Science.gov (United States)

    Chamma, Carolina Maria de Oliveira; Bargut, Thereza Cristina Lonzetti; Mandarim-de-Lacerda, Carlos Alberto; Aguila, Marcia Barbosa

    2017-02-22

    We investigated the increasing amounts of medium-chain triacylglycerol (MCT) in the diet on hepatic lipid metabolism. Mature C57BL/6 male mice were randomly divided into five groups (n = 10/group). The animals received their diet for 12 weeks, as a control (C group, 10% of energy from lipids); high-fat lard (HF group, isoenergetic diet, 50% of energy from lipids with lard); a mixture of lard and MCT oil (with a gradual replacement of lard by MCT: HF-MCT25%, HF-MCT75%, and HF-MCT100% groups). At euthanasia, we collected blood and dissected the liver for analyses (glucose, insulin, HOMA-IR, QUICK index, and triacylglycerol, light microscopy, western blotting, and RT-qPCR). The HF diet groups showed a greater body mass gain compared to the C group, but the HF-MCT100% group showed diminished adiposity and amelioration of insulin resistance. All the HF groups also showed a clear increase in hepatic lipid accumulation, increased lipogenesis and decreased PPAR-alpha expression, although HF-MCT groups showed improved local insulin signaling. Lastly, the HF-MCT100% group had raised markers of beta-oxidation (UCP3 and MCAD) and mitochondrial biogenesis (PGC1-alpha and NRF1). In conclusion, the findings demonstrated that a high amount of MCT (HF-MCT100% group) added to an HF diet reduces the body fat accumulation and insulin resistance. However, the lipid accumulation as well as the lipid metabolism is altered in the liver of animals fed with a very high MCT diet, indicating that higher doses of MCT may be harmful in a long-term.

  3. Emodin Induces Apoptotic Death in Murine Myelomonocytic Leukemia WEHI-3 Cells In Vitro and Enhances Phagocytosis in Leukemia Mice In Vivo

    Directory of Open Access Journals (Sweden)

    Yuan-Chang Chang

    2011-01-01

    Full Text Available Emodin is one of major compounds in rhubarb (Rheum palmatum L., a plant used as herbal medicine in Chinese population. Although many reports have shown that emodin exhibits anticancer activity in many tumor cell types, there is no available information addressing emodin-affected apoptotic responses in the murine leukemia cell line (WEHI-3 and modulation of the immune response in leukemia mice. We investigated that emodin induced cytotoxic effects in vitro and affected WEHI-3 cells in vivo. This study showed that emodin decreased viability and induced DNA fragmentation in WEHI-3 cells. Cells after exposure to emodin for 24 h have shown chromatin condensation and DNA damage. Emodin stimulated the productions of ROS and Ca2+ and reduced the level of ΔΨm by flow cytometry. Our results from Western blotting suggest that emodin triggered apoptosis of WEHI-3 cells through the endoplasmic reticulum (ER stress, caspase cascade-dependent and -independent mitochondrial pathways. In in vivo study, emodin enhanced the levels of B cells and monocytes, and it also reduced the weights of liver and spleen compared with leukemia mice. Emodin promoted phagocytic activity by monocytes and macrophages in comparison to the leukemia mice group. In conclusions, emodin induced apoptotic death in murine leukemia WEHI-3 cells and enhanced phagocytosis in the leukemia animal model.

  4. Phagocytosis of cholesteryl ester is amplified in diabetic mouse macrophages and is largely mediated by CD36 and SR-A.

    Directory of Open Access Journals (Sweden)

    Christopher B Guest

    Full Text Available Type 2 diabetes (T2D is associated with accelerated atherosclerosis, which accounts for approximately 75% of all diabetes-related deaths. Here we investigate the link between diabetes and macrophage cholesteryl ester accumulation. When diabetic (db/db mice are given cholesteryl ester intraperitoneally (IP, peritoneal macrophages (PerMPhis recovered from these animals showed a 58% increase in intracellular cholesteryl ester accumulation over PerMPhis from heterozygote control (db/+ mice. Notably, PerMPhi fluid-phase endocytosis and large particle phagocytosis was equivalent in db/+and db/db mice. However, IP administration of CD36 and SR-A blocking antibodies led to 37% and 25% reductions in cholesteryl ester accumulation in PerMPhi. Finally, in order to determine if these scavenger receptors (SRs were part of the mechanism responsible for the increased accumulation of cholesteryl esters observed in the diabetic mouse macrophages, receptor expression was quantified by flow cytometry. Importantly, db/db PerMPhis showed a 43% increase in CD36 expression and an 80% increase in SR-A expression. Taken together, these data indicate that direct cholesteryl ester accumulation in mouse macrophages is mediated by CD36 and SR-A, and the magnitude of accumulation is increased in db/db macrophages due to increased scavenger receptor expression.

  5. Complement Receptor 3-Mediated Inhibition of Inflammasome Priming by Ras GTPase-Activating Protein During Francisella tularensis Phagocytosis by Human Mononuclear Phagocytes

    Directory of Open Access Journals (Sweden)

    Ky V. Hoang

    2018-03-01

    Full Text Available Francisella tularensis is a remarkably infectious facultative intracellular bacterium of macrophages that causes tularemia. Early evasion of host immune responses contributes to the success of F. tularensis as a pathogen. F. tularensis entry into human monocytes and macrophages is mediated by the major phagocytic receptor, complement receptor 3 (CR3, CD11b/CD18. We recently determined that despite a significant increase in macrophage uptake following C3 opsonization of the virulent Type A F. tularensis spp. tularensis Schu S4, this phagocytic pathway results in limited pro-inflammatory cytokine production. Notably, MAP kinase/ERK activation is suppressed immediately during C3-opsonized Schu S4-CR3 phagocytosis. A mathematical model of CR3-TLR2 crosstalk predicted early involvement of Ras GTPase-activating protein (RasGAP in immune suppression by CR3. Here, we link CR3-mediated uptake of opsonized Schu S4 by human monocytes and macrophages with inhibition of early signal 1 inflammasome activation, evidenced by limited caspase-1 cleavage and IL-18 release. This inhibition is due to increased RasGAP activity, leading to a reduction in the Ras-ERK signaling cascade upstream of the early inflammasome activation event. Thus, our data uncover a novel signaling pathway mediated by CR3 following engagement of opsonized virulent F. tularensis to limit inflammasome activation in human phagocytic cells, thereby contributing to evasion of the host innate immune system.

  6. CD14(hi)CD16+ monocytes phagocytose antibody-opsonised Plasmodium falciparum infected erythrocytes more efficiently than other monocyte subsets, and require CD16 and complement to do so.

    Science.gov (United States)

    Zhou, Jingling; Feng, Gaoqian; Beeson, James; Hogarth, P Mark; Rogerson, Stephen J; Yan, Yan; Jaworowski, Anthony

    2015-07-07

    With more than 600,000 deaths from malaria, mainly of children under five years old and caused by infection with Plasmodium falciparum, comes an urgent need for an effective anti-malaria vaccine. Limited details on the mechanisms of protective immunity are a barrier to vaccine development. Antibodies play an important role in immunity to malaria and monocytes are key effectors in antibody-mediated protection by phagocytosing antibody-opsonised infected erythrocytes (IE). Eliciting antibodies that enhance phagocytosis of IE is therefore an important potential component of an effective vaccine, requiring robust assays to determine the ability of elicited antibodies to stimulate this in vivo. The mechanisms by which monocytes ingest IE and the nature of the monocytes which do so are unknown. Purified trophozoite-stage P. falciparum IE were stained with ethidium bromide, opsonised with anti-erythrocyte antibodies and incubated with fresh whole blood. Phagocytosis of IE and TNF production by individual monocyte subsets was measured by flow cytometry. Ingestion of IE was confirmed by imaging flow cytometry. CD14(hi)CD16+ monocytes phagocytosed antibody-opsonised IE and produced TNF more efficiently than CD14(hi)CD16- and CD14(lo)CD16+ monocytes. Blocking experiments showed that Fcγ receptor IIIa (CD16) but not Fcγ receptor IIa (CD32a) or Fcγ receptor I (CD64) was necessary for phagocytosis. CD14(hi)CD16+ monocytes ingested antibody-opsonised IE when peripheral blood mononuclear cells were reconstituted with autologous serum but not heat-inactivated autologous serum. Antibody-opsonised IE were rapidly opsonised with complement component C3 in serum (t1/2 = 2-3 minutes) and phagocytosis of antibody-opsonised IE was inhibited in a dose-dependent manner by an inhibitor of C3 activation, compstatin. Compared to other monocyte subsets, CD14(hi)CD16+ monocytes expressed the highest levels of complement receptor 4 (CD11c) and activated complement receptor 3 (CD11b) subunits

  7. The AgI/II family adhesin AspA is required for respiratory infection by Streptococcus pyogenes.

    Directory of Open Access Journals (Sweden)

    Linda Franklin

    Full Text Available Streptococcus pyogenes (GAS is a human pathogen that causes pharyngitis and invasive diseases such as toxic shock syndrome and sepsis. The upper respiratory tract is the primary reservoir from which GAS can infect new hosts and cause disease. The factors involved in colonisation are incompletely known however. Previous evidence in oral streptococci has shown that the AgI/II family proteins are involved. We hypothesized that the AspA member of this family might be involved in GAS colonization. We describe a novel mouse model of GAS colonization of the nasopharynx and lower respiratory tract to elucidate these interactions. We used two clinical M serotypes expressing AspA, and their aspA gene deletant isogenic mutants in experiments using adherence assays to respiratory epithelium, macrophage phagocytosis and neutrophil killing assays and in vivo models of respiratory tract colonisation and infection. We demonstrated the requirement for AspA in colonization of the respiratory tract. AspA mutants were cleared from the respiratory tract and were deficient in adherence to epithelial cells, and susceptible to phagocytosis. Expression of AspA in the surrogate host Lactococcus lactis protected bacteria from phagocytosis. Our results suggest that AspA has an essential role in respiratory infection, and may function as a novel anti-phagocytic factor.

  8. Synthesis of structured triacylglycerols containing medium-chain and long-chain fatty acids by interesterification with a stereoespecific lipase from Mucor miehei.

    Directory of Open Access Journals (Sweden)

    Nieto, Susana

    1999-06-01

    Full Text Available The preparation of structured triacylglycerols sn-1, sn-3 dilauryl, sn-2 eicosapentaenoyl glycerol and sn-1, sn-3 dilauryl, sn-2 docosahexaenoyl glycerol by enzymatic interesterification under restricted water availability is described. Laurie acid, one of the substrates for interesterification, was obtained by the controlled hydrolysis of coconut oil by a non-specific lipase obtained from Candida cylindracea. The fatty acid was separated from the hydrolysis products by silverresin column chromatography and converted to methyl ester, sn-2 Eicosapentaenoyl glycerol and sn-2 docosahexaenoyl glycerol were prepared by the hydrolysis of fish oil by the sn-1, sn-3 stereospecific immobilized lipase Lipozyme IM-20 obtained from Mucor miehei as described in the accompanying paper. The interesterification was carried out in a water jacketed glass reactor and the triacylglycerol products were separated and recovered through aluminum oxide column chromatography The interesterification procedure described allows to obtain In laboratory scale structured triacylglycerols containing medium-chain fatty acids at the sn-1 and sn-3 positions and long-chain polyunsaturated fatty acid from marine origin at the sn-2 glycerol position.

    Se describe la preparación de triacilgliceroles estructurados sn-1, sn-3 dilauril, sn-2 ecosapentaenoil glicerol y sn-1, sn-3 diiauril, sn-2 docosahexaenoil glicerol por interesterificación enzimática bajo disponibilidad de agua reducida. Acido láurico, uno de los sustratos para la interesterificación, se obtuvo mediante hidrólisis controlada del aceite de coco por una lipasa no-específica obtenida de Candida cylindracea. Los ácidos grasos se separaron de los productos de hidrólisis mediante cromatografía en columna de resina de plata y convertidos en sus esteres metílicos, sn-2 Eicosapentaenoil glicerol y sn-2 docosahexaenoil glicerol se prepararon mediante hidrólisis de aceite de pescado por la sn-1, sn

  9. Accurate Mass GC/LC-Quadrupole Time of Flight Mass Spectrometry Analysis of Fatty Acids and Triacylglycerols of Spicy Fruits from the Apiaceae Family

    Directory of Open Access Journals (Sweden)

    Thao Nguyen

    2015-12-01

    Full Text Available The triacylglycerol (TAG structure and the regio-stereospecific distribution of fatty acids (FA of seed oils from most of the Apiaceae family are not well documented. The TAG structure ultimately determines the final physical properties of the oils and the position of FAs in the TAG molecule affects the digestion; absorption and metabolism; and physical and technological properties of TAGs. Fixed oils from the fruits of dill (Anethum graveolens, caraway (Carum carvi, cumin (Cuminum cyminum, coriander (Coriandrum sativum, anise (Pimpinella anisum, carrot (Daucus carota, celery (Apium graveolens, fennel (Foeniculum vulgare, and Khella (Ammi visnaga, all from the Apiaceae family, were extracted at room temperature in chloroform/methanol (2:1 v/v using percolators. Crude lipids were fractionated by solid phase extraction to separate neutral triacylglycerols (TAGs from other lipids components. Neutral TAGs were subjected to transesterification process to convert them to their corresponding fatty acids methyl esters (FAMES using 1% boron trifluoride (BF3 in methanol. FAMES were analyzed by gas chromatography-quadrupole time of flight (GC-QTOF mass spectrometry. Triglycerides were analyzed using high performance liquid chromatography-quadrupole time of flight (LC-QTOF mass spectrometry. Petroselinic acid was the major fatty acid in all samples ranging from 57% of the total fatty acids in caraway up to 82% in fennel. All samples contained palmitic (16:0, palmitoleic (C16:1n-9, stearic (C18:0, petroselinic (C18:1n-12, linoleic (C18:2n-6, linolinic (18:3n-3, and arachidic (C20:0 acids. TAG were analyzed using LC-QTOF for accurate mass identification and mass spectrometry/mass spectrometry (MS/MS techniques for regiospesific elucidation of the identified TAGs. Five major TAGs were detected in all samples but with different relative concentrations in all of the tested samples. Several other TAGs were detected as minor components and were present in some

  10. High-performance liquid chromatography-atmospheric pressure chemical ionization mass spectrometry and gas chromatography-flame ionization detection chracterization of delta 5-polyenoic fatty acids in triacylglycerols from conifer seed oils

    Czech Academy of Sciences Publication Activity Database

    Lísa, M.; Holčapek, M.; Řezanka, Tomáš; Kabátová, N.

    2007-01-01

    Roč. 1146, - (2007), s. 67-77 ISSN 0021-9673 R&D Projects: GA ČR GA203/06/0219 Grant - others:GA ČR(CZ) GA203/05/2106 Program:GA Institutional research plan: CEZ:AV0Z50200510 Keywords : conifer seed oil * triacylglycerol * triglyceride Subject RIV: EE - Microbiology, Virology Impact factor: 3.641, year: 2007

  11. Relevance of splenic tissue preservation to bacteria phagocytosis; Importancia da preservacao de tecido esplenico para a fagocitose bacteriana

    Energy Technology Data Exchange (ETDEWEB)

    Marques, Ruy Garcia [Universidade do Estado do Rio de Janeiro (UERJ), RJ (Brazil). Faculdade de Ciencias Medicas. Dept. de Cirurgia Geral]. E-mail: rmarques@uerj.br; Petroianu, Andy [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Faculdade de Medicina. Dept. de Cirurgia; Oliveira, Marcia Betania Nunes de [Universidade do Estado do Rio de Janeiro (UERJ), RJ (Brazil). Inst. de Biologia Roberto Alcantara Gomes. Lab.de Radiofarmacia; Bernardo Filho, Mario [Universidade do Estado do Rio de Janeiro (UERJ), RJ (Brazil). Inst. de Biologia Roberto Alcantara Gomes. Dept. de Biofisica e Biometria

    2002-07-01

    Objective: Being the major lymphoid organ of human body, spleen performs critical immunological functions such as bacteria depuration from bloodstream and precocious antibody production against various antigenic particles. Spleen, liver and lung accounts for more than 95% of phagocytic activity in humans. Methods: We used intravenous inoculation of Escherichia coli AB 1157 in Wistar rats in order to evaluate this function. Young and adult rats of both sexes were submitted to total splenectomy and compared to animals not submitted to any surgical procedure. Sixteen weeks later, rats of both groups were challenged with a suspension of E. coli labeled with Tc-{sup 99}m, and, after twenty minutes, they were killed. Liver, lung, spleen and a blood clot sample were removed for radioactivity determination. The statistical study was performed with Student.s t test. Results: There were no significant differences in the radioactivity uptake obtained from young and adult splenectomized rats. However, these rats were associated with higher levels of blood counts than animals of the control group (p<0.0001) due to a larger bacteria remnant in the bloodstream. Conclusion: This finding suggests that some failure in mononuclear phagocytic system occurs in spleen absence adaptation, ratifying the necessity of development of alternative surgical techniques to total splenectomy in patients requiring spleen removal. (author)

  12. FcγRII-binding Centyrins mediate agonism and antibody-dependent cellular phagocytosis when fused to an anti-OX40 antibody.

    Science.gov (United States)

    Zhang, Di; Whitaker, Brian; Derebe, Mehabaw G; Chiu, Mark L

    2018-04-01

    Immunostimulatory antibodies against the tumor necrosis factor receptors (TNFR) are emerging as promising cancer immunotherapies. The agonism activity of such antibodies depends on crosslinking to Fc gamma RIIB receptor (FcγRIIB) to enable the antibody multimerization that drives TNFR activation. Previously, Fc engineering was used to enhance the binding of such antibodies to Fcγ receptors. Here, we report the identification of Centyrins as alternative scaffold proteins with binding affinities to homologous FcγRIIB and FcγRIIA, but not to other types of Fcγ receptors. One Centyrin, S29, was engineered at distinct positions of an anti-OX40 SF2 antibody to generate bispecific and tetravalent molecules named as mAbtyrins. Regardless of the position of S29 on the SF2 antibody, SF2-S29 mAbtyrins could bind FcγRIIB and FcγRIIA specifically while maintaining binding to OX40 receptors. In a NFκB reporter assay, attachment of S29 Centyrin molecules at the C-termini, but not the N-termini, resulted in SF2 antibodies with increased agonism owing to FcγRIIB crosslinking. The mAbtyrins also showed agonism in T-cell activation assays with immobilized FcγRIIB and FcγRIIA, but this activity was confined to mAbtyrins with S29 specifically at the C-termini of antibody heavy chains. Furthermore, regardless of the position of the molecule, S29 Centyrin could equip an otherwise Fc-silent antibody with antibody-dependent cellular phagocytosis activity without affecting the antibody's intrinsic antibody-dependent cell-meditated cytotoxicity and complement-dependent cytotoxicity. In summary, the appropriate adoption FcγRII-binding Centyrins as functional modules represents a novel strategy to engineer therapeutic antibodies with improved functionalities.

  13. Humanised IgG1 antibody variants targeting membrane-bound carcinoembryonic antigen by antibody-dependent cellular cytotoxicity and phagocytosis.

    Science.gov (United States)

    Ashraf, S Q; Umana, P; Mössner, E; Ntouroupi, T; Brünker, P; Schmidt, C; Wilding, J L; Mortensen, N J; Bodmer, W F

    2009-11-17

    The effect of glycoengineering a membrane specific anti-carcinoembryonic antigen (CEA) (this paper uses the original term CEA for the formally designated CEACAM5) antibody (PR1A3) on its ability to enhance killing of colorectal cancer (CRC) cell lines by human immune effector cells was assessed. In vivo efficacy of the antibody was also tested. The antibody was modified using EBNA cells cotransfected with beta-1,4-N-acetylglucosaminyltransferase III and the humanised hPR1A3 antibody genes. The resulting alteration of the Fc segment glycosylation pattern enhances the antibody's binding affinity to the FcgammaRIIIa receptor on human immune effector cells but does not alter the antibody's binding capacity. Antibody-dependent cellular cytotoxicity (ADCC) is inhibited in the presence of anti-FcgammaRIII blocking antibodies. This glycovariant of hPR1A3 enhances ADCC 10-fold relative to the parent unmodified antibody using either unfractionated peripheral blood mononuclear or natural killer (NK) cells and CEA-positive CRC cells as targets. NK cells are far more potent in eliciting ADCC than either freshly isolated monocytes or granulocytes. Flow cytometry and automated fluorescent microscopy have been used to show that both versions of hPR1A3 can induce antibody-dependent cellular phagocytosis (ADCP) by monocyte-derived macrophages. However, the glycovariant antibody did not mediate enhanced ADCP. This may be explained by the relatively low expression of FcgammaRIIIa on cultured macrophages. In vivo studies show the efficacy of glycoengineered humanised IgG1 PR1A3 in significantly improving survival in a CRC metastatic murine model. The greatly enhanced in vitro ADCC activity of the glycoengineered version of hPR1A3 is likely to be clinically beneficial.

  14. Characterisation of Fecal Soap Fatty Acids, Calcium Contents, Bacterial Community and Short-Chain Fatty Acids in Sprague Dawley Rats Fed with Different sn-2 Palmitic Triacylglycerols Diets.

    Science.gov (United States)

    Wan, Jianchun; Hu, Songyou; Ni, Kefeng; Chang, Guifang; Sun, Xiangjun; Yu, Liangli

    2016-01-01

    The structure of dietary triacylglycerols is thought to influence fatty acid and calcium absorption, as well as intestinal microbiota population of the host. In the present study, we investigated the impact of palmitic acid (PA) esterified at the sn-2 position on absorption of fatty acid and calcium and composition of intestinal microorganisms in rats fed high-fat diets containing either low sn-2 PA (12.1%), medium sn-2 PA (40.4%) or high sn-2 PA (56.3%), respectively. Fecal fatty acid profiles in the soaps were measured by gas chromatography (GC), while fecal calcium concentration was detected by ICP-MS. The fecal microbial composition was assessed using a 16S rRNA high-throughput sequencing technology and fecal short-chain fatty acids were detected by ion chromatograph. Dietary supplementation with a high sn-2 PA fat significantly reduced total fecal contents of fatty acids soap and calcium compared with the medium or low sn-2 PA fat groups. Diet supplementation with sn-2 PA fat did not change the entire profile of the gut microbiota community at phylum level and the difference at genera level also were minimal in the three treatment groups. However, high sn-2 PA fat diet could potentially improve total short-chain fatty acids content in the feces, suggesting that high dietary sn-2 PA fat might have a beneficial effect on host intestinal health.

  15. Acquired intestinal lymphangiectasia successfully treated with a low-fat and medium-chain triacylglycerol-enriched diet in a patient with liver transplantation.

    Science.gov (United States)

    Biselli, Maurizio; Andreone, Pietro; Gramenzi, Annagiulia; Cursaro, Carmela; Lorenzini, Stefania; Bonvicini, Fiorenza; Bernardi, Mauro

    2006-05-01

    Intestinal lymphangiectasia is defined as a dilatation of small bowel lymphatic capillaries and a loss of lymph into the bowel lumen. Clinically it is characterized by hypoproteinaemia and oedema. We present here a case of protein-losing enteropathy due to intestinal lymphangiectasia after liver transplantation in a 57-year-old man who was transplanted for hepatitis C virus. Four years after liver transplantation, the patient developed hypoalbuminaemia and ascites associated with recurrence of cirrhosis. The sudden fall in serum albumin led us to look for a cause of reduction other than or in addition to cirrhosis. Duodenal biopsies showed tall villi with dilated lymphatic vessels and widening of the villi caused by oedema, demonstrating intestinal lymphangiectasia. In this case a low-fat diet supplemented with medium-chain triacylglycerols achieved an early clinical improvement with increased serum albumin levels and ascites disappearance. Intestinal lymphangiectasia should be suspected in liver-transplanted patients developing hypoproteinaemia and hypoalbuminaemia after the recurrence of cirrhosis.

  16. Influence of dietary triacylglycerol structure and level of n-3 fatty acids administered during development on brain phospholipids and memory and learning ability of rats

    DEFF Research Database (Denmark)

    Hartvigsen, M.S.; Mu, Huiling; Hougaard, K.S.

    2004-01-01

    of the nervous system. Methods: Pregnant rats were fed experimental diets from the 8th day of pregnancy throughout lactation. After weaning and until 13 weeks of age, the pups were fed the same diet as their dams. The experimental diets contained either a structured oil, a linseed oil, or a fish oil...... and 22:6n-3 adding up to a total of 2 mol% n-3 fatty acids. The effects of the experimental diets were compared to the effect of a chow diet. Results: The amount of 22:6n-3 in brain phosphatidyl ethanolamine (PE) and phosphatidyl serine (PS) of dams and offspring (3 and 13 weeks of age) was not affected......The objective of this study was to examine the effects of triacylglycerol (TAG) structure and level of n-3 fatty acids on fatty acid profile of brain phospholipids (PL) of dams and offspring, and the memory and learning ability of the offspring, when administered during initial development...

  17. Murine Diacylglycerol Acyltransferase-2 (DGAT2) Can Catalyze Triacylglycerol Synthesis and Promote Lipid Droplet Formation Independent of Its Localization to the Endoplasmic Reticulum*

    Science.gov (United States)

    McFie, Pamela J.; Banman, Shanna L.; Kary, Steven; Stone, Scot J.

    2011-01-01

    Triacylglycerol (TG) is the major form of stored energy in eukaryotic organisms and is synthesized by two distinct acyl-CoA:diacylglycerol acyltransferase (DGAT) enzymes, DGAT1 and DGAT2. Both DGAT enzymes reside in the endoplasmic reticulum (ER), but DGAT2 also co-localizes with mitochondria and lipid droplets. In this report, we demonstrate that murine DGAT2 is part of a multimeric complex consisting of several DGAT2 subunits. We also identified the region of DGAT2 responsible for its localization to the ER. A DGAT2 mutant lacking both its transmembrane domains, although still associated with membranes, was absent from the ER and instead localized to mitochondria. Unexpectedly, this mutant was still active and capable of interacting with lipid droplets to promote TG storage. Additional experiments indicated that the ER targeting signal was present in the first transmembrane domain (TMD1) of DGAT2. When fused to a fluorescent reporter, TMD1, but not TMD2, was sufficient to target mCherry to the ER. Finally, the interaction of DGAT2 with lipid droplets was dependent on the C terminus of DGAT2. DGAT2 mutants, in which regions of the C terminus were either truncated or specific regions were deleted, failed to co-localize with lipid droplets when cells were oleate loaded to stimulate TG synthesis. Our findings demonstrate that DGAT2 is capable of catalyzing TG synthesis and promote its storage in cytosolic lipid droplets independent of its localization in the ER. PMID:21680734

  18. Phenolics from Winemaking By-Products Better Decrease VLDL-Cholesterol and Triacylglycerol Levels than Those of Red Wine in Wistar Rats.

    Science.gov (United States)

    de Oliveira, Walkia Polliana; Biasoto, Aline Camarão Telles; Marques, Valquíria Fernanda; Dos Santos, Ieda Maria; Magalhães, Kedma; Correa, Luiz Claudio; Negro-Dellacqua, Melissa; Miranda, Maria Spínola; de Camargo, Adriano Costa; Shahidi, Fereidoon

    2017-10-01

    Winemaking by-products account for more than 30% of the grape production, but this inexpensive feedstock has not yet been fully exploited. Accordingly, we evaluated the potential biological activity of winemaking by-products produced with Syrah grapes in comparison with those of the wine produced using the same grape cultivar. Winemaking by-products showed higher contents of total anthocyanins, flavonols, stilbenes, and flavanols than red wine as evaluated by HPLC-DAD-FD (on a dry weight basis). In contrast, red wine was a better source of phenolic acids. However, the contribution of phenolic acids was minor for both samples. Furthermore, equivalent concentration of winemaking by-products (100 mg/kg/d) showed greater biological activity by than that of red wine by decreasing the levels of VLDL-cholesterol and triacylglycerols in Wistar rats. Therefore, this study supports the use of winemaking by-products as an economical source of bioactive phenolics with potential use in the food and nutraceutical industries. © 2017 Institute of Food Technologists®.

  19. Profiling of Polar Lipids in Marine Oleaginous Diatom Fistulifera solaris JPCC DA0580: Prediction of the Potential Mechanism for Eicosapentaenoic Acid-Incorporation into Triacylglycerol

    Directory of Open Access Journals (Sweden)

    Yue Liang

    2014-05-01

    Full Text Available The marine oleaginous diatom Fistulifera solaris JPCC DA0580 is a candidate for biodiesel production because of its high lipid productivity. However, the substantial eicosapentaenoic acid (EPA content in this strain would affect the biodiesel quality. On the other hand, EPA is also known as the essential health supplement for humans. EPAs are mainly incorporated into glycerolipids in the microalgal cell instead of the presence as free fatty acids. Therefore, the understanding of the EPA biosynthesis including the incorporation of the EPA into glycerolipids especially triacylglycerol (TAG is fundamental for regulating EPA content for different purposes. In this study, in order to identify the biosynthesis pathway for the EPA-containing TAG species, a lipidomic characterization of the EPA-enriched polar lipids was performed by using direct infusion electrospray ionization (ESI-Q-TRAP-MS and MS/MS analyses. The determination of the fatty acid positional distribution showed that the sn-2 position of all the chloroplast lipids and part of phosphatidylcholine (PC species was occupied by C16 fatty acids. This result suggested the critical role of the chloroplast on the lipid synthesis in F. solaris. Furthermore, the exclusive presence of C18 fatty acids in PC highly indicated the biosynthesis of EPA on PC. Finally, the PC-based acyl-editing and head group exchange processes were proposed to be essential for the incorporation of EPA into TAG and chloroplast lipids.

  20. The critical micelle concentration of lecithin in bulk oils and medium chain triacylglycerol is influenced by moisture content and total polar materials.

    Science.gov (United States)

    Kim, JiSu; Kim, Mi-Ja; Lee, JaeHwan

    2018-09-30

    Effects of different moisture contents and oxidised compounds on the critical micelle concentration (CMC) of lecithin were determined in bulk oils and in medium-chain triacylglycerols (MCT). CMC of lecithin in MCT was significantly higher than that in other vegetable oils including olive, soybean, corn, and rapeseed oils (p < 0.05). Presence of moisture significantly affected the CMC of lecithin in MCT (p < 0.05). CMC of lecithin was high when the moisture content was below 900 ppm, whereas at a moisture content of 1000 ppm, CMC of lecithin decreased significantly (p < 0.05), and then started to increase. Addition of total polar materials (TPM), which are oxidation products, at 3 and 5% concentrations, decreased CMC of lecithin significantly (p < 0.05) in MCT, compared to when 0, 1, and 1.5% of TPM was added to MCT. As the degree of oxidation increased in corn oil, CMC of lecithin gradually decreased. Additionally, under different moisture contents, corn oils showed a similar pattern of CMC of lecithin in MCT, whereas oxidised corn oil had a little lower CMC of lecithin than unoxidised corn oil. The results clearly showed that the concentration of lecithin for the formation of micelles is greatly influenced by the presence of oxidation products and the moisture content in bulk oils. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. Comparison of the Incorporation of DHA in Circulatory and Neural Tissue When Provided as Triacylglycerol (TAG), Monoacylglycerol (MAG) or Phospholipids (PL) Provides New Insight into Fatty Acid Bioavailability.

    Science.gov (United States)

    Destaillats, Frédéric; Oliveira, Manuel; Bastic Schmid, Viktoria; Masserey-Elmelegy, Isabelle; Giuffrida, Francesca; Thakkar, Sagar K; Dupuis, Lénaïck; Gosoniu, Maria Laura; Cruz-Hernandez, Cristina

    2018-05-15

    Phospholipids (PL) or partial acylglycerols such as sn -1(3)-monoacylglycerol (MAG) are potent dietary carriers of long-chain polyunsaturated fatty acids (LC-PUFA) and have been reported to provide superior bioavailability when compared to conventional triacylglycerol (TAG). The main objective of the present study was to compare the incorporation of docosahexaenoic acid (DHA) in plasma, erythrocytes, retina and brain tissues in adult rats when provided as PL (PL-DHA) and MAG (MAG-DHA). Conventional dietary DHA oil containing TAG (TAG-DHA) as well as control chow diet were used to evaluate the potency of the two alternative DHA carriers over a 60-day feeding period. Fatty acid profiles were determined in erythrocytes and plasma lipids at time 0, 7, 14, 28, 35 and 49 days of the experimental period and in retina, cortex, hypothalamus, and hippocampus at 60 days. The assessment of the longitudinal evolution of DHA in erythrocyte and plasma lipids suggest that PL-DHA and MAG-DHA are efficient carriers of dietary DHA when compared to conventional DHA oil (TAG-DHA). Under these experimental conditions, both PL-DHA and MAG-DHA led to higher incorporations of DHA erythrocytes lipids compared to TAG-DHA group. After 60 days of supplementation, statistically significant increase in DHA level incorporated in neural tissues analyzed were observed in the DHA groups compared with the control. The mechanism explaining hypothetically the difference observed in circulatory lipids is discussed.

  2. Transport of C-13-labelled linoleic and C-13-labelled caprylic acid in rat plasma after administration of specific structured triacylglycerols

    DEFF Research Database (Denmark)

    Vistisen, Bodil; Høy, Carl-Erik

    2004-01-01

    the transport of dietary C-13-labelled fatty acids in rat plasma to compare the chylomicron fatty acid metabolism after administration of specific structured, long chain and medium chain triacylglycerols. Rats were fed ML*M, M*LM*, L*L*L* or M*M*M* (L=linoleic acid, 18:2n-6, M=caprylic acid, 8:0, * = C-13......-labelled fatty acid) by gavage. A maximum transport of 0.5% of the administered C-13-labelled 18:2n-6 was observed in 1mL rat plasma both after administration of L*L*L* and ML*M, while approximately 0.04% of the administered C-13-labelled 8:0 was detected in 1mL plasma following administration of M......*M*M* or M*LM*. After L*L*L* administration C-13-labelled 20:4n-6 was observed in plasma, probably formed by elongation and desaturation of 18:2n-6 in the enterocyte or liver cells. Furthermore, C-13-labelled 16:0, 48:0, 18: 1n-9 and 20:4n-6 were observed in plasma of rats fed M*M*M* and M*LM* due...

  3. Fungal Screening on Olive Oil for Extracellular Triacylglycerol Lipases: Selection of a Trichoderma harzianum Strain and Genome Wide Search for the Genes

    Science.gov (United States)

    Canseco-Pérez, Miguel Angel; Castillo-Avila, Genny Margarita; Islas-Flores, Ignacio; Apolinar-Hernández, Max M.; Rivera-Muñoz, Gerardo; Gamboa-Angulo, Marcela; Couoh-Uicab, Yeny

    2018-01-01

    A lipolytic screening with fungal strains isolated from lignocellulosic waste collected in banana plantation dumps was carried out. A Trichoderma harzianum strain (B13-1) showed good extracellular lipolytic activity (205 UmL−1). Subsequently, functional screening of the lipolytic activity on Rhodamine B enriched with olive oil as the only carbon source was performed. The successful growth of the strain allows us to suggest that a true lipase is responsible for the lipolytic activity in the B13-1 strain. In order to identify the gene(s) encoding the protein responsible for the lipolytic activity, in silico identification and characterization of triacylglycerol lipases from T. harzianum is reported for the first time. A survey in the genome of this fungus retrieved 50 lipases; however, bioinformatic analyses and putative functional descriptions in different databases allowed us to choose seven lipases as candidates. Suitability of the bioinformatic screening to select the candidates was confirmed by reverse transcription polymerase chain reaction (RT-PCR). The gene codifying 526309 was expressed when the fungus grew in a medium with olive oil as carbon source. This protein shares homology with commercial lipases, making it a candidate for further applications. The success in identifying a lipase gene inducible with olive oil and the suitability of the functional screening and bioinformatic survey carried out herein, support the premise that the strategy can be used in other microorganisms with sequenced genomes to search for true lipases, or other enzymes belonging to large protein families. PMID:29370083

  4. Positional Distribution of Fatty Acids in Triacylglycerols and Phospholipids from Fillets of Atlantic Salmon (Salmo Salar) Fed Vegetable and Fish Oil Blends.

    Science.gov (United States)

    Ruiz-Lopez, Noemi; Stubhaug, Ingunn; Ipharraguerre, Ignacio; Rimbach, Gerald; Menoyo, David

    2015-07-10

    The nutritional and functional characteristics of dietary fat are related to the fatty acid (FA) composition and its positional distribution in the triacylglycerol (TAG) fraction. Atlantic salmon is an important source of healthy long chain omega 3 FA (particularly, eicosapentaenoic (EPA) and docoxahexaenoic (DHA) acids). However, the impact of lipid sources in salmon feeds on the regiospecificity of FA in the fish TAG remains to be explored. The present study determines the effect of feeding salmon with blends of palm, rapeseed, and fish oil, providing two different EPA + DHA concentrations (high: H-ED 10.3% and low: L-ED 4.6%) on the fillet lipid class composition and the positional distribution of FA in TAG and phospholipids. The regiospecific analysis of fillet TAG showed that around 50% of the EPA and around 80% of DHA was located in the sn-2 position. The positional distribution of FA in phosphatidylcholine (PC), showed that around 80% of the EPA and around 90% of DHA were located in the sn-2. Fish fed the vegetable-rich diets showed higher EPA in the sn-2 position in PC (77% vs. 83% in the H-ED and L-ED diets, respectively) but similar DHA concentrations. It is concluded that feeding salmon with different EPA + DHA concentrations does not affect their positional distribution in the fillet TAG.

  5. Positional Distribution of Fatty Acids in Triacylglycerols and Phospholipids from Fillets of Atlantic Salmon (Salmo Salar Fed Vegetable and Fish Oil Blends

    Directory of Open Access Journals (Sweden)

    Noemi Ruiz-Lopez

    2015-07-01

    Full Text Available The nutritional and functional characteristics of dietary fat are related to the fatty acid (FA composition and its positional distribution in the triacylglycerol (TAG fraction. Atlantic salmon is an important source of healthy long chain omega 3 FA (particularly, eicosapentaenoic (EPA and docoxahexaenoic (DHA acids. However, the impact of lipid sources in salmon feeds on the regiospecificity of FA in the fish TAG remains to be explored. The present study determines the effect of feeding salmon with blends of palm, rapeseed, and fish oil, providing two different EPA + DHA concentrations (high: H-ED 10.3% and low: L-ED 4.6% on the fillet lipid class composition and the positional distribution of FA in TAG and phospholipids. The regiospecific analysis of fillet TAG showed that around 50% of the EPA and around 80% of DHA was located in the sn-2 position. The positional distribution of FA in phosphatidylcholine (PC, showed that around 80% of the EPA and around 90% of DHA were located in the sn-2. Fish fed the vegetable-rich diets showed higher EPA in the sn-2 position in PC (77% vs. 83% in the H-ED and L-ED diets, respectively but similar DHA concentrations. It is concluded that feeding salmon with different EPA + DHA concentrations does not affect their positional distribution in the fillet TAG.

  6. Precolumn Derivatization with Bromine to Improve Separation and Detection Sensitivity of Triacylglycerols in Edible Oil by Reversed-Phase High Performance Liquid Chromatography.

    Science.gov (United States)

    Shan, Xiao-Lin; Liu, Xiao-Ting; Gong, Can; Xu, Xu

    2018-01-01

    The complexity of triacylglycerols (TAGs) in edible oils is largely due to the many similar unsaturated TAG compounds, which makes profiling TAGs difficult. In this study, precolumn derivatization with bromine (Br 2 ) was used to improve the separation and detection sensitivity of TAGs in edible oils by RP-HPLC. Oil samples dissolved in n-hexane and TAGs were derived by reaction with a Br2-CCl 4 (1:1, v/v) solution for 3 h at room temperature. The derivate product solution was stable and was best separated and detected by RP-HPLC using a C18 column, with a mobile phase of methanol-n-hexane (91.5:8.5, v/v) at 25°C. A detection wavelength of 230 nm was used. The results showed that the approach enabled the separation and detection of more similar TAGs by RP-HPLC. The method was applied to profile 20 types of edible oil, and the results presented the differences in the TAG profiles of various edible oils, which may be useful in the identification of edible oils.

  7. Application of selected ion monitoring to the analysis of triacylglycerols in olive oil by high temperature-gas chromatography/mass spectrometry.

    Science.gov (United States)

    Ruiz-Samblás, C; González-Casado, A; Cuadros-Rodríguez, L; García, F P Rodríguez

    2010-06-30

    The analysis of the triacylglycerol (TAG) composition of oils is a very challenging task, since the TAGs have very similar physico-chemical properties. In this work, a high temperature-gas chromatographic method coupled to electron ionization-mass spectrometry (HT-GC/EI-MS), in the Selected Ion Monitoring (SIM) mode, method was developed for the analysis of TAGs in the olive oil; this is a method suitable for routine analysis. This method was developed using commercially available standard TAGs. The TAGs studied were separated according to their equivalent carbon number and degree of unsaturation. The peak assignment was carried out by locating the characteristic fragment ions having the same retention time on the SIM profile such as [RCO+74](+) and [RCO+128](+) ions, due to the fatty acyl residues on sn-1, sn-2 and sn-3 positions of the TAG molecule and the [M-OCOR](+) ions corresponding to the acyl ions. The developed method was very useful to eliminate the interferences that appeared in the mass spectrum since electron ionization can prevent satisfactory interpretation of spectra. Copyright 2010 Elsevier B.V. All rights reserved.

  8. Preparation of triacylglycerols rich in omega-3 fatty acids from sardine oil using a Rhizomucor miehei lipase: focus in the EPA/DHA ratio.

    Science.gov (United States)

    Bispo, Paulo; Batista, Irineu; Bernardino, Raul J; Bandarra, Narcisa Maria

    2014-02-01

    The increasing evidence on the differential biochemical effects of eicosapentaenoic acid (EPA)/docosahexaenoic acid (DHA) raises the need of n-3 highly unsaturated fatty acid concentrates with different amounts of these fatty acids. In the present work, physicochemical and enzymatic techniques were combined to obtain acylglycerols, mainly triacylglycerols (TAG), rich in n-3 fatty acids. Sardine oil was obtained by washing sardine (Sardina pilchardus) mince with a NaHCO3 solution, hydrolyzed in a KOH-ethanol solution, and concentrated with urea. The esterification reaction was performed in the stoichiometric proportion of substrates for re-esterification to TAG, with 10 % level of Rhizomucor miehei lipase based on the weight of substrates, without any solvent, during 48 h. This procedure led to approximately 88 % of acylglycerols, where more than 66 % were TAG and the concentration of n-3 fatty acids was higher than 60 %, the EPA and DHA ratio (EPA/DHA) was 4:1. The content of DHA in the unesterifed fraction (free fatty acids) increased from 20 to 54 %, while the EPA level in the same fraction decreased from 33 to 12.5 % (EPA/DHA ratio ≈1:4). Computational methods (density functional theory calculations) have been carried out at the B3LYP/6-31G(d,p) level to explain some of the experimental results.

  9. Docosahexaenoic acid (DHA) at the sn-2 position of triacylglycerols increases DHA incorporation in brown, but not in white adipose tissue, of hamsters.

    Science.gov (United States)

    Lopes, Paula A; Bandarra, Narcisa M; Martins, Susana V; Madeira, Marta S; Ferreira, Júlia; Guil-Guerrero, José L; Prates, José A M

    2018-06-01

    We hypothesised that the incorporation of docosahexaenoic acid (DHA) across adipose tissues will be higher when it is ingested as triacylglycerols (TAG) structured at the sn-2 position. Ten-week old male hamsters were allocated to 4 dietary treatments (n = 10): linseed oil (LSO-control group), fish oil (FO), fish oil ethyl esters (FO-EE) and structured DHA at the sn-2 position of TAG (DHA-SL) during 12 weeks. In opposition to the large variations found for fatty acid composition in retroperitoneal white adipose tissue (WAT), brown adipose tissue (BAT) was less responsive to diets. DHA was not found in subcutaneous and retroperitoneal WAT depots but it was successfully incorporated in BAT reaching the highest percentage in DHA-SL. The PCA on plasma hormones (insulin, leptin, adiponectin) and fatty acids discriminated BAT from WATs pointing towards an individual signature on fatty acid deposition, but did not allow for full discrimination of dietary treatments within each adipose tissue.

  10. Lysophosphatidic Acid Acyltransferase from Coconut Endosperm Mediates the Insertion of Laurate at the sn-2 Position of Triacylglycerols in Lauric Rapeseed Oil and Can Increase Total Laurate Levels

    Science.gov (United States)

    Knutzon, Deborah S.; Hayes, Thomas R.; Wyrick, Annette; Xiong, Hui; Maelor Davies, H.; Voelker, Toni A.

    1999-01-01

    Expression of a California bay laurel (Umbellularia californica) 12:0-acyl-carrier protein thioesterase, bay thioesterase (BTE), in developing seeds of oilseed rape (Brassica napus) led to the production of oils containing up to 50% laurate. In these BTE oils, laurate is found almost exclusively at the sn-1 and sn-3 positions of the triacylglycerols (T.A. Voelker, T.R. Hayes, A.C. Cranmer, H.M. Davies [1996] Plant J 9: 229–241). Coexpression of a coconut (Cocos nucifera) 12:0-coenzyme A-preferring lysophosphatitic acid acyltransferase (D.S. Knutzon, K.D. Lardizabal, J.S. Nelsen, J.L. Bleibaum, H.M. Davies, J.G. Metz [1995] Plant Physiol 109: 999–1006) in BTE oilseed rape seeds facilitates efficient laurate deposition at the sn-2 position, resulting in the acccumulation of trilaurin. The introduction of the coconut protein into BTE oilseed rape lines with laurate above 50 mol % further increases total laurate levels. PMID:10398708

  11. Safety and lipid-altering efficacy of a new omega-3 fatty acid and antioxidant-containing medical food in men and women with elevated triacylglycerols.

    Science.gov (United States)

    Maki, K C; Geohas, J G; Dicklin, M R; Huebner, M; Udani, J K

    2015-08-01

    This randomized, double-blind, placebo-controlled multi-center trial investigated the lipid-altering effects of a medical food (PDL-0101) providing 1.8 g/d eicosapentaenoic acid; 12 mg/d astaxanthin, a marine algae-derived carotenoid; and 100 mg/d tocopherol-free gamma/delta tocotrienols enriched with geranylgeraniol, extracted from annatto, on triacylglycerols (TAG), other lipoprotein lipids, and oxidized low-density lipoprotein (LDL) in 102 subjects with TAG 150-499 mg/dL (1.69-5.63 mmol/L) and LDL cholesterol (LDL-C) ≥70 mg/dL (1.81 mmol/L). Compared to placebo, after eight weeks of treatment, PDL-0101 significantly reduced median TAG (-9.5% vs. 10.6%, p<0.001), while not significantly altering mean LDL-C (-3.0% vs. -8.0% for PDL-0101 and placebo, respectively, p=0.071), mean high-density lipoprotein cholesterol (~3% decrease in both groups, p=0.732), or median oxidized LDL concentrations (5% vs. -5% for PDL-0101 and placebo, respectively, p=0.112). These results demonstrate that PDL-0101 is an effective medical food for the management of elevated TAG. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. Two different pathways of phosphatidylcholine synthesis, the Kennedy Pathway and the Lands Cycle, differentially regulate cellular triacylglycerol storage

    DEFF Research Database (Denmark)

    Moessinger, Christine; Klizaite, Kristina; Steinhagen, Almut

    2014-01-01

    BackgroundLipids are stored within cells in lipid droplets (LDs). They consist of a core of neutral lipids surrounded by a monolayer of phospholipids, predominantly phosphatidylcholine (PC). LDs are very dynamic and can rapidly change in size upon lipid uptake or release. These dynamics require a...

  13. Treatment of serum with supernatants from cultures of Candida albicans reduces its serum-dependent phagocytosis Tratamento de soro com sobrenadante de cultura de Candida albicans reduz a fagocitose soro-dependente

    Directory of Open Access Journals (Sweden)

    Aderbal Antonio dos Santos

    2002-01-01

    Full Text Available Candida albicans is a potent activator of the complement system, and heat labile opsonins produced by activation of C3 (C3b and iC3b enhance phagocytosis of C. albicans mediated by complement receptors. In this study we treated mouse serum with supernatants from cultures of a protease producer strain of C. albicans and evaluated the ability of this serum to enhance phagocytosis of C. albicans. Cell-free supernatants from cultures of C. albicans were concentrated 5 fold and added to mouse serum for 30 min at 37ºC, before using this serum for opsonization of glutaraldehyde-fixed yeast cells. We observed that normal mouse serum increased about 3 fold the phagocytosis of C. albicans by mice peritoneal macrophages, whereas supernatant-treated serum did not increase phagocytosis. This effect of supernatants on serum was prevented by addition of pepstatin (5 µg/ ml; an inhibitor of C. albicans acid proteases to the medium. Serum treated with supernatants from cultures of a protease-deficient mutant of C. albicans also increased about 3 fold phagocytosis of the yeast. These results suggest that a protease produced by C. albicans causes proteolysis of serum opsonins, thereby reducing the phagocytosis of the yeast.Candida albicans é um potente ativador do sistema complemento, e opsoninas lábeis ao calor produzidas por ativação de C3 (C3b e iC3b aumentam a fagocitose de C. albicans mediada por receptores de complemento. Neste estudo, tratamos o soro de camundongo com sobrenadante de culturas de uma cepa de C. albicans produtora de proteases e avaliamos a capacidade deste soro reduzir a fagocitose de C. albicans. Sobrenadantes livres de células obtidos de cultura de C. albicans foram concentrados 5 vezes e adicionados ao soro de camundongo por 30 minutos a 37ºC, antes deste soro ser usado para opsonização de C. albicans na forma de levedura e fixadas em glutaraldeido. Nós observamos que soro normal aumentou 3 vezes a fagocitose de C. albicans por

  14. Stages of in vitro phagocytosis of Plasmodium falciparum-infected erythrocytes by human monocytes Estágios da fagocitose in vitro por monócitos humanos de eritrócitos infectados por Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Maria Imaculada Muniz-Junqueira

    2009-04-01

    Full Text Available Monocytes/macrophages play a critical role in the defense mechanisms against malaria parasites, and are the main cells responsible for the elimination of malaria parasites from the blood circulation. We carried out a microscope-aided evaluation of the stages of in vitro phagocytosis of Plasmodium falciparum-infected erythrocytes, by human monocytes. These cells were obtained from healthy adult individuals by means of centrifugation through a cushion of Percoll density medium and were incubated with erythrocytes infected with Plasmodium falciparum that had previously been incubated with a pool of anti-plasmodial immune serum. We described the stages of phagocytosis, starting from adherence of infected erythrocytes to the phagocyte membrane and ending with their destruction within the phagolisosomes of the monocytes. We observed that the different erythrocytic forms of the parasite were ingested by monocytes, and that the process of phagocytosis may be completed in around 30 minutes. Furthermore, we showed that phagocytosis may occur continuously, such that different phases of the process were observed in the same phagocyte.Monócitos/macrófagos desempenham uma função crítica nos mecanismos de defesa antiplasmódio e constituem as principais células responsáveis pela eliminação das formas eritrocitárias do plasmódio da circulação sangüínea. Realizamos uma avaliação microscópica dos estágios da fagocitose in vitro de eritrócitos infectados por Plasmodium falciparum por monócitos humanos. Essas células foram obtidas de indivíduos adultos sadios por centrifugação em Percoll e incubadas com eritrócitos infectados por Plasmodium falciparum previamente incubados com um pool de soro imune contra plasmódio. Descrevemos os estágios da fagocitose, desde a aderência dos eritrócitos infectados até sua destruição nos fagolisossomas dos monócitos. Observou-se que eritrócitos infectados por todos os diferentes est

  15. Bovine S protein (vitronectin increases phagocytosis of Streptococcus dysgalactiae Aumento na fagocitose de Streptococcus dysgalactiae pela ação da proteína S bovina (vitronectina

    Directory of Open Access Journals (Sweden)

    Laerte Francisco Filippsen

    1999-01-01

    Full Text Available The effects of bovine S protein (vitronectin on phagocytosis of Streptococcus dysgalactiae strains isolated from cattle with mastitis were investigated. Phagocytized streptococci were determined by a fluorometric microassay using glass adherent polymorphonuclear neutrophils (PMN. Preincubation of S. dysgalactiae with bovine S protein significantly increased their phagocytosis by PMN. Bovine S protein had no effect on phagocytic killing of non-S protein binding S. pyogenes cultures. Enzymatic digestion of the bovine S protein binding sites on S. dysgalactiae with pronase resulted in a significative reduction of the effects of S protein on phagocytosis. It could thus be concluded that in addition to its role as a promoter of cellular adhesion and complement inhibitor, bovine S protein may also influence the phagocytosis of S. dysgalactiae during inflammatory processes.Foram investigados os efeitos da proteína S bovina (vitronectina na fagocitose de amostras de Streptococcus dysgalactiae isoladas de bovinos com mastite. A determinação do número de estreptococos fagocitados foi realizada pelo método fluorométrico utilizando neutrófilos polimorfonucleares (NPM aderidos em lâminas de vidro. A pré-incubação do S. dysgalactiae com a proteína S bovina aumentou significativamente a sua fagocitose por NPM. A proteína S bovina não causou efeito na fagocitose de culturas de S. pyogenes, já que não apresentam sítios de ligação para esta proteína. A digestão enzimática com pronase dos sítios de ligação S. dysgalactiae para a proteína S bovina resultou numa significativa redução do efeito da proteína S na fagocitose. Pode-se concluir que além do papel como promotor da adesão celular e inibidor do complemento, a proteína S bovina pode também influir na fagocitose do S. dysgalactiae durante os processos inflamatórios.

  16. Phagocytosis of haemozoin (malarial pigment enhances metalloproteinase-9 activity in human adherent monocytes: Role of IL-1beta and 15-HETE

    Directory of Open Access Journals (Sweden)

    Giribaldi Giuliana

    2008-08-01

    Full Text Available Abstract Background It has been shown previously that human monocytes fed with haemozoin (HZ or trophozoite-parasitized RBCs displayed increased matrix metalloproteinase-9 (MMP-9 enzyme activity and protein/mRNA expression and increased TNF production, and showed higher matrix invasion ability. The present study utilized the same experimental model to analyse the effect of phagocytosis of: HZ, delipidized HZ, beta-haematin (lipid-free synthetic HZ and trophozoites on production of IL-1beta and MMP-9 activity and expression. The second aim was to find out which component of HZ was responsible for the effects. Methods Native HZ freshly isolated from Plasmodium falciparum (Palo Alto strain, Mycoplasma-free, delipidized HZ, beta-haematin (lipid-free synthetic HZ, trophozoites and control meals such as opsonized non-parasitized RBCs and inert latex particles, were fed to human monocytes. The production of IL-1beta by differently fed monocytes, in presence or absence of specific MMP-9 inhibitor or anti-hIL-1beta antibodies, was quantified in supernatants by ELISA. Expression of IL-1beta was analysed by quantitative real-time RT-PCR. MMP-9 activity and protein expression were quantified by gelatin zymography and Western blotting. Results Monocytes fed with HZ or trophozoite-parasitized RBCs generated increased amounts of IL-1beta and enhanced enzyme activity (in cell supernatants and protein/mRNA expression (in cell lysates of monocyte MMP-9. The latter appears to be causally related to enhanced IL-1beta production, as enhancement of both expression and enzyme activity were abrogated by anti-hIL-1beta Abs. Upregulation of IL-1beta and MMP-9 were absent in monocytes fed with beta-haematin or delipidized HZ, indicating a role for HZ-attached or HZ-generated lipid components. 15-HETE (15(S,R-hydroxy-6,8,11,13-eicosatetraenoic acid a potent lipoperoxidation derivative generated by HZ from arachidonic acid via haem-catalysis was identified as one mediator

  17. Non-immune binding of human IgG to M-related proteins confers resistance to phagocytosis of group A streptococci in blood.

    Directory of Open Access Journals (Sweden)

    Harry S Courtney

    Full Text Available The non-immune binding of immunoglobulins by bacteria is thought to contribute to the pathogenesis of infections. M-related proteins (Mrp are group A streptococcal (GAS receptors for immunoglobulins, but it is not known if this binding has any impact on virulence. To further investigate the binding of immunoglobulins to Mrp, we engineered mutants of an M type 4 strain of GAS by inactivating the genes for mrp, emm, enn, sof, and sfbX and tested these mutants in IgG-binding assays. Inactivation of mrp dramatically decreased the binding of human IgG, whereas inactivation of emm, enn, sof, and sfbx had only minor effects, indicating that Mrp is a major IgG-binding protein. Binding of human immunoglobulins to a purified, recombinant form of Mrp indicated that it selectively binds to the Fc domain of human IgG, but not IgA or IgM and that it preferentially bound subclasses IgG₁>IgG₄>IgG₂>IgG₃. Recombinant proteins encompassing different regions of Mrp were engineered and used to map its IgG-binding domain to its A-repeat region and a recombinant protein with 3 A-repeats was a better inhibitor of IgG binding than one with a single A-repeat. A GAS mutant expressing Mrp with an in-frame deletion of DNA encoding the A-repeats had a dramatically reduced ability to bind human IgG and to grow in human blood. Mrp exhibited host specificity in binding IgG; human IgG was the best inhibitor of the binding of IgG followed by pig, horse, monkey, and rabbit IgG. IgG from goat, mouse, rat, cow, donkey, chicken, and guinea pig were poor inhibitors of binding. These findings indicate that Mrp preferentially binds human IgG and that this binding contributes to the ability of GAS to resist phagocytosis and may be a factor in the restriction of GAS infections to the human host.

  18. Trichoderma asperelloides Spores Downregulate dectin1/2 and TLR2 Receptors of Mice Macrophages and Decrease Candida parapsilosis Phagocytosis Independent of the M1/M2 Polarization

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    Andréa G. dos Santos

    2017-09-01

    Full Text Available The intensive use of pesticides to control pests in agriculture has promoted several issues relating to environment. As chemical pesticides remain controversial, biocontrol agents originating from fungi could be an alternative. Among them, we highlight biocontrol agents derived from the fungi genus Trichoderma, which have been documented in limiting the growth of other phytopathogenic fungus in the roots and leaves of several plant species. An important member of this genus is Trichoderma asperelloides, whose biocontrol agents have been used to promote plant growth while also treating soil diseases caused by microorganisms in both greenhouses and outdoor crops. To evaluate the safety of fungal biological agents for human health, tests to detect potentially adverse effects, such as allergenicity, toxicity, infectivity and pathogenicity, are crucial. In addition, identifying possible immunomodulating properties of fungal biocontrol agents merits further investigation. Thus, the aim of this study was to evaluate the effects of T. asperelloides spores in the internalization of Candida parapsilosis yeast by mice phagocytes, in order to elucidate the cellular and molecular mechanism of this interaction, as a model to understand possible in vivo effects of this fungus. For this, mice were exposed to a fungal spore suspension through-intraperitoneal injection, euthanized and cells from the peripheral blood and peritoneal cavity were collected for functional, quantitative and phenotypic analysis, throughout analysis of membrane receptors gene expression, phagocytosis ability and cells immunophenotyping M1 (CCR7 and CD86 and M2 (CCR2 and CD206. Our analyses showed that phagocytes exposed to fungal spores had reduced phagocytic capacity, as well as a decrease in the quantity of neutrophils and monocytes in the peripheral blood and peritoneal cavity. Moreover, macrophages exposed to T. asperelloides spores did not display the phenotypic profile M1/M2, and

  19. Phagocytosis and Epithelial Cell Invasion by Crohn’s Disease-Associated Adherent-Invasive Escherichia coli Are Inhibited by the Anti-inflammatory Drug 6-Mercaptopurine

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    Federica Migliore

    2018-05-01

    Full Text Available Adherent-invasive Escherichia coli (AIEC strains are overrepresented in the dysbiotic microbiota of Crohn’s disease (CD patients, and contribute to the onset of the chronic inflammation typical of the disease. However, the effects of anti-inflammatory drugs used for CD treatment on AIEC virulence have not yet been investigated. In this report, we show that exposure of AIEC LF82 strain to amino-6-mercaptopurine (6-MP riboside, one of the most widely used anti-inflammatory drugs in CD, impairs its ability to adhere to, and consequently to invade, human epithelial cells. Notably, phagocytosis of LF82 treated with 6-MP by human macrophages is also reduced, suggesting that 6-MP affects AIEC cell surface determinants involved both in interaction with epithelial cells and in uptake by macrophages. Since a main target of 6-MP in bacterial cells is the inhibition of the important signal molecule c-di-GMP, we also tested whether perturbations in cAMP, another major signaling pathway in E. coli, might have similar effects on interactions with human cells. To this aim, we grew LF82 in the presence of glucose, which leads to inhibition of cAMP synthesis. Growth in glucose-supplemented medium resulted in a reduction in AIEC adhesion to epithelial cells and uptake by macrophages. Consistent with these results, both 6-MP and glucose can affect expression of cell adhesion-related genes, such as the csg genes, encoding thin aggregative fimbriae (curli. In addition, glucose strongly inhibits expression of the fim operon, encoding type 1 pili, a known AIEC determinant for adhesion to human cells. To further investigate whether 6-MP can indeed inhibit c-di-GMP signaling in AIEC, we performed biofilm and motility assays and determination of extracellular polysaccharides. 6-MP clearly affected biofilm formation and cellulose production, but also, unexpectedly, reduced cell motility, itself an important virulence factor for AIEC. Our results provide strong evidence

  20. Trastuzumab mediates antibody-dependent cell-mediated cytotoxicity and phagocytosis to the same extent in both adjuvant and metastatic HER2/neu breast cancer patients.

    Science.gov (United States)

    Petricevic, Branka; Laengle, Johannes; Singer, Josef; Sachet, Monika; Fazekas, Judit; Steger, Guenther; Bartsch, Rupert; Jensen-Jarolim, Erika; Bergmann, Michael

    2013-12-12

    Monoclonal antibodies (mAb), such as trastuzumab are a valuable addition to breast cancer therapy. Data obtained from neoadjuvant settings revealed that antibody-dependent cell-mediated cytotoxicity (ADCC) is a major mechanism of action for the mAb trastuzumab. Conflicting results still call into question whether disease progression, prolonged treatment or concomitant chemotherapy influences ADCC and related immunological phenomena. We analyzed the activity of ADCC and antibody-dependent cell-mediated phagocytosis (ADCP) of peripheral blood mononuclear cells (PBMCs) from human epidermal growth factor receptor 2 (HER2/neu) positive breast cancer patients receiving trastuzumab therapy either in an adjuvant (n = 13) or metastatic (n = 15) setting as well as from trastuzumab treatment-naive (t-naive) HER2/neu negative patients (n = 15). PBMCs from healthy volunteers (n = 24) were used as controls. ADCC and ADCP activity was correlated with the expression of antibody binding Fc-gamma receptor (FcγR)I (CD64), FcγRII (CD32) and FcγRIII (CD16) on CD14+ (monocytes) and CD56+ (NK) cells, as well as the expression of CD107a+ (LAMP-1) on CD56+ cells and the total amount of CD4+CD25+FOXP3+ (Treg) cells. In metastatic patients, markers were correlated with progression-free survival (PFS). ADCC activity was significantly down regulated in metastatic, adjuvant and t-naive patient cohorts as compared to healthy controls. Reduced ADCC activity was inversely correlated with the expression of CD107a on CD56+ cells in adjuvant patients. ADCC and ADCP activity of the patient cohorts were similar, regardless of treatment duration or additional chemotherapy. PFS in metastatic patients inversely correlated with the number of peripheral Treg cells. The reduction of ADCC in patients as compared to healthy controls calls for adjuvant strategies, such as immune-enhancing agents, to improve the activity of trastuzumab. However, efficacy of trastuzumab-specific ADCC and ADCP appears not to

  1. Long-term intake of a high protein diet increases liver triacylglycerol deposition pathways and hepatic signs of injury in rats

    KAUST Repository

    Diaz Rua, Ruben

    2017-04-19

    Intake of high-protein (HP) diets has increased over the last years, mainly due to their popularity for body weight control. Liver is the main organ handling ingested macronutrients and it is associated with the beginning of different pathologies. We aimed to deepen our knowledge on molecular pathways affected by long-term intake of an HP diet. We performed a transcriptome analysis on liver of rats chronically fed with a casein-rich HP diet and analyzed molecular parameters related to liver injury. Chronic increase in the dietary protein/carbohydrate ratio up-regulated processes related with amino acid uptake/metabolism and lipid synthesis, promoting a molecular environment indicative of hepatic triacylglycerol (TG) deposition. Moreover, changes in expression of genes involved in acid–base maintenance and oxidative stress indicate alterations in the pH balance due to the high acid load of the diet, which has been linked to liver/health damage. Up-regulation of immune-related genes was also observed. In concordance with changes at gene expression level, we observed increased liver TG content and increased serum markers of hepatic injury/inflammation (aspartate transaminase, C-reactive protein and TNF-alpha). Moreover, the HP diet strongly increased hepatic mRNA and protein levels of HSP90, a marker of liver injury. Thus, we show for the first time that long-term consumption of an HP diet, resulting in a high acid load, results in a hepatic transcriptome signature reflecting increased TG deposition and increased signs of health risk (increased inflammation, alterations in the acid–base equilibrium and oxidative stress). Persistence of this altered metabolic status could have unhealthy consequences.

  2. Long-term intake of a high protein diet increases liver triacylglycerol deposition pathways and hepatic signs of injury in rats

    KAUST Repository

    Diaz Rua, Ruben; Keijer, Jaap; Palou, Andreu; van Schothorst, Evert M.; Oliver, Paula

    2017-01-01

    Intake of high-protein (HP) diets has increased over the last years, mainly due to their popularity for body weight control. Liver is the main organ handling ingested macronutrients and it is associated with the beginning of different pathologies. We aimed to deepen our knowledge on molecular pathways affected by long-term intake of an HP diet. We performed a transcriptome analysis on liver of rats chronically fed with a casein-rich HP diet and analyzed molecular parameters related to liver injury. Chronic increase in the dietary protein/carbohydrate ratio up-regulated processes related with amino acid uptake/metabolism and lipid synthesis, promoting a molecular environment indicative of hepatic triacylglycerol (TG) deposition. Moreover, changes in expression of genes involved in acid–base maintenance and oxidative stress indicate alterations in the pH balance due to the high acid load of the diet, which has been linked to liver/health damage. Up-regulation of immune-related genes was also observed. In concordance with changes at gene expression level, we observed increased liver TG content and increased serum markers of hepatic injury/inflammation (aspartate transaminase, C-reactive protein and TNF-alpha). Moreover, the HP diet strongly increased hepatic mRNA and protein levels of HSP90, a marker of liver injury. Thus, we show for the first time that long-term consumption of an HP diet, resulting in a high acid load, results in a hepatic transcriptome signature reflecting increased TG deposition and increased signs of health risk (increased inflammation, alterations in the acid–base equilibrium and oxidative stress). Persistence of this altered metabolic status could have unhealthy consequences.

  3. Analysis of 13C-mixed triacylglycerol in stool by bulk (EA-IRMS) and compound specific (GC/MS) methods.

    Science.gov (United States)

    Slater, C; Ling, S C; Preston, T; Weaver, L T

    2002-06-01

    This paper was presented in poster form at the 17th International Congress of Nutrition, August 27-31, Vienna, Austria (Annals of Nutrition & Metabolism 2001; 45(Suppl.1):349). Some of the data were also presented in poster form at the British Society of Gastroenterology Meeting, March 18-21, Glasgow, UK (Gut 2001; 48(Suppl.1):A91). The 13C-mixed triacylglycerol (MTG) breath test is used to measure intraluminal fat digestion. In normal digestion, 20-40% of the ingested 13C label is recovered in breath CO2. We aimed to identify the proportions of ingested label excreted in stool, as well as breath following ingestion of 13C-MTG by children with impaired exocrine pancreatic function and healthy controls. 13C enrichment of breath samples was measured by continuous flow isotope ratio mass spectrometry (IRMS) and cumulative percent dose recovered (cPDR) in 10 h was calculated. Total 13C of a faecal fat extract from each stool was measured by elemental analyser-IRMS, and 13C enrichment and concentration of the TBDMS derivative of octanoic acid was measured by GC/MS after hydrolysis of the fat extract. Stool 5-day cPDR was calculated. Mean breath cPDR was 35%. Mean cPDR in stool by combustion-IRMS and GC/ MS, respectively, was 0.8% and 1.0%. Therefore, the remaining 64% of the 13C label must remain in the body and variability in breath cPDR is due to postabsorptive rather than predigestive factors.

  4. Comparison of the Distribution of Unsaturated Fatty Acids at the Sn-2 Position of Phospholipids and Triacylglycerols in Marine Fishes and Mammals.

    Science.gov (United States)

    Beppu, Fumiaki; Yasuda, Keiko; Okada, Ayako; Hirosaki, Yoshitsugu; Okazaki, Masako; Gotoh, Naohiro

    2017-11-01

    Highly unsaturated fatty acid (HUFA) binding at the sn-2 position of phospholipids (PL) becomes a resource for prostaglandin, leukotriene, resolvin, and protectin synthesis. Both triacylglycerol (TAG) and PL synthesis pathways in vivo are via phosphatidic acid; therefore, the distribution of fatty acid species at the sn-2 position must theoretically be the same for TAG and PL if rearrangement does not occur. However, it is known that little HUFA is located at the sn-2 position of TAG in marine mammals. Therefore, distribution of fatty acid species at the sn-2 position of TAG and PL was compared between marine fishes and mammals in this study. The composition of fatty acids binding at the sn-2 or sn-1,3 position of PL and TAG was analyzed via hydrolysis with enzymes and GC-FID. The results showed that 20:4n-6, 20:5n-3, 22:5n-3, and 22:6n-3 were primarily located at the sn-1,3 positions of TAG in marine mammals. Comparison of the binding positions of HUFA and 16:0 in PL and TAG suggested the existence of Lands' cycle in marine fishes and mammals. In conclusion, both marine fishes and mammals condensed HUFA as a source of eicosanoid at the sn-2 position of PL. Furthermore, abundance ratios for 22:5n-3 or 22:6n-3 at the sn-2 position (sn-2 ratio) in TAG and PL (calculated by the equation: [abundance ratio at sn-2 position of TAG]/[abundance ratio at sn-2 position of PL]) was less than 0.35 in marine mammals; however, it was greater than 0.80 in marine fishes. These differences suggested that the HUFA consisted of 22 carbon atoms and had different roles in marine fishes and mammals.

  5. From Trioleoyl glycerol to extra virgin olive oil through multicomponent triacylglycerol mixtures: Crystallization and polymorphic transformation examined with differential scanning calorimetry and X-ray diffration techniques.

    Science.gov (United States)

    Bayés-García, L; Calvet, T; Cuevas-Diarte, M A; Ueno, S

    2017-09-01

    The polymorphic crystallization and transformation behavior of extra virgin olive oil (EVOO) was examined by using differential scanning calorimetry (DSC) and X-ray diffraction with both laboratory-scale (XRD) and synchrotron radiation source (SR-XRD). The complex behavior observed was studied by previously analyzing mixtures composed by its main 2 to 6 triacylglycerol (TAG) components. Thus, component TAGs were successively added to simulate EVOO composition, until reaching a 6 TAGs mixture, composed by trioleoyl glycerol (OOO), 1-palmitoyl-2,3-dioleoyl glycerol (POO), 1,2-dioleoyl-3-linoleoyl glycerol (OOL), 1-palmitoyl-2-oleoyl-3-linoleoyl glycerol (POL), 1,2-dipalmitoyl-3-oleoyl glycerol (PPO) and 1-stearoyl-2,3-dioleoyl glycerol (SOO). Molten samples were cooled from 25°C to -80°C at a controlled rate of 2°C/min and subsequently heated at the same rate. The polymorphic behavior observed in multicomponent TAG mixtures was interpreted by considering three main groups of TAGs with different molecular structures: triunsaturated OOO and OOL, saturated-unsaturated-unsaturated POO, POL and SOO, and saturated-saturated-unsaturated PPO. As confirmed by our previous work, TAGs belonging to the same structural group displayed a highly similar polymorphic behavior. EVOO exhibited two different β'-2L polymorphic forms (β' 2 -2L and β' 1 -2L), which transformed into β'-3L when heated. Equivalent polymorphic pathways were detected when the same experimental conditions were applied to the 6 TAG components mixture. Hence, minor components may not exert a strong influence in this case. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Krill protein hydrolysate reduces plasma triacylglycerol level with concurrent increase in plasma bile acid level and hepatic fatty acid catabolism in high-fat fed mice

    Directory of Open Access Journals (Sweden)

    Marie S. Ramsvik

    2013-11-01

    Full Text Available Background: Krill powder, consisting of both lipids and proteins, has been reported to modulate hepatic lipid catabolism in animals. Fish protein hydrolysate diets have also been reported to affect lipid metabolism and to elevate bile acid (BA level in plasma. BA interacts with a number of nuclear receptors and thus affects a variety of signaling pathways, including very low density lipoprotein (VLDL secretion. The aim of the present study was to investigate whether a krill protein hydrolysate (KPH could affect lipid and BA metabolism in mice. Method: C57BL/6 mice were fed a high-fat (21%, w/w diet containing 20% crude protein (w/w as casein (control group or KPH for 6 weeks. Lipids and fatty acid composition were measured from plasma, enzyme activity and gene expression were analyzed from liver samples, and BA was measured from plasma. Results: The effect of dietary treatment with KPH resulted in reduced levels of plasma triacylglycerols (TAG and non-esterified fatty acids (NEFAs. The KPH treated mice had also a marked increased plasma BA concentration. The increased plasma BA level was associated with induction of genes related to membrane canalicular exporter proteins (Abcc2, Abcb4 and to BA exporters to blood (Abcc3 and Abcc4. Of note, we observed a 2-fold increased nuclear farnesoid X receptor (Fxr mRNA levels in the liver of mice fed KPH. We also observed increased activity of the nuclear peroxiosme proliferator-activated receptor alpha (PPARα target gene carnitine plamitoyltransferase 2 (CPT-2. Conclusion: The KPH diet showed to influence lipid and BA metabolism in high-fat fed mice. Moreover, increased mitochondrial fatty acid oxidation and elevation of BA concentration may regulate the plasma level of TAGs and NEFAs.

  7. Long-term intake of a high-protein diet increases liver triacylglycerol deposition pathways and hepatic signs of injury in rats.

    Science.gov (United States)

    Díaz-Rúa, Rubén; Keijer, Jaap; Palou, Andreu; van Schothorst, Evert M; Oliver, Paula

    2017-08-01

    Intake of high-protein (HP) diets has increased over the last years, mainly due to their popularity for body weight control. Liver is the main organ handling ingested macronutrients and it is associated with the beginning of different pathologies. We aimed to deepen our knowledge on molecular pathways affected by long-term intake of an HP diet. We performed a transcriptome analysis on liver of rats chronically fed with a casein-rich HP diet and analyzed molecular parameters related to liver injury. Chronic increase in the dietary protein/carbohydrate ratio up-regulated processes related with amino acid uptake/metabolism and lipid synthesis, promoting a molecular environment indicative of hepatic triacylglycerol (TG) deposition. Moreover, changes in expression of genes involved in acid-base maintenance and oxidative stress indicate alterations in the pH balance due to the high acid load of the diet, which has been linked to liver/health damage. Up-regulation of immune-related genes was also observed. In concordance with changes at gene expression level, we observed increased liver TG content and increased serum markers of hepatic injury/inflammation (aspartate transaminase, C-reactive protein and TNF-alpha). Moreover, the HP diet strongly increased hepatic mRNA and protein levels of HSP90, a marker of liver injury. Thus, we show for the first time that long-term consumption of an HP diet, resulting in a high acid load, results in a hepatic transcriptome signature reflecting increased TG deposition and increased signs of health risk (increased inflammation, alterations in the acid-base equilibrium and oxidative stress). Persistence of this altered metabolic status could have unhealthy consequences. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Effect of a fat spread enriched with medium-chain triacylglycerols and a special fatty acid-micronutrient combination on cardiometabolic risk factors in overweight patients with diabetes

    Directory of Open Access Journals (Sweden)

    Bitterlich Norman

    2011-04-01

    Full Text Available Abstract Background Medium-chain triacylglycerols (MCT, omega-3 polyunsaturated fatty acids (n-3-PUFA and micronutrients may be useful for weight and cardiometabolic risk management. However, studies analyzing the effect of a combination of both in individuals at increased cardiometabolic risk are lacking. Therefore, this randomized, controlled, double-blind study investigated the effect of a fat spread enriched with two different doses of MCT and a special long-chain fatty acid-micronutrient combination on cardiometabolic risk factors in overweight diabetic patients. Methods Fifty-four patients received either a fat spread with 6 g/d MCT (MCT30% or 1.2 g/d (MCT6%. Forty-three completed the study. Analysis was performed according to the median of MCT intake (supplemented and food-derived MCT. Clinical, anthropometric, blood, 24 h-urine parameters and dietary intake were assessed at baseline and after 12 weeks. Results Total MCT intake > 7 g/d (MCT > 7 group significantly reduced waist circumference (WC by 1.81 ± 2.69 cm, whereas ≤ 7 g/d MCT (MCT ≤ 7 group increased WC by 0.32 ± 3.03 cm (p = 0.027, which was supported by a change in waist-to-height ratio (WHtR (p = 0.018. Fasting serum triglycerides (TG increased in both groups over time due to dietary habits. In contrast, diabetic metabolic situation and urinary albumin excretion did not alter. Urinary pH differed significantly between groups after 12 weeks. Conclusion An intake of >7 g/d MCT reduced WC in overweight diabetics, whereas the increase in the intake of fatty acids may have worsened fasting TG. Therefore, the suitability of a fat for nutrient enrichment remains to be challenged, and further studies in low-fat matrices are desirable.

  9. Software requirements

    CERN Document Server

    Wiegers, Karl E

    2003-01-01

    Without formal, verifiable software requirements-and an effective system for managing them-the programs that developers think they've agreed to build often will not be the same products their customers are expecting. In SOFTWARE REQUIREMENTS, Second Edition, requirements engineering authority Karl Wiegers amplifies the best practices presented in his original award-winning text?now a mainstay for anyone participating in the software development process. In this book, you'll discover effective techniques for managing the requirements engineering process all the way through the development cy

  10. n-3 PUFA added to high-fat diets affect differently adiposity and inflammation when carried by phospholipids or triacylglycerols in mice

    Directory of Open Access Journals (Sweden)

    Awada Manar

    2013-02-01

    Full Text Available Abstract Background Dietary intake of n-3 polyunsaturated fatty acids (PUFA is primarily recognized to protect against cardiovascular diseases, cognitive dysfunctions and the onset of obesity and associated metabolic disorders. However, some of their properties such as bioavailability can depend on their chemical carriers. The objective of our study was to test the hypothesis that the nature of n-3 PUFA carrier results in different metabolic effects related to adiposity, oxidative stress and inflammation. Methods 4 groups of C57BL/6 mice were fed for 8 weeks low fat (LF diet or high-fat (HF, 20% diets. Two groups of high-fat diets were supplemented with long-chain n-3 PUFA either incorporated in the form of phospholipids (HF-ω3PL or triacylglycerols (HF-ω3TG. Results Both HF-ω3PL and HF-ω3TG diets reduced the plasma concentrations of (i inflammatory markers such as monocyte chemoattractant protein-1 (MCP-1 and interleukin 6 (IL-6, (ii leptin and (iii 4-hydroxy-2-nonenal (4-HNE, a marker of n-6 PUFA-derived oxidative stress compared with the control HF diet. Moreover, in both HF-ω3PL and HF-ω3TG groups, MCP-1 and IL-6 gene expressions were decreased in epididymal adipose tissue and the mRNA level of gastrointestinal glutathione peroxidase GPx2, an antioxidant enzyme, was decreased in the jejunum compared with the control HF diet. The type of n-3 PUFA carrier affected other outcomes. The phospholipid form of n-3 PUFA increased the level of tocopherols in epididymal adipose tissue compared with HF-ω3TG and resulted in smaller adipocytes than the two others HF groups. Adipocytes in the HF-ω3PL and LF groups were similar in size distribution. Conclusion Supplementation of mice diet with long-chain n-3 PUFA during long-term consumption of high-fat diets had the same lowering effects on inflammation regardless of triacyglycerol or phospholipid carrier, whereas the location of these fatty acids on a PL carrier had a major effect on decreasing

  11. Effects of a meal rich in 1,3-diacylglycerol on postprandial cardiovascular risk factors and the glucose-dependent insulinotropic polypeptide in subjects with high fasting triacylglycerol concentrations.

    Science.gov (United States)

    Shoji, Kentaro; Mizuno, Tomohito; Shiiba, Daisuke; Kawagoe, Tadanobu; Mitsui, Yuuki

    2012-03-14

    It was previously reported that compared to triacylglycerol (TAG) oil, diacylglycerol (DAG) oil improves postprandial lipid response. However, the effects of DAG oil on postprandial hyperglycemia and incretin response have not yet been determined. In this study, the effects of DAG oil on both postprandial hyperlipidemia and hyperglycemia and the response to the glucose-dependent insulinotropic polypeptide (GIP) were studied. This randomized, double-blind, crossover study analyzed data for 41 individuals with high fasting triacylglycerol concentrations. The subjects ingested test meals (30.3 g of protein, 18.6 g of fat, and 50.1 g of carbohydrate) containing 10 g of DAG oil (DAG meal) or TAG oil (TAG meal) after fasting for at least 12 h. Blood samples were collected prior to and 0.5, 2, 3, 4, and 6 h after ingestion of the test meal. Postprandial TAG concentrations were significantly lower after the DAG meal compared with the TAG meal. Postprandial TAG, insulin, and GIP concentrations were significantly lower after the DAG meal compared with the TAG meal in 26 subjects with fasting serum TAG levels between 1.36 and 2.83 mmol/L. DAG-oil-based meals, as a replacement for TAG oil, may provide cardiovascular benefits in high-risk individuals by limiting lipid and insulin excursions.

  12. Phagocytosis and production of reactive oxygen species by peripheral blood phagocytes in patients with different stages of alcohol-induced liver disease: effect of acute exposure to low ethanol concentrations

    DEFF Research Database (Denmark)

    Parlesak, Alexandr; Schäfer, C.; Paulus, S. B.

    2003-01-01

    BACKGROUND: In rodents, the development of alcoholic liver disease (ALD) after chronic alcohol feeding was shown to depend on the activity of enzymes that are necessary for production of reactive oxygen species (ROS) in phagocytes. The aim of this study was to determine the formation of ROS...... by resting and challenged phagocytes of patients with different stages of ALD in the presence of ethanol concentrations commonly found in the blood of alcohol abusers. PATIENTS AND METHODS: The release of ROS and the phagocytosis of bacteria by neutrophils and monocytes obtained from 60 patients, who were...... produced significantly more ROS than those of healthy controls. Basal values of ROS production from neutrophils correlated closely to markers of the severity of ALD. ROS formation was depressed dose-dependently by ethanol in the healthy controls but not in alcohol abusers. CONCLUSIONS: Changes in the ROS...

  13. EFFECT OF DRY COW THERAPY DRUGS ON PHAGOCYTOSIS RATES EFEITO DE MEDICAMENTOS INDICADOS PARA O TRATAMENTO DE MASTITE BOVINA NO PERÍODO SECO SOBRE OS ÍNDICES DE FAGOCITOSE

    Directory of Open Access Journals (Sweden)

    Alice Maria Melville Paiva Della Libera

    2009-07-01

    Full Text Available The dry period is considered to be a phase when cows are very vulnerable to mastitis and phagocytosis mechanisms are specially important at this moment. The objective of the present trial was to evaluate the influence of dry cow therapy drugs available in the market on the phagocytic function of leukocytes obtained from cow milk. In order to do that, paired milk samples negative in bacteriological examination were submitted to in vitro phagocytosis test, after they were exposed to solutions of the following drugs: anhydrous cephalonium (M1, Gentamicin (M2, Benzylpenicillin procaine associated with Nafcillin and Dihydrostreptomycin (M3 and Cloxacillin benzathine (M4. Mean phagocytosis rates of control samples and of cells submitted to treatments M1 and M3 were, respectively, (58.88% ± 12.04, (64.87% ± 15.36 and (65.14% ± 17.96 there were no differences among these samples, as well as in the comparison with the control group. Mean phagocytosis rate obtained in cells submitted to treatment M4 (46.90% ± 22.08 was lower than that of the control group, and than that of cells submitted to treatments M1 and M3. On the other hand, mean phagocytosis rate of cells submitted to treatment M2 (0.68% ± 1.63 was lower than that observed in the control group, and than that of all other treatments. These results show that the use of drugs available for dry cow therapy may no be totally innocuous for the host.

    KEY WORDS: Dry period, mastitis, phagocytosis, treatment.
    O período seco é considerado uma fase da lactação muito vulnerável à mastite, e os mecanismos de fagocitose são especialmente importantes nessa etapa. Para avaliar a influência de medicamentos indicados para o tratamento de mastite no período seco sobre a função fagocítica de leucócitos obtidos do leite de vacas, foram utilizadas amostras pareadas de leite de vacas, negativas ao exame bacteriológico. Submeteram-se essas células à prova de fagocitose in vitro, ap

  14. White tea (Camellia sinensis extract reduces oxidative stress and triacylglycerols in obese mice Extrato de chá branco reduz extresse oxidativo e triacilglicerois em camundongos obesos

    Directory of Open Access Journals (Sweden)

    Lílian Gonçalves Teixeira

    2012-12-01

    Full Text Available White tea is an unfermented tea made from young shoots of Camellia sinensis protected from sunlight to avoid polyphenol degradation. Although its levels of catechins are higher than those of green tea (derived from the same plant, there are no studies addressing the relationship between this tea and obesity associated with oxidative stress.The objective of this study was to evaluate the effect of white tea on obesity and its complications using a diet induced obesity model. Forty male C57BL/6 mice were fed a high-fat diet to induce obesity (Obese group or the same diet supplemented with 0.5% white tea extract (Obese + WTE for 8 weeks. Adipose tissue, serum lipid profile, and oxidative stress were studied. White tea supplementation was not able to reduce food intake, body weight, or visceral adiposity. Similarly, there were no changes in cholesterol rich lipoprotein profile between the groups. A reduction in blood triacylglycerols associated with increased cecal lipids was observed in the group fed the diet supplemented with white tea. White tea supplementation also reduced oxidative stress in liver and adipose tissue. In conclusion, white tea extract supplementation (0.5% does not influence body weight or adiposity in obese mice. Its benefits are restricted to the reduction in oxidative stress associated with obesity and improvement of hypertriacylglycerolemia.O chá branco é um chá não fermentando feito a partir de brotos jovens da Camellia sinensis protegidos da luz solar para evitar a degradação de polifenóis. Embora os níveis de catequinas sejam mais elevados que os do chá verde (mesma planta, nenhum estudo foi realizado sobre estresse oxidativo relacionado obesidade. Nosso objetivo foi avaliar o efeito do chá verde na obesidade e suas complicações, usando um modelo de obesidade induzida por dieta. Quarenta camundongos C57BL/6 machos foram alimentados com dieta hiperlipídica para indução da obesidade ou mesma dieta suplementada

  15. Two bifunctional enzymes from the marine protist Thraustochytrium roseum: biochemical characterization of wax ester synthase/acyl-CoA:diacylglycerol acyltransferase activity catalyzing wax ester and triacylglycerol synthesis.

    Science.gov (United States)

    Zhang, Nannan; Mao, Zejing; Luo, Ling; Wan, Xia; Huang, Fenghong; Gong, Yangmin

    2017-01-01

    Triacylglycerols (TAGs) and wax esters (WEs) are important neutral lipids which serve as energy reservoir in some plants and microorganisms. In recent years, these biologically produced neutral lipids have been regarded as potential alternative energy sources for biofuel production because of the increased interest on developing renewable and environmentally benign alternatives for fossil fuels. In bacteria, the final step in TAG and WE biosynthetic pathway is catalyzed by wax ester synthase/acyl coenzyme A (acyl-CoA):diacylglycerol acyltransferase (WS/DGAT). This bifunctional WS/DGAT enzyme is also a key enzyme in biotechnological production of liquid WE via engineering of plants and microorganisms. To date, knowledge about this class of biologically and biotechnologically important enzymes is mainly from biochemical characterization of WS/DGATs from Arabidopsis, jojoba and some bacteria that can synthesize both TAGs and WEs intracellularly, whereas little is known about WS/DGATs from eukaryotic microorganisms. Here, we report the identification and characterization of two bifunctional WS/DGAT enzymes (designated TrWSD4 and TrWSD5) from the marine protist Thraustochytrium roseum . Both TrWSD4 and TrWSD5 comprise a WS-like acyl-CoA acyltransferase domain and the recombinant proteins purified from Escherichia coli Rosetta (DE3) have substantial WS and lower DGAT activity. They exhibit WS activity towards various-chain-length saturated and polyunsaturated acyl-CoAs and fatty alcohols ranging from C 10 to C 18 . TrWSD4 displays WS activity with the lowest K m value of 0.14 μM and the highest k cat / K m value of 1.46 × 10 5  M -1  s -1 for lauroyl-CoA (C 12:0 ) in the presence of 100 μM hexadecanol, while TrWSD5 exhibits WS activity with the lowest K m value of 0.96 μM and the highest k cat / K m value of 9.83 × 10 4  M -1  s -1 for decanoyl-CoA (C 10:0 ) under the same reaction condition. Both WS/DGAT enzymes have the highest WS activity at 37 and 47

  16. Closure requirements

    International Nuclear Information System (INIS)

    Hutchinson, I.P.G.; Ellison, R.D.

    1992-01-01

    Closure of a waste management unit can be either permanent or temporary. Permanent closure may be due to: economic factors which make it uneconomical to mine the remaining minerals; depletion of mineral resources; physical site constraints that preclude further mining and beneficiation; environmental, regulatory or other requirements that make it uneconomical to continue to develop the resources. Temporary closure can occur for a period of several months to several years, and may be caused by factors such as: periods of high rainfall or snowfall which prevent mining and waste disposal; economic circumstances which temporarily make it uneconomical to mine the target mineral; labor problems requiring a cessation of operations for a period of time; construction activities that are required to upgrade project components such as the process facilities and waste management units; and mine or process plant failures that require extensive repairs. Permanent closure of a mine waste management unit involves the provision of durable surface containment features to protect the waters of the State in the long-term. Temporary closure may involve activities that range from ongoing maintenance of the existing facilities to the installation of several permanent closure features in order to reduce ongoing maintenance. This paper deals with the permanent closure features

  17. Secretive production of long-chain fatty acids, triacylglycerols and n-alkane-2-ones by fermentation processes; Hakkoho ni yoru ekitai nenryo no seisan wa kanoka (shishitsu no bunpi seisan)

    Energy Technology Data Exchange (ETDEWEB)

    Fukui, S. [University of Fukuyama, Hiroshima (Japan). Faculty of Engineering

    1995-10-20

    Secretive production of lipids, which are useful source for engine-driving fuel, by microbial process using carbohydrate biomasses as substrate has been investigated in our laboratory. This review consists of four parts concerning breedings and selection of lipid-secretive microorganisms : (1) breedings of long-chain fatty acid-secretive strains from Candida lipolytica L-1 by a step-wise mutagenesis process, (2) selection of a triacylglycerol (TG)-secretive and accumulative yeast strain Trichosporon sp. SH45Y, (3) breedings of mutants, SH45Y-derivatieves, having potent ability to produce TG secretively from glucose, a typical biomass charbohydrate, and (4) selection of microorganisms which produce liquid-n-alkane in culture medium by utilizing long-chain fatty acids and TG; a strain of Penicillium decumbens can produce liquid alkalis with a yield of approximately 60 % in weight from palm kernel oil, a commercial TG. 10 refs., 7 figs., 7 tabs.

  18. Investigation of Lipid Metabolism by a New Structured Lipid with Medium- and Long-Chain Triacylglycerols from Cinnamomum camphora Seed Oil in Healthy C57BL/6J Mice.

    Science.gov (United States)

    Hu, Jiang-Ning; Shen, Jin-Rong; Xiong, Chao-Yue; Zhu, Xue-Mei; Deng, Ze-Yuan

    2018-02-28

    In the present study, a new structured lipid with medium- and long-chain triacylglycerols (MLCTs) was synthesized from camellia oil (CO) and Cinnamomum camphora seed oil (CCSO) by enzymatic interesterification. Meanwhile, the antiobesity effects of structured lipid were investigated through observing the changes of enzymes related to lipid mobilization in healthy C57BL/6J mice. Results showed that after synthesis, the major triacylgeride (TAG) species of intesterificated product changed to LaCC/CLaC (12.6 ± 0.46%), LaCO/LCL (21.7 ± 0.76%), CCO/LaCL (14.2 ± 0.55%), COO/OCO (10.8 ± 0.43%), and OOO (18.6 ± 0.64%). Through second-stage molecular distillation, the purity of interesterified product (MLCT) achieved 95.6%. Later, male C57BL/6J mice were applied to study whether the new structured lipid with MLCT has the efficacy of preventing the formation of obesity or not. After feeding with different diets for 6 weeks, MLCTs could reduce body weight and fat deposition in adipose tissue, lower plasma triacylglycerols (TG) (0.89 ± 0.16 mmol/L), plasma total cholesterol (TC) (4.03 ± 0.08 mmol/L), and hepatic lipids (382 ± 34.2 mg/mice) by 28.8%, 16.0%, and 30.5%, respectively, when compared to the control 2 group. This was also accompanied by increasing fecal lipids (113%) and the level of enzymes including cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), hormone-sensitive lipase (HSL), and adipose triglyceride lipase (ATGL) related to lipid mobilization in MLCT group. From the results, it can be concluded that MLCT reduced body fat deposition probably by modulating enzymes related to lipid mobilization in C57BL/6J mice.

  19. A conserved tryptophan within the WRDPLVDID domain of yeast Pah1 phosphatidate phosphatase is required for its in vivo function in lipid metabolism.

    Science.gov (United States)

    Park, Yeonhee; Han, Gil-Soo; Carman, George M

    2017-12-01

    PAH1 -encoded phosphatidate phosphatase, which catalyzes the dephosphorylation of phosphatidate to produce diacylglycerol at the endoplasmic reticulum membrane, plays a major role in controlling the utilization of phosphatidate for the synthesis of triacylglycerol or membrane phospholipids. The conserved N-LIP and haloacid dehalogenase-like domains of Pah1 are required for phosphatidate phosphatase activity and the in vivo function of the enzyme. Its non-conserved regions, which are located between the conserved domains and at the C terminus, contain sites for phosphorylation by multiple protein kinases. Truncation analyses of the non-conserved regions showed that they are not essential for the catalytic activity of Pah1 and its physiological functions ( e.g. triacylglycerol synthesis). This analysis also revealed that the C-terminal region contains a previously unrecognized WRDPLVDID domain (residues 637-645) that is conserved in yeast, mice, and humans. The deletion of this domain had no effect on the catalytic activity of Pah1 but caused the loss of its in vivo function. Site-specific mutational analyses of the conserved residues within WRDPLVDID indicated that Trp-637 plays a crucial role in Pah1 function. This work also demonstrated that the catalytic activity of Pah1 is required but is not sufficient for its in vivo functions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Some inferences from in vivo experiments with metal and metal oxide nanoparticles: the pulmonary phagocytosis response, subchronic systemic toxicity and genotoxicity, regulatory proposals, searching for bioprotectors (a self-overview

    Directory of Open Access Journals (Sweden)

    Katsnelson BA

    2015-04-01

    Full Text Available Boris A Katsnelson,1 Larisa I Privalova,1 Marina P Sutunkova,1 Vladimir B Gurvich,1 Nadezhda V Loginova,1 Ilzira A Minigalieva,1 Ekaterina P Kireyeva,1 Vladimir Y Shur,2 Ekaterina V Shishkina,2 Ya B Beikin,3 Oleg H Makeyev,4 Irene E Valamina4 1The Medical Research Center for Prophylaxis and Health Protection in Industrial Workers, Ekaterinburg, Russia; 2The Institute of Natural Sciences, The Ural Federal University, Ekaterinburg, Russia; 3The City Clinical Diagnostics Centre, Ekaterinburg, Russia; 4The Ural State Medical University, Ekaterinburg, Russia Abstract: The purpose of this paper is to overview and summarize previously published results of our experiments on white rats exposed to either a single intratracheal instillation or repeated intraperitoneal injections of silver, gold, iron oxide, copper oxide, nickel oxide, and manganese oxide nanoparticles (NPs in stable water suspensions without any chemical additives. Based on these results and some corroborating data of other researchers we maintain that these NPs are much more noxious on both cellular and systemic levels as compared with their 1 µm or even submicron counterparts. However, within the nanometer range the dependence of systemic toxicity on particle size is intricate and non-unique due to complex and often contra-directional relationships between the intrinsic biological aggressiveness of the specific NPs, on the one hand, and complex mechanisms that control their biokinetics, on the other. Our data testify to the high activity of the pulmonary phagocytosis of NPs deposited in airways. This fact suggests that safe levels of exposure to airborne NPs are possible in principle. However, there are no reliable foundations for establishing different permissible exposure levels for particles of different size within the nanometric range. For workroom air, such permissible exposure levels of metallic NP can be proposed at this stage, even if tentatively, based on a sufficiently

  1. Mechanism of hyperinsulinemia after reticuloendothelial system phagocytosis.

    Science.gov (United States)

    Filkins, J P; Yelich, M R

    1982-02-01

    Endocytic loading of the reticuloendothelial system (RES) results in acute hyperinsulinemia and functional hyperinsulinism. Colloidal carbon blockade of the RES in rats resulted in elevations of both portal vein and systemic serum immunoreactive insulin and increases in the hepatic portal vein insulin glucose ratios. Two mechanisms for the hyperinsulinemia were evaluated: 1) decreased removal of insulin by the postendocytic liver and 2) increased secretion of insulin by the isolated perfused pancreas. Colloidal carbon blockade did not alter removal of 125I-insulin as evaluated in the isolated perfused rat liver. Pancreases from postendocytic donor rats when perfused according to the technique of Grodsky manifested enhanced insulin secretion. Macrophage culture-conditioned media enhanced glucose-mediated insulin secretion both as assayed in vivo and in the isolated perfused rat pancreas. The data suggest that postendocytic activated macrophages secrete a monokine that alters insulin release and thus produces the hyperinsulinemia of RES blockade. The acronym MIRA for macrophage insulin-releasing activity is proposed for the monokine.

  2. Curvature recognition and force generation in phagocytosis

    Directory of Open Access Journals (Sweden)

    Prassler Jana

    2010-12-01

    Full Text Available Abstract Background The uptake of particles by actin-powered invagination of the plasma membrane is common to protozoa and to phagocytes involved in the immune response of higher organisms. The question addressed here is how a phagocyte may use geometric cues to optimize force generation for the uptake of a particle. We survey mechanisms that enable a phagocyte to remodel actin organization in response to particles of complex shape. Results Using particles that consist of two lobes separated by a neck, we found that Dictyostelium cells transmit signals concerning the curvature of a surface to the actin system underlying the plasma membrane. Force applied to a concave region can divide a particle in two, allowing engulfment of the portion first encountered. The phagosome membrane that is bent around the concave region is marked by a protein containing an inverse Bin-Amphiphysin-Rvs (I-BAR domain in combination with an Src homology (SH3 domain, similar to mammalian insulin receptor tyrosine kinase substrate p53. Regulatory proteins enable the phagocyte to switch activities within seconds in response to particle shape. Ras, an inducer of actin polymerization, is activated along the cup surface. Coronin, which limits the lifetime of actin structures, is reversibly recruited to the cup, reflecting a program of actin depolymerization. The various forms of myosin-I are candidate motor proteins for force generation in particle uptake, whereas myosin-II is engaged only in retracting a phagocytic cup after a switch to particle release. Thus, the constriction of a phagocytic cup differs from the contraction of a cleavage furrow in mitosis. Conclusions Phagocytes scan a particle surface for convex and concave regions. By modulating the spatiotemporal pattern of actin organization, they are capable of switching between different modes of interaction with a particle, either arresting at a concave region and applying force in an attempt to sever the particle there, or extending the cup along the particle surface to identify the very end of the object to be ingested. Our data illustrate the flexibility of regulatory mechanisms that are at the phagocyte's disposal in exploring an environment of irregular geometry.

  3. PHAGOCYTOSIS AND REMODELING OF COLLAGEN MATRICES

    OpenAIRE

    Abraham, Leah C.; Dice, J Fred.; Lee, Kyongbum; Kaplan, David L.

    2007-01-01

    The biodegradation of collagen and the deposition of new collagen-based extracellular matrices are of central importance in tissue remodeling and function. Similarly, for collagen-based biomaterials used in tissue engineering, the degradation of collagen scaffolds with accompanying cellular infiltration and generation of new extracellular matrix is critical for integration of in vitro grown tissues in vivo. In earlier studies we observed significant impact of collagen structure on primary lun...

  4. Influence of radiographic contrast media on phagocytosis

    International Nuclear Information System (INIS)

    Rasmussen, F.; Georgsen, J.; Grunnet, N.; Aalborg Sygehus

    1988-01-01

    To evaluate the influence of radiographic contrast media (CM) on human polymorphonuclear leucocytes (PML), the ability of these cells to ingest latex particles after in vitro exposure to five different radiographic contrast media was investigated. All CM inhibited the phagocytic properties of PML. The inhibition was dose dependent. The inhibitory effect was partly due to hyperosmolality but CM specific inhibition was also evident. (orig.)

  5. Enhanced Phagocytosis and Antibody Production by Tinospora ...

    African Journals Online (AJOL)

    SERVER

    2008-01-18

    Jan 18, 2008 ... antibody production through in vitro and in vivo studies. MATERIALS AND METHODS. Collection ..... components with candidicidal activity in human, rabbit and guinea pig leukocytes. Infect. Immun., 11: 1226-1234. Manjrekar ...

  6. Differential effects of low-fat and high-fat diets on fed-state hepatic triacylglycerol secretion, hepatic fatty acid profiles, and DGAT-1 protein expression in obese-prone Sprague–Dawley rats

    Science.gov (United States)

    Heden, Timothy D.; Morris, E. Matthew; Kearney, Monica L.; Liu, Tzu-Wen; Park, Young-min; Kanaley, Jill A.; Thyfault, John P.

    2015-01-01

    The purpose of this study was to compare the effects of short-term low-fat (LF) and high-fat (HF) diets on fed-state hepatic triacylglycerol (TAG) secretion, the content of proteins involved in TAG assembly and secretion, fatty acid oxidation (FAO), and the fatty acid profile of stored TAG. Using selectively bred obese-prone Sprague–Dawley rats, we directly measured fed-state hepatic TAG secretion, using Tyloxapol (a lipoprotein lipase inhibitor) and a standardized oral mixed meal (45% carbohydrate, 40% fat, 15% protein) bolus in animals fed a HF or LF diet for 2 weeks, after which the rats were maintained on their respective diet for 1 week (washout) prior to the liver being excised to measure protein content, FAO, and TAG fatty acid profiles. Hepatic DGAT-1 protein expression was ~27% lower in HF- than in LF-fed animals (p < 0.05); the protein expression of all other molecules was similar in the 2 diets. The fed-state hepatic TAG secretion rate was ~39% lower (p < 0.05) in HF- (4.62 ± 0.18 mmol·h−1) than in LF- (7.60 ± 0.57 mmol·h−1) fed animals. Hepatic TAG content was ~2-fold higher (p < 0.05) in HF- (1.07 ± 0.15 nmol·g−1 tissue) than in LF- (0.50 ± 0.16 nmol·g−1 tissue) fed animals. In addition, the fatty acid profile of liver TAG in HF-fed animals closely resembled the diet, whereas in LF-fed animals, the fatty acid profile consisted of mostly de novo synthesized fatty acids. FAO was not altered by diet. LF and HF diets differentially alter fed-state hepatic TAG secretion, hepatic fatty acid profiles, and DGAT-1 protein expression. PMID:24669989

  7. Differential effects of low-fat and high-fat diets on fed-state hepatic triacylglycerol secretion, hepatic fatty acid profiles, and DGAT-1 protein expression in obese-prone Sprague-Dawley rats.

    Science.gov (United States)

    Heden, Timothy D; Morris, E Matthew; Kearney, Monica L; Liu, Tzu-Wen; Park, Young-Min; Kanaley, Jill A; Thyfault, John P

    2014-04-01

    The purpose of this study was to compare the effects of short-term low-fat (LF) and high-fat (HF) diets on fed-state hepatic triacylglycerol (TAG) secretion, the content of proteins involved in TAG assembly and secretion, fatty acid oxidation (FAO), and the fatty acid profile of stored TAG. Using selectively bred obese-prone Sprague-Dawley rats, we directly measured fed-state hepatic TAG secretion, using Tyloxapol (a lipoprotein lipase inhibitor) and a standardized oral mixed meal (45% carbohydrate, 40% fat, 15% protein) bolus in animals fed a HF or LF diet for 2 weeks, after which the rats were maintained on their respective diet for 1 week (washout) prior to the liver being excised to measure protein content, FAO, and TAG fatty acid profiles. Hepatic DGAT-1 protein expression was ∼27% lower in HF- than in LF-fed animals (p < 0.05); the protein expression of all other molecules was similar in the 2 diets. The fed-state hepatic TAG secretion rate was ∼39% lower (p < 0.05) in HF- (4.62 ± 0.18 mmol·h(-1)) than in LF- (7.60 ± 0.57 mmol·h(-1)) fed animals. Hepatic TAG content was ∼2-fold higher (p < 0.05) in HF- (1.07 ± 0.15 nmol·g(-1) tissue) than in LF- (0.50 ± 0.16 nmol·g(-1) tissue) fed animals. In addition, the fatty acid profile of liver TAG in HF-fed animals closely resembled the diet, whereas in LF-fed animals, the fatty acid profile consisted of mostly de novo synthesized fatty acids. FAO was not altered by diet. LF and HF diets differentially alter fed-state hepatic TAG secretion, hepatic fatty acid profiles, and DGAT-1 protein expression.

  8. Chronic alcohol exposure disturbs lipid homeostasis at the adipose tissue-liver axis in mice: analysis of triacylglycerols using high-resolution mass spectrometry in combination with in vivo metabolite deuterium labeling.

    Directory of Open Access Journals (Sweden)

    Xiaoli Wei

    Full Text Available A method of employing high-resolution mass spectrometry in combination with in vivo metabolite deuterium labeling was developed in this study to investigate the effects of alcohol exposure on lipid homeostasis at the white adipose tissue (WAT-liver axis in a mouse model of alcoholic fatty liver. In order to differentiate the liver lipids synthesized from the fatty acids that were transported back from adipose tissue and the lipids synthesized from other sources of fatty acids, a two-stage mouse feeding experiment was performed to incorporate deuterium into metabolites. Hepatic lipids extracted from mouse liver, epididymal white adipose tissue (eWAT and subcutaneous white adipose tissue (sWAT were analyzed. It was found that 13 and 10 triacylglycerols (TGs incorporated with a certain number of deuterium were significantly increased in alcohol induced fatty liver at two and four weeks of alcohol feeding periods, respectively. The concentration changes of these TGs ranged from 1.7 to 6.3-fold increase. A total of 14 deuterated TGs were significantly decreased in both eWAT and sWAT at the two and four weeks and the fold-change ranged from 0.19 to 0.77. The increase of deuterium incorporated TGs in alcohol-induced fatty liver and their decrease in both eWAT and sWAT indicate that alcohol exposure induces hepatic influx of fatty acids which are released from WATs. The results of time course analysis further indicate a mechanistic link between adipose fat loss and hepatic fat gain in alcoholic fatty liver.

  9. Melatonin-Stimulated Triacylglycerol Breakdown and Energy Turnover under Salinity Stress Contributes to the Maintenance of Plasma Membrane H+-ATPase Activity and K+/Na+ Homeostasis in Sweet Potato.

    Science.gov (United States)

    Yu, Yicheng; Wang, Aimin; Li, Xiang; Kou, Meng; Wang, Wenjun; Chen, Xianyang; Xu, Tao; Zhu, Mingku; Ma, Daifu; Li, Zongyun; Sun, Jian

    2018-01-01

    Melatonin (MT) is a multifunctional molecule in animals and plants and is involved in defense against salinity stress in various plant species. In this study, MT pretreatment was simultaneously applied to the roots and leaves of sweet potato seedlings [ Ipomoea batatas (L.) Lam.], which is an important food and industry crop worldwide, followed by treatment of 150 mM NaCl. The roles of MT in mediating K + /Na + homeostasis and lipid metabolism in salinized sweet potato were investigated. Exogenous MT enhanced the resistance to NaCl and improved K + /Na + homeostasis in sweet potato seedlings as indicated by the low reduced K + content in tissues and low accumulation of Na + content in the shoot. Electrophysiological experiments revealed that exogenous MT significantly suppressed NaCl-induced K + efflux in sweet potato roots and mesophyll tissues. Further experiments showed that MT enhanced the plasma membrane (PM) H + -ATPase activity and intracellular adenosine triphosphate (ATP) level in the roots and leaves of salinized sweet potato. Lipidomic profiling revealed that exogenous MT completely prevented salt-induced triacylglycerol (TAG) accumulation in the leaves. In addition, MT upregulated the expression of genes related to TAG breakdown, fatty acid (FA) β-oxidation, and energy turnover. Chemical inhibition of the β-oxidation pathway led to drastic accumulation of lipid droplets in the vegetative tissues of NaCl-stressed sweet potato and simultaneously disrupted the MT-stimulated energy state, PM H + -ATPase activity, and K + /Na + homeostasis. Results revealed that exogenous MT stimulated TAG breakdown, FA β-oxidation, and energy turnover under salinity conditions, thereby contributing to the maintenance of PM H + -ATPase activity and K + /Na + homeostasis in sweet potato.

  10. In situ crystallization and transformation kinetics of polymorphic forms of saturated-unsaturated-unsaturated triacylglycerols: 1-palmitoyl-2,3-dioleoyl glycerol, 1-stearoyl-2,3-dioleoyl glycerol, and 1-palmitoyl-2-oleoyl-3-linoleoyl glycerol.

    Science.gov (United States)

    Bayés-García, L; Calvet, T; Cuevas-Diarte, M A; Ueno, S

    2016-07-01

    We examined the influence of dynamic thermal treatment (variation of cooling/heating rates) on the polymorphic crystallization and transformation pathways of 1-palmitoyl-2,3-dioleoyl glycerol (POO), 1-stearoyl-2,3-dioleoyl glycerol (SOO), and 1-palmitoyl-2-oleoyl-3-linoleoyl glycerol (POL), which are major saturated-unsaturated-unsaturated (SUU) triacylglycerols (TAGs) of vegetable oils and animal fats (e.g., palm oil, olive oil, and Iberian ham fat). Using mainly a combination of differential scanning calorimetry (DSC) and synchrotron radiation X-ray diffraction (SR-XRD), we analyzed the polymorphic behavior of TAGs when high (15°Cmin -1 ), intermediate (2°Cmin -1 ), and low (0.5°Cmin -1 ) cooling and heating rates were applied. Multiple polymorphic forms were detected in POO, SOO, and POL (sub-α, α, β' 2 , and β' 1 ). Transient disordered phases, defined as kinetic liquid crystal (KLC) phases, were determined in POO and SOO for the first time. The results demonstrated that more stable forms were directly obtained from the melt by decreasing the cooling rates, whereas less stable forms predominated at high cooling rates, as confirmed in our previous work. Regarding heating rate variation, we confirmed that the nature of the polymorphic transformations observed (solid-state, transformation through KLC phase, or melt-mediation) depended largely on the heating rate. These results were discussed considering the activation energies involved in each process and compared with previous studies on TAGs with different saturated-unsaturated structures (1,3-dioleoyl-2-palmitoylglycerol, 1,3-dipalmitoyl-2-oleoyl-glycerol, trioleoyl glycerol, and 1,2-dioleoyl-3-linoleoyl glycerol). Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Melatonin-Stimulated Triacylglycerol Breakdown and Energy Turnover under Salinity Stress Contributes to the Maintenance of Plasma Membrane H+–ATPase Activity and K+/Na+ Homeostasis in Sweet Potato

    Directory of Open Access Journals (Sweden)

    Yicheng Yu

    2018-02-01

    Full Text Available Melatonin (MT is a multifunctional molecule in animals and plants and is involved in defense against salinity stress in various plant species. In this study, MT pretreatment was simultaneously applied to the roots and leaves of sweet potato seedlings [Ipomoea batatas (L. Lam.], which is an important food and industry crop worldwide, followed by treatment of 150 mM NaCl. The roles of MT in mediating K+/Na+ homeostasis and lipid metabolism in salinized sweet potato were investigated. Exogenous MT enhanced the resistance to NaCl and improved K+/Na+ homeostasis in sweet potato seedlings as indicated by the low reduced K+ content in tissues and low accumulation of Na+ content in the shoot. Electrophysiological experiments revealed that exogenous MT significantly suppressed NaCl-induced K+ efflux in sweet potato roots and mesophyll tissues. Further experiments showed that MT enhanced the plasma membrane (PM H+–ATPase activity and intracellular adenosine triphosphate (ATP level in the roots and leaves of salinized sweet potato. Lipidomic profiling revealed that exogenous MT completely prevented salt-induced triacylglycerol (TAG accumulation in the leaves. In addition, MT upregulated the expression of genes related to TAG breakdown, fatty acid (FA β-oxidation, and energy turnover. Chemical inhibition of the β-oxidation pathway led to drastic accumulation of lipid droplets in the vegetative tissues of NaCl-stressed sweet potato and simultaneously disrupted the MT-stimulated energy state, PM H+–ATPase activity, and K+/Na+ homeostasis. Results revealed that exogenous MT stimulated TAG breakdown, FA β-oxidation, and energy turnover under salinity conditions, thereby contributing to the maintenance of PM H+–ATPase activity and K+/Na+ homeostasis in sweet potato.

  12. Assessment of genetic integrity, splenic phagocytosis and cell death potential of (Z-4-((1,5-dimethyl-3-oxo-2-phenyl-2,3dihydro-1H-pyrazol-4-yl amino-4-oxobut-2-enoic acid and its effect when combined with commercial chemotherapeutics

    Directory of Open Access Journals (Sweden)

    Rodrigo Juliano Oliveira

    2018-02-01

    Full Text Available Abstract The increased incidence of cancer and its high treatment costs have encouraged the search for new compounds to be used in adjuvant therapies for this disease. This study discloses the synthesis of (Z-4-((1,5-dimethyl-3-oxo-2-phenyl-2,3dihydro-1H-pyrazol-4-yl amino-4-oxobut-2-enoic acid (IR-01 and evaluates not only the action of this compound on genetic integrity, increase in splenic phagocytosis and induction of cell death but also its effects in combination with the commercial chemotherapeutic agents doxorubicin, cisplatin and cyclophosphamide. IR-01 was designed and synthesized based on two multifunctionalyzed structural fragments: 4-aminoantipyrine, an active dipyrone metabolite, described as an antioxidant and anti-inflammatory agent; and the pharmacophore fragment 1,4-dioxo-2-butenyl, a cytotoxic agent. The results indicated that IR-01 is an effective chemoprotector because it can prevent clastogenic and/or aneugenic damage, has good potential to prevent genomic damage, can increase splenic phagocytosis and lymphocyte frequency and induces cell death. However, its use as an adjuvant in combination with chemotherapy is discouraged since IR-01 interferes in the effectiveness of the tested chemotherapeutic agents. This is a pioneer study as it demonstrates the chemopreventive effects of IR-01, which may be associated with the higher antioxidant activity of the precursor structure of 4-aminoantipyrine over the effects of the 1,4-dioxo-2-butenyl fragment.

  13. Feed tank transfer requirements

    International Nuclear Information System (INIS)

    Freeman-Pollard, J.R.

    1998-01-01

    This document presents a definition of tank turnover; DOE responsibilities; TWRS DST permitting requirements; TWRS Authorization Basis (AB) requirements; TWRS AP Tank Farm operational requirements; unreviewed safety question (USQ) requirements; records and reporting requirements, and documentation which will require revision in support of transferring a DST in AP Tank Farm to a privatization contractor for use during Phase 1B

  14. Effect of pressure on the selectivity of polymeric C18 and C30 stationary phases in reversed-phase liquid chromatography. Increased separation of isomeric fatty acid methyl esters, triacylglycerols, and tocopherols at high pressure.

    Science.gov (United States)

    Okusa, Kensuke; Iwasaki, Yuki; Kuroda, Ikuma; Miwa, Shohei; Ohira, Masayoshi; Nagai, Toshiharu; Mizobe, Hoyo; Gotoh, Naohiro; Ikegami, Tohru; McCalley, David V; Tanaka, Nobuo

    2014-04-25

    A high-density, polymeric C18 stationary phase (Inertsil ODS-P) or a polymeric C30 phase (Inertsil C30) provided improved resolution of the isomeric fatty acids (FAs), FA methyl esters (FAMEs), triacylglycerols (TAGs), and tocopherols with an increase in pressure of 20-70MPa in reversed-phase HPLC. With respect to isomeric C18 FAMEs with one cis-double bond, ODS-P phase was effective for recognizing the position of a double bond among petroselinic (methyl 6Z-octadecenoate), oleic (methyl 9Z-octadecenoate), and cis-vaccenic (methyl 11Z-octadecenoate), especially at high pressure, but the differentiation between oleic and cis-vaccenic was not achieved by C30 phase regardless of the pressure. A monomeric C18 phase (InertSustain C18) was not effective for recognizing the position of the double bond in monounsaturated FAME, while the separation of cis- and trans-isomers was achieved by any of the stationary phases. The ODS-P and C30 phases provided increased separation for TAGs and β- and γ-tocopherols at high pressure. The transfer of FA, FAME, or TAG molecules from the mobile phase to the ODS-P stationary phase was accompanied by large volume reduction (-30∼-90mL/mol) resulting in a large increase in retention (up to 100% for an increase of 50MPa) and improved isomer separation at high pressure. For some isomer pairs, the ODS-P and C30 provided the opposite elution order, and in each case higher pressure improved the separation. The two stationary phases showed selectivity for the isomers having rigid structures, but only the ODS-P was effective for differentiating the position of a double bond in monounsaturated FAMEs. The results indicate that the improved isomer separation was provided by the increased dispersion interactions between the solute and the binding site of the stationary phase at high pressure. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Muc5b is required for airway defence

    Science.gov (United States)

    Roy, Michelle G.; Livraghi-Butrico, Alessandra; Fletcher, Ashley A.; McElwee, Melissa M.; Evans, Scott E.; Boerner, Ryan M.; Alexander, Samantha N.; Bellinghausen, Lindsey K.; Song, Alfred S.; Petrova, Youlia M.; Tuvim, Michael J.; Adachi, Roberto; Romo, Irlanda; Bordt, Andrea S.; Bowden, M. Gabriela; Sisson, Joseph H.; Woodruff, Prescott G.; Thornton, David J.; Rousseau, Karine; de La Garza, Maria M.; Moghaddam, Seyed J.; Karmouty-Quintana, Harry; Blackburn, Michael R.; Drouin, Scott M.; Davis, C. William; Terrell, Kristy A.; Grubb, Barbara R.; O'Neal, Wanda K.; Flores, Sonia C.; Cota-Gomez, Adela; Lozupone, Catherine A.; Donnelly, Jody M.; Watson, Alan M.; Hennessy, Corinne E.; Keith, Rebecca C.; Yang, Ivana V.; Barthel, Lea; Henson, Peter M.; Janssen, William J.; Schwartz, David A.; Boucher, Richard C.; Dickey, Burton F.; Evans, Christopher M.

    2014-01-01

    Respiratory surfaces are exposed to billions of particulates and pathogens daily. A protective mucus barrier traps and eliminates them through mucociliary clearance (MCC). However, excessive mucus contributes to transient respiratory infections and to the pathogenesis of numerous respiratory diseases. MUC5AC and MUC5B are evolutionarily conserved genes that encode structurally related mucin glycoproteins, the principal macromolecules in airway mucus. Genetic variants are linked to diverse lung diseases, but specific roles for MUC5AC and MUC5B in MCC, and the lasting effects of their inhibition, are unknown. Here we show that mouse Muc5b (but not Muc5ac) is required for MCC, for controlling infections in the airways and middle ear, and for maintaining immune homeostasis in mouse lungs, whereas Muc5ac is dispensable. Muc5b deficiency caused materials to accumulate in upper and lower airways. This defect led to chronic infection by multiple bacterial species, including Staphylococcus aureus, and to inflammation that failed to resolve normally. Apoptotic macrophages accumulated, phagocytosis was impaired, and interleukin-23 (IL-23) production was reduced in Muc5b-/- mice. By contrast, in mice that transgenically overexpress Muc5b, macrophage functions improved. Existing dogma defines mucous phenotypes in asthma and chronic obstructive pulmonary disease (COPD) as driven by increased MUC5AC, with MUC5B levels either unaffected or increased in expectorated sputum. However, in many patients, MUC5B production at airway surfaces decreases by as much as 90%. By distinguishing a specific role for Muc5b in MCC, and by determining its impact on bacterial infections and inflammation in mice, our results provide a refined framework for designing targeted therapies to control mucin secretion and restore MCC.

  16. Extracellular requirements for the endocytosis of carcinogenic crystalline nickel sulfide particles by facultative phagocytes

    International Nuclear Information System (INIS)

    Heck, J.D.; Costa, M.

    1982-01-01

    Various culture medium components were examined for their effect upon the phagocytosis of carcinogenic crystalline and non-carcinogenic amorphous NiS by cultured fibroblastic cells using both a visual and radioactive assay for phagocytosis. Crystalline 63 NiS was phagocytosed by cells in a simple salts/glucose maintenance medium to an extent similar to that observed in complex culture medium fortified with 10% fetal bovine serum (FBS), suggesting that serum proteins and other components in complex culture medium exert little influence upon the uptake of these heavy metal particles. Phagocytosis of crystalline NiS was shown to be highly dependent upon Ca 2+ since omission of Ca 2+ from the salts/glucose medium substantially reduced phagocytosis, while readdition of Ca 2+ stimulated uptake in a concentration-dependent manner. The uptake of the NiS particles was inhibited by trifluoperazine, a calmodulin antagonist, implicating intracellular Ca 2+ in this phagocytosis process. Since the opposite surface charge of crystalline and amorphous NiS has been related to their different phagocytic uptake by cells whose primary function is not phagocytosis (facultative phagocytes), these results show that the culture medium components do not modify the surface charge of these particles in a way that significantly influences their uptake. (Auth.)

  17. Feed tank transfer requirements

    International Nuclear Information System (INIS)

    Freeman-Pollard, J.R.

    1998-01-01

    This document presents a definition of tank turnover. Also, DOE and PC responsibilities; TWRS DST permitting requirements; TWRS Authorization Basis (AB) requirements; TWRS AP Tank Farm operational requirements; unreviewed safety question (USQ) requirements are presented for two cases (i.e., tank modifications occurring before tank turnover and tank modification occurring after tank turnover). Finally, records and reporting requirements, and documentation which will require revision in support of transferring a DST in AP Tank Farm to a privatization contractor are presented

  18. Feed tank transfer requirements

    Energy Technology Data Exchange (ETDEWEB)

    Freeman-Pollard, J.R.

    1998-09-16

    This document presents a definition of tank turnover. Also, DOE and PC responsibilities; TWRS DST permitting requirements; TWRS Authorization Basis (AB) requirements; TWRS AP Tank Farm operational requirements; unreviewed safety question (USQ) requirements are presented for two cases (i.e., tank modifications occurring before tank turnover and tank modification occurring after tank turnover). Finally, records and reporting requirements, and documentation which will require revision in support of transferring a DST in AP Tank Farm to a privatization contractor are presented.

  19. Determinação de substitutos da manteiga de cacau em coberturas de chocolate através da análise de triacilgliceróis Determination of cocoa butter substitutes in coating chocolate.by analysis the triacylglycerol composition

    Directory of Open Access Journals (Sweden)

    Valéria Paula Rodrigues MINIM

    1999-05-01

    Full Text Available A legislação brasileira proíbe a adição de substitutos da manteiga de cacau ao chocolate. O presente trabalho teve como objetivo verificar o padrão de qualidade de coberturas comercializadas na região de Campinas. Para isso, foram analisadas cinco marcas de cobertura de chocolate ao leite e quatro de cobertura de chocolate amargo. Para verificar a possível adição de substitutos determinou-se, por cromatografia gasosa a alta temperatura (CGAT, a composição em triacilgliceróis da gordura extraída e os resultados foram analisados pelo método matemático de Padley & Timms. Não foi detectada a presença de substitutos da manteiga de cacau nas amostras de cobertura de chocolate analisadas.Brazilian regulations prohibit the addition of cocoa butter replacements to chocolate, in total or partial substitution. The objective of the present work was to check the quality standards of four of coating bitter Brasilian chocolate bars and five of coating milk chocolate bars, commercialized in Campinas. In order to check the possible addition of substitutes, the triacylglycerol composition was determined, and the results were analysed by Padley & Timms mathematical method. The triacylglycerol composition of each sample was determined by high temperature gas chromatography (HTGC. The presence of cocoa butter replacements was not detected in the brands of coating chocolate.

  20. Production of structured triacylglycerols from microalgae

    Czech Academy of Sciences Publication Activity Database

    Řezanka, Tomáš; Lukavský, Jaromír; Nedbalová, Linda; Sigler, Karel

    2014-01-01

    Roč. 104, AUG 2014 (2014), s. 95-104 ISSN 0031-9422 R&D Projects: GA ČR(CZ) GAP503/11/0215 Institutional support: RVO:61388971 ; RVO:67985939 Keywords : microalgae * enantiomers * chiral LC Subject RIV: EE - Microbiology, Virology Impact factor: 2.547, year: 2014

  1. Triacylglycerol: nourishing molecule in endurance of Mycobacterium ...

    Indian Academy of Sciences (India)

    Pratap C Mali

    2018-01-24

    Jan 24, 2018 ... solution, which serves as energy source by the production of acetyl-coA (Walker et .... dormant phase and become resistant to antibiotics (Gomez and Mckinney 2004). .... animals, plants and microorganisms. Prog. Lipid Res.

  2. Assessing Requirements Quality through Requirements Coverage

    Science.gov (United States)

    Rajan, Ajitha; Heimdahl, Mats; Woodham, Kurt

    2008-01-01

    In model-based development, the development effort is centered around a formal description of the proposed software system the model. This model is derived from some high-level requirements describing the expected behavior of the software. For validation and verification purposes, this model can then be subjected to various types of analysis, for example, completeness and consistency analysis [6], model checking [3], theorem proving [1], and test-case generation [4, 7]. This development paradigm is making rapid inroads in certain industries, e.g., automotive, avionics, space applications, and medical technology. This shift towards model-based development naturally leads to changes in the verification and validation (V&V) process. The model validation problem determining that the model accurately captures the customer's high-level requirements has received little attention and the sufficiency of the validation activities has been largely determined through ad-hoc methods. Since the model serves as the central artifact, its correctness with respect to the users needs is absolutely crucial. In our investigation, we attempt to answer the following two questions with respect to validation (1) Are the requirements sufficiently defined for the system? and (2) How well does the model implement the behaviors specified by the requirements? The second question can be addressed using formal verification. Nevertheless, the size and complexity of many industrial systems make formal verification infeasible even if we have a formal model and formalized requirements. Thus, presently, there is no objective way of answering these two questions. To this end, we propose an approach based on testing that, when given a set of formal requirements, explores the relationship between requirements-based structural test-adequacy coverage and model-based structural test-adequacy coverage. The proposed technique uses requirements coverage metrics defined in [9] on formal high-level software

  3. Postmarket Requirements and Commitments

    Data.gov (United States)

    U.S. Department of Health & Human Services — Provides information to the public on postmarket requirements and commitments. The phrase postmarket requirements and commitments refers to studies and clinical...

  4. Requirements for existing buildings

    DEFF Research Database (Denmark)

    Thomsen, Kirsten Engelund; Wittchen, Kim Bjarne

    This report collects energy performance requirements for existing buildings in European member states by June 2012.......This report collects energy performance requirements for existing buildings in European member states by June 2012....

  5. Replacing reserve requirements

    OpenAIRE

    Edward J. Stevens

    1993-01-01

    An examination of the fading significance of the Federal Reserve System's reserve requirements and the recent flowering of required clearing balances, a rapidly growing feature of Reserve Bank operations.

  6. PIT Coating Requirements Analysis

    International Nuclear Information System (INIS)

    MINTEER, D.J.

    2000-01-01

    This study identifies the applicable requirements for procurement and installation of a coating intended for tank farm valve and pump pit interior surfaces. These requirements are intended to be incorporated into project specification documents and design media. This study also evaluates previously recommended coatings and identifies requirement-compliant coating products

  7. PIT Coating Requirements Analysis

    Energy Technology Data Exchange (ETDEWEB)

    MINTEER, D.J.

    2000-10-20

    This study identifies the applicable requirements for procurement and installation of a coating intended for tank farm valve and pump pit interior surfaces. These requirements are intended to be incorporated into project specification documents and design media. This study also evaluates previously recommended coatings and identifies requirement-compliant coating products.

  8. Future Home Network Requirements

    DEFF Research Database (Denmark)

    Charbonnier, Benoit; Wessing, Henrik; Lannoo, Bart

    This paper presents the requirements for future Home Area Networks (HAN). Firstly, we discuss the applications and services as well as their requirements. Then, usage scenarios are devised to establish a first specification for the HAN. The main requirements are an increased bandwidth (towards 1...

  9. User Requirements for Wireless

    DEFF Research Database (Denmark)

    in the elicitation process. Cases and user requirement elements discussed in the book include: User requirements elicitation processes for children, construction workers, and farmers User requirements for personalized services of a broadcast company Variations in user involvement Practical elements of user...

  10. TRANSPORTATION SYSTEM REQUIREMENTS DOCUMENT

    International Nuclear Information System (INIS)

    2004-01-01

    This document establishes the Transportation system requirements for the U.S. Department of Energy's (DOE's) Civilian Radioactive Waste Management System (CRWMS). These requirements are derived from the Civilian Radioactive Waste Management System Requirements Document (CRD). The Transportation System Requirements Document (TSRD) was developed in accordance with LP-3.1Q-OCRWM, Preparation, Review, and Approval of Office of National Transportation Level-2 Baseline Requirements. As illustrated in Figure 1, the TSRD forms a part of the DOE Office of Civilian Radioactive Waste Management (OCRWM) Technical Baseline

  11. Transportation System Requirements Document

    International Nuclear Information System (INIS)

    1993-09-01

    This Transportation System Requirements Document (Trans-SRD) describes the functions to be performed by and the technical requirements for the Transportation System to transport spent nuclear fuel (SNF) and high-level radioactive waste (HLW) from Purchaser and Producer sites to a Civilian Radioactive Waste Management System (CRWMS) site, and between CRWMS sites. The purpose of this document is to define the system-level requirements for Transportation consistent with the CRWMS Requirement Document (CRD). These requirements include design and operations requirements to the extent they impact on the development of the physical segments of Transportation. The document also presents an overall description of Transportation, its functions, its segments, and the requirements allocated to the segments and the system-level interfaces with Transportation. The interface identification and description are published in the CRWMS Interface Specification

  12. Distribuição estereoespecífica de lipídios estruturados a partir de gorduras de palma, palmiste e triacilgliceróis de cadeia média Stereospecific distribution of structured lipids obtained from palm oil, palm kernel oil, and medium chain triacylglycerols

    Directory of Open Access Journals (Sweden)

    Denise D'Agostini

    2002-09-01

    Full Text Available Por meio de interesterificação química foram sintetizados lipídios estruturados a partir das gorduras de palma, palmiste e triacilgliceróis de cadeia média. O objetivo deste trabalho foi verificar a distribuição estereoespecífica dos ácidos graxos nos lipídios estruturados. Foi possível comprovar a ocorrência da interesterificação através da hidrólise enzimática, que permitiu conhecer a composição dos ácidos graxos em posições específicas dos triacilgliceróis. Foram estudadas 10 amostras, representadas por 3 amostras individuais, 3 misturas binárias e 4 misturas ternárias. As amostras foram submetidas à hidrólise com lipase pancreática suína à temperatura de 40 ºC e posteriormente analisadas por cromatografia gasosa quanto à composição em ácidos graxos na posição sn-2. A partir dos resultados foram calculados os grupos de triacilgliceróis nas amostras individuais e nas misturas antes e após a reação de interesterificação, utilizando as teorias 1,3-random 2-random e 1,2,3-random. Os resultados demonstraram que antes do rearranjo ao acaso houve preferência do ácido oléico pela posição sn-2, enquanto que os ácidos palmítico e esteárico distribuíram-se principalmente pelas posições sn-1 e sn-3. Nos lipídios estruturados, os ácidos graxos saturados aumentaram sua participação na posição central do triacilglicerol, enquanto que os ácidos graxos insaturados apresentaram diminuição nesta mesma posição.Structured lipids were synthesized by chemical interesterification from palm oil, palm kernel oil, and medium chain triacylglycerols. The objective of this study was to verify the fatty acids positional distribution in the structured lipids. It was possible to confirm the interesterification occurrence through enzymatic hydrolysis, which allowed to know the fatty acids composition in specific positions of the triacylglycerols. Ten samples composed by three individual samples, three binary

  13. Environmental Requirements Management

    Energy Technology Data Exchange (ETDEWEB)

    Cusack, Laura J.; Bramson, Jeffrey E.; Archuleta, Jose A.; Frey, Jeffrey A.

    2015-01-08

    CH2M HILL Plateau Remediation Company (CH2M HILL) is the U.S. Department of Energy (DOE) prime contractor responsible for the environmental cleanup of the Hanford Site Central Plateau. As part of this responsibility, the CH2M HILL is faced with the task of complying with thousands of environmental requirements which originate from over 200 federal, state, and local laws and regulations, DOE Orders, waste management and effluent discharge permits, Comprehensive Environmental Response, Compensation, and Liability Act (CERCLA) response and Resource Conservation and Recovery Act (RCRA) corrective action documents, and official regulatory agency correspondence. The challenge is to manage this vast number of requirements to ensure they are appropriately and effectively integrated into CH2M HILL operations. Ensuring compliance with a large number of environmental requirements relies on an organization’s ability to identify, evaluate, communicate, and verify those requirements. To ensure that compliance is maintained, all changes need to be tracked. The CH2M HILL identified that the existing system used to manage environmental requirements was difficult to maintain and that improvements should be made to increase functionality. CH2M HILL established an environmental requirements management procedure and tools to assure that all environmental requirements are effectively and efficiently managed. Having a complete and accurate set of environmental requirements applicable to CH2M HILL operations will promote a more efficient approach to: • Communicating requirements • Planning work • Maintaining work controls • Maintaining compliance

  14. An Amoebal Grazer of Cyanobacteria Requires Cobalamin Produced by Heterotrophic Bacteria.

    Science.gov (United States)

    Ma, Amy T; Beld, Joris; Brahamsha, Bianca

    2017-05-15

    Amoebae are unicellular eukaryotes that consume microbial prey through phagocytosis, playing a role in shaping microbial food webs. Many amoebal species can be cultivated axenically in rich media or monoxenically with a single bacterial prey species. Here, we characterize heterolobosean amoeba LPG3, a recent natural isolate, which is unable to grow on unicellular cyanobacteria, its primary food source, in the absence of a heterotrophic bacterium, a Pseudomonas species coisolate. To investigate the molecular basis of this requirement for heterotrophic bacteria, we performed a screen using the defined nonredundant transposon library of Vibrio cholerae , which implicated genes in corrinoid uptake and biosynthesis. Furthermore, cobalamin synthase deletion mutations in V. cholerae and the Pseudomonas species coisolate do not support the growth of amoeba LPG3 on cyanobacteria. While cyanobacteria are robust producers of a corrinoid variant called pseudocobalamin, this variant does not support the growth of amoeba LPG3. Instead, we show that it requires cobalamin that is produced by the Pseudomonas species coisolate. The diversity of eukaryotes utilizing corrinoids is poorly understood, and this amoebal corrinoid auxotroph serves as a model for examining predator-prey interactions and micronutrient transfer in bacterivores underpinning microbial food webs. IMPORTANCE Cyanobacteria are important primary producers in aquatic environments, where they are grazed upon by a variety of phagotrophic protists and, hence, have an impact on nutrient flux at the base of microbial food webs. Here, we characterize amoebal isolate LPG3, which consumes cyanobacteria as its primary food source but also requires heterotrophic bacteria as a source of corrinoid vitamins. Amoeba LPG3 specifically requires the corrinoid variant produced by heterotrophic bacteria and cannot grow on cyanobacteria alone, as they produce a different corrinoid variant. This same corrinoid specificity is also

  15. Requirements for Ion Sources

    International Nuclear Information System (INIS)

    Scrivens, R

    2013-01-01

    Ion sources produce beams for a large variety of different physical experiments, industrial processes and medical applications. In order to characterize the beam delivered by them, a list of requirements is necessary. In this chapter the list of principal requirements is specified and definitions for them are given. (author)

  16. Entrepreneurial learning requires action

    DEFF Research Database (Denmark)

    Brink, Tove; Madsen, Svend Ole

    2014-01-01

    that is enhanced by essential large-scale industry players and other SME managers are required to create action and value in learning. An open-mindedness to new learning approaches by SME managers and an open-mindedness to multi- and cross-disciplinary collaboration with SME managers by facilitators is required....

  17. Writing testable software requirements

    Energy Technology Data Exchange (ETDEWEB)

    Knirk, D. [Sandia National Labs., Albuquerque, NM (United States)

    1997-11-01

    This tutorial identifies common problems in analyzing requirements in the problem and constructing a written specification of what the software is to do. It deals with two main problem areas: identifying and describing problem requirements, and analyzing and describing behavior specifications.

  18. Engineering Requirements for crowds

    Directory of Open Access Journals (Sweden)

    Rogeiro Silva

    2015-12-01

    Full Text Available In the software project the interested parts are highly distributed and form numerous and heterogeneous groups, online or face, constituting what could be called crowds. The development of social applications and cloud computing and mobile has generated a marked increase in environments based requirements in crowds. Technical Requirements Engineering (RE traditional face these scalability issues, and require the co-presence of interested and engineers in joint meetings that can not be made in common physical environments. While different approaches have been introduced to partially automate RE in these contexts, still is required a multi-method approach to semi-automate all activities related to work with crowds. In this paper is propose an approach that integrates existing elicitation techniques and requirements analysis and is complemented by introducing new concepts. The information is collected through direct interaction and social collaboration, and through data mining techniques.

  19. Triacilglicerol intramuscular: um importante substrato energético para o exercício de endurance Triacilglicerol intramuscular: un importante substrato energético para el ejercicio de endurance Intramuscular triacylglycerol: an important energetic substrate for endurance exercise

    Directory of Open Access Journals (Sweden)

    Mônica Aparecida Belmonte

    2005-04-01

    de endurance. El objetivo del trabajo presente fue lograr una revisión de la extensión de la literatura en la importancia de los TAG intramusculares como substrato de energía. La revisión de la literatura sugiere que la contribución endógena de las acciones de TAG durante el desarrollo del ejercicio de endurance es bastante importante. Además, puede concluirse que una adaptación inducida por el entrenamiento de endurance es el aumento de los depósitos intramusculares de los TAG. Después del entrenar endurance, se observa también un aumento en la capacidad de uso de estos depósitos. A pesar de parecer importante, la contribución del intramuscular de la TAG es todavía razón de controversia en la literatura. Esa diferencia de resultados se relaciona a las metodologías empleadas para estimar su oxidación por el ejercicio. Para culminar este asunto de manera apropiado, más investigaciones, con nuevos métodos (por ejemplo, el uso de isótopos, resonancia magnética nuclear y microscopia electrónica, necesitarán ser manejados.Free fatty acids represent an important source of energy during endurance exercise. Plasmatic fatty acids are delivered to skeletal muscle as free fatty acids bounded to albumin or associated to triacylglycerol (TAG found in lipoproteins. However, besides these plasmatic sources, hydrolysis of intramuscular TAG also contributes to increase fatty acids availability during endurance exercise. The objective of the present study was to access the role of intramuscular TAG as an energetic substrate to endurance exercise. The present data suggests that the contribution of endogenous intramuscular TAG supplies is quite relevant. Furthermore, it is possible to conclude that endurance training induces an increase of these intramuscular supplies. After endurance training, it is also observed an increase in the utilization of intramuscular TAG. Although the intramuscular TAG seems to be an important energetic substrate, there is a controversy

  20. Requirements in engineering projects

    CERN Document Server

    Fernandes, João M

    2016-01-01

    This book focuses on various topics related to engineering and management of requirements, in particular elicitation, negotiation, prioritisation, and documentation (whether with natural languages or with graphical models). The book provides methods and techniques that help to characterise, in a systematic manner, the requirements of the intended engineering system.  It was written with the goal of being adopted as the main text for courses on requirements engineering, or as a strong reference to the topics of requirements in courses with a broader scope. It can also be used in vocational courses, for professionals interested in the software and information systems domain.   Readers who have finished this book will be able to: - establish and plan a requirements engineering process within the development of complex engineering systems; - define and identify the types of relevant requirements in engineering projects; - choose and apply the most appropriate techniques to elicit the requirements of a giv...

  1. Creativity in Requirement Identification

    DEFF Research Database (Denmark)

    Sørensen, Lene Tolstrup; Olesen, Henning

    Traditional requirements engineering focuses mainly on analysis and elicitation. However, current trends in new system, device and software are towards involving all stakeholders in the early stages of the engineering process to define the user requirements. Creativity is here seen as a major...... keystone in this process in order to open up stakeholder's mind to new technologies, which do not yet exist. This paper dis-cusses the application of creativity in the requirements process and illustrate through cases from the MAGNET and MAGNET Beyond projects....

  2. Polarization of the epithelial layer and apical localization of integrins are required for engulfment of apoptotic cells in the Drosophila ovary

    Directory of Open Access Journals (Sweden)

    Tracy L. Meehan

    2015-12-01

    Full Text Available Inefficient clearance of dead cells or debris by epithelial cells can lead to or exacerbate debilitating conditions such as retinitis pigmentosa, macular degeneration, chronic obstructive pulmonary disease and asthma. Despite the importance of engulfment by epithelial cells, little is known about the molecular changes that are required within these cells. The misregulation of integrins has previously been associated with disease states, suggesting that a better understanding of the regulation of receptor trafficking could be key to treating diseases caused by defects in phagocytosis. Here, we demonstrate that the integrin heterodimer αPS3/βPS becomes apically enriched and is required for engulfment by the epithelial follicle cells of the Drosophila ovary. We found that integrin heterodimer localization and function is largely directed by the α-subunit. Moreover, proper cell polarity promotes asymmetric integrin enrichment, suggesting that αPS3/βPS trafficking occurs in a polarized fashion. We show that several genes previously known for their roles in trafficking and cell migration are also required for engulfment. Moreover, as in mammals, the same α-integrin subunit is required by professional and non-professional phagocytes and migrating cells in Drosophila. Our findings suggest that migrating and engulfing cells use common machinery, and demonstrate a crucial role for integrin function and polarized trafficking of integrin subunits during engulfment. This study also establishes the epithelial follicle cells of the Drosophila ovary as a powerful model for understanding the molecular changes required for engulfment by a polarized epithelium.

  3. FAA Financial Requirements

    Science.gov (United States)

    1997-06-04

    In June 1995, the FAA developed a "total requirements" estimate for the period : FY 97-FY 02 to help explain the difficulty of supporting a dynamic, growing : aviation industry under a federal budget picture which projected flat or reduced : funding ...

  4. TWRSview system requirements specification

    International Nuclear Information System (INIS)

    Caldwell, J.A.; Lee, A.K.

    1995-12-01

    This document provides the system requirements specification for the TWRSview software system. The TWRSview software system is being developed to integrate electronic data supporting the development of the TWRS technical baseline

  5. Estimating ISABELLE shielding requirements

    International Nuclear Information System (INIS)

    Stevens, A.J.; Thorndike, A.M.

    1976-01-01

    Estimates were made of the shielding thicknesses required at various points around the ISABELLE ring. Both hadron and muon requirements are considered. Radiation levels at the outside of the shield and at the BNL site boundary are kept at or below 1000 mrem per year and 5 mrem/year respectively. Muon requirements are based on the Wang formula for pion spectra, and the hadron requirements on the hadron cascade program CYLKAZ of Ranft. A muon shield thickness of 77 meters of sand is indicated outside the ring in one area, and hadron shields equivalent to from 2.7 to 5.6 meters in thickness of sand above the ring. The suggested safety allowance would increase these values to 86 meters and 4.0 to 7.2 meters respectively. There are many uncertainties in such estimates, but these last figures are considered to be rather conservative

  6. Utility requirements for fusion

    International Nuclear Information System (INIS)

    Vondrasek, R.J.

    1982-02-01

    This report describes work done and results obtained during performance of Task 1 of a study of Utility Requirements and Criteria for Fusion Options. The work consisted of developing a list of utility requirements for fusion optics containing definition of the requirements and showing their relative importance to the utility industry. The project team members developed a preliminary list which was refined by discussions and literature searches. The refined list was recast as a questionnaire which was sent to a substantial portion of the utility industry in this country. Forty-three questionnaire recipients responded including thirty-two utilities. A workshop was held to develop a revised requirements list using the survey responses as a major input. The list prepared by the workshop was further refined by a panel consisting of vice presidents of the three project team firms. The results of the study indicate that in addition to considering the cost of energy for a power plant, utilities consider twenty-three other requirements. Four of the requirements were judged to be vital to plant acceptability: Plant Capital Cost, Financial Liability, Plant Safety and Licensability

  7. Triacylglycerol Profile of Summer and Winter Bovine Milk Fat and the Feasibility of Triacylglycerol Fragmentation

    NARCIS (Netherlands)

    Tzompa-Sosa, Daylan A.; Meurs, Pim P.; Valenberg, van Hein J.F.

    2018-01-01

    Bovine milk fat (MF) is considered to be one of the most complex fats, as it can consist of 400 FA which are non-randomly esterified into the TAG molecule. This study aims to determine specific differences on MF TAG profile between summer and winter cow's milk, and to study the feasibility of TAG

  8. Range Flight Safety Requirements

    Science.gov (United States)

    Loftin, Charles E.; Hudson, Sandra M.

    2018-01-01

    The purpose of this NASA Technical Standard is to provide the technical requirements for the NPR 8715.5, Range Flight Safety Program, in regards to protection of the public, the NASA workforce, and property as it pertains to risk analysis, Flight Safety Systems (FSS), and range flight operations. This standard is approved for use by NASA Headquarters and NASA Centers, including Component Facilities and Technical and Service Support Centers, and may be cited in contract, program, and other Agency documents as a technical requirement. This standard may also apply to the Jet Propulsion Laboratory or to other contractors, grant recipients, or parties to agreements to the extent specified or referenced in their contracts, grants, or agreements, when these organizations conduct or participate in missions that involve range flight operations as defined by NPR 8715.5.1.2.2 In this standard, all mandatory actions (i.e., requirements) are denoted by statements containing the term “shall.”1.3 TailoringTailoring of this standard for application to a specific program or project shall be formally documented as part of program or project requirements and approved by the responsible Technical Authority in accordance with NPR 8715.3, NASA General Safety Program Requirements.

  9. NP Science Network Requirements

    Energy Technology Data Exchange (ETDEWEB)

    Dart, Eli [Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States); Rotman, Lauren [Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States); Tierney, Brian [Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)

    2011-08-26

    The Energy Sciences Network (ESnet) is the primary provider of network connectivity for the U.S. Department of Energy (DOE) Office of Science (SC), the single largest supporter of basic research in the physical sciences in the United States. To support SC programs, ESnet regularly updates and refreshes its understanding of the networking requirements of the instruments, facilities, scientists, and science programs it serves. This focus has helped ESnet to be a highly successful enabler of scientific discovery for over 20 years. In August 2011, ESnet and the Office of Nuclear Physics (NP), of the DOE SC, organized a workshop to characterize the networking requirements of the programs funded by NP. The requirements identified at the workshop are summarized in the Findings section, and are described in more detail in the body of the report.

  10. Quantifying requirements volatility effects

    NARCIS (Netherlands)

    Kulk, G.P.; Verhoef, C.

    2008-01-01

    In an organization operating in the bancassurance sector we identified a low-risk IT subportfolio of 84 IT projects comprising together 16,500 function points, each project varying in size and duration, for which we were able to quantify its requirements volatility. This representative portfolio

  11. Requirements for Xenon International

    Energy Technology Data Exchange (ETDEWEB)

    Hayes, James C.; Ely, James H.

    2013-09-26

    This document defines the requirements for the new Xenon International radioxenon system. The output of this project will be a Pacific Northwest National Laboratory (PNNL) developed prototype and a manufacturer-developed production prototype. The two prototypes are intended to be as close to matching as possible; this will be facilitated by overlapping development cycles and open communication between PNNL and the manufacturer.

  12. Requirements for Xenon International

    Energy Technology Data Exchange (ETDEWEB)

    Hayes, James C. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Ely, James H. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Haas, Derek A. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Harper, Warren W. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Heimbigner, Tom R. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Hubbard, Charles W. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Humble, Paul H. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Madison, Jill C. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Morris, Scott J. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Panisko, Mark E. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Ripplinger, Mike D. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Stewart, Timothy L. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2015-12-30

    This document defines the requirements for the new Xenon International radioxenon system. The output of this project will be a Pacific Northwest National Laboratory (PNNL) developed prototype and a manufacturer-developed production prototype. The two prototypes are intended to be as close to matching as possible; this will be facilitated by overlapping development cycles and open communication between PNNL and the manufacturer.

  13. Requirements for enrichment tools

    NARCIS (Netherlands)

    Boer, A.; Winkels, R.; Trompper, M.

    2016-01-01

    This report gives a high level overview of requirements for Enrichment tools in the Openlaws.eu project. Openlaws.eu aims to initiate a platform and develop a vision for Big Open Legal Data (BOLD): an open framework for legislation, case law, and legal literature from across Europe.

  14. Data Crosscutting Requirements Review

    Energy Technology Data Exchange (ETDEWEB)

    Kleese van Dam, Kerstin [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Shoshani, Arie [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Plata, Charity [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

    2013-04-01

    In April 2013, a diverse group of researchers from the U.S. Department of Energy (DOE) scientific community assembled to assess data requirements associated with DOE-sponsored scientific facilities and large-scale experiments. Participants in the review included facilities staff, program managers, and scientific experts from the offices of Basic Energy Sciences, Biological and Environmental Research, High Energy Physics, and Advanced Scientific Computing Research. As part of the meeting, review participants discussed key issues associated with three distinct aspects of the data challenge: 1) processing, 2) management, and 3) analysis. These discussions identified commonalities and differences among the needs of varied scientific communities. They also helped to articulate gaps between current approaches and future needs, as well as the research advances that will be required to close these gaps. Moreover, the review provided a rare opportunity for experts from across the Office of Science to learn about their collective expertise, challenges, and opportunities. The "Data Crosscutting Requirements Review" generated specific findings and recommendations for addressing large-scale data crosscutting requirements.

  15. Ongoing experiments: diagnostics requirements

    International Nuclear Information System (INIS)

    Dickerman, C.E.

    1976-01-01

    The paper reviews the fuel motion diagnostics needs for ongoing LMFBR safety experiments over approximately the next five years, with the discussion centered on TREAT. Brief comments on the direction in which clad motion diagnostics requirements are expected to develop are also presented

  16. Human Systems Integration Requirements

    Science.gov (United States)

    2009-09-01

    52 Stratofortress, C/ KC -135 Stratotanker, E-3 Sentry and contractor logistics support aircraft; as well as a substantial jet engine inventory...www.hqda.army.mil/ teo /Sutability%20Terms%20MOA%20Oct%2005.pdf q. ISO 9241-11, Ergonomic requirements for office work with visual display terminals (VDTs

  17. Spent fuel storage requirements

    International Nuclear Information System (INIS)

    Fletcher, J.

    1982-06-01

    Spent fuel storage requirements, as projected through the year 2000 for U.S. LWRs, were calculated using information supplied by the utilities reflecting plant status as of December 31, 1981. Projections through the year 2000 combined fuel discharge projections of the utilities with the assumed discharges of typical reactors required to meet the nuclear capacity of 165 GWe projected by the Energy Information Administration (EIA) for the year 2000. Three cases were developed and are summarized. A reference case, or maximum at-reactor (AR) capacity case, assumes that all reactor storage pools are increased to their maximum capacities as estimated by the utilities for spent fuel storage utilizing currently licensed technologies. The reference case assumes no transshipments between pools except as currently licensed by the Nuclear Regulatory Commission (NRC). This case identifies an initial requirement for 13 MTU of additional storage in 1984, and a cumulative requirement for 14,490 MTU additional storage in the year 2000. The reference case is bounded by two alternative cases. One, a current capacity case, assumes that only those pool storage capacity increases currently planned by the operating utilities will occur. The second, or maximum capacity with transshipment case, assumes maximum development of pool storage capacity as described above and also assumes no constraints on transshipment of spent fuel among pools of reactors of like type (BWR, PWR) within a given utility. In all cases, a full core discharge capability (full core reserve or FCR) is assumed to be maintained for each reactor, except that only one FCR is maintained when two reactors share a common pool. For the current AR capacity case the indicated storage requirements in the year 2000 are indicated to be 18,190 MTU; for the maximum capacity with transshipment case they are 11,320 MTU

  18. Induction of functional Fc receptors in P388 leukemia cells. Requirement for multiple differentiation signals.

    Science.gov (United States)

    Cohen, D A; Stotelmyer, N L; Kaplan, A M

    1985-04-01

    The development of functional Fc receptors (FcR) during induced differentiation with the tumor promoter, phorbol myristate acetate (PMA), was studied in the murine tumor cell line, P388. PMA induced the appearance of FcR on the membranes of P388 cells as indicated by the binding of IgG-coated sheep red blood cells (IgG-SRBC). Concentrations of PMA as low as 1 ng/ml were sufficient to induce the expression of FcR as well as to inhibit cellular division and to induce adherence in the P388 tumor cell line; however, optimal FcR induction occurred at PMA concentrations of 10-100 ng/ml. Immunofluorescent analysis with heat-aggregated myeloma proteins indicated that PMA induced FcR which were capable of binding IgG2a and IgG2b immunoglobulins, but not IgG1. Adherence to a substratum was determined to be a second required signal for expression of FcR, since PMA induction of P388 tumor cells in teflon dishes failed to fully develop FcR and adherence of P388 cells to poly-L-lysine-coated culture dishes in the absence of PMA was insufficient for FcR expression. FcR which appeared after PMA induction were non-functional in the sense that membrane-bound IgG-SRBC were not ingested to any significant extent by the tumor cells. However, if FcR induction occurred in the presence conA-induced rat spleen cell culture supernatants, phagocytosis of membrane-bound erythrocytes occurred. These findings suggest that for the expression of FcR which are capable of particle internalization, at least three identifiable membrane-transmitted signals are required during differentiation.

  19. Next Generation Microbiology Requirements

    Science.gov (United States)

    Ott, C. M.; Oubre, C. M.; Elliott, T. F.; Castro, V. A.; Pierson, D. L.

    2012-01-01

    As humans continue to explore deep into space, microorganisms will travel with them. The primary means to mitigate the risk of infectious disease are a combination of prudent spacecraft design and rigorous operational controls. The effectiveness of these methods are evaluated by microbiological monitoring of spacecraft, food, water, and the crew that is performed preflight, in-flight, and post-flight. Current NASA requirements associated with microbiological monitoring are based on culture-based methodology where microorganisms are grown on a semi-solid growth medium and enumerated. Subsequent identification of the organisms requires specialized labor and large equipment, which historically has been performed on Earth. Requirements that rely strictly on culture-based units limit the use of non-culture based monitoring technology. Specifically, the culture-based "measurement criteria" are Colony Forming Units (CFU, representing the growth of one microorganism at a single location on the agar medium) per a given volume, area, or sample size. As the CFU unit by definition is culture-based, these requirements limit alternative technologies for spaceflight applications. As spaceflight missions such as those to Mars extend further into space, culture-based technology will become difficult to implement due to the (a) limited shelf life of the culture media, (b) mass/volume necessary to carry these consumables, and (c) problems associated with the production of biohazardous material in the habitable volume of the spacecraft. In addition, an extensive amount of new knowledge has been obtained during the Space Shuttle, NASA-Mir, and International Space Station Programs, which gave direction for new or modified microbial control requirements for vehicle design and mission operations. The goal of this task is to develop and recommend a new set of requirements for vehicle design and mission operations, including microbiological monitoring, based upon "lessons learned" and new

  20. Optical CDMA components requirements

    Science.gov (United States)

    Chan, James K.

    1998-08-01

    Optical CDMA is a complementary multiple access technology to WDMA. Optical CDMA potentially provides a large number of virtual optical channels for IXC, LEC and CLEC or supports a large number of high-speed users in LAN. In a network, it provides asynchronous, multi-rate, multi-user communication with network scalability, re-configurability (bandwidth on demand), and network security (provided by inherent CDMA coding). However, optical CDMA technology is less mature in comparison to WDMA. The components requirements are also different from WDMA. We have demonstrated a video transport/switching system over a distance of 40 Km using discrete optical components in our laboratory. We are currently pursuing PIC implementation. In this paper, we will describe the optical CDMA concept/features, the demonstration system, and the requirements of some critical optical components such as broadband optical source, broadband optical amplifier, spectral spreading/de- spreading, and fixed/programmable mask.

  1. Preanalytical requirements of urinalysis

    Science.gov (United States)

    Delanghe, Joris; Speeckaert, Marijn

    2014-01-01

    Urine may be a waste product, but it contains an enormous amount of information. Well-standardized procedures for collection, transport, sample preparation and analysis should become the basis of an effective diagnostic strategy for urinalysis. As reproducibility of urinalysis has been greatly improved due to recent technological progress, preanalytical requirements of urinalysis have gained importance and have become stricter. Since the patients themselves often sample urine specimens, urinalysis is very susceptible to preanalytical issues. Various sampling methods and inappropriate specimen transport can cause important preanalytical errors. The use of preservatives may be helpful for particular analytes. Unfortunately, a universal preservative that allows a complete urinalysis does not (yet) exist. The preanalytical aspects are also of major importance for newer applications (e.g. metabolomics). The present review deals with the current preanalytical problems and requirements for the most common urinary analytes. PMID:24627718

  2. Users' requirements for IFMIF

    International Nuclear Information System (INIS)

    Noda, K.; Jitsukawa, S.; Ehrlich, K.; Moeslang, A.

    1998-01-01

    The International Fusion Materials Irradiation Facility (IFMIF) is a high energy neutron irradiation facility which generates an intense neutron flux with D-Li stripping reactions for fusion materials testing. The role of IFMIF is (1) development of various fusion reactor materials, (2) determination of design-relevant engineering databases for the DEMO fusion reactor, (3) calibration and validation of data generated from fission reactor irradiations and the other simulation experiments, etc. The conceptual design activity (CDA) of IFMIF was initiated in February 1995 as an IEA collaborative activity to complete a reference conceptual design of IFMIF in December 1996. Users' requirements for the conceptual design of IFMIF were developed for materials to be tested, types of experiments, small specimen test technology and irradiation conditions. Furthermore, the neutron irradiation field characteristics (spectrum, flux/volume, etc.) of IFMIF were evaluated for the conceptual design parameters and were shown to meet the essential requirements of the users. (orig.)

  3. LHCb Online Networking Requirements

    CERN Document Server

    Jost, B

    2003-01-01

    This document describes the networking requirements of the LHCb online installation. It lists both quantitative aspects such as the number of required switch ports, as well as some qualitative features of the equipment, such as minimum buffer sizes in switches. The document comprises both the data acquisition network and the controls/general-purpose network. While the numbers represent our best current knowledge and are intended to give (in particular) network equipment manufacturers an overview of our needs, this document should not be confused with a market survey questionnaire or a formal tendering document. However the information contained in this document will be the input of any such document. A preliminary schedule for procurement and installation is also given.

  4. TANK FARM ENVIRONMENTAL REQUIREMENTS

    International Nuclear Information System (INIS)

    TIFFT, S.R.

    2003-01-01

    Through regulations, permitting or binding negotiations, Regulators establish requirements, limits, permit conditions and Notice of Construction (NOC) conditions with which the Office of River Protection (ORP) and the Tank Farm Contractor (TFC) must comply. Operating Specifications are technical limits which are set on a process to prevent injury to personnel, or damage to the facility or environment, The main purpose of this document is to provide specification limits and recovery actions for the TFC Environmental Surveillance Program at the Hanford Site. Specification limits are given for monitoring frequencies and permissible variation of readings from an established baseline or previous reading. The requirements in this document are driven by environmental considerations and data analysis issues, rather than facility design or personnel safety issues. This document is applicable to all single-shell tank (SST) and double-shell tank (DST) waste tanks, and the associated catch tanks and receiver tanks, and transfer systems. This Tank Farm Environmental Specifications Document (ESD) implements environmental-regulatory limits on the configuration and operation of the Hanford Tank Farms facility that have been established by Regulators. This ESD contains specific field operational limits and recovery actions for compliance with airborne effluent regulations and agreements, liquid effluents regulations and agreements, and environmental tank system requirements. The scope of this ESD is limited to conditions that have direct impact on Operations/Projects or that Operations Projects have direct impact upon. This document does not supercede or replace any Department of Energy (DOE) Orders, regulatory permits, notices of construction, or Regulatory agency agreements binding on the ORP or the TFC. Refer to the appropriate regulation, permit, or Notice of Construction for an inclusive listing of requirements

  5. Utility requirements for HTGRs

    International Nuclear Information System (INIS)

    Nicholls, D.R.

    1997-01-01

    Eskom, the state utility of South Africa, is currently evaluating the technical and economic feasibility of the helium cooled Pebble Bed Modular Reactor with a closed cycle gas turbine power conversion system for future power generating additions to its electric system. This paper provides an overview of the Eskom system including the needs of the utility for future generation capacity and the key performance requirements necessary for incorporation of this gas cooled reactor plant. (author)

  6. ENVIRONMENTAL SPECIFICATION REQUIREMENTS

    International Nuclear Information System (INIS)

    TIFFT, S.R.

    2003-01-01

    Through regulations, permitting or binding negotiations, Regulators establish requirements, limits, permit conditions and Notice of Construction (NOC) conditions with which the Office of River Protection (ORP) and the Tank Farm Contractor (TFC) must comply. Operating Specifications are technical limits which are set on a process to prevent injury to personnel, or damage to the facility or environment. The main purpose of this document is to provide specification limits and recovery actions for the TFC Environmental Surveillance Program at the Hanford Site. Specification limits are given for monitoring frequencies and permissible variation of readings from an established baseline or previous reading. The requirements in this document are driven by environmental considerations and data analysis issues, rather than facility design or personnel safety issues. This document is applicable to all single-shell tank (SST) and double-shell tank (DST) waste tanks, and the associated catch tanks and receiver tanks, and transfer systems. This Tank Farm Environmental Specifications Document (ESD) implements environmental-regulatory limits on the configuration and operation of the Hanford Tank Farms facility that have been established by Regulators. This ESD contains specific field operational limits and recovery actions for compliance with airborne effluent regulations and agreements, liquid effluents regulations and agreements, and environmental tank system requirements. The scope of this ESD is limited to conditions that have direct impact on Operations Projects or that Operations/Projects have direct impact upon. This document does not supercede or replace any DOE Orders, regulatory permits, notices of construction, or Regulatory agency agreements binding on the ORP or the TFC. Refer to the appropriate regulation, permit, or NOC for an inclusive listing of requirements

  7. ENVIRONMENTAL SPECIFICATION REQUIREMENTS

    International Nuclear Information System (INIS)

    TIFFT, S.R.

    2003-01-01

    Through regulations, permitting or binding negotiations, Regulators establish requirements, limits, permit conditions and Notice of Construction (NOC) conditions with which the Office of River Protection (ORP) and the Tank Farm Contractor (TFC) must comply. Operating Specifications are technical limits which are set on a process to prevent injury to personnel, or damage to the facility or environment. The main purpose of this document is to provide specification limits and recovery actions for the TFC Environmental Surveillance Program at the Hanford Site. Specification limits are given for monitoring frequencies and permissible variation of readings from an established baseline or previous reading. The requirements in this document are driven by environmental considerations and data analysis issues, rather than facility design or personnel safety issues. This document is applicable to all SST and DST waste tanks, and the associated catch tanks and receiver tanks, and transfer systems. This Tank Farm ESD implements environmental-regulatory limits on the configuration and operation of the Hanford Tank Farms facility that have been established by Regulators. This ESD contains specific field operational limits and recovery actions for compliance with airborne effluent regulations and agreements, liquid effluents regulations and agreements, and environmental tank system requirements. The scope of this ESD is limited to conditions that have direct impact on Operations/Projects or that Operations/Projects have direct impact upon. This document does not supercede or replace any DOE Orders, regulatory permits, notices of construction, or Regulatory agency agreements binding on the ORP or the TFC. Refer to the appropriate regulation, permit, or NOC for an inclusive listing of requirements

  8. BER Science Network Requirements

    Energy Technology Data Exchange (ETDEWEB)

    Alapaty, Kiran; Allen, Ben; Bell, Greg; Benton, David; Brettin, Tom; Canon, Shane; Dart, Eli; Cotter, Steve; Crivelli, Silvia; Carlson, Rich; Dattoria, Vince; Desai, Narayan; Egan, Richard; Tierney, Brian; Goodwin, Ken; Gregurick, Susan; Hicks, Susan; Johnston, Bill; de Jong, Bert; Kleese van Dam, Kerstin; Livny, Miron; Markowitz, Victor; McGraw, Jim; McCord, Raymond; Oehmen, Chris; Regimbal, Kevin; Shipman, Galen; Strand, Gary; Flick, Jeff; Turnbull, Susan; Williams, Dean; Zurawski, Jason

    2010-11-01

    The Energy Sciences Network (ESnet) is the primary provider of network connectivity for the US Department of Energy Office of Science, the single largest supporter of basic research in the physical sciences in the United States. In support of the Office of Science programs, ESnet regularly updates and refreshes its understanding of the networking requirements of the instruments, facilities, scientists, and science programs that it serves. This focus has helped ESnet to be a highly successful enabler of scientific discovery for over 20 years. In April 2010 ESnet and the Office of Biological and Environmental Research, of the DOE Office of Science, organized a workshop to characterize the networking requirements of the science programs funded by BER. The requirements identified at the workshop are summarized and described in more detail in the case studies and the Findings section. A number of common themes emerged from the case studies and workshop discussions. One is that BER science, like many other disciplines, is becoming more and more distributed and collaborative in nature. Another common theme is that data set sizes are exploding. Climate Science in particular is on the verge of needing to manage exabytes of data, and Genomics is on the verge of a huge paradigm shift in the number of sites with sequencers and the amount of sequencer data being generated.

  9. LEGACY MANAGEMENT REQUIRES INFORMATION

    International Nuclear Information System (INIS)

    CONNELL, C.W.; HILDEBRAND, R.D.

    2006-01-01

    ''Legacy Management Requires Information'' describes the goal(s) of the US Department of Energy's Office of Legacy Management (LM) relative to maintaining critical records and the way those goals are being addressed at Hanford. The paper discusses the current practices for document control, as well as the use of modern databases for both storing and accessing the data to support cleanup decisions. In addition to the information goals of LM, the Hanford Federal Facility Agreement and Consent Order, known as the ''Tri-Party Agreement'' (TPA) is one of the main drivers in documentation and data management. The TPA, which specifies discrete milestones for cleaning up the Hanford Site, is a legally binding agreement among the US Department of Energy (DOE), the Washington State Department of Ecology (Ecology), and the US Environmental Protection Agency (EPA). The TPA requires that DOE provide the lead regulatory agency with the results of analytical laboratory and non-laboratory tests/readings to help guide them in making decisions. The Agreement also calls for each signatory to preserve--for at least ten years after the Agreement has ended--all of the records in its or its contractors, possession related to sampling, analysis, investigations, and monitoring conducted. The tools used at Hanford to meet TPA requirements are also the tools that can satisfy the needs of LM

  10. BES Science Network Requirements

    Energy Technology Data Exchange (ETDEWEB)

    Biocca, Alan; Carlson, Rich; Chen, Jackie; Cotter, Steve; Tierney, Brian; Dattoria, Vince; Davenport, Jim; Gaenko, Alexander; Kent, Paul; Lamm, Monica; Miller, Stephen; Mundy, Chris; Ndousse, Thomas; Pederson, Mark; Perazzo, Amedeo; Popescu, Razvan; Rouson, Damian; Sekine, Yukiko; Sumpter, Bobby; Dart, Eli; Wang, Cai-Zhuang -Z; Whitelam, Steve; Zurawski, Jason

    2011-02-01

    The Energy Sciences Network (ESnet) is the primary provider of network connectivityfor the US Department of Energy Office of Science (SC), the single largest supporter of basic research in the physical sciences in the United States. In support of the Office ofScience programs, ESnet regularly updates and refreshes its understanding of the networking requirements of the instruments, facilities, scientists, and science programs that it serves. This focus has helped ESnet to be a highly successful enabler of scientific discovery for over 20 years.

  11. BES Science Network Requirements

    International Nuclear Information System (INIS)

    Dart, Eli; Tierney, Brian; Biocca, A.; Carlson, R.; Chen, J.; Cotter, S.; Dattoria, V.; Davenport, J.; Gaenko, A.; Kent, P.; Lamm, M.; Miller, S.; Mundy, C.; Ndousse, T.; Pederson, M.; Perazzo, A.; Popescu, R.; Rouson, D.; Sekine, Y.; Sumpter, B.; Wang, C.-Z.; Whitelam, S.; Zurawski, J.

    2011-01-01

    The Energy Sciences Network (ESnet) is the primary provider of network connectivity for the US Department of Energy Office of Science (SC), the single largest supporter of basic research in the physical sciences in the United States. In support of the Office of Science programs, ESnet regularly updates and refreshes its understanding of the networking requirements of the instruments, facilities, scientists, and science programs that it serves. This focus has helped ESnet to be a highly successful enabler of scientific discovery for over 20 years.

  12. Section 4: Requirements Intertwining

    Science.gov (United States)

    Loucopoulos, Pericles

    Business analysts are being asked to develop increasingly complex and varied business systems that need to cater to the changing and dynamic market conditions of the new economy. This is particularly acute in today’s turbulent business environment where powerful forces such as deregulation, globalisation, mergers, advances in information and telecommunications technologies, and increasing education of people provide opportunities for organising work in ways that have never before been possible. Enterprises attempt to create wealth either by getting better at improving their products and services or by harnessing creativity and human-centred management to create innovative solutions. In these business settings, requirements become critical in bridging system solutions to organisational and societal problems. They intertwine organisational, social, cognitive, and implementation considerations and they can provide unique insights to change in systems and their business context. Such design situations often involve multiple stakeholders from different participating organisations, subcontractors, divisions, etc., who may have a diversity of expertise, come from different organisational cultures and often have competing goals. The success or failure of many projects depends, to a large extent, on understanding the contextual setting of requirements and their interaction amongst a diverse population of stakeholders.

  13. Beauty Requires Thought.

    Science.gov (United States)

    Brielmann, Aenne A; Pelli, Denis G

    2017-05-22

    The experience of beauty is a pleasure, but common sense and philosophy suggest that feeling beauty differs from sensuous pleasures such as eating or sex. Immanuel Kant [1, 2] claimed that experiencing beauty requires thought but that sensuous pleasure can be enjoyed without thought and cannot be beautiful. These venerable hypotheses persist in models of aesthetic processing [3-7] but have never been tested. Here, participants continuously rated the pleasure felt from a nominally beautiful or non-beautiful stimulus and then judged whether they had experienced beauty. The stimuli, which engage various senses, included seeing images, tasting candy, and touching a teddy bear. The observer reported the feelings that the stimulus evoked. The time course of pleasure, across stimuli, is well-fit by a model with one free parameter: pleasure amplitude. Pleasure amplitude increases linearly with the feeling of beauty. To test Kant's claim of a need for thought, we reduce cognitive capacity by adding a "two-back" task to distract the observer's thoughts. The distraction greatly reduces the beauty and pleasure experienced from stimuli that otherwise produce strong pleasure and spares that of less-pleasant stimuli. We also find that strong pleasure is always beautiful, whether produced reliably by beautiful stimuli or just occasionally by sensuous stimuli. In sum, we confirm Kant's claim that only the pleasure associated with feeling beauty requires thought and disprove his claim that sensuous pleasures cannot be beautiful. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. Display Parameters and Requirements

    Science.gov (United States)

    Bahadur, Birendra

    The following sections are included: * INTRODUCTION * HUMAN FACTORS * Anthropometry * Sensory * Cognitive * Discussions * THE HUMAN VISUAL SYSTEM - CAPABILITIES AND LIMITATIONS * Cornea * Pupil and Iris * Lens * Vitreous Humor * Retina * RODS - NIGHT VISION * CONES - DAY VISION * RODS AND CONES - TWILIGHT VISION * VISUAL PIGMENTS * MACULA * BLOOD * CHOROID COAT * Visual Signal Processing * Pathways to the Brain * Spatial Vision * Temporal Vision * Colour Vision * Colour Blindness * DICHROMATISM * Protanopia * Deuteranopia * Tritanopia * ANOMALOUS TRICHROMATISM * Protanomaly * Deuteranomaly * Tritanomaly * CONE MONOCHROMATISM * ROD MONOCHROMATISM * Using Colour Effectively * COLOUR MIXTURES AND THE CHROMATICITY DIAGRAM * Colour Matching Functions and Chromaticity Co-ordinates * CIE 1931 Colour Space * CIE PRIMARIES * CIE COLOUR MATCHING FUNCTIONS AND CHROMATICITY CO-ORDINATES * METHODS FOR DETERMINING TRISTIMULUS VALUES AND COLOUR CO-ORDINATES * Spectral Power Distribution Method * Filter Method * CIE 1931 CHROMATICITY DIAGRAM * ADDITIVE COLOUR MIXTURE * CIE 1976 Chromaticity Diagram * CIE Uniform Colour Spaces and Colour Difference Formulae * CIELUV OR L*u*v* * CIELAB OR L*a*b* * CIE COLOUR DIFFERENCE FORMULAE * Colour Temperature and CIE Standard Illuminants and source * RADIOMETRIC AND PHOTOMETRIC QUANTITIES * Photopic (Vλ and Scotopic (Vλ') Luminous Efficiency Function * Photometric and Radiometric Flux * Luminous and Radiant Intensities * Incidence: Illuminance and Irradiance * Exitance or Emittance (M) * Luminance and Radiance * ERGONOMIC REQUIREMENTS OF DISPLAYS * ELECTRO-OPTICAL PARAMETERS AND REQUIREMENTS * Contrast and Contrast Ratio * Luminance and Brightness * Colour Contrast and Chromaticity * Glare * Other Aspects of Legibility * SHAPE AND SIZE OF CHARACTERS * DEFECTS AND BLEMISHES * FLICKER AND DISTORTION * ANGLE OF VIEW * Switching Speed * Threshold and Threshold Characteristic * Measurement Techniques For Electro-optical Parameters * RADIOMETRIC

  15. Knowing requires data

    Science.gov (United States)

    Naranjo, Ramon C.

    2017-01-01

    Groundwater-flow models are often calibrated using a limited number of observations relative to the unknown inputs required for the model. This is especially true for models that simulate groundwater surface-water interactions. In this case, subsurface temperature sensors can be an efficient means for collecting long-term data that capture the transient nature of physical processes such as seepage losses. Continuous and spatially dense network of diverse observation data can be used to improve knowledge of important physical drivers, conceptualize and calibrate variably saturated groundwater flow models. An example is presented for which the results of such analysis were used to help guide irrigation districts and water management decisions on costly upgrades to conveyance systems to improve water usage, farm productivity and restoration efforts to improve downstream water quality and ecosystems.

  16. SOFG: Standards requirements

    International Nuclear Information System (INIS)

    Gerganov, T.; Grigorov, S.; Kozhukharov, V.; Brashkova, N.

    2005-01-01

    It is well-known that Solid Oxide Fuel Cells will have industrial application in the nearest future. In this context, the problem of SOFC materials and SOFC systems standardization is of high level of priority. In the present study the attention is focused on the methods for physical and chemical characterization of the materials for SOFC components fabrication and about requirements on single SOFC cells tests. The status of the CEN, ISO, ASTM (ANSI, ASSN) and JIS class of standards has been verified. Standards regarding the test methods for physical-chemical characterization of vitreous materials (as sealing SOFC component), ceramic materials (as electrodes and electrolyte components, including alternative materials used) and metallic materials (interconnect components) are subject of overview. It is established that electrical, mechanical, surface and interfacial phenomena, chemical durability and thermal corrosion behaviour are the key areas for standardization of the materials for SOFC components

  17. p85α recruitment by the CD300f phosphatidylserine receptor mediates apoptotic cell clearance required for autoimmunity suppression

    Science.gov (United States)

    Tian, Linjie; Choi, Seung-Chul; Murakami, Yousuke; Allen, Joselyn; Morse, Herbert C., III; Qi, Chen-Feng; Krzewski, Konrad; Coligan, John E.

    2014-01-01

    Apoptotic cell (AC) clearance is essential for immune homeostasis. Here we show that mouse CD300f (CLM-1) recognizes outer membrane-exposed phosphatidylserine, and regulates the phagocytosis of ACs. CD300f accumulates in phagocytic cups at AC contact sites. Phosphorylation within CD300f cytoplasmic tail tyrosine-based motifs initiates signals that positively or negatively regulate AC phagocytosis. Y276 phosphorylation is necessary for enhanced CD300f-mediated phagocytosis through the recruitment of the p85α regulatory subunit of phosphatidylinositol-3-kinase (PI3K). CD300f-PI3K association leads to activation of downstream Rac/Cdc42 GTPase and mediates changes of F-actin that drive AC engulfment. Importantly, primary macrophages from CD300f-deficient mice have impaired phagocytosis of ACs. The biological consequence of CD300f deficiency is predisposition to autoimmune disease development, as FcγRIIB-deficient mice develop a systemic lupus erythematosus-like disease at a markedly accelerated rate if CD300f is absent. In this report we identify the mechanism and role of CD300f in AC phagocytosis and maintenance of immune homeostasis.

  18. Equipment Operational Requiremen